Co-targeting Dysregulated Ocular and Cochlear Blood Flow via Ophthalmic Nerve Stimulation for the Treatment of Type 2 Usher Syndrome: Prospective Case Series

Co-targeting Dysregulated Ocular and Cochlear Blood Flow via Ophthalmic Nerve Stimulation for the Treatment of Type 2 Usher Syndrome: Prospective Case Series | Research Square window.SnipcartSettings = { analytics: { enabled: false } }; (function() { var accessVector = localStorage.getItem('access_vector') || ''; window.dataLayer = window.dataLayer || []; if (accessVector) { window.dataLayer.push({ user: { profile: { profileInfo: { snid: accessVector } } } }); } })(); (function(w,d,s,l,i){w[l]=w[l]||[];w[l].push({'gtm.start':new Date().getTime(),event:'gtm.js'});var f=d.getElementsByTagName(s)[0],j=d.createElement(s),dl=l!='dataLayer'?'&l='+l:'';j.async=true;j.src='https://www.googletagmanager.com/gtm.js?id='+i+dl;f.parentNode.insertBefore(j,f);})(window,document,'script','dataLayer','GTM-K279D39R'); Browse Preprints In Review Journals COVID-19 Preprints AJE Video Bytes Research Tools Research Promotion AJE Professional Editing AJE Rubriq About Preprint Platform In Review Editorial Policies Our Team Advisory Board Help Center Sign In Submit a Preprint Cite Share Download PDF Research Article Co-targeting Dysregulated Ocular and Cochlear Blood Flow via Ophthalmic Nerve Stimulation for the Treatment of Type 2 Usher Syndrome: Prospective Case Series Ismail M Musallam This is a preprint; it has not been peer reviewed by a journal. https://doi.org/ 10.21203/rs.3.rs-6190348/v1 This work is licensed under a CC BY 4.0 License Status: Posted Version 1 posted You are reading this latest preprint version Abstract Reduced dysregulated ocular and cochlear blood flow are proposed as common pathways in the pathogenesis of type 2 Usher syndrome (USH2). The purpose of the study was to evaluate the safety and efficacy of ophthalmic nerve stimulation (ONS) combined with ascorbic acid (AA) in treatment of USH2. Nine participants with USH 2, were enrolled in a prospective interventional case series. All participants were daily treated with ONS sessions and intravenous AA for two weeks. The primary efficiency endpoint was 6 months’ changes in scotopic vision as measured by a Low Luminance Questionnaire-10 (LLQ-10) with a maximum score of 100 points. Rod responders were defined by ≥ 25points increment of LLQ-10 score. The results showed that ONS-based therapy significantly improved scotopic vision by 42.1 + 11.3 points ( p = 0.0001 ) and 7 (77.8%) of the participants were identified as rod responders. Additionally, clinically significant improvement visual acuity (≥ 0.2 logMAR) and contrast sensitivity (≥ 0.3 log unit) were noticed in 22.2% of the left eyes. Furthermore, a significant improvement of hearing was subjectively reported by one third of the participants. In conclusion, ONS-based therapy significantly improved night vision in patients with USH2. Additionally, a clinically significant improvement of hearing was noticed in one third of patients. Ophthalmology Usher syndrome retinitis pigmentosa sensorineural hearing loss ocular neuromodulation ophthalmic nerve stimulation Low luminance questionnaire-10 rod responders ascorbic acid substance P Figures Figure 1 Figure 2 1. Introduction Usher syndrome (USH) represents a group of autosomal recessive disorders characterized by sensorineural hearing loss (SNHL), vision loss, and vestibular dysfunction ( 1 ). It primarily affects the auditory hair cells followed by light-sensitive photoreceptor cells in the retina ( 2 ). USH is the leading genetic cause of deaf-blindness (50%of all cases), with a prevalence in the range of 4 to 17 per 100,000 people. ( 3 ). Three clinical subtypes of USH have been defined – Type 1, Type 2, and Type 3 – based upon the severity, and progression of SNHL. Type 2 (USH2) is the most common subtype, accounting for more than half of all USH cases worldwide ( 4 ). USH2 patients present with moderate-to-severe SNHL, normal vestibular function, and retinitis pigmentosa (RP) that begins generally during puberty. There is no cure for USH2. Several treatment options are available to help patients manage hearing problems. This includes auditory rehabilitation using hearing aid devices or cochlear implants. Unfortunately, these technologies do not treat the underlying cause of hearing loss and are far from providing quality natural hearing. Moreover, there is no cure for RP. Currently, there are a range of new therapeutic strategies being under investigation that utilize new technologies such as gene and cell-based therapies. Given that USH2 is a syndromic condition affecting multiple senses, it is important to note that current therapeutic trials generally focus on one sense rather than the full spectrum of the disease. Therefore, a comprehensive therapeutic strategy is needed that simultaneously targets the underlying pathophysiology in both the retina and the inner ear regardless of the genetic background of the disease The human inner ear depends on a vascular supply to maintain fluid homeostasis, ion balance, and metabolic supply. Disruption of cochlear blood flow (CoBF) leads to instantaneous pathological changes in the inner ear ( 5 ) ( 6 ). A growing body of evidence suggests that microvascular changes may contribute to hearing loss. ( 7 ) ( 8 ) ( 9 ) ( 10 ) ( 11 ). Histopathological examination of the temporal bone of patients with USH revealed atrophy of the stria vascularis ( 12 ). This is thought to cause degeneration of hair cells by alteration of the endolymph composition. ( 13 ). The stria vascularis, a highly vascularized region located in the cochlear microcirculation, is the metabolic energy source underlying cochlear function ( 14 ). Circulatory changes have also been observed in the retina and choroid of patients with Usher’s syndrome. Vascular density was significantly decreased in the retinal circulation of patients with USH2A and MYO7A mutations compared with controls ( 15 ). Changes were observed in both the superficial and deep capillary plexus. Vascular density in the deep capillary plexus showed strong association with macular sensitivity and ellipsoid zoon width ( 15 ). Choroidal thickness is reduced in people with USH2 if compared to healthy subjects., which might be considered as a sign of blood flow reduction ( 16 ). Evaluating choroidal hemodynamic changes has shown the reduction in choroidal blood flow (ChBF) is proportional to the progression of RP ( 17 ). Recently, relative choroidal ischemia and reduced ChBF have been demonstrated in RP by measuring the choroidal vascularity index via Spectral Domain Optical coherence tomography (SD-OCT) ( 18 ). The ocular blood flow (OBF) is reduced not only in the retina and choroid but also in the retro-ocular vessels ( 19 ). The eye receives its blood supply through the anterior cerebral circulation (Internal carotid arteries) via ophthalmic arteries. The retina has a dual blood supply, with the retinal circulation supplying the inner layers and the choroidal circulation supplying the outer layers of the retina including photoreceptors. The blood supply to the choroid in mammals arises from the ophthalmic artery via the long and short ciliary arteries. Like the eye, the inner ear is supplied with an end artery. The blood supply to the inner ear is derived from the posterior cerebral circulation (Basilar artery), mostly from labyrinthine artery, a branch of the anterior inferior cerebellar artery. Ophthalmic artery, central retinal artery, posterior ciliary arteries, and choroid are richly innervated by parasympathetic, sympathetic and trigeminal sensory nerve fibers. The parasympathetic innervation has been shown to cause vasodilation and increase ChBF, the sympathetic input has been shown to induce vasoconstriction and decrease ChBF. In contrast, autonomic control for cochlear blood vessels is limited to sympathetic innervation that arises from superior cervical ganglion, whereas parasympathetic innervation of cochlear blood vessels has not been found ( 20 ). The majority of the intracranial blood vessels including those contributing to both the ocular and cochlear circulation are innervated by the ophthalmic division of the trigeminal nerve ( 20 ) ( 21 ). Direct stimulation of the trigeminal ganglion in experimental models has been found to lead to increased cerebral blood flow ( 22 ). Stimulation of the ophthalmic nerve may be topographically oriented, with specific branches leading to increased blood flow in specific regions of the eye, ear, and brain arterial tree. The ophthalmic nerve plays a pivotal role in the regulation of ocular and CoBF (Fig. 1 ), by inducing vasodilation, as a result of the release of substance P (SP), calcitonin gene-related peptide (CGRP), and neuropeptide Y as mediators of vasodilative mechanisms ( 23 , 24 ). Stimulation of sensory afferents from the ophthalmic nerve results in the activation of the trigeminovascular system via antidromic impulse ( 25 ) ( 26 ) ( 27 ). The nasociliary nerve, which originates from the ophthalmic nerve, contains major vasodilatory innervation ( 28 , 29 ), and its stimulation results in the release of vasoactive neuropeptides from the free nerve endings, such as SP and CGRP ( 23 , 24 ). Consistent with this, stimulation of the ophthalmic nerve in experimental animals was found to increase OBF ( 30 ), decrease carotid arterial resistance, ( 31 ) and enhance uveal release of SP ( 32 ). Infusion of the internal carotid artery or the anterior inferior cerebellar artery with SP induces increase in CoBF ( 33 ). There is extensive evidence that excessive free radical formation along with diminished CoBF are essential factors involved in the pathogenesis of SNHL ( 34 ) ( 35 ). Anti-oxidant interventions have been reported to substantially delay the death of photoreceptor cells of the retina ( 36 ) and hair cells of the inner ear ( 37 ). Interestingly, high dose of vitamin C has been shown to enhance hearing recovery in idiopathic sudden SNHL patients, which suggests that ascorbate treatment reduces levels of reactive oxygen metabolites produced by inner ear ischemia or inflammation. ( 37 ). Therefore, it is reasonable to suggest that supra-physiological doses of intravenous AA could be beneficial in a perfusion-compromised retina or inner ear as in USH2. Preclinical studies show that high-dose of AA can prevent or restore microcirculatory flow impairment. AA can restore vascular responsiveness to vasoconstrictors and preserve endothelial barrier ( 38 ). Additionally, AA exerts sympatholytic effects via central action ( 39 ). Pathogenesis of retinal and inner ear disorders in USH2 reflects a summation of primary reduced dysregulated OBF/CoBF, and consecutive unstable oxygen supply, hair cell/ photoreceptors starvation, oxidative stress and inflammation of the retina and the inner ear. These vascular and cellular processes are ultimately combined to produce damage of cochlear hair cells followed by retinal rods and finally cone photoreceptors apoptosis. The mechanisms responsible for reduced dysregulated ocular and CoBF in patients with USH2 uncertain. It is also uncertain if blood flow decline stems from disturbed neural control of ocular and cochlear circulation or vascular pathology, or both. The reduced dysregulated CoBF/ OBF is proposed as a common pathway in the pathogenesis of SNHL and blindness, which can be a target for a novel therapeutic strategy in USH2. Because of complexity of this syndrome, it is unlikely that optimal protection of the inner ear mechanoreceptor and retinal photoreceptors can be achieved through a single therapy, suggesting that an alternative approach based on combination therapy to be considered. Up-regulation of trigeminovascular system and parasympathetic system via ONS along with anti-oxidation, sympathetic inhibition and restoration of vascular endothelial function via systemic administration of AA might form the basis of neuromodulation therapy for treating this non-curable multi-sense damaging syndrome. Here we report the safety and efficacy results of ONS paired with systemic ascorbate for treating participants with USH2. 2. Materials and Methods The study was a prospective, consecutive, open-label, interventional study aimed at investigating the safety and efficacy of ONS combined with AA on patients with USH2. The protocol was reviewed and approved by the Ethics Committee of the Musallam Specialty Hospital, Palestinian Authority, and it adhered to the tenets of the Declaration of Helsinki. Written informed consent was obtained from all participants or legal guardians after an explanation of the purpose and possible outcome of therapy. 2.1 Patient Selection Patient enrollment started in April 2018, and cases were recruited from a single tertiary ophthalmic center. The inclusion criteria of the study included patients with an established diagnosis of USH2 with BCVA ≥ 20/400. Exclusion criteria were pregnancy, glaucoma, central retinal vein occlusion, retinal detachment. bradycardia, malignancy, and active infection. Following recruitment, all participants with USH2 completed baseline vision testing, slit lamp examination and fundoscopy. Spectral Domain Optical coherence tomography (SD-OCT) was performed within 2–4 weeks prior to treatment. The severity of the RP associated with Usher’s syndrome was clinically graded into six stages based on fundoscopy and SD-OCT findings ( 40 ). The primary efficiency endpoints were 6-month changes in scotopic vision and adverse effects of the therapy. The scotopic vision was assessed with a Low luminance questionnaire-10 (LLQ-10) ( 40 ). The secondary efficiency points include best corrected visual acuity (BCVA), contrast sensitivity (CS), and the change of hearing acuity. BCVA was recorded in Snellen equivalents and then transformed to logMAR unit. CS was measured using a Pelli- Robson chart under standardized photopic illumination at one meter in each eye, using best correction spectacles. To characterize the severity of hearing loss at baseline a subjective numerical rating scale (NRS) of 1 (normal hearing) to 6 (complete deafness) was used ( 41 ). Improvement of an NRS score by 2 or more points was considered a clinically significant improvement of hearing. To evaluate the therapy outcome, LLQ-10 score changes from baseline to 6 months after treatment were calculated (The primary efficacy endpoint). LLQ-10 has been used to quantify the participant’s visual dysfunction under low luminance, integrating aspects of visual acuity, mobility, visual field, depth of perception, reading, color vision, dependency on others’ help, social functions, and mental health ( 40 ). Patient-centered outcome measures have been increasingly viewed as necessary in clinical trials, serving as primary endpoints( 40 ) ( 42 ) ( 43 ) ( 44 ). Participants rated how much difficulty they had performing each of the activities under low luminance on a 5-point Likert scale. Rod responders were identified as those in whom the LLQ-10 score increased by ≥ 25 points at 6 months’ post-treatment. For safety purposes, the vital signs and electrocardiogram were monitored before, during, and after each session of neuromodulation. Ocular and non-ocular adverse events were also reported (Primary safety endpoint) 2.2 The Treatment Protocol ONS-based therapy protocol included the administration of AA followed by ONS over a period of two weeks. AA was given intravenously in a dose of 3 gm dissolved in 100 ml of saline infused at 5 ml min − 1 for 20 min on the first day, followed by a drip infusion of 1 g daily during the rest of the treatment period. A modulated low magnitude, low-frequency vibration, with a frequency of approximately 60 Hz- 90 Hz and stimulation amplitude range of 1.5 µm- 3.5 µm were used for ONS. Modulated frequency and amplitude waveforms were used to avoid adaptation to mechanical stimulus. A head-mounted prototype of ophthalmic nerve stimulators was applied to different application sites by using modified commercially available micro-vibrators, along with different types of intranasal and extra-nasal application heads. Bilateral ONS was applied to the subjects over a session of 30 minutes per day, 6 sessions per week, and one day off. Each session included intra-nasal vibrochemical, and extra-nasal vibrotactile stimulation over the nasal bridge and the supraorbital region in both sides, using appropriate application heads. During intra-nasal vibrochemical stimulation, the nasal application head was covered by a rubber cap impregnated by 2% menthol cream (Dermacool plus R ) as a transient receptor potential cation channel subfamily M (melastatin) member 8 (TRPM8) agonist. Participants were assessed clinically, and by SD-OCT, and ILQ-10 at baseline and at 1, 6 months after treatment 2.3 Statistical analysis The statistical analysis was performed using SPSS R , V. 21. The change in the mean value of visual function parameters between baseline and different post-treatment points was evaluated using a paired sample t-test. The BCVA and CS at baseline, and at post-treatment visits were recorded for the left eye in the whole cohort. All tests were two-tailed and a p -value less than 0.05 was considered significant. The proportion of left eyes that showed clinically significant improvement in visual function, among the whole cohort was also evaluated. Additionally, the proportion of patients who experienced clinically significant improvement in hearing was also determined 3. Results Baseline Demographics Between April 14, 2018, and April 29, 2020, 9 participants, aged between 8–55 years (median; 23 years), mean (24.8 ± 13.9) among them 7 females (77.8%) were recruited. The demographics and baseline characteristics of participants are presented in Table. At 6 months’ visits, 7 out of 9 (77.7%) patients were defined as rod responders. In the whole cohort, the night vision score was improved from 21.4 ± 11.3 points at baseline to a level of 59.2 ± 19.8 points ( P = 0.0001 ) at 1 month and, 56.4 ± 18.7 points (P = 0.0001 ) at 6 months (Fig. 2 ). In rod responders, the mean change in LLQ-10 score was 45.4 ± 8.1 points (p = 0.0001) at one month and (42.1 ± 11.3 points) ( p = 0.0001 ) at 6 months. The two non-responders have stage III and V RP and the duration of night blindness was 12 and 20 years (16 ± 5.7). The BCVA of the left eye was improved from (0.38 ± 0.43 logMAR) at baseline to (0.26 ± 0.33 Log MAR) ( P = 0.19 ) at 1 month and (0.28 ± 0.34 Log MAR) ( p = 0.24 ) at 6 months which was found statistically not significant. In two out of 9 patients (22.2%), a clinically significant improvement of BCVA (improvement of ≥ 0.2 logMAR) was noticed. Regarding CS, the mean CS of the left eye was improved from 0.64 ± 0.60 log unit to 0.79 ± 0.57 Log unit at 1 month and to a value of 0.76 ± 0.54 log unit at 6 months. However, this change was not statistically significant. At 6 months’ visits, two eyes (22.2%) of the whole cohort had a clinically significant improvement of CS (improvement of ≥ 0.3 log unit). Interestingly, a clinically significant improvement in hearing was observed in 3 patients (33.3%). No serious adverse effects were reported and headache in one patient was the only encountered side effect in this study. Table Baseline characteristics Baseline Characteristics Total cohort (9 patients; 9 eyes) Age, Mean ± SD Rang Median age 24.8 ± 13.9 yrs. 8–55 yrs. 23 M/F, (male %) 2/7 (22.2%) Rang (yrs.) of night blindness Mean duration of night blindness (yrs.) Proportion of patients with night blindness 10 years or less 5–20 yrs. 11.8 ± 6.2 yrs. 4 (44.4%) Stage of the Disease * II, no. of patients (%) III, no. of patients/ (%) IV, no. of patients (%) V, no. of patients/ (%) 4 (44.4%) 2 (22.2%) 2 (22.2%) 1 (11.1%) LLQ-10 Score at baseline (mean ± SD) 22.8 ± 12.4 4. Discussion Up-to-date, there is no treatment option available to cure sensorineural hearing loss or vision loss in USH2, and only prostheses such as hearing aids and cochlear implants offer some help. In the current study, we have reported successful use of ONS-based therapy as a novel treatment for USH2 syndrome. It has led to a clinically meaningful and statistically significant improvement of scotopic vision. Additionally, clinically significant improvement in BCVA, contrast sensitivity and hearing was also noticed in some patients. In this study we found that the most robust improvement in visual function for the subjects with USH2 was night vision as measured by LLQ-10. Recovery of night vision was likely to be related to reactivation of dormant/starving rods via enhanced oxygen and glucose delivery and shunting metabolites toward aerobic glucose metabolism ( 45 ). There are four clinico-pathological overlapping phases that characterize USH2; an early hair cell degeneration phase, rod degeneration phase, transitional phase of rod/cone photoreceptors damage and late cone degeneration phase. SNHL occurs prior to visual symptoms in all subtypes of USH syndrome including USH2 ( 2 ) probably because the inner ear is more vulnerable to hypoperfusion. This might be explained at least in part by early development of dysregulated CoBF as a result of dominant sympathetic innervation of cochlear blood vessels and absence parasympathetic input. ( 20 ). Interestingly, cochlear circulation is innervated by branches of ophthalmic nerve ( 20 ) that produce robust dilatation of labyrinthine arteries and their branches, increasing CoBF, as result of release of vasoactive neuropeptides. Therefore, the upregulation of trigeminovascular system may counteract the dysregulated CoBF induced by increased sympathetic tone. The vascular alteration in the inner ear and the retina might explain the sequence of pathological events in USH2. A highly vulnerable cochlear circulation compared to ocular circulation might explain the early development of SNHL. Additionally, the atrophy of the stria vascularis ( 12 ) supports the notion that auditory hair cell damage is primarily of vascular origin. Stria vascularis, a key structure for producing and maintaining the endocochlear potential. Morphologic alterations in the stria vascularis decrease the endocochlear potential and consequently, affect the cochlear amplification of acoustic signals leading to an increase in auditory thresholds, even in the absence of hair cell death ( 34 ). On the other hand, the development of annular scotoma at the mid-periphery opposite the choroidal watershed zone between the anterior and posterior ciliary arteries ( 46 ), provides further support to the vascular theory of USH2. SNHL including Meiners disease, sudden SNHL, senile deafness and inherited NSHL were attributed at least in part to underlying vascular causes and reduced CoBF. Nevertheless, the unmet oxygen and glucose requirements of mutant hair cells and rod photoreceptors might be detrimental for the disease process and its sequence of occurrence in USH2. A key finding in the present study was the substantial improvement of hearing in one third of the participants. Such an improvement is unlikely to be due to chance, and mostly related to the effect of treatment as this phenomenon is not normally seen in untreated patients with USH2. Hearing improvement might be related to one of the following mechanisms, these include recovery of dormant hair cells, regeneration of hair cells, or trans differentiation of supporting cells into hair cells. These effects might occur as a result of recruitment of endogenous stem cell in response to overexpression of SP ( 47 ). It is reasonable to suggest that the SNHL seen in the early stages of USH2 is explained, at least in part by hair cell dormancy. The increased CoBF and glucose delivery to the inner ear in response to ONS might restore the function of these dormant hair cells, resulting in improved hearing. Hair cell dormancy has been demonstrated in organ culture, wherein the hair cells completely loss their bundles. These dormant bundleless hair cells are viable cells that can grow back to the apical surface of the epithelium, resynthesize their bundles, and thereby restore their function ( 48 ). Although the exact mechanism of action of ONS-based therapy in USH2 is not fully understood, the distinctive effect of ONS-based therapy could be due to its capability to target diverse pathophysiologies of USH2. It is known that antidromic activation of the ophthalmic nerve especially its nasociliary branch leads to the release of neuropeptides such as SP from nerve terminals. SP, an 11-amino acid neuropeptide that belongs to the tachykinin family of peptides, has a preferential affinity to the neurokinin-1 receptor (NK-1R) ( 49 ). Several studies have indicated that SP is capable of boosting both the ocular/CoBF ( 25 ) ( 26 ) ( 30 ) ( 33 ) and tissue regeneration by the recruitment of endogenous stem cells ( 47 ). Moreover SP can protect diverse types of cells including retinal pigment epithelium ( 50 ), and ganglion cells ( 51 ). It also contribute to the prevention of apoptosis ( 52 ) ( 53 ), suppression of both inflammation ( 54 ) ( 55 ) and oxidative stress ( 56 ) ( 57 ). SP was reported to modulate Akt/GSK-3β signaling, inhibit reactive oxygen species-induced cell death, preserve cell viability, and block cellular alterations ( 56 ) ( 57 ) ( 58 ).SP exhibited a protective effect on the cochlea from noise damage in guinea pigs exposed to noise after an infusion of SP into the inner ear ( 59 ). However, the therapeutic applications of SP have been limited by its low stability. SP tends to be degraded by various proteases, including neutral endopeptidase and angiotensin-converting enzymes ( 60 ) ( 61 ) ( 62 ). The half-life time of SP is very short, from seconds to minutes ( 63 ) ( 64 ) ( 65 ) If endogenous SP is to be therapeutically effective, then the sustained release of this neuropeptide from the nerve terminals of the ophthalmic nerve is essential and its stability should be enhanced. SP has been identified at several locations in the auditory pathway ( 66 ) ( 67 ), including the inner ear ( 68 ) ( 69 ) ( 70 ). Immunohistochemical and electrophysiological studies confirm that SP may act as a neuromodulator at the synapses of the inner hair cells in the guinea pig ( 71 ). SP immunolabeling has been observed on the labyrinthine and spiral modiolar arteries ( 68 ) ( 72 ). Similarly, SP is located primarily in sensory nerves, surrounding ophthalmic and its branches. The perivascular location of SP in the ocular and inner ear blood vessels makes the sustained release of this potent vasoactive neuropeptide by up-regulation of the trigeminovascular system an innovative therapeutic strategy for USH2. Concomitant infusion of AA and administration of ONS might significantly correlated with the clinical outcome of this therapeutic strategy. AA is an efficient inhibitor for both angiotensin-converting enzyme, and neutral endopeptidase enzyme, enzymes that are commonly involved in SP degradation ( 73 ) ( 74 ) ( 75 ) . The underlying mechanisms of retinal degeneration and hair cell loss in USH are highly complex, oxidative stress and dysregulated blood flow may be common to retinal degeneration as well as to hair cell damage. Previous studies have demonstrated that an excess of free radical formation and blood reduction in the cochlea occur in noise, drug-induced hearing loss, and senile SNHL ( 76 ). Then it is reasonable to suggest that dysregulated CoBF and oxidative stress may be critical factors in triggering hearing loss associated with USH2. Production of oxidative stress via free radical generation has been implicated by a variety of insults that can result in SNHL ( 77 ). The role of antioxidants in the prevention of hearing loss has been reported in a number of studies. ( 37 ) ( 78 ) ( 79 ). Interestingly, high doses of vitamin C have been shown to enhance hearing recovery in idiopathic sudden SNHL patients, which suggests that vitamin C reduces levels of reactive oxygen metabolites produced by inner ear ischemia or inflammation. Therefore, it is reasonable to suggest that high-dose of intravenous AA could be beneficial in Usher syndrome. On the other hand, photoreceptor cells have a rich complement of mitochondria to accommodate the high metabolic rate and are more sensitive to oxidative stress than other cell types ( 80 ). The development of RP, for instance, is tightly linked to oxidative damage ( 81 ), and antioxidant therapy can delay cell loss in retinal degenerative diseases and lead to reduced photoreceptor cell death in experimental animals ( 82 ) and in clinical trials ( 36 ). It has been demonstrated that SP could stimulate the recovery of RPE cells under oxidative stress, possibly by promoting cell proliferation and inhibiting apoptosis through the activation of Akt/GSK-3β signaling ( 56 ). The interpretation of study findings should be considered within the context of a few limitations. These include a small number of participants, short duration of follow up, lack of information regarding the genotype, electrophysiology of the retina, and objective hearing tests. Nevertheless, this is an open-labeled single-armed intervention and was intended to be an exploratory one. In the future, a proper control group should be considered in a large multicenter double-blind prospective study with stratified randomization 5. Conclusions This study introduces ONS-based therapy as a novel method for treatment of USH2, a disease that currently has no treatment. It demonstrates the efficacy and safety of ONS-based therapy as a comprehensive treatment for various sense organs involved in this syndrome irrespective of genetic background. The neuroprotective effects of ONS-based therapy are probably mediated by modification of the neural circuit of the ocular and cochlear circulation. Moreover, auditory hair cells and retinal photoreceptor death could be prevented by strengthening endogenous pro-survival mechanisms or by directly blocking cell death via the expression of endogenous SP by noninvasive method of ONS. Additionally, the oxidative damage for hair cells and photoreceptors can be minimized by use of supraphysiological doses of intravenous ascorbate. 6. Patents Ismail Musallam has US and Israeli patents licensed to himself for which he has waived financial interest Abbreviations The following abbreviations are used in this manuscript: AA Ascorbic acid Ach Acetylcholine Akt Protein kinase B (Akt = PKB) BCVA Best corrected visual acuity CGRP Calcitonin gene-related peptide ChBF Choroidal blood flow CoBF Cochlear blood flow CS Contrast sensitivity GSK-3 beta Glycogen synthase kinase-3 beta LLQ-10 Low Luminance Questionnaire-10 NA Noradrenaline NK-1R Neurokinin-1 receptor NO Nitric oxide NPY Neuropeptide Y ONS Ophthalmic nerve stimulation OBF Ocular blood flow PPG: Pterygopalatine ganglion RP Retinitis pigmentosa RVLM Rostro-ventrolateral medulla RPE Retinal pigment Epithelium SNHL Sensorineural hearing loss SD-OCT Spectral Domain Optical coherence tomography SP Substance P SCG Superior cervical ganglion SSN Superior salivatory nucleus TG Trigeminal ganglion USH2 Type 2 Usher syndrome VAP Vasoactive intestinal polypeptide Declarations Author Contributions: I.M. (Musallam I). conceptualized and designed the study. analyzed the data, wrote the manuscript, supervised the project and have read and agreed to the published version of the manuscript. Funding: none. Institutional Review Board Statement This study was approved by the Institutional Review Board, and it adhered to the tenets of the Declaration of Helsinki. Written informed consent was obtained from all participants or legal guardians after an explanation of the purpose and possible outcome of therapy. Data Availability The datasets used and/or analyzed during the current study are available from the corresponding author upon reasonable request. Conflicts of Interest: Ismail Musallam has US and Israeli patents licensed to himself. Study Registration: The study has not been registered in advance References Mathur P, Yang J. Usher syndrome: Hearing loss, retinal degeneration and associated abnormalities. Biochimica et biophysica acta. 2015;1852(3):406-20. 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Release of vasoactive peptides in the extracerebral circulation of humans and the cat during activation of the trigeminovascular system. Annals of neurology. 1988;23(2):193-6. Suzuki N, Hardebo JE, Owman C. Trigeminal fibre collaterals storing substance P and calcitonin gene-related peptide associate with ganglion cells containing choline acetyltransferase and vasoactive intestinal polypeptide in the sphenopalatine ganglion of the rat. An axon reflex modulating parasympathetic ganglionic activity? Neuroscience. 1989;30(3):595-604. Hosaka F, Yamamoto M, Cho KH, Jang HS, Murakami G, Abe S. Human nasociliary nerve with special reference to its unique parasympathetic cutaneous innervation. Anatomy & cell biology. 2016;49(2):132-7. Stjernschantz J, Geijer C, Bill A. Electrical stimulation of the fifth cranial nerve in rabbits: effects on ocular blood flow, extravascular albumin content and intraocular pressure. Experimental eye research. 1979;28(2):229-38. 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Campochiaro PA, Iftikhar M, Hafiz G, Akhlaq A, Tsai G, Wehling D, et al. Oral N-acetylcysteine improves cone function in retinitis pigmentosa patients in phase I trial. The Journal of clinical investigation. 2020;130(3):1527-41. Kang HS, Park JJ, Ahn SK, Hur DG, Kim HY. Effect of high dose intravenous vitamin C on idiopathic sudden sensorineural hearing loss: a prospective single-blind randomized controlled trial. European archives of oto-rhino-laryngology : official journal of the European Federation of Oto-Rhino-Laryngological Societies (EUFOS) : affiliated with the German Society for Oto-Rhino-Laryngology - Head and Neck Surgery. 2013;270(10):2631-6. Oudemans-van Straaten HM, Spoelstra-de Man AM, de Waard MC. Vitamin C revisited. Critical care (London, England). 2014;18(4):460. Bruno RM, Daghini E, Ghiadoni L, Sudano I, Rugani I, Varanini M, et al. Effect of acute administration of vitamin C on muscle sympathetic activity, cardiac sympathovagal balance, and baroreflex sensitivity in hypertensive patients. The American journal of clinical nutrition. 2012;96(2):302-8. Musallam IM. Ocular Neuromodulation as a Novel Treatment for Retinitis Pigmentosa: Identifying Rod Responders and Predictors of Visual Improvement. Preprints: Preprints; 2024. Ungar OJ, Cavel O, Oron Y, Wengier A, Wasserzug O, Handzel O. A Subjective Rating Scale for Initial Assessment of Sudden Unilateral Sensorineural Hearing Loss. Audiology and Neurotology. 2017;22(3):154-9. Guidance for industry: patient-reported outcome measures: use in medical product development to support labeling claims: draft guidance. Health and quality of life outcomes. 2006;4:79. Heneghan C, Goldacre B, Mahtani KR. Why clinical trial outcomes fail to translate into benefits for patients. Trials. 2017;18(1):122. Weldring T, Smith SM. Patient-Reported Outcomes (PROs) and Patient-Reported Outcome Measures (PROMs). Health services insights. 2013;6:61-8. Wang W, Lee SJ, Scott PA, Lu X, Emery D, Liu Y, et al. Two-Step Reactivation of Dormant Cones in Retinitis Pigmentosa. Cell reports. 2016;15(2):372-85. Hayreh SS. In vivo choroidal circulation and its watershed zones. Eye (London, England). 1990;4 ( Pt 2):273-89. Kim JH, Jung Y, Kim BS, Kim SH. Stem cell recruitment and angiogenesis of neuropeptide substance P coupled with self-assembling peptide nanofiber in a mouse hind limb ischemia model. Biomaterials. 2013;34(6):1657-68. Gale JE, Meyers JR, Periasamy A, Corwin JT. Survival of bundleless hair cells and subsequent bundle replacement in the bullfrog's saccule. Journal of neurobiology. 2002;50(2):81-92. Saito R, Nonaka S, Konishi H, Takano Y, Shimohigashi Y, Matsumoto H, et al. Pharmacological properties of the tachykinin receptor subtype in the endothelial cell and vasodilation. Annals of the New York Academy of Sciences. 1991;632:457-9. Lee D, Hong HS. Substance P Alleviates Retinal Pigment Epithelium Dysfunction Caused by High Glucose-Induced Stress. Life (Basel, Switzerland). 2023;13(5). Ou K, Mertsch S, Theodoropoulou S, Wu J, Liu J, Copland DA, et al. Restoring retinal neurovascular health via substance P. Experimental cell research. 2019;380(2):115-23. Backman LJ, Eriksson DE, Danielson P. Substance P reduces TNF-α-induced apoptosis in human tenocytes through NK-1 receptor stimulation. British journal of sports medicine. 2014;48(19):1414-20. Yang J-H, Guo Z, Zhang T, Meng XX, Xie L-S. Restoration of endogenous substance P is associated with inhibition of apoptosis of retinal cells in diabetic rats. Regulatory Peptides. 2013;187:12-6. Yoo K, Son BK, Kim S, Son Y, Yu S-Y, Hong HS. Substance P prevents development of proliferative vitreoretinopathy in mice by modulating TNF-α. Mol Vis. 2017;23:933-43. Lim JE, Chung E, Son Y. A neuropeptide, Substance-P, directly induces tissue-repairing M2 like macrophages by activating the PI3K/Akt/mTOR pathway even in the presence of IFNγ. Scientific reports. 2017;7(1):9417. Baek SM, Yu SY, Son Y, Hong HS. Substance P promotes the recovery of oxidative stress-damaged retinal pigmented epithelial cells by modulating Akt/GSK-3β signaling. Mol Vis. 2016;22:1015-23. Kim DY, Piao J, Hong HS. Substance-P Inhibits Cardiac Microvascular Endothelial Dysfunction Caused by High Glucose-Induced Oxidative Stress. Antioxidants (Basel, Switzerland). 2021;10(7). Piao J, Hong HS, Son Y. Substance P ameliorates tumor necrosis factor-alpha-induced endothelial cell dysfunction by regulating eNOS expression in vitro. Microcirculation (New York, NY : 1994). 2018;25(3):e12443. Kanagawa E, Sugahara K, Hirose Y, Mikuriya T, Shimogori H, Yamashita H. Effects of substance P during the recovery of hearing function after noise-induced hearing loss. Brain Res. 2014;1582:187-96. Scholzen TE, Luger TA. Neutral endopeptidase and angiotensin-converting enzyme -- key enzymes terminating the action of neuroendocrine mediators. Experimental dermatology. 2004;13 Suppl 4:22-6. Diekmann O, Tschesche H. Degradation of kinins, angiotensins and substance P by polymorphonuclear matrix metalloproteinases MMP 8 and MMP 9. Brazilian journal of medical and biological research = Revista brasileira de pesquisas medicas e biologicas. 1994;27(8):1865-76. Pernow B. Inactivation of substance P by proteolytic enzymes. Acta physiologica Scandinavica. 1955;34(4):295-302. Mashaghi A, Marmalidou A, Tehrani M, Grace PM, Pothoulakis C, Dana R. Neuropeptide substance P and the immune response. Cellular and molecular life sciences : CMLS. 2016;73(22):4249-64. Schaffalitzky De Muckadell OB, Aggestrup S, Stentoft P. Flushing and plasma substance P concentration during infusion of synthetic substance P in normal man. Scandinavian journal of gastroenterology. 1986;21(4):498-502. Saidi M, Kamali S, Beaudry F. Characterization of Substance P processing in mouse spinal cord S9 fractions using high-resolution Quadrupole-Orbitrap mass spectrometry. Neuropeptides. 2016;59:47-55. Mulders WH, Robertson D. Morphological relationships of peptidergic and noradrenergic nerve terminals to olivocochlear neurones in the rat. Hearing research. 2000;144(1-2):53-64. Wynne B, Robertson D. Somatostatin and substance P-like immunoreactivity in the auditory brainstem of the adult rat. Journal of chemical neuroanatomy. 1997;12(4):259-66. Lyon MJ, Payman RN. Comparison of the vascular innervation of the rat cochlea and vestibular system. Hearing research. 2000;141(1-2):189-98. Qiu J. [Localization of substance P in middle ear mucosa and peripheral auditory pathways in guinea pigs]. Zhonghua er bi yan hou ke za zhi. 1991;26(3):148-50, 88. Ylikoski J, Pirvola U, Häppölä O, Panula P, Virtanen I. Immunohistochemical demonstration of neuroactive substances in the inner ear of rat and guinea pig. Acta oto-laryngologica. 1989;107(5-6):417-23. Schickinger B, Ehrenberger K, Felix D, Heiniger-Bürki C, Imboden H, Davies WE, et al. Substance P in the auditory hair cells in the guinea pig. ORL; journal for oto-rhino-laryngology and its related specialties. 1996;58(3):121-6. Uddman R, Ninoyu O, Sundler F. Adrenergic and peptidergic innervation of cochlear blood vessels. Archives of oto-rhino-laryngology. 1982;236(1):7-14. Amssayef A, Bouadid I, Eddouks M. Vitamin C Inhibits Angiotensin-Converting Enzyme-2 in Isolated Rat Aortic Ring. Cardiovascular & hematological disorders drug targets. 2021;21(4):235-42. Schmid C, Ghirlanda-Keller C, Gosteli-Peter M. Ascorbic acid decreases neutral endopeptidase activity in cultured osteoblastic cells. Regulatory peptides. 2005;130(1):57-66. Zuo Y, Zheng Z, Huang Y, He J, Zang L, Ren T, et al. Vitamin C promotes ACE2 degradation and protects against SARS-CoV-2 infection. EMBO reports. 2023;24(4):e56374. Fujimoto C, Yamasoba T. Oxidative stresses and mitochondrial dysfunction in age-related hearing loss. Oxid Med Cell Longev. 2014;2014:582849. Kamogashira T, Fujimoto C, Yamasoba T. Reactive oxygen species, apoptosis, and mitochondrial dysfunction in hearing loss. Biomed Res Int. 2015;2015:617207. Kopke R, Bielefeld E, Liu J, Zheng J, Jackson R, Henderson D, et al. Prevention of impulse noise-induced hearing loss with antioxidants. Acta oto-laryngologica. 2005;125(3):235-43. Molina SJ, Miceli M, Guelman LR. Noise exposure and oxidative balance in auditory and extra-auditory structures in adult and developing animals. Pharmacological approaches aimed to minimize its effects. Pharmacological research. 2016;109:86-91. Campochiaro PA, Mir TA. The mechanism of cone cell death in Retinitis Pigmentosa. Progress in retinal and eye research. 2018;62:24-37. Martínez-Fernández de la Cámara C, Salom D, Sequedo MD, Hervás D, Marín-Lambíes C, Aller E, et al. Altered antioxidant-oxidant status in the aqueous humor and peripheral blood of patients with retinitis pigmentosa. PloS one. 2013;8(9):e74223. Komeima K, Rogers BS, Lu L, Campochiaro PA. Antioxidants reduce cone cell death in a model of retinitis pigmentosa. Proceedings of the National Academy of Sciences of the United States of America. 2006;103(30):11300-5. Additional Declarations The authors declare potential competing interests as follows: Ismail Musallam has US and Israeli patents licensed to himself Cite Share Download PDF Status: Posted Version 1 posted You are reading this latest preprint version Research Square lets you share your work early, gain feedback from the community, and start making changes to your manuscript prior to peer review in a journal. As a division of Research Square Company, we’re committed to making research communication faster, fairer, and more useful. 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Also discoverable on Platform About Our Team In Review Editorial Policies Advisory Board Help Center Resources Author Services Accessibility API Access RSS feed Manage Cookie Preferences © Research Square 2026 | ISSN 2693-5015 (online) Privacy Policy Terms of Service Do Not Sell My Personal Information {"props":{"pageProps":{"initialData":{"identity":"rs-6190348","acceptedTermsAndConditions":true,"allowDirectSubmit":true,"archivedVersions":[],"articleType":"Research Article","associatedPublications":[],"authors":[{"id":426293097,"identity":"8e004043-8bc7-4ba5-9c2b-f2a4ba8be919","order_by":0,"name":"Ismail M Musallam","email":"data:image/png;base64,iVBORw0KGgoAAAANSUhEUgAAAZAAAAAyAQMAAABI0h/eAAAABlBMVEX///8AAABVwtN+AAAACXBIWXMAAA7EAAAOxAGVKw4bAAABCUlEQVRIiWNgGAWjYLCCBwVwpk0CmEoowKEUBhIM4My0BAY2VBGCWg5DtDDg0aI7I/3hhwQDu2gG9h7DBww15/P45bsTPzwwYJDnFzuAVYvZjRxjiQSD5NwGnjPGBgzHbhdLtvFuBoowGM6cnYBLCwNQAXNug0SOmQRjw+3EDcd4N4C0JBjcxqUl/fGPBIP63Ab5N+Y/GBvOgbRs/oFfS4IZ0MzDQFt4zBgYGw6AtGzDb8uZN2YWCQbHgX5JK5ZgOJacOLMtdxtQRAK3X46nP77xoaI6t4H98MYPDDV2if3MZzff/FFhI88vjV0LHNgf4DBg/oPgS+BXDgHsD4hRNQpGwSgYBSMQAADl7F6V9SIX9gAAAABJRU5ErkJggg==","orcid":"https://orcid.org/0000-0001-5981-1623","institution":"Musallam Eye Center of Jerusalem","correspondingAuthor":true,"prefix":"","firstName":"Ismail","middleName":"M","lastName":"Musallam","suffix":""}],"badges":[],"createdAt":"2025-03-09 20:32:08","currentVersionCode":1,"declarations":{"humanSubjects":true,"vertebrateSubjects":false,"conflictsOfInterestStatement":true,"humanSubjectEthicalGuidelines":true,"humanSubjectConsent":true,"humanSubjectClinicalTrial":true,"humanSubjectCaseReport":true,"vertebrateSubjectEthicalGuidelines":false},"doi":"10.21203/rs.3.rs-6190348/v1","doiUrl":"https://doi.org/10.21203/rs.3.rs-6190348/v1","draftVersion":[],"editorialEvents":[],"editorialNote":"","failedWorkflow":false,"files":[{"id":78516704,"identity":"d2a2bc4b-1abd-465e-ade0-7d0e33c6cc49","added_by":"auto","created_at":"2025-03-14 11:00:16","extension":"png","order_by":1,"title":"Figure 1","display":"","copyAsset":false,"role":"figure","size":129521,"visible":true,"origin":"","legend":"\u003cp\u003eSchematic of neural pathways that control ocular (OBF) and cochlear blood flow (CoBF).ONS; ophthalmic nerve stimulation, SNS; sympathetic nervous system, PNS; parasympathetic nervous system, TVS: trigeminovascular system, TG: trigeminal ganglion, PPG: pterygopalatine ganglion, SSN: superior salivatory nucleus, SCG: superior cervical ganglion, RVLM: rostro-ventrolateral medulla, SP: substance P, CGRP; calcitonin gene-related peptide, Ach: acetylcholine, NO: nitric oxide, VAP: vasoactive intestinal polypeptide, NA: noradrenaline, NPY; neuropeptide Y.\u003c/p\u003e","description":"","filename":"1.png","url":"https://assets-eu.researchsquare.com/files/rs-6190348/v1/b628bef9f1a9cf80e516e587.png"},{"id":78515378,"identity":"ae5c32c8-0c13-4341-b918-87932ca27c19","added_by":"auto","created_at":"2025-03-14 10:44:16","extension":"png","order_by":2,"title":"Figure 2","display":"","copyAsset":false,"role":"figure","size":46223,"visible":true,"origin":"","legend":"\u003cp\u003eChanges in scotopic vision; Mean value and standard deviation of LLQ-10 score among the whole cohort at baseline and at post-treatment visits. * (* p≤ 0.05); statistically significant differences in the mean value of ILQ-10 score at different post-treatment visits compared to baseline respectively.\u003c/p\u003e","description":"","filename":"2.png","url":"https://assets-eu.researchsquare.com/files/rs-6190348/v1/931fe3424cfe7b3654521079.png"},{"id":78517173,"identity":"e0c5ba3c-904d-4411-860f-802b71e46f51","added_by":"auto","created_at":"2025-03-14 11:08:16","extension":"pdf","order_by":0,"title":"","display":"","copyAsset":false,"role":"manuscript-pdf","size":862470,"visible":true,"origin":"","legend":"","description":"","filename":"manuscript.pdf","url":"https://assets-eu.researchsquare.com/files/rs-6190348/v1/96b0002e-6e8a-4a90-9619-d04a557a54ad.pdf"}],"financialInterests":"The authors declare potential competing interests as follows: Ismail Musallam has US and Israeli patents licensed to himself","formattedTitle":"\u003cp\u003e\u003cstrong\u003eCo-targeting Dysregulated Ocular and Cochlear Blood Flow via Ophthalmic Nerve Stimulation for the Treatment of Type 2 Usher Syndrome: Prospective Case Series\u003c/strong\u003e\u003c/p\u003e","fulltext":[{"header":"1. Introduction","content":"\u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eUsher syndrome (USH) represents a group of autosomal recessive disorders characterized by sensorineural hearing loss (SNHL), vision loss, and vestibular dysfunction (\u003cspan citationid=\"CR1\" class=\"CitationRef\"\u003e1\u003c/span\u003e). It primarily affects the auditory hair cells followed by light-sensitive photoreceptor cells in the retina (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e). USH is the leading genetic cause of deaf-blindness (50%of all cases), with a prevalence in the range of 4 to 17 per 100,000 people. (\u003cspan citationid=\"CR3\" class=\"CitationRef\"\u003e3\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThree clinical subtypes of USH have been defined \u0026ndash; Type 1, Type 2, and Type 3 \u0026ndash; based upon the severity, and progression of SNHL. Type 2 (USH2) is the most common subtype, accounting for more than half of all USH cases worldwide (\u003cspan citationid=\"CR4\" class=\"CitationRef\"\u003e4\u003c/span\u003e). USH2 patients present with moderate-to-severe SNHL, normal vestibular function, and retinitis pigmentosa (RP) that begins generally during puberty.\u003c/p\u003e \u003cp\u003eThere is no cure for USH2. Several treatment options are available to help patients manage hearing problems. This includes auditory rehabilitation using hearing aid devices or cochlear implants. Unfortunately, these technologies do not treat the underlying cause of hearing loss and are far from providing quality natural hearing. Moreover, there is no cure for RP. Currently, there are a range of new therapeutic strategies being under investigation that utilize new technologies such as gene and cell-based therapies. Given that USH2 is a syndromic condition affecting multiple senses, it is important to note that current therapeutic trials generally focus on one sense rather than the full spectrum of the disease. Therefore, a comprehensive therapeutic strategy is needed that simultaneously targets the underlying pathophysiology in both the retina and the inner ear regardless of the genetic background of the disease\u003c/p\u003e \u003cp\u003eThe human inner ear depends on a vascular supply to maintain fluid homeostasis, ion balance, and metabolic supply. Disruption of cochlear blood flow (CoBF) leads to instantaneous pathological changes in the inner ear (\u003cspan citationid=\"CR5\" class=\"CitationRef\"\u003e5\u003c/span\u003e) (\u003cspan citationid=\"CR6\" class=\"CitationRef\"\u003e6\u003c/span\u003e). A growing body of evidence suggests that microvascular changes may contribute to hearing loss. (\u003cspan citationid=\"CR7\" class=\"CitationRef\"\u003e7\u003c/span\u003e) (\u003cspan citationid=\"CR8\" class=\"CitationRef\"\u003e8\u003c/span\u003e) (\u003cspan citationid=\"CR9\" class=\"CitationRef\"\u003e9\u003c/span\u003e) (\u003cspan citationid=\"CR10\" class=\"CitationRef\"\u003e10\u003c/span\u003e) (\u003cspan citationid=\"CR11\" class=\"CitationRef\"\u003e11\u003c/span\u003e). Histopathological examination of the temporal bone of patients with USH revealed atrophy of the stria vascularis (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e). This is thought to cause degeneration of hair cells by alteration of the endolymph composition. (\u003cspan citationid=\"CR13\" class=\"CitationRef\"\u003e13\u003c/span\u003e). The stria vascularis, a highly vascularized region located in the cochlear microcirculation, is the metabolic energy source underlying cochlear function (\u003cspan citationid=\"CR14\" class=\"CitationRef\"\u003e14\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eCirculatory changes have also been observed in the retina and choroid of patients with Usher\u0026rsquo;s syndrome. Vascular density was significantly decreased in the retinal circulation of patients with \u003cem\u003eUSH2A\u003c/em\u003e and \u003cem\u003eMYO7A\u003c/em\u003e mutations compared with controls (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e). Changes were observed in both the superficial and deep capillary plexus. Vascular density in the deep capillary plexus showed strong association with macular sensitivity and ellipsoid zoon width (\u003cspan citationid=\"CR15\" class=\"CitationRef\"\u003e15\u003c/span\u003e). Choroidal thickness is reduced in people with USH2 if compared to healthy subjects., which might be considered as a sign of blood flow reduction (\u003cspan citationid=\"CR16\" class=\"CitationRef\"\u003e16\u003c/span\u003e). Evaluating choroidal hemodynamic changes has shown the reduction in choroidal blood flow (ChBF) is proportional to the progression of RP (\u003cspan citationid=\"CR17\" class=\"CitationRef\"\u003e17\u003c/span\u003e). Recently, relative choroidal ischemia and reduced ChBF have been demonstrated in RP by measuring the choroidal vascularity index via Spectral Domain Optical coherence tomography (SD-OCT) (\u003cspan citationid=\"CR18\" class=\"CitationRef\"\u003e18\u003c/span\u003e). The ocular blood flow (OBF) is reduced not only in the retina and choroid but also in the retro-ocular vessels (\u003cspan citationid=\"CR19\" class=\"CitationRef\"\u003e19\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe eye receives its blood supply through the anterior cerebral circulation (Internal carotid arteries) via ophthalmic arteries. The retina has a dual blood supply, with the retinal circulation supplying the inner layers and the choroidal circulation supplying the outer layers of the retina including photoreceptors. The blood supply to the choroid in mammals arises from the ophthalmic artery via the long and short ciliary arteries. Like the eye, the inner ear is supplied with an end artery. The blood supply to the inner ear is derived from the posterior cerebral circulation (Basilar artery), mostly from labyrinthine artery, a branch of the anterior inferior cerebellar artery.\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eOphthalmic artery, central retinal artery, posterior ciliary arteries, and choroid are richly innervated by parasympathetic, sympathetic and trigeminal sensory nerve fibers. The parasympathetic innervation has been shown to cause vasodilation and increase ChBF, the sympathetic input has been shown to induce vasoconstriction and decrease ChBF. In contrast, autonomic control for cochlear blood vessels is limited to sympathetic innervation that arises from superior cervical ganglion, whereas parasympathetic innervation of cochlear blood vessels has not been found (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe majority of the intracranial blood vessels including those contributing to both the ocular and cochlear circulation are innervated by the ophthalmic division of the trigeminal nerve (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e) (\u003cspan citationid=\"CR21\" class=\"CitationRef\"\u003e21\u003c/span\u003e). Direct stimulation of the trigeminal ganglion in experimental models has been found to lead to increased cerebral blood flow (\u003cspan citationid=\"CR22\" class=\"CitationRef\"\u003e22\u003c/span\u003e). Stimulation of the ophthalmic nerve may be topographically oriented, with specific branches leading to increased blood flow in specific regions of the eye, ear, and brain arterial tree.\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003cp\u003eThe ophthalmic nerve plays a pivotal role in the regulation of ocular and CoBF (Fig.\u0026nbsp;\u003cspan refid=\"Fig1\" class=\"InternalRef\"\u003e1\u003c/span\u003e), by inducing vasodilation, as a result of the release of substance P (SP), calcitonin gene-related peptide (CGRP), and neuropeptide Y as mediators of vasodilative mechanisms (\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e). Stimulation of sensory afferents from the ophthalmic nerve results in the activation of the trigeminovascular system via antidromic impulse (\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e) (\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e) (\u003cspan citationid=\"CR27\" class=\"CitationRef\"\u003e27\u003c/span\u003e). The nasociliary nerve, which originates from the ophthalmic nerve, contains major vasodilatory innervation (\u003cspan citationid=\"CR28\" class=\"CitationRef\"\u003e28\u003c/span\u003e, \u003cspan citationid=\"CR29\" class=\"CitationRef\"\u003e29\u003c/span\u003e), and its stimulation results in the release of vasoactive neuropeptides from the free nerve endings, such as SP and CGRP (\u003cspan citationid=\"CR23\" class=\"CitationRef\"\u003e23\u003c/span\u003e, \u003cspan citationid=\"CR24\" class=\"CitationRef\"\u003e24\u003c/span\u003e). Consistent with this, stimulation of the ophthalmic nerve in experimental animals was found to increase OBF (\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e), decrease carotid arterial resistance, (\u003cspan citationid=\"CR31\" class=\"CitationRef\"\u003e31\u003c/span\u003e) and enhance uveal release of SP (\u003cspan citationid=\"CR32\" class=\"CitationRef\"\u003e32\u003c/span\u003e). Infusion of the internal carotid artery or the anterior inferior cerebellar artery with SP induces increase in CoBF (\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e).\u003cdiv class=\"BlockQuote\"\u003e\u003cp\u003eThere is extensive evidence that excessive free radical formation along with diminished CoBF are essential factors involved in the pathogenesis of SNHL (\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e) (\u003cspan citationid=\"CR35\" class=\"CitationRef\"\u003e35\u003c/span\u003e). Anti-oxidant interventions have been reported to substantially delay the death of photoreceptor cells of the retina (\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e) and hair cells of the inner ear (\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e). Interestingly, high dose of vitamin C has been shown to enhance hearing recovery in idiopathic sudden SNHL patients, which suggests that ascorbate treatment reduces levels of reactive oxygen metabolites produced by inner ear ischemia or inflammation. (\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e). Therefore, it is reasonable to suggest that supra-physiological doses of intravenous AA could be beneficial in a perfusion-compromised retina or inner ear as in USH2. Preclinical studies show that high-dose of AA can prevent or restore microcirculatory flow impairment. AA can restore vascular responsiveness to vasoconstrictors and preserve endothelial barrier (\u003cspan citationid=\"CR38\" class=\"CitationRef\"\u003e38\u003c/span\u003e). Additionally, AA exerts sympatholytic effects via central action (\u003cspan citationid=\"CR39\" class=\"CitationRef\"\u003e39\u003c/span\u003e).\u003c/p\u003e\u003cp\u003ePathogenesis of retinal and inner ear disorders in USH2 reflects a summation of primary reduced dysregulated OBF/CoBF, and consecutive unstable oxygen supply, hair cell/ photoreceptors starvation, oxidative stress and inflammation of the retina and the inner ear. These vascular and cellular processes are ultimately combined to produce damage of cochlear hair cells followed by retinal rods and finally cone photoreceptors apoptosis. The mechanisms responsible for reduced dysregulated ocular and CoBF in patients with USH2 uncertain. It is also uncertain if blood flow decline stems from disturbed neural control of ocular and cochlear circulation or vascular pathology, or both.\u003c/p\u003e\u003cp\u003eThe reduced dysregulated CoBF/ OBF is proposed as a common pathway in the pathogenesis of SNHL and blindness, which can be a target for a novel therapeutic strategy in USH2. Because of complexity of this syndrome, it is unlikely that optimal protection of the inner ear mechanoreceptor and retinal photoreceptors can be achieved through a single therapy, suggesting that an alternative approach based on combination therapy to be considered. Up-regulation of trigeminovascular system and parasympathetic system via ONS along with anti-oxidation, sympathetic inhibition and restoration of vascular endothelial function via systemic administration of AA might form the basis of neuromodulation therapy for treating this non-curable multi-sense damaging syndrome. Here we report the safety and efficacy results of ONS paired with systemic ascorbate for treating participants with USH2.\u003c/p\u003e\u003c/div\u003e\u003c/p\u003e"},{"header":"2. Materials and Methods","content":"\u003cp\u003e\u003cdiv class=\"BlockQuote\"\u003e\u003cp\u003eThe study was a prospective, consecutive, open-label, interventional study aimed at investigating the safety and efficacy of ONS combined with AA on patients with USH2. The protocol was reviewed and approved by the Ethics Committee of the Musallam Specialty Hospital, Palestinian Authority, and it adhered to the tenets of the Declaration of Helsinki. Written informed consent was obtained from all participants or legal guardians after an explanation of the purpose and possible outcome of therapy.\u003c/p\u003e\u003c/div\u003e\u003c/p\u003e \u003cdiv id=\"Sec3\" class=\"Section2\"\u003e \u003ch2\u003e2.1 Patient Selection\u003c/h2\u003e \u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003ePatient enrollment started in April 2018, and cases were recruited from a single tertiary ophthalmic center. The inclusion criteria of the study included patients with an established diagnosis of USH2 with BCVA\u0026thinsp;\u0026ge;\u0026thinsp;20/400. Exclusion criteria were pregnancy, glaucoma, central retinal vein occlusion, retinal detachment. bradycardia, malignancy, and active infection.\u003c/p\u003e \u003cp\u003eFollowing recruitment, all participants with USH2 completed baseline vision testing, slit lamp examination and fundoscopy. Spectral Domain Optical coherence tomography (SD-OCT) was performed within 2\u0026ndash;4 weeks prior to treatment. The severity of the RP associated with Usher\u0026rsquo;s syndrome was clinically graded into six stages based on fundoscopy and SD-OCT findings (\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e).\u003c/p\u003e \u003cp\u003eThe primary efficiency endpoints were 6-month changes in scotopic vision and adverse effects of the therapy. The scotopic vision was assessed with a Low luminance questionnaire-10 (LLQ-10) (\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e). The secondary efficiency points include best corrected visual acuity (BCVA), contrast sensitivity (CS), and the change of hearing acuity. BCVA was recorded in Snellen equivalents and then transformed to logMAR unit. CS was measured using a Pelli- Robson chart under standardized photopic illumination at one meter in each eye, using best correction spectacles. To characterize the severity of hearing loss at baseline a subjective numerical rating scale (NRS) of 1 (normal hearing) to 6 (complete deafness) was used (\u003cspan citationid=\"CR41\" class=\"CitationRef\"\u003e41\u003c/span\u003e). Improvement of an NRS score by 2 or more points was considered a clinically significant improvement of hearing.\u003c/p\u003e \u003cp\u003eTo evaluate the therapy outcome, LLQ-10 score changes from baseline to 6 months after treatment were calculated (The primary efficacy endpoint). LLQ-10 has been used to quantify the participant\u0026rsquo;s visual dysfunction under low luminance, integrating aspects of visual acuity, mobility, visual field, depth of perception, reading, color vision, dependency on others\u0026rsquo; help, social functions, and mental health (\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e). Patient-centered outcome measures have been increasingly viewed as necessary in clinical trials, serving as primary endpoints(\u003cspan citationid=\"CR40\" class=\"CitationRef\"\u003e40\u003c/span\u003e) (\u003cspan citationid=\"CR42\" class=\"CitationRef\"\u003e42\u003c/span\u003e) (\u003cspan citationid=\"CR43\" class=\"CitationRef\"\u003e43\u003c/span\u003e) (\u003cspan citationid=\"CR44\" class=\"CitationRef\"\u003e44\u003c/span\u003e). Participants rated how much difficulty they had performing each of the activities under low luminance on a 5-point Likert scale. Rod responders were identified as those in whom the LLQ-10 score increased by \u0026ge;\u0026thinsp;25 points at 6 months\u0026rsquo; post-treatment.\u003c/p\u003e \u003cp\u003eFor safety purposes, the vital signs and electrocardiogram were monitored before, during, and after each session of neuromodulation. Ocular and non-ocular adverse events were also reported (Primary safety endpoint)\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec4\" class=\"Section2\"\u003e \u003ch2\u003e2.2 The Treatment Protocol\u003c/h2\u003e \u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eONS-based therapy protocol included the administration of AA followed by ONS over a period of two weeks. AA was given intravenously in a dose of 3 gm dissolved in 100 ml of saline infused at 5 ml min\u003csup\u003e\u0026minus;\u0026thinsp;1\u003c/sup\u003e for 20 min on the first day, followed by a drip infusion of 1 g daily during the rest of the treatment period.\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003cp\u003eA modulated low magnitude, low-frequency vibration, with a frequency of approximately 60 Hz- 90 Hz and stimulation amplitude range of 1.5 \u0026micro;m- 3.5 \u0026micro;m were used for ONS. Modulated frequency and amplitude waveforms were used to avoid adaptation to mechanical stimulus. A head-mounted prototype of ophthalmic nerve stimulators was applied to different application sites by using modified commercially available micro-vibrators, along with different types of intranasal and extra-nasal application heads.\u003cdiv class=\"BlockQuote\"\u003e\u003cp\u003eBilateral ONS was applied to the subjects over a session of 30 minutes per day, 6 sessions per week, and one day off. Each session included intra-nasal vibrochemical, and extra-nasal vibrotactile stimulation over the nasal bridge and the supraorbital region in both sides, using appropriate application heads. During intra-nasal vibrochemical stimulation, the nasal application head was covered by a rubber cap impregnated by 2% menthol cream (Dermacool plus \u003csup\u003eR\u003c/sup\u003e) as a transient receptor potential cation channel subfamily M (melastatin) member 8 (TRPM8) agonist. Participants were assessed clinically, and by SD-OCT, and ILQ-10 at baseline and at 1, 6 months after treatment\u003c/p\u003e\u003c/div\u003e\u003c/p\u003e \u003c/div\u003e \u003cdiv id=\"Sec5\" class=\"Section2\"\u003e \u003ch2\u003e2.3 Statistical analysis\u003c/h2\u003e \u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eThe statistical analysis was performed using SPSS\u003csup\u003eR\u003c/sup\u003e, V. 21. The change in the mean value of visual function parameters between baseline and different post-treatment points was evaluated using a paired sample t-test. The BCVA and CS at baseline, and at post-treatment visits were recorded for the left eye in the whole cohort. All tests were two-tailed and a \u003cem\u003ep\u003c/em\u003e-value less than 0.05 was considered significant.\u003c/p\u003e \u003cp\u003eThe proportion of left eyes that showed clinically significant improvement in visual function, among the whole cohort was also evaluated. Additionally, the proportion of patients who experienced clinically significant improvement in hearing was also determined\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e \u003c/div\u003e"},{"header":"3. Results","content":"\u003cdiv class=\"BlockQuote\"\u003e\n \u003cp\u003e\u003cstrong\u003eBaseline Demographics\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003eBetween April 14, 2018, and April 29, 2020, 9 participants, aged between 8\u0026ndash;55 years (median; 23 years), mean (24.8\u0026thinsp;\u0026plusmn;\u0026thinsp;13.9) among them 7 females (77.8%) were recruited. The demographics and baseline characteristics of participants are presented in Table.\u003c/p\u003e\n \u003cp\u003eAt 6 months\u0026rsquo; visits, 7 out of 9 (77.7%) patients were defined as rod responders. In the whole cohort, the night vision score was improved from 21.4\u0026thinsp;\u0026plusmn;\u0026thinsp;11.3 points at baseline to a level of 59.2\u0026thinsp;\u0026plusmn;\u0026thinsp;19.8 points (\u003cem\u003eP\u0026thinsp;=\u0026thinsp;0.0001\u003c/em\u003e) at 1 month and, 56.4\u0026thinsp;\u0026plusmn;\u0026thinsp;18.7 points (P\u0026thinsp;=\u0026thinsp;\u003cem\u003e0.0001\u003c/em\u003e) at 6 months (Fig.\u0026nbsp;\u003cspan class=\"InternalRef\"\u003e2\u003c/span\u003e). In rod responders, the mean change in LLQ-10 score was 45.4\u0026thinsp;\u0026plusmn;\u0026thinsp;8.1 points (p\u0026thinsp;=\u0026thinsp;0.0001) at one month and (42.1\u0026thinsp;\u0026plusmn;\u0026thinsp;11.3 points) (\u003cem\u003ep\u0026thinsp;=\u0026thinsp;0.0001\u003c/em\u003e) at 6 months. The two non-responders have stage III and V RP and the duration of night blindness was 12 and 20 years (16\u0026thinsp;\u0026plusmn;\u0026thinsp;5.7).\u003c/p\u003e\n \u003cp\u003eThe BCVA of the left eye was improved from (0.38\u0026thinsp;\u0026plusmn;\u0026thinsp;0.43 logMAR) at baseline to (0.26\u0026thinsp;\u0026plusmn;\u0026thinsp;0.33 Log MAR) (\u003cem\u003eP\u0026thinsp;=\u0026thinsp;0.19\u003c/em\u003e) at 1 month and (0.28\u0026thinsp;\u0026plusmn;\u0026thinsp;0.34 Log MAR) (\u003cem\u003ep\u0026thinsp;=\u0026thinsp;0.24\u003c/em\u003e) at 6 months which was found statistically not significant. In two out of 9 patients (22.2%), a clinically significant improvement of BCVA (improvement of \u0026ge;\u0026thinsp;0.2 logMAR) was noticed.\u003c/p\u003e\n \u003cp\u003eRegarding CS, the mean CS of the left eye was improved from 0.64\u0026thinsp;\u0026plusmn;\u0026thinsp;0.60 log unit to 0.79\u0026thinsp;\u0026plusmn;\u0026thinsp;0.57 Log unit at 1 month and to a value of 0.76\u0026thinsp;\u0026plusmn;\u0026thinsp;0.54 log unit at 6 months. However, this change was not statistically significant. At 6 months\u0026rsquo; visits, two eyes (22.2%) of the whole cohort had a clinically significant improvement of CS (improvement of \u0026ge;\u0026thinsp;0.3 log unit).\u003c/p\u003e\n \u003cp\u003eInterestingly, a clinically significant improvement in hearing was observed in 3 patients (33.3%). No serious adverse effects were reported and headache in one patient was the only encountered side effect in this study.\u003c/p\u003e\n\u003c/div\u003e\n\u003cdiv class=\"gridtable\"\u003e\n \u003cdiv align=\"left\" class=\"colspec\"\u003e\u003cstrong\u003eTable Baseline characteristics\u003c/strong\u003e\u003c/div\u003e\u0026nbsp;\u003ctable id=\"Taba\" border=\"1\"\u003e\n \u003ccolgroup cols=\"2\"\u003e\u003c/colgroup\u003e\n \u003cthead\u003e\n \u003ctr\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eBaseline Characteristics\u003c/p\u003e\n \u003c/th\u003e\n \u003cth align=\"left\"\u003e\n \u003cp\u003eTotal cohort\u003c/p\u003e\n \u003cp\u003e(9 patients; 9 eyes)\u003c/p\u003e\n \u003c/th\u003e\n \u003c/tr\u003e\n \u003c/thead\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eAge, Mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eRang\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eMedian age\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e24.8\u0026thinsp;\u0026plusmn;\u0026thinsp;13.9 yrs.\u003c/p\u003e\n \u003cp\u003e8\u0026ndash;55 yrs.\u003c/p\u003e\n \u003cp\u003e23\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eM/F, (male %)\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e2/7 (22.2%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eRang (yrs.) of night blindness\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eMean duration of night blindness (yrs.)\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003e\u003cstrong\u003eProportion of patients with night blindness 10 years or less\u003c/strong\u003e\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e5\u0026ndash;20 yrs.\u003c/p\u003e\n \u003cp\u003e11.8\u0026thinsp;\u0026plusmn;\u0026thinsp;6.2 yrs.\u003c/p\u003e\n \u003cp\u003e4 (44.4%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eStage of the Disease *\u003c/strong\u003e\u003c/p\u003e\n \u003cp\u003eII, no. of patients (%)\u003c/p\u003e\n \u003cp\u003eIII, no. of patients/ (%)\u003c/p\u003e\n \u003cp\u003eIV, no. of patients (%)\u003c/p\u003e\n \u003cp\u003eV, no. of patients/ (%)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e4 (44.4%)\u003c/p\u003e\n \u003cp\u003e2 (22.2%)\u003c/p\u003e\n \u003cp\u003e2 (22.2%)\u003c/p\u003e\n \u003cp\u003e1 (11.1%)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e\u003cstrong\u003eLLQ-10 Score at baseline\u003c/strong\u003e (mean\u0026thinsp;\u0026plusmn;\u0026thinsp;SD)\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd align=\"left\"\u003e\n \u003cp\u003e22.8\u0026thinsp;\u0026plusmn;\u0026thinsp;12.4\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n \u003c/table\u003e\n\u003c/div\u003e"},{"header":"4. Discussion","content":"\u003cp\u003e\u003cdiv class=\"BlockQuote\"\u003e\u003cp\u003eUp-to-date, there is no treatment option available to cure sensorineural hearing loss or vision loss in USH2, and only prostheses such as hearing aids and cochlear implants offer some help. In the current study, we have reported successful use of ONS-based therapy as a novel treatment for USH2 syndrome. It has led to a clinically meaningful and statistically significant improvement of scotopic vision. Additionally, clinically significant improvement in BCVA, contrast sensitivity and hearing was also noticed in some patients.\u003c/p\u003e\u003cp\u003eIn this study we found that the most robust improvement in visual function for the subjects with USH2 was night vision as measured by LLQ-10. Recovery of night vision was likely to be related to reactivation of dormant/starving rods via enhanced oxygen and glucose delivery and shunting metabolites toward aerobic glucose metabolism (\u003cspan citationid=\"CR45\" class=\"CitationRef\"\u003e45\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eThere are four clinico-pathological overlapping phases that characterize USH2; an early hair cell degeneration phase, rod degeneration phase, transitional phase of rod/cone photoreceptors damage and late cone degeneration phase. SNHL occurs prior to visual symptoms in all subtypes of USH syndrome including USH2 (\u003cspan citationid=\"CR2\" class=\"CitationRef\"\u003e2\u003c/span\u003e) probably because the inner ear is more vulnerable to hypoperfusion. This might be explained at least in part by early development of dysregulated CoBF as a result of dominant sympathetic innervation of cochlear blood vessels and absence parasympathetic input. (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e). Interestingly, cochlear circulation is innervated by branches of ophthalmic nerve (\u003cspan citationid=\"CR20\" class=\"CitationRef\"\u003e20\u003c/span\u003e) that produce robust dilatation of labyrinthine arteries and their branches, increasing CoBF, as result of release of vasoactive neuropeptides. Therefore, the upregulation of trigeminovascular system may counteract the dysregulated CoBF induced by increased sympathetic tone.\u003c/p\u003e\u003cp\u003eThe vascular alteration in the inner ear and the retina might explain the sequence of pathological events in USH2. A highly vulnerable cochlear circulation compared to ocular circulation might explain the early development of SNHL. Additionally, the atrophy of the stria vascularis (\u003cspan citationid=\"CR12\" class=\"CitationRef\"\u003e12\u003c/span\u003e) supports the notion that auditory hair cell damage is primarily of vascular origin. Stria vascularis, a key structure for producing and maintaining the endocochlear potential. Morphologic alterations in the stria vascularis decrease the endocochlear potential and consequently, affect the cochlear amplification of acoustic signals leading to an increase in auditory thresholds, even in the absence of hair cell death (\u003cspan citationid=\"CR34\" class=\"CitationRef\"\u003e34\u003c/span\u003e). On the other hand, the development of annular scotoma at the mid-periphery opposite the choroidal watershed zone between the anterior and posterior ciliary arteries (\u003cspan citationid=\"CR46\" class=\"CitationRef\"\u003e46\u003c/span\u003e), provides further support to the vascular theory of USH2. SNHL including Meiners disease, sudden SNHL, senile deafness and inherited NSHL were attributed at least in part to underlying vascular causes and reduced CoBF. Nevertheless, the unmet oxygen and glucose requirements of mutant hair cells and rod photoreceptors might be detrimental for the disease process and its sequence of occurrence in USH2.\u003c/p\u003e\u003cp\u003e A key finding in the present study was the substantial improvement of hearing in one third of the participants. Such an improvement is unlikely to be due to chance, and mostly related to the effect of treatment as this phenomenon is not normally seen in untreated patients with USH2. Hearing improvement might be related to one of the following mechanisms, these include recovery of dormant hair cells, regeneration of hair cells, or trans differentiation of supporting cells into hair cells. These effects might occur as a result of recruitment of endogenous stem cell in response to overexpression of SP (\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e). It is reasonable to suggest that the SNHL seen in the early stages of USH2 is explained, at least in part by hair cell dormancy. The increased CoBF and glucose delivery to the inner ear in response to ONS might restore the function of these dormant hair cells, resulting in improved hearing. Hair cell dormancy has been demonstrated in organ culture, wherein the hair cells completely loss their bundles. These dormant bundleless hair cells are viable cells that can grow back to the apical surface of the epithelium, resynthesize their bundles, and thereby restore their function (\u003cspan citationid=\"CR48\" class=\"CitationRef\"\u003e48\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eAlthough the exact mechanism of action of ONS-based therapy in USH2 is not fully understood, the distinctive effect of ONS-based therapy could be due to its capability to target diverse pathophysiologies of USH2. It is known that antidromic activation of the ophthalmic nerve especially its nasociliary branch leads to the release of neuropeptides such as SP from nerve terminals. SP, an 11-amino acid neuropeptide that belongs to the tachykinin family of peptides, has a preferential affinity to the neurokinin-1 receptor (NK-1R) (\u003cspan citationid=\"CR49\" class=\"CitationRef\"\u003e49\u003c/span\u003e). Several studies have indicated that SP is capable of boosting both the ocular/CoBF (\u003cspan citationid=\"CR25\" class=\"CitationRef\"\u003e25\u003c/span\u003e) (\u003cspan citationid=\"CR26\" class=\"CitationRef\"\u003e26\u003c/span\u003e) (\u003cspan citationid=\"CR30\" class=\"CitationRef\"\u003e30\u003c/span\u003e) (\u003cspan citationid=\"CR33\" class=\"CitationRef\"\u003e33\u003c/span\u003e) and tissue regeneration by the recruitment of endogenous stem cells (\u003cspan citationid=\"CR47\" class=\"CitationRef\"\u003e47\u003c/span\u003e). Moreover SP can protect diverse types of cells including retinal pigment epithelium (\u003cspan citationid=\"CR50\" class=\"CitationRef\"\u003e50\u003c/span\u003e), and ganglion cells (\u003cspan citationid=\"CR51\" class=\"CitationRef\"\u003e51\u003c/span\u003e). It also contribute to the prevention of apoptosis (\u003cspan citationid=\"CR52\" class=\"CitationRef\"\u003e52\u003c/span\u003e) (\u003cspan citationid=\"CR53\" class=\"CitationRef\"\u003e53\u003c/span\u003e), suppression of both inflammation (\u003cspan citationid=\"CR54\" class=\"CitationRef\"\u003e54\u003c/span\u003e) (\u003cspan citationid=\"CR55\" class=\"CitationRef\"\u003e55\u003c/span\u003e) and oxidative stress (\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e) (\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e). SP was reported to modulate Akt/GSK-3β signaling, inhibit reactive oxygen species-induced cell death, preserve cell viability, and block cellular alterations (\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e) (\u003cspan citationid=\"CR57\" class=\"CitationRef\"\u003e57\u003c/span\u003e) (\u003cspan citationid=\"CR58\" class=\"CitationRef\"\u003e58\u003c/span\u003e).SP exhibited a protective effect on the cochlea from noise damage in guinea pigs exposed to noise after an infusion of SP into the inner ear (\u003cspan citationid=\"CR59\" class=\"CitationRef\"\u003e59\u003c/span\u003e). However, the therapeutic applications of SP have been limited by its low stability. SP tends to be degraded by various proteases, including neutral endopeptidase and angiotensin-converting enzymes (\u003cspan citationid=\"CR60\" class=\"CitationRef\"\u003e60\u003c/span\u003e) (\u003cspan citationid=\"CR61\" class=\"CitationRef\"\u003e61\u003c/span\u003e) (\u003cspan citationid=\"CR62\" class=\"CitationRef\"\u003e62\u003c/span\u003e). The half-life time of SP is very short, from seconds to minutes (\u003cspan citationid=\"CR63\" class=\"CitationRef\"\u003e63\u003c/span\u003e) (\u003cspan citationid=\"CR64\" class=\"CitationRef\"\u003e64\u003c/span\u003e) (\u003cspan citationid=\"CR65\" class=\"CitationRef\"\u003e65\u003c/span\u003e)\u003c/p\u003e\u003cp\u003eIf endogenous SP is to be therapeutically effective, then the sustained release of this neuropeptide from the nerve terminals of the ophthalmic nerve is essential and its stability should be enhanced. SP has been identified at several locations in the auditory pathway (\u003cspan citationid=\"CR66\" class=\"CitationRef\"\u003e66\u003c/span\u003e) (\u003cspan citationid=\"CR67\" class=\"CitationRef\"\u003e67\u003c/span\u003e), including the inner ear (\u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e) (\u003cspan citationid=\"CR69\" class=\"CitationRef\"\u003e69\u003c/span\u003e) (\u003cspan citationid=\"CR70\" class=\"CitationRef\"\u003e70\u003c/span\u003e). Immunohistochemical and electrophysiological studies confirm that SP may act as a neuromodulator at the synapses of the inner hair cells in the guinea pig (\u003cspan citationid=\"CR71\" class=\"CitationRef\"\u003e71\u003c/span\u003e). SP immunolabeling has been observed on the labyrinthine and spiral modiolar arteries (\u003cspan citationid=\"CR68\" class=\"CitationRef\"\u003e68\u003c/span\u003e) (\u003cspan citationid=\"CR72\" class=\"CitationRef\"\u003e72\u003c/span\u003e). Similarly, SP is located primarily in sensory nerves, surrounding ophthalmic and its branches. The perivascular location of SP in the ocular and inner ear blood vessels makes the sustained release of this potent vasoactive neuropeptide by up-regulation of the trigeminovascular system an innovative therapeutic strategy for USH2. Concomitant infusion of AA and administration of ONS might significantly correlated with the clinical outcome of this therapeutic strategy. AA is an efficient inhibitor for both angiotensin-converting enzyme, and neutral endopeptidase enzyme, enzymes that are commonly involved in SP degradation (\u003cspan citationid=\"CR73\" class=\"CitationRef\"\u003e73\u003c/span\u003e) (\u003cspan citationid=\"CR74\" class=\"CitationRef\"\u003e74\u003c/span\u003e) (\u003cspan citationid=\"CR75\" class=\"CitationRef\"\u003e75\u003c/span\u003e) .\u003c/p\u003e\u003cp\u003eThe underlying mechanisms of retinal degeneration and hair cell loss in USH are highly complex, oxidative stress and dysregulated blood flow may be common to retinal degeneration as well as to hair cell damage. Previous studies have demonstrated that an excess of free radical formation and blood reduction in the cochlea occur in noise, drug-induced hearing loss, and senile SNHL (\u003cspan citationid=\"CR76\" class=\"CitationRef\"\u003e76\u003c/span\u003e). Then it is reasonable to suggest that dysregulated CoBF and oxidative stress may be critical factors in triggering hearing loss associated with USH2. Production of oxidative stress via free radical generation has been implicated by a variety of insults that can result in SNHL (\u003cspan citationid=\"CR77\" class=\"CitationRef\"\u003e77\u003c/span\u003e). The role of antioxidants in the prevention of hearing loss has been reported in a number of studies. (\u003cspan citationid=\"CR37\" class=\"CitationRef\"\u003e37\u003c/span\u003e) (\u003cspan citationid=\"CR78\" class=\"CitationRef\"\u003e78\u003c/span\u003e) (\u003cspan citationid=\"CR79\" class=\"CitationRef\"\u003e79\u003c/span\u003e). Interestingly, high doses of vitamin C have been shown to enhance hearing recovery in idiopathic sudden SNHL patients, which suggests that vitamin C reduces levels of reactive oxygen metabolites produced by inner ear ischemia or inflammation. Therefore, it is reasonable to suggest that high-dose of intravenous AA could be beneficial in Usher syndrome.\u003c/p\u003e\u003cp\u003eOn the other hand, photoreceptor cells have a rich complement of mitochondria to accommodate the high metabolic rate and are more sensitive to oxidative stress than other cell types (\u003cspan citationid=\"CR80\" class=\"CitationRef\"\u003e80\u003c/span\u003e). The development of RP, for instance, is tightly linked to oxidative damage (\u003cspan citationid=\"CR81\" class=\"CitationRef\"\u003e81\u003c/span\u003e), and antioxidant therapy can delay cell loss in retinal degenerative diseases and lead to reduced photoreceptor cell death in experimental animals (\u003cspan citationid=\"CR82\" class=\"CitationRef\"\u003e82\u003c/span\u003e) and in clinical trials (\u003cspan citationid=\"CR36\" class=\"CitationRef\"\u003e36\u003c/span\u003e). It has been demonstrated that SP could stimulate the recovery of RPE cells under oxidative stress, possibly by promoting cell proliferation and inhibiting apoptosis through the activation of Akt/GSK-3β signaling (\u003cspan citationid=\"CR56\" class=\"CitationRef\"\u003e56\u003c/span\u003e).\u003c/p\u003e\u003cp\u003eThe interpretation of study findings should be considered within the context of a few limitations. These include a small number of participants, short duration of follow up, lack of information regarding the genotype, electrophysiology of the retina, and objective hearing tests. Nevertheless, this is an open-labeled single-armed intervention and was intended to be an exploratory one. In the future, a proper control group should be considered in a large multicenter double-blind prospective study with stratified randomization\u003c/p\u003e\u003c/div\u003e\u003c/p\u003e"},{"header":"5. Conclusions","content":"\u003cp\u003e \u003cdiv class=\"BlockQuote\"\u003e \u003cp\u003eThis study introduces ONS-based therapy as a novel method for treatment of USH2, a disease that currently has no treatment. It demonstrates the efficacy and safety of ONS-based therapy as a comprehensive treatment for various sense organs involved in this syndrome irrespective of genetic background. The neuroprotective effects of ONS-based therapy are probably mediated by modification of the neural circuit of the ocular and cochlear circulation. Moreover, auditory hair cells and retinal photoreceptor death could be prevented by strengthening endogenous pro-survival mechanisms or by directly blocking cell death via the expression of endogenous SP by noninvasive method of ONS. Additionally, the oxidative damage for hair cells and photoreceptors can be minimized by use of supraphysiological doses of intravenous ascorbate.\u003c/p\u003e \u003c/div\u003e \u003c/p\u003e"},{"header":"6. Patents","content":"\u003cp\u003eIsmail Musallam has US and Israeli patents licensed to himself for which he has waived financial interest\u003c/p\u003e"},{"header":"Abbreviations","content":"\u003cp\u003eThe following abbreviations are used in this manuscript:\u003c/p\u003e\n\u003ctable border=\"0\" cellspacing=\"0\" cellpadding=\"0\" width=\"524\"\u003e\n \u003ctbody\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eAA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eAscorbic acid\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eAch\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eAcetylcholine\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003e\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eAkt\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eProtein kinase B (Akt = PKB)\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eBCVA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eBest corrected visual acuity\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eCGRP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eCalcitonin gene-related peptide\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eChBF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eChoroidal blood flow\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eCoBF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eCochlear blood flow\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eCS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eContrast sensitivity\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eGSK-3 beta\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eGlycogen synthase kinase-3\u0026nbsp;beta\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eLLQ-10\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eLow Luminance Questionnaire-10\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eNA\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eNoradrenaline\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eNK-1R\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eNeurokinin-1 receptor\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eNO\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eNitric oxide\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eNPY\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eNeuropeptide Y\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eONS\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eOphthalmic nerve stimulation\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eOBF\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eOcular blood flow\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003ePPG:\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003ePterygopalatine ganglion\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eRP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eRetinitis pigmentosa\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eRVLM\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eRostro-ventrolateral medulla\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eRPE\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eRetinal pigment Epithelium\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eSNHL\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eSensorineural hearing loss\u0026nbsp;\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eSD-OCT\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eSpectral Domain Optical\u0026nbsp;coherence tomography\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eSP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eSubstance P\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eSCG\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eSuperior cervical ganglion\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eSSN\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eSuperior salivatory nucleus\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eTG\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eTrigeminal ganglion\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eUSH2\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eType 2 Usher syndrome\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003ctr\u003e\n \u003ctd style=\"width: 81px;\"\u003e\n \u003cp\u003eVAP\u003c/p\u003e\n \u003c/td\u003e\n \u003ctd style=\"width: 442px;\"\u003e\n \u003cp\u003eVasoactive intestinal polypeptide\u003c/p\u003e\n \u003c/td\u003e\n \u003c/tr\u003e\n \u003c/tbody\u003e\n\u003c/table\u003e"},{"header":"Declarations","content":"\u003cp\u003e\u003cstrong\u003eAuthor Contributions:\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eI.M. (Musallam I). conceptualized and designed the study. analyzed the data, wrote the manuscript, supervised the project and have read and agreed to the published version of the manuscript.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eFunding:\u003c/strong\u003e\u0026nbsp; none.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eInstitutional Review Board Statement\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThis study was approved by the Institutional Review Board, and it adhered to the tenets of the Declaration of Helsinki. Written informed consent was obtained from all participants or legal guardians after an explanation of the purpose and possible outcome of therapy.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eData Availability\u003c/strong\u003e\u003c/p\u003e\n\u003cp\u003eThe datasets used and/or analyzed during the current study are available from the corresponding author upon reasonable request.\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eConflicts of Interest:\u0026nbsp;\u003c/strong\u003eIsmail Musallam has US and Israeli patents licensed to himself.\u0026nbsp;\u003c/p\u003e\n\u003cp\u003e\u003cstrong\u003eStudy Registration:\u0026nbsp;\u003c/strong\u003eThe study has not been registered in advance\u003c/p\u003e"},{"header":"References","content":"\u003col\u003e\n\u003cli\u003eMathur P, Yang J. Usher syndrome: Hearing loss, retinal degeneration and associated abnormalities. 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Proceedings of the National Academy of Sciences of the United States of America. 2006;103(30):11300-5.\u003c/li\u003e\n\u003c/ol\u003e"}],"fulltextSource":"","fullText":"","funders":[],"hasAdminPriorityOnWorkflow":false,"hasManuscriptDocX":true,"hasOptedInToPreprint":true,"hasPassedJournalQc":"","hasAnyPriority":true,"hideJournal":true,"highlight":"","institution":"Musallam Specialty Hospital ","isAcceptedByJournal":false,"isAuthorSuppliedPdf":false,"isDeskRejected":"","isHiddenFromSearch":false,"isInQc":false,"isInWorkflow":false,"isPdf":false,"isPdfUpToDate":true,"isWithdrawnOrRetracted":false,"journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true},"keywords":"Usher syndrome, retinitis pigmentosa, sensorineural hearing loss, ocular neuromodulation, ophthalmic nerve stimulation, Low luminance questionnaire-10, rod responders, ascorbic acid, substance P","lastPublishedDoi":"10.21203/rs.3.rs-6190348/v1","lastPublishedDoiUrl":"https://doi.org/10.21203/rs.3.rs-6190348/v1","license":{"name":"CC BY 4.0","url":"https://creativecommons.org/licenses/by/4.0/"},"manuscriptAbstract":"\u003cp\u003eReduced dysregulated ocular and cochlear blood flow are proposed as common pathways in the pathogenesis of type 2 Usher syndrome (USH2). The purpose of the study was to evaluate the safety and efficacy of ophthalmic nerve stimulation (ONS) combined with ascorbic acid (AA) in treatment of USH2. Nine participants with USH 2, were enrolled in a prospective interventional case series. All participants were daily treated with ONS sessions and intravenous AA for two weeks. The primary efficiency endpoint was 6 months\u0026rsquo; changes in scotopic vision as measured by a Low Luminance Questionnaire-10 (LLQ-10) with a maximum score of 100 points. Rod responders were defined by \u0026ge;\u0026thinsp;25points increment of LLQ-10 score. The results showed that ONS-based therapy significantly improved scotopic vision by 42.1\u0026thinsp;+\u0026thinsp;11.3 points (\u003cem\u003ep\u0026thinsp;=\u0026thinsp;0.0001\u003c/em\u003e) and 7 (77.8%) of the participants were identified as rod responders. Additionally, clinically significant improvement visual acuity (\u0026ge;\u0026thinsp;0.2 logMAR) and contrast sensitivity (\u0026ge;\u0026thinsp;0.3 log unit) were noticed in 22.2% of the left eyes. Furthermore, a significant improvement of hearing was subjectively reported by one third of the participants. In conclusion, ONS-based therapy significantly improved night vision in patients with USH2. Additionally, a clinically significant improvement of hearing was noticed in one third of patients.\u003c/p\u003e","manuscriptTitle":"Co-targeting Dysregulated Ocular and Cochlear Blood Flow via Ophthalmic Nerve Stimulation for the Treatment of Type 2 Usher Syndrome: Prospective Case Series","msid":"","msnumber":"","nonDraftVersions":[{"code":1,"date":"2025-03-14 10:44:11","doi":"10.21203/rs.3.rs-6190348/v1","editorialEvents":[{"type":"communityComments","content":0}],"status":"published","journal":{"display":true,"email":"[email protected]","identity":"researchsquare","isNatureJournal":false,"hasQc":true,"allowDirectSubmit":true,"externalIdentity":"","sideBox":"","snPcode":"","submissionUrl":"/submission","title":"Research Square","twitterHandle":"researchsquare","acdcEnabled":true,"dfaEnabled":false,"editorialSystem":"","reportingPortfolio":"","inReviewEnabled":false,"inReviewRevisionsEnabled":true}}],"origin":"","ownerIdentity":"5cdf3ac7-697f-4980-af5d-bd473dc82011","owner":[],"postedDate":"March 14th, 2025","published":true,"recentEditorialEvents":[],"rejectedJournal":[],"revision":"","amendment":"","status":"posted","subjectAreas":[{"id":45423876,"name":"Ophthalmology"}],"tags":[],"updatedAt":"2025-03-14T10:44:11+00:00","versionOfRecord":[],"versionCreatedAt":"2025-03-14 10:44:11","video":"","vorDoi":"","vorDoiUrl":"","workflowStages":[]},"version":"v1","identity":"rs-6190348","journalConfig":"researchsquare"},"__N_SSP":true},"page":"/article/[identity]/[[...version]]","query":{"redirect":"/article/rs-6190348","identity":"rs-6190348","version":["v1"]},"buildId":"XKTyCvWXoU3ODBz1xrDgd","isFallback":false,"isExperimentalCompile":false,"dynamicIds":[84888],"gssp":true,"scriptLoader":[]}