{"paper_id":"edc83e3c-b530-45e5-a023-da2a2f4543c6","body_text":"140 \nCell and Organ Transplantology   2019 November; Vol. 7, No. 2\nEndometriosis is a chronic benign hormone-dependent condition \nwhen the endometrial tissue, identical with the endometrium by its mor-\nphological and functional properties, grows outside the borders of the \nuterine mucous membrane. It leads to clinical symptoms able to affect the \nphysical condition, psychological status and social status of the patient \n[1-4]. According to research data endometriosis is diagnosed in 5-10 % \nof the female population. There are approximately 176 million women \nwith endometriosis in the world, mainly of a reproductive age.\nNowadays, certain isolated reports in scientific literature are found \nconcerning proliferation suppression, as well as regression of tumors and \nendometrioid heterotopias under the influence of some dopamine ago-\nnists [5], as well as COX-2 inhibitors [6, 7]. Pathogenetic theory of retro-\ngrade outflow of endometrial cells into the peritoneal cavity is gaining an \nincreasing support [8, 9]. And the peritoneal fluid, namely the state of the \nThe effects of certain angioneogenesis \ninhibitors in experimental  \nendometriosis in rats\nCell and Organ Transplantology. 2019; 7(2):140-147\nDOI: 10.22494/cot.v7i2.100\nBarbe A., Berbets A., Davydenko I., Yuzko V ., Yuzko O.\ne-mail: adryanbarbe@gmail.com \nBukovinian State Medical University, Chernivtsi, Ukraine\nABSTRACT\nThe pathogenetic theory of retrograde outflow of endometrial cells into the peritoneal cavity at endometriosis is gaining an increasing support. \nAdequate blood supply and angioneogenesis play an important role in successful implantation and occurrence of ectopic foci. VEGF plays an \nimportant role in this process. Determination of the pathways affecting the activity of this factor seems to be promising in terms of its effect \nproduced on the early pathogenetic links of endometriosis.\nOBJECTIVES: to determine the effectiveness of dopamine agonist cabergoline and highly selective COX-2 inhibitor celecoxib in an experimental \nmodel of external genital endometriosis in rats.\nMETHODS. 83 outbred white female rats of Rattus Norvegicus Wistar were involved in the experiment. Experimental induction of endometriosis \nwas performed by surgery and implantation of the autologous uterine fragments. The type and volume of experimental endometriosis lesions \non the peritoneum of experimental animals were evaluated macroscopically as well as their hystologic examination were performed. \nRESULTS. Administration of a dopamine receptor agonist as a VEGF inhibitor separately was found to produce a pronounced inhibitory effect \non ectopic endometrioid formation. However, the use of a dopamine receptor agonist in combination with a highly selective COX-2 inhibitor \ndoes not lead to potentiation or summation of their effects. At the same time, the use of COX-2 inhibitor alone has shown significantly lower \nefficacy than using the dopamine receptor agonist as a VEGF inhibitor.\nCONCLUSION. At experimental endometriosis in rats, dopamine agonists and highly selective COX-2 inhibitors were found to be one of VEGF \ninhibitors available.\nKEY WORDS: experimental endometriosis; angioneogenesis; VEGF; dopamine agonist; COX-2 inhibitor; cabergoline; celecoxib\nORIGINAL RESEARCH\npro- and anti-inflammatory cytokines of the peritoneal fluid, plays a key \nrole. The implantation of endometrium cells results from balance disorder \nand excretion of growth factors. Adequate blood supply and neoangio-\ngenesis play an important role in successful implantation and occurrence \nof ectopic foci [10-12]. \nConsidering the above, it can be suggested that one of the promising \nareas of conservative treatment of external genital endometriosis is the \neffect produced on one of the pathogenetic links, namely, inhibition of \nangiogenesis by the use of dopamine agonists and COX-2 inhibitors [13].\nAngiogenesis is a complex process of formation of new blood vessels \nfrom the vessels existed before. This process plays a fundamental role in \nthe reproduction, development and healing of wounds. In adults, endo-\nthelial proliferation is a strictly regulated process based on the balance \nbetween angiogenic and angiostatic factors which are activated in case \n\nwww.transplantology.org \n141 \nCell and Organ Transplantology   2019 November; Vol. 7, No. 2\nof necessity and subsequently completely inhibited when necessary [14, \n15]. Cases of increased endothelial cell proliferation rate are often associ-\nated with tumors and their development [16], which are known to be de-\npendent on angiogenesis for growth and metastasis [17]. The survival of \nendometrioid implants in the abdominal cavity depends on the formation \nof blood supply to provide oxygen and nutrients to the developing lesions. \nPathomorphological examination of endometriosis foci determines their \ndense vascularization [15, 18]. Similar to the process of tumor vascula-\nrization, endometriosis can utilize the mechanisms of angiogenesis and \nvasculogenesis to form its own vascular network to maintain viability.\nIt should be supposed that endometrial fragments separated from the \nuterine endometrium may be the carriers of angiogenic potential. The hu-\nman endometrium, which consists of the functional and basal layer, is a \nunique organ that undergoes proliferation, differentiation and regeneration \nat each menstrual cycle under the regulation of steroid hormones of the \novaries – estrogens and progesterone. In addition to endometrial growth, \nthe vascular bed of the endometrium undergoes proliferation and regen-\neration at every cycle under the influence of ovarian steroids, especially \nestradiol (E2). Shifren et al. [19] identified the increased expression of \nvascular endothelial growth factor (VEGF) mRNA in the functional layer \nof the endometrium in the proliferative and secretory phase of the men-\nstrual cycle, indicating the involvement of angiogenesis for proliferation \nand regeneration. The same study found that E2 is responsible for stimu-\nlating VEGF expression in isolated human endometrial cells, and E2 ad-\nministration increased VEGF mRNA expression compared to endometrial \ncells without E2 stimulus. Endometriosis can be suggested to result from \nimplantation of endometrial fragments into the abdominal cavity with sub-\nsequent activation of cellular aggression and proliferation mechanisms.\nVEGF appears to play an important role in supporting angiogenesis \nat endometriosis. It is a vasoactive substance which is involved in vari-\nous normal physiological processes, including wound healing and endo-\nmetrial revascularization, mediating endothelial proliferation and migration. \nIn oncogenesis VEGF concentration usually correlates with increased blood \nsupply in various tissue types that are associated with the tumor [20]. In \nnormal endometrium, elevation of VEGF mRNA levels and expression of \nthe corresponding proteins can be caused by hypoxia [21]. No wonder that \nthe peritoneal fluid of women with the last stages of endometriosis con-\ntains higher VEGF concentrations than that of women with initial stages of \nendometriosis or healthy women [22]. Various sources of VEGF have been \nidentified, including endometrioid lesions [23] and peritoneal fluid macro-\nphages that increase VEGF expression when treated with ovarian steroids \nsuch as E2 and progesterone [24], transforming the notion that VEGF is \ninvolved in angiogenesis associated with endometrial lesions [25].\nVEGF, also known as VEGF-A, is a secretory cytokine structurally  \nrelated to platelet-derived growth factor (PDGF), which mediates physio-\nlogical and tumor angiogenesis [26-29]. In mice a complete damage of \nthis factor is lethal and causes severe cardiovascular abnormalities [30, \n31]. In vitro it promotes endothelial cell proliferation, migration, and vas-\ncular tube formation [32]. In various in vivo models it induces a strong \nangiogenic response [33]. At adulthood VEGF is involved in wound hea-\nling, menstruation, pregnancy and blood pressure support [29]. During \nphysiological angiogenesis, it is produced by various cell types, including \nneutrophils, platelets, and macrophages [29]. \nCurrently there are the following ways of VEGF inhibition: (1) neutra-\nlizing monoclonal antibodies against VEGF and VEGFR; (2) small-mole-\ncule VEGF receptor tyrosine kinase inhibitors; (3) soluble VEGF receptors \n(VEGF-Trap) and (4) ribozymes [34]. In addition to the above mentioned \nVEGF inhibition pathways which are widely discussed in clinical onco-\nlogy, there is the evidence of possibile inhibition of VEGF-induced an-\ngiogenesis by using dopamine neurotransmitter [35]. Bacic et al [36] \nreported that dopaminergic receptors are linked to adenylyl cyclase in the \nhuman cerebral microvascular endothelium. Dopamine is a catecholamic \nneurotransmitter involved in pathogenesis of both Parkinson’s disease \nand development of schizophrenia [37-40]. Dopamine and its derivative \nmolecules have shown inhibitory potential in several types of malignant \ntumors in mice, and its effects have been explained by inhibition of tumor \ncell proliferation, stimulation of immunity etc. [41-44]. Recent studies \ndemonstrate the presence of D2 dopamine receptors on endothelial cells \n[45, 46], which may affect angiogenesis. It is this mechanism that is likely \nto be important in inhibiting tumorigenesis [47, 48], as VEGF is the most \nimportant angiogenic cytokine in tumors and in other types of pathologi-\ncal angiogenesis [49, 50]. \nVEGF is believed to induce angiogenesis by involving VEGF receptor \ntype 2 (VEGFR-2), which leads to its phosphorylation and downstream \nsignaling events series [51]. 1 μM dopamine was found to inhibit VEGF-\ninduced phosphorylation of VEGFR-2 in vitro. D2 bromocriptine and \nquinpyrrole agonists similarly inhibit VEGF-induced phosphorylation of \nVEGFR-2 [52-54]. Dopamine is indicated to stimulate internalization of \nVEGFR-2 surface, probably by endocytosis, leaving less VEGFR-2 on the \nsurface to interact with VEGF [52-54]. Thus, the inhibitory effect of do-\npamine on VEGF-induced angiogenesis is explained by its action at the \nearly signaling stage. It is important to note that dopamine inhibits an-\ngiogenesis mediated by VEGF, but probably does not affect angiogenesis \nmediated by other mechanisms [54]. \nVEGF is not the only way to activate angiogenesis. There are alterna-\ntive activation pathways, such as through COX-2. COX-2 converts arachi-\ndonic acid into prostaglandin H2, which can be enzymatically converted \ninto five major prostanoids: prostaglandin E2, prostaglandin D2, prosta-\nglandin I2 (prostacyclin), prostaglandin F2a and thromboxane. COX-2 and \nprostaglandin E2 have been previously demonstrated to induce angioge-\nnesis, but most studies indicate an indirect role by means of stimulating \nother proangiogenic growth factors [55]. In addition to indirect effects, \nhypothetically, the COX-2 / PGE2 pathway may play a direct role in stimu-\nlating angiogenesis through VEGF-independent paracrine mechanisms \nthat directly affect endothelial cells [56]. Certain studies have shown that \nCOX-2 regulates VEGF-induced angiogenesis, and also that VEGF con-\ntrols COX-2-induced angiogenesis, indicating reciprocity between these \npathways [57, 58]. The ability of COX-2/PGE2 pathway to induce VEGF-\nindependent angiogenesis suggests that this pathway may promote an-\ngiogenesis when VEGF pathway is blocked. \nMoreover, scientific literature contains certain information concer-\nning suppression of VEGF-induced phosphorylation of type 2 VEGF re-\nceptor by dopamine [59-61], occurring due to phosphorylation of tyrosine \nphosphatase 2 [62], and therefore, its inactivation. \nOBJECTIVE OF THE STUDY: to determine the effectiveness of dopa-\nmine agonist cabergoline and highly selective COX-2 inhibitor celecoxib in \nan experimental model of external genital endometriosis in rats.\nThe experiment involved 83 outbred, sexually mature, nulliparous \nwhite Rattus Norvegicus Wistar rats weighing 170-200 g, 14-17 weeks of \nage. All the manipulations on animals were carried out in accordance with \nthe recommendations of the UNESCO Intergovernmental Committee on \nBioethics (IGBC). All the procedures and experiments of this study respect \nthe ethical standards in the Helsinki Declaration of 1975, as revised in \n2008 [5], as well as the national law. All institutional and national guide-\nlines for the care and use of laboratory animals were followed.\nAt the end of the adaptation period, starting from the first day of the \nexperiment, rats were injected subcutaneously in the back with 0.06 mg/kg \nbody weight of estradiol valerate. Estradiol valerate injection at the dose \nof 0.06 mg/kg was repeated on the third day of the experiment.\nOn the fourth day of the experiment, experimental induction of en-\ndometriosis was performed by means of surgery and implantation of an \nautologous uterine fragment using the method described by A. Golan et \nal. [63] and T. Hirata et al. [64]. After awakening from ketamine-xylazine \nintraperitoneal anesthesia, rats were randomly and evenly divided into \ngroups. Rats of all the groups were treated with estrogenic hormonal \nMATERIAL AND METHODS\n\n142 \nCell and Organ Transplantology   2019 November; Vol. 7, No. 2\nsupport in the form of daily subcutaneous injections of 0.03 mg/kg body \nweight of estradiol valerate until the last day of the experiment.\nIn the first experimental group comprising 21 rats, dopamine recep-\ntors agonist cabergoline (ATC: G02CB03; Dostinex ® , Pfizer Italia S.r.l.) \nwas used as a subcutaneous injection in the dose of 0.075 mg/kg body \nweight daily. In the second experimental group including 19 rats, a do-\npamine receptor agonist and a COX-2 inhibitor were administered as  \na single subcutaneous injection of cabergoline  in the dose of 0.075 mg/kg  \nof animal body weight and celecoxib (АТХ: L01XX33, M01AH01; Cele-\nbrex\n® , Pfizer Inc.) in the dose of 30 mg/kg of animal body weight daily. In \nthe third experimental group of 20 rats, the COX-2 inhibitor celecoxib was \nadministered as a subcutaneous injection in the dose of 30 mg/kg of ani-\nmal body weight daily. In the fourth, the control group, which comprised \n20 rats, nothing more than estrogen support has been applied.\nIn the first, second and third experimental groups, drugs were admi-\nnistered from the 12\nth  day of the experiment (from the 8 th  day after sur-\ngery) to the end of the experiment (the 25 th  day of the experiment, the \n21 st  day after surgery). On the 26 th  day of the experiment (22 nd  day after \nsurgery) euthanasia was performed by decapitation using thiopental an-\nesthesia in the dose of 5.7 mg/kg of the body weight.\nThe presence and type of lesions were evaluated macroscopically: \ncysts filled with dark fluid, cysts filled with light fluid, lesions in the form \nof solid tissue, or the absence of macroscopic signs of lesions.\nUsing the office transparent millimeter ruler the smallest and largest \ndiameters of lesions were measured. The following formula was used to \ndetermine the volume of lesions: where d\n1  and d 2  – are the smallest and \nlargest diameters of lesion respectively. \nORIGINAL RESEARCH\nSampling of the material for morphological studies was performed \naccording to standard requirements for making histological preparations. \nThe tissue fragments after washing in distilled water were fixed in 10-12 % \nsolution of neutral formalin, after which they were transferred to 3-5 % \nsolution of neutral formalin, where they were stored. The preparations \nwere dehydrated by sequentially running the objects through ethanol of \nincreasing concentration. \nSubsequently, paraffin embedded blocks were made and 3 to 5 μm \nthick histologic sections were perfomed using a microtome. To obtain \ndifferentiated polychromy, the tissue was stained with hematoxylin-eosin. \nHistological sections were examined, analyzed and photographed using a \ncomputer-based image analysis system consisting of an CX-21 light mi-\ncroscope (Olympus, Japan) and C450 digital camera (Olympus, Japan). \nEstimation of optical density and the area calculations of microscopic \nstructures was performed using free-licensed software ImageJ, ver. 1.50b \n(NIH, USA). The optical density of cytoplasm and nuclei of secretory epi-\nthelial cells was estimated using 0-255 RGB gradation (0 – black color, \n255 – white color).\nThe results were statistically processed by MedCalc 15.8 software \n(MedCalc Software bvba, Belgium) using Mann-Whitney U-test and de-\nscriptive statistics analysis. Data were presented as mean +/- standard \ndeviation (SD). Graph data were presented as clustered bar chart and box \nand whisker charts.\nFig. 1. Macroscopic view of ectopic uterine tissue on the peritoneum when using cabergoline only ( A), celecoxib (B) or combination of cabergoline and \ncelecoxib (C) compared to control group (D). Arrows indicate implant growth sites.\nD\nB\nC\nА\n\nwww.transplantology.org \n143 \nCell and Organ Transplantology   2019 November; Vol. 7, No. 2\nMost current theories of endometriosis are based on the theory of \nSampson’s retrograde menstruation. However, an open question remains \nwhy retrograde menstruation occurs in 90 % of women of reproductive \nage, while the incidence of endometriosis is several times less. Among \nthe contributing factors for the implantation of eutopic endometrium to \nthe peritoneum, we have identified VEGF as one of the major contributors \nto the growth of heterotopias. Dopamine agonists and highly selective \nCOX-2 inhibitors have been identified as one of the available VEGF inhibi-\ntors. In order to determine the effectiveness of their use, we created an \nexperimental model of endometriosis in rats. \nMacroscopic examination of endometrioid foci was performed imme-\ndiately after euthanasia, after opening the abdominal cavity of a rat using \nU-shaped incision. Considering the fact that the only reliable method of \ndiagnosis of endometriosis is visual identification of foci during laparo-\nscopy with subsequent histological verification, it is advisable to visualize \nand perform morphometry in an experimental model of endometriosis. \nBased on morphometry, data the volume of lesions was determined, \nwhich is another parameter of objectification when conducting statistical \nresearch. Due to the fact that two flaps of their own uterine horn were \ntransplanted to each animal, the number of observed cases was doubled \nin relation to the number of animals in this group.\nThe morphological data we have obtained coincide with those of \nother researchers who used a similar technique. Thus, Elgamal et al. [65] \ndescribes ectopic endometrioid lesions macroscopically in the form of \ncysts filled with fluid, and the histological picture obtained by us com-\npletely corresponds to the described one by Rezende et al. [66], Neto et \nal. [67] and Amaral et al. [68] (Fig. 1).\nWhile comparing the types of lesions (Fig. 2), we determined that the \nleast cystic endometrioid formation occurred when cabergoline was used. \nThe use of celecoxib alone and in combination with cabergoline showed \nless effectiveness concerning this parameter.\nWhile comparing the volume of lesions (Fig. 3), we found that the best \nresults were obtained when cabergoline was used separately. The volume \nof lesions after cabergoline administration is almost 9 times smaller than \nthat in the control group. It should be noted that the volume of lesions after \nadministration of cabergoline + celecoxib combination is almost 3.5 times \nsmaller than that in the control. When celecoxib is used separately, there is \nalso a significant difference in the volume of lesions (celecoxib reduces the \nsize of lesions), but less pronounced (slightly more than 2 times).\nOne of the indirect signs of vascular and capillary proliferation, as \nwell as a sign of successful implantation of ectopic endometrium, is the \ngrowth of the glandular epithelium and its secretory activity.\nDuring microscopic examination of endometrioid lesions, we ob-\nserved glandular secreting epithelium oriented by the secretory pole to-\nwards the cystic cavity. The stroma of the glands is found lower, which \nrespectively lie on the myometrial basis (which was preserved during \ntransplantation). We observed the viability and functional activity of the \nglandular epithelium of ectopic transplants (Fig. 4).\nSeveral morphometric parameters were used to objectify microscop-\nic examination and functional activity of the cells. They include determi-\nnation of the glandular epithelium cellular height, determination of the \nheight of the secretory part of glandular epithelium cells, determination \nof the nucleus area, carrying out densitometry of the nucleus and cells in \ngeneral. \nStatistical processing and analysis of morphometric examination of \nhistological specimens resulted in the following. The height of secretory \nendometrioid epitheliocytes was significantly lower (p < 0.0001) only when \ncabergoline was used. No significant difference was found in case of caber-\ngoline and celecoxib combination, and with celecoxib alone (Fig. 5).\nThe following regularity was found after examination of a secretory \npart of epitheliocytes: a reliable decrease was observed only with the use \nof cabergoline. No significant difference was found in the use of cele-\nRESULTS AND DISCUSSION\nFig. 3. Average volume of experimental endometriosis lesions on the \nperitoneum in experimental groups, mm 3 . \nNote: * – p < 0.0001 compared to control group;  \n▼▲ – extreme outliers.\nFig. 2. Types of experimental endometriosis lesions on the peritoneum \nin experimental groups.\n700\n600\n500\n400\n300\n200\n100\n0\nCabergolin*\nCabergolin+Celecoxib\nCelecoxib\nControl group\nNo cystic structures Cysts with dark contentsCysts with light contents\nTypes of lesions\n2 20\n20\n20\n29\n30 40\n10\n10\n10\n7\n8 12\n4\n0\n18\n18\nCabergolin\nCelecoxib* Cabergolin & Celecoxib Control Cabergolin* Celecoxib* Cabergolin & Celecoxib Control\n90\n80\n70\n60\n50\n40\n30\n0\n10\n20\nCabergolin* Celecoxib* Cabergolin & Celecoxib Control\n140\n120\n60\n60\n100\n40\n0\n20\n700\n600\n500\n400\n300\n200\n100\n0\nCabergolin*\nCabergolin+Celecoxib\nCelecoxib\nControl group\nвідсутність кіст кісти з темним вмістомкісти з світлим вмістом\nТипи уражень\n2 20\n20\n20\n29\n30 40\n10\n10\n10\n7\n8 12\n4\n0\n18\n18\nCabergolin\nCelecoxib* Cabergolin & Celecoxib* Control Cabergolin* Celecoxib* Cabergolin & Celecoxib* Control\n90\n80\n70\n60\n50\n40\n30\n0\n10\n20\nCabergolin* Celecoxib*** Cabergolin & Celecoxib*** Control\n140\n120\n60\n60\n100\n40\n0\n20\ncoxib, and in case of cabergoline and celecoxib combination, a significant \nincrease in the secretory portion of epitheliocytes was detected.\nThe determined area of the epitheliocyte nucleus (Fig. 6) was sig-\nnificantly smaller in all the experimental groups (without a significant \ndifference between the experimental groups) compared to the control. \nMorphometry of samples from the control group of animals revealed  \na smaller nucleus compared to that of the eutopic endometrium.\nA relative densitometric density of epitheliocyte nuclei was signifi-\ncantly (p < 0.05) higher (indicating less compacting of nuclei) in all the \nexperimental groups compared to that of the control one. However, in \nfact, it was equal to a relative nucleus density of eutopic epitheliocytes. \nDetermination of the relative density of epitheliocytes did not reveal  \na significant difference (p > 0.05) between the experimental groups and \nthe control group.\nAccording to current literature data, we suggest that the effectiveness \n(according to the above mentioned indices) of cabergoline administration \nin experimental endometriosis compared with that of the control group \nis caused by its direct effect on dopamine D2 receptors. In its turn, it \nactivates phosphorylation of VEGF receptors and their internalization via \nthe adenylate cyclase pathway, which directly blocks the action of VEGF \nfurther leading to inhibition of angiogenesis. Due to inability to form new \nvessels, with nutrient and oxygen deficiency, the ectopic endometrioid \n\n144 \nCell and Organ Transplantology   2019 November; Vol. 7, No. 2\nORIGINAL RESEARCH\nFig. 4. Microscopic views of ectopic uterine tissue on the peritoneum in rats when using cabergoline ( A), celecoxib (B) or combination of cabergoline \nand celecoxib (C) compared to control group (D). Arrow indicates secretory epithelium. Hematoxylin and eosin stain. Ob. 10х (A, C) and 40х (B, D),  \nOc. 10х.\nD\nB\nC\nА\nFig. 5. Height of epitheliocytes in ectopic endometrium on experimental \nendometriosis, μm. \nNotes: * – p < 0.0001 compared to control group; *** – p > 0.05 \ncompared to control group; ▲ – extreme outliers.\nFig. 6. Nucleus area of epitheliocytes in ectopic endometrium on \nexperimental endometriosis, μm\n2 . \nNote: * – p < 0.0001 compared to control group; ▲ – extreme outliers\n700\n600\n500\n400\n300\n200\n100\n0\nCabergolin*\nCabergolin+Celecoxib\nCelecoxib\nControl group\nвідсутність кіст кісти з темним вмістомкісти з світлим вмістом\nТипи уражень\n2 20\n20\n20\n29\n30 40\n10\n10\n10\n7\n8 12\n4\n0\n18\n18\nCabergolin\nCelecoxib* Cabergolin & Celecoxib* Control Cabergolin* Celecoxib* Cabergolin & Celecoxib* Control\n90\n80\n70\n60\n50\n40\n30\n0\n10\n20\nCabergolin* Celecoxib*** Cabergolin & Celecoxib*** Control\n140\n120\n60\n60\n100\n40\n0\n20\n700\n600\n500\n400\n300\n200\n100\n0\nCabergolin*\nCabergolin+Celecoxib\nCelecoxib\nControl group\nвідсутність кіст кісти з темним вмістомкісти з світлим вмістом\nТипи уражень\n2 20\n20\n20\n29\n30 40\n10\n10\n10\n7\n8 12\n4\n0\n18\n18\nCabergolin\nCelecoxib* Cabergolin & Celecoxib* Control Cabergolin* Celecoxib* Cabergolin & Celecoxib* Control\n90\n80\n70\n60\n50\n40\n30\n0\n10\n20\nCabergolin* Celecoxib*** Cabergolin & Celecoxib*** Control\n140\n120\n60\n60\n100\n40\n0\n20\n\nwww.transplantology.org \n145 \nCell and Organ Transplantology   2019 November; Vol. 7, No. 2\nimplant has no potential for further development and secretory activity. \nThis is confirmed by the data indicating decrease in epitheliocytes and \ntheir secretory poles in size, as well as the compacting of the epitheliocyte \nnuclei.\nAccording to the theoretical data reported in the relevant publications, \nCOX-2 inhibitors potentially produce an inhibitory effect on angiogenesis \nby deactivating PGE2 formation, which, in the opinion of many authors, \nis responsible for an alternative (non-VEGF) pathway of angiogenesis. \nHowever, as indicated by our study, the use of celecoxib in experimen-\ntal endometriosis in rats did not produce the expected efficacy. Despite  \na significant reduction of lesions in size (although less significant than \nafter cabergoline use), the size of epitheliocytes, their secretory part re-\nmained unchanged. Moreover, endometrioid lesions predominate in this \ngroup in the form of cystic structures, including almost 1/3 of them with \na dark content confirming their activity. Apparently blocking non-VEGF \npathways of angiogenesis is insufficient to suppress the growth of endo-\nmetrioid lesions in the experiment. A similar statement is found in Xu et \nal. [69], where the authors did not obtain the expected results when using \nhighly selective COX-2 inhibitors to inhibit tumor angiogenesis. Obvious-\nly, a possible role and time-related effects of the use of highly selective \nCOX-2 inhibitors in endometriosis should be investigated.\nA complex administration of cabergoline, a dopamine receptor ago-\nnist, and celecoxib, a highly selective COX-2 inhibitor, similar to that of \nthe previous group (using celecoxib alone) did not produce the expected \nefficacy, despite a noticeable efficacy of cabergoline alone. Although an \naverage volume of lesions is significantly smaller than that in the control \ngroup, the size of epitheliocytes did not decrease significantly compared \nto the control, and their secretory part is even larger than in the control \ngroup. After administration of both cabergoline in combination with ce-\nlecoxib, and celecoxib alone, vacuolation of epitheliocytes was observed, \nas it is evidenced by an increased rate of relative cell density. At the same \ntime, compaction of epitheliocyte nuclei was observed. These data in-\ndicate insufficient inhibition of endometrial cell functional activity and, \ntherefore, insufficient regression of endometrioid ectopic foci.\nDespite a theoretically expected synergistic effect of cabergoline and \ncelecoxib, the results of our study are indicative of its lack. Similar conclu-\nsions were also drawn by Xu et al. (2015) [69]. In this study, bevacizumab \n(a direct specific VEGF blocker) in combination with celecoxib was shown \nto have much less effectiveness in blocking tumor angiogenesis than in \na single administration. At the same time, in a separate administration, \ncelecoxib showed less efficacy in blocking angiogenesis in tumors than \nbevacizumab. \nTherefore, the use of a dopamine receptor agonist as a VEGF inhibi-\ntor in combination with a highly selective COX-2 inhibitor does not lead \nto potentiation or summation of their effects. 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Sci Transl Med. 2015; 6 (242):1-12. http://doi.org/10.1126/scitranslmed.3008455.\nThe authors declared no potential conflicts of interest with respect to the \nresearch, authorship, and/or publication of this article.\nReceived: October 30, 2019\nAccepted: November 30, 2019\nARTICLE ON THE SITE  \nTRANSPLANTOLOGY.ORG","source_license":"CC0","license_restricted":false}