Regulation of aquaporin-2 using traditional Chinese medicine in water balance disorders: a literature review.

Aquaporins (AQPs) are a family of membrane proteins that are specifically responsible for facilitating water transport across cellular membranes, with some subtypes permitting the passage of small molecules such as glycerol and urea ( 1 ). To date, at least 13 AQPs (AQP0–AQP12) have been identified in mammals. These proteins are distributed across various organs and tissues, significantly enhancing water permeability in the cellular membrane and performing critical physiological functions through their involvement in intracellular and extracellular water balance regulation ( 2 ). Among these, aquaporin-2 (AQP2) is predominantly localized in the apical plasma membrane and intracellular vesicles of renal collecting duct cells. AQP2 is uniquely responsive to arginine vasopressin (AVP), facilitating water transport in and out of renal cells, thereby enhancing water reabsorption from urine ( 3 ). Due to its critical role in renal water balance regulation, AQP2 dysfunction is closely associated with various kidney pathologies. Recent studies have revealed that, beyond renal function, AQP2 is also involved in water metabolism disorders related to other organ systems, notably the cardiovascular system, liver, inner ear, and uterus ( 4 ). In particular, emerging evidence highlights the distinctive role of AQP2 in cardiovascular diseases through its modulation of fluid balance and blood pressure regulation, underscoring the importance of exploring its potential as a therapeutic target. Traditional Chinese medicine (TCM) has historically provided comprehensive diagnostic and therapeutic approaches to managing conditions arising from abnormal water metabolism, such as pathological states involving water dampness and phlegm fluid retention ( 5 ). Clinical practice demonstrates that TCM interventions frequently yield substantial therapeutic effects when used to treat these disorders. At the molecular level, the function of AQP2 is primarily regulated through the vasopressin type 2 receptor (V2R) signaling pathway. In this classical pathway, AVP binds to V2R located on the basolateral membrane of renal collecting duct principal cells, leading to the activation of adenylate cyclase (AC) via Gs protein signaling ( 6 ). This activation increases intracellular cyclic adenosine monophosphate (cAMP) levels, which subsequently activate protein kinase A (PKA). Activated PKA then promotes the phosphorylation and trafficking of AQP2-bearing vesicles to the apical membrane, thereby increasing water reabsorption by enhancing membrane water permeability ( 6 ). At present, TCM research is increasingly focused on this signaling pathway, providing novel insights and therapeutic targets for managing water metabolism disorders. This review aims to systematically summarize the current research regarding TCM interventions that modulate AQP2 expression and activity in diseases related to water metabolism. Moreover, we aim to elucidate the underlying molecular mechanisms, thereby providing robust theoretical support and guidance for future clinical applications of TCM in treating water metabolism disorders.

In addition to modulating AQP2 expression to regulate water metabolism, TCM can also influence the pharmacokinetics and pharmacodynamics of conventional cardiovascular drugs, including diuretics and antihypertensive agents. Several TCM formulations and single herbs possess active compounds that interact with key regulatory pathways, such as the renin-angiotensin-aldosterone system ( 61 ) and the sympathetic nervous system, both of which are critical in blood pressure regulation and fluid balance. For instance, herbs such as Ephedra and Glycyrrhiza have been shown to affect not only water reabsorption mechanisms via AQP modulation but also the activity of cytochrome P450 enzymes involved in drug metabolism ( 62 ). Such interactions alter the bioavailability or efficacy of diuretics and hypertensive medications, potentially leading to synergistic effects. When TCMs are used in conjunction with diuretics, by targeting different points (such as AQP2 expression and ion channels/pumps), they can enhance the excretion of water and electrolytes, potentially increasing the diuretic effect and aiding in the alleviation of edema or hypertension caused by sodium and water retention ( 63 – 65 ). However, combined use, if not properly matched in dose or formula, can increase risks such as electrolyte disturbances and dehydration, necessitating close monitoring of patient electrolyte levels. However, some TCMs that have tonic effects or regulate kidney yang may promote water reabsorption under certain stages or specific pathogenic conditions, which could theoretically reduce the efficacy of diuretics if combined inappropriately. Thus, a comprehensive consideration of patient diagnosis and pharmacological properties is required. Moreover, by modulating neurohormonal pathways, TCM enhances the therapeutic outcomes of conventional therapies or reduces their side effects, offering a complementary approach in managing complex conditions such as heart failure and hypertension ( 66 ). Given these possibilities, further research is warranted to systematically explore the molecular mechanisms behind these interactions, which may ultimately facilitate the rational design of integrated treatment regimens that combine TCM with standard cardiovascular drugs for optimized patient care.

Relevant studies were identified through systematic searches of multiple databases, including PubMed, Web of Science, Embase, China National Knowledge Infrastructure (CNKI), and Wanfang. The search terms used were combinations of “Aquaporin-2,” “AQP2,” “Traditional Chinese Medicine,” “TCM,” “water metabolism disorders,” “renal diseases,” “cardiovascular diseases,” “edema,” and the names of specific Chinese medicinal formulas and herbs known to regulate water metabolism. Articles published within the past 20 years were selected based on their relevance to the regulatory effects and mechanisms of TCM on AQP2 expression and activity. In addition, references from key articles were reviewed to ensure comprehensive coverage of the relevant literature. Most of the included studies comprised in vivo animal models (primarily rats and mice) and explored the regulatory effects of TCM formulas or compounds on AQP2 and related aquaporins in pathological conditions such as heart failure, renal dysfunction, and edema.

Water and sodium metabolism disorders in the kidneys are the leading mechanisms of salt-sensitive hypertension. Wulingsan has a great effect when used to treat hypertension resulting from water metabolic disorders in the kidneys. Researchers have shown that Wulingsan can significantly relieve water reabsorption by decreasing the expression of AQP2 protein and AVP-V2R in kidney tissue from rats with hypertension, preliminarily illustrating the hypotensive mechanism of Wulingsan on a molecular level ( 31 ). Containing Polyporus umbellatus , Poria cocos , and Rhizoma alismatis , Wulingsan has a similar regulation mechanism to Zhuling decoction, proving the reliability of the study ( 32 ). Scholars have demonstrated that acupuncture with Zusanli (ST36) and Quchi (LI11), combined with acupoint application on Pishu (BL20), regulates the abnormal expression of AQP1 and AQP2 in the kidneys, thus correcting the water metabolism disorder to decrease blood pressure and control the occurrence and progression of hypertension ( 33 ). Studies on AQP1 and AQP2 will provide new ideas for elucidating the pathogenesis of salt-sensitive hypertension. Acupuncture combined with acupoint application can modulate the expression of AQP1 and AQP2, which may be one of the mechanisms by which acupuncture lowers blood pressure, with great significance for the prevention and treatment of hypertension. Vasopressin is considered the main mechanism of water retention in chronic heart failure (CHF) ( 34 ). During heart failure, the sensitivity of the atrial stretch receptors decreases, giving rise to a weakened inhibition effect on AVP, which increases the concentration of AVP in circulating blood. AQP2 is then transferred from the cytoplasm to the membrane through the AVP-V2R-AQP2 pathway, resulting in water retention in the body by increasing water reabsorption ( 35 ). The water retention resulting from the abnormal increase in AQP2 is a fundamental cause of the poor prognosis of CHF. Therefore, AQP2 has attracted attention as an emerging target in research into water metabolism imbalance and has become an entry point for the treatment of cardiac edema ( Table 1 ). Regulation of aquaporin expression by traditional Chinese medicine in cardiovascular diseases. NF-κB, nuclear factor kappa B; IL-6, interleukin 6; TNF-α, tumor necrosis factor alpha; CHF, chronic heart failure; CHD, coronary heart disease. There are two regulatory mechanisms of AQP2, namely short-term and long-term regulation. According to the differences in their principles, scholars have carried out studies on the effects of drugs on AQP2. It has been reported that the Yiqi Huoxue formula inhibits the increase in AQP2 protein expression induced by long-term continuous elevated AVP levels, in which cAMP/exchange protein is directly activated by the cyclic adenosine monophosphate (Epac)/extracellular signal-regulated protein kinase (ERK) signaling pathway involved in long-term regulation ( 47 ). Experiments have shown that the short-term application of water-draining medicinal herbs [ Poria cocos , plantain seeds (Cheqianzi), and pepperweed seeds (Tinglizi)] decreases AQP2 expression in the urine of rats with heart failure, and the conclusion is consistent with previous studies ( 48 ). However, the effect of blood-activating medicinal herbs [ Radix salviae miltiorrhizae (Danshen), Carthamus tinctorius (Honghua), and Radix paeoniae rubra (Chishao)], qi-tonifying medicinal herbs [ Radix ginseng rubra (Renshen) and Radix astragali (Huangqi)], and interior-warming medicinal herbs [ Cassia twig (Guizhi) and Aconite (Fuzi)] was not obvious under the same conditions. With increasing administration time, the effects of the latter three sorts of drugs on AQP2 continuously increased, while the impact of water-draining medicinal herbs did not change significantly. R. salviae miltiorrhizae is associated with a reduction of AQP2, leading to a redistribution of AQP2 in renal collecting ducts ( 49 ). Moreover, the therapeutic effect of R. astragali on heart failure was partly related to an improvement in the AVP system and the abnormal expression of AQP2 in rats. The difference in the results may be related to the short-term and long-term regulatory mechanisms of different drugs, which are worthy of further study. In addition, the Bushen Huoxue recipe was found to decrease the level of BNP in serum and AQP2 expression in renal tissue of CHF model rats, and its regulatory effect was similar to captopril tablets ( 38 ). Therefore, it can be inferred that the Bushen Huoxue recipe may be used to treat CHF by lessening the expression of AQP2 in the kidneys, but this needs to be confirmed by further experimental data ( Figure 1 ). Mechanisms by which traditional Chinese medicine regulates AQP2 in water metabolism disorders. AQP2-mediated regulation of renal water balance is intimately linked to the development and progression of heart failure. Angiotensin II (Ang II) engages AT1 receptors to trigger the ENaC/NHE3 pathway, leading to VP release and increased aldosterone production. These hormonal signals synergistically elevate AQP2 expression in renal collecting duct cells, resulting in fluid retention and hyponatremia that worsen heart failure symptoms. In contrast, the serine protease Corin converts pro-ANP into its active form, ANP, which suppresses AQP2 expression, and ultimately improving the clinical manifestations of heart failure. Ascites is one of the complications of liver cirrhosis and hepatocellular carcinoma. In decompensated cirrhosis, increased portal venous pressure leads to renal insufficiency and water retention, resulting in ascites that seriously affects the patient's quality of life ( 50 ). AVP and AQP2 are thought to play an essential role in ascites formation. It has been proven that when the expression of AQP2 and V2R is downregulated, liver ascites in mice with hepatocellular carcinoma is significantly alleviated and ascites symptoms are relieved. Danggui Shaoyao San may decrease the expression of AQP2 to treat ascites by inhibiting the release of AVP and the binding of AVP and V2R ( 51 ). Scholars found that the protein expression levels of AQP2 and V2R were considerably increased in a hepatocellular carcinoma rat model with ascites after 12 days of ascites inoculation, while Euphorbia pekinensis (Daji) decoction effectively moderated abnormally elevated AQP2 and V2R expression levels ( 52 ). The inhibitory effect of E. pekinensis was further enhanced when combined with twice the amount of Glycyrrhiza (Gancao). Preliminary mechanism exploration verified that Glycyrrhiza combined with E. pekinensis , when used to treat ascites in hepatocellular carcinoma, may regulate renal AQP2 and V2R ( 53 ). Researchers have found that 3-O-(2'E,4'Z-decadienoyl)-20-O-acetylingenol (3-O-EZ), a major diterpenoid of EK, exhibits a specific therapeutic effect on ascites in mice with hepatoma ( 54 ). In conclusion, AQP2 inhibition may be a therapeutic strategy for treating ascites in hepatoma. In a study on water metabolism-related diseases of the inner ear, all the downstream factors of AVP in the AVP-V2R-AQP2 system that regulate AQP2 expression were found to be present in the inner ear. The AQP2 protein is mainly distributed in the stria vascularis, Corti's organ, and spiral ganglion cells ( 55 ). Previous animal experiments showed that the classical Linggui Zhu Gan decoction and Zexie decoction formulae lower the degree of endolymphatic hydrops induced by intraperitoneal injections of desmopressin acetate in guinea pigs ( 56 ). One of the mechanisms may be the downregulation of AQP2 expression in the vestibular membrane. Electroacupuncture may modulate the AVP-V2R-AQP2 system and lower the expression of AQP2 in the whole body, which in turn decreases the opening of water channels to weaken endolymphatic hydrops symptoms. However, electroacupuncture at GV20 can reverse the decrease in V2R expression caused by AVP ( 57 ). Whether the effect of electroacupuncture on V2R expression is directly caused by electroacupuncture or due to the decrease in plasma AVP still needs to be studied further. AQP2 expression is periodic as it is upregulated in the endometrial epithelium during the secretory phase and plays a vital role in the periodic changes of the endometrium ( 58 ). Shaofu zhuyu decoction was found to decrease the AQP1 and AQP5 mRNA levels and upregulate the AQP2mRNA expression level in the abdominal fluid of rats with endometriosis (EM), thus affecting the disease course ( 59 ). In this case, the drug works by restraining the promotion of AQP2 to endometrial debris infiltration, which may be a function of AQP2. AQP2 expression in the vaginal epithelium at different stages of fertility may play a role in regulating the production and quantity of vaginal secretions. Clinical research indicates that the use of TCM treatment after cervical columnar epithelium ectopic physical therapy can alleviate an increase in postoperative vaginal secretions and abnormal water drainage and shorten the vaginal drainage duration ( 60 ). Moreover, scholars found that AQP2 expression diminished simultaneously after TCM treatment, indicating that there was an apparent connection between the decreasing expression of AQP2 and the decrease in vagina drainage.

Studies have demonstrated that a single Chinese herb can also be effective in treating water metabolism-related diseases. Urine production is regulated by renal autoregulation and humoral regulation, and urine volume chiefly depends on the amount of water reabsorbed by distal convoluted tubules and the renal collecting duct. This is affected by changes in the aquaporin expression in this segment. The humoral factors involved in the regulation of urine production are manifold, among which ADH, ALD, and atrial natriuretic peptide (ANP) are the most significant. A study discovered that Phytolacca acinosa Roxb (Shanglu), when processed with vinegar, can significantly decrease the AQP2 expression in rat kidneys, act as a vasopressin-sensitive water channel, significantly decrease serum ADH and ALD, and increase the content of ANP ( 22 ). There is speculation that the regulating effect may be associated with further changes in the regulation of water channel protein expression levels by kidney hormones. Lagopsis supina and its four soluble parts were found to have potential diuretic activity in salt-loaded rats and significantly inhibited renin angiotensin aldosterone system (RAAS) activity, including lowering serum angiotensin II (Ang II), ADH, and ALD levels and increasing serum ANP levels ( 23 ). In particular, two components of Lagopsis supina , stachysoside A and acteoside, have practical diuretic effects through the inhibition of AQPs and RAAS and upregulation of atriopeptin expression in salt-loaded rats, which strongly supports the potential use of Lagopsis supina as a new diuretic ( 24 ). Previous studies have revealed that Ang II may play a meaningful role during AQP2 transport to chief cells of the medullary collecting duct ( 25 ). Further quantitative research needs to be explicit about whether AQP-2 is directly related to RAAS in regulating urine volume. In chronic renal failure (CRF), TCM can also play an adjunctive role by regulating AQP2. Researchers have shown that quercetin, the main component of Shengqing Jiangzhuo capsule, may regulate AQP1 and AQP2 in the kidneys of rats with chronic renal failure, thereby moderating water retention and toxin accumulation to relieve CRF ( 26 ). Ephedra (Mahuang), a water-draining and swelling-dispersing medicinal herb, has a therapeutic effect in renal edema. Experts found that a water extract of Ephedra and its alkaloid components can inhibit the expression of AQP1 and AQP2 in rat kidneys. They speculate that Ephedra and its alkaloid fractions may lower the expression of AQP1 and AQP2 to achieve the effect of inducing diuresis to alleviate edema ( 27 ). In further research, a water extract of Ephedra and its alkaloid components increased the urine volume of rats and reduced the plasma ADH level and the expression of AQP2 protein in the kidneys. This research concluded that its diuretic effect may be related to the level of ADH. However, a different result was observed in another experiment, as a water extract of Ephedra and its alkaloid components had no effect on AQP1 and AQP2 in the kidneys of a rat model with a water retention pattern in the upper energizer ( 28 ). This contradiction with previous research results may be attributed to the differences in the models, as they focused on the kidneys that belonged to the lower energizer and the upper energizer, respectively. In addition to renal AQP2, Ephedra and Epimedium (Yinyanghuo) can suppress the expression of AQP3 in the lungs for the treatment of edema, while a high dose of Epimedium alone can significantly reduce the levels of AQP2 in the kidneys ( 29 , 30 ), regulating the secretion of relevant body fluids from various organs of rats under water load to induce diuresis. This conclusion is consistent with the theory that Ephedra enters the lung meridian and Epimedium enters the kidney meridian, which may explain the insignificant effect of Ephedra on renal AQP2 in some specific disease models. This series of studies indicated that the regulatory effect of traditional Chinese medicine on AQP2 may be related to medicinal meridian tropism and pattern identification, including visceral pattern identification, triple energizer pattern identification, and six-meridian pattern identification in TCM, which may guide studies on Chinese medicinal targets in the future.

As can be seen from the above studies, AQP2 is a critical protein in the regulation of water metabolism and has become a valuable target in the study of TCM treatment of water metabolism imbalance. However, as AQPs are widely distributed in the body, it should be considered whether TCM affects the expression of other aquaporins while regulating AQP2. At the same time, the relationship between the effect of TCM on AQP2 expression and neural and humoral regulation requires further research. Moreover, TCM has fewer side effects and can be used for a long duration. However, due to the lack of detailed research on long-term and short-term regulation of AQP2, the limits of its effect are not clear, and the specific role of TCM still requires further study. In conclusion, an in-depth study on the mechanism of AQP2 and its influence on diseases may provide new ideas for TCM treatment of related diseases ( Supplementary Table S1) .

This study primarily focused on the regulation of AQP2 by traditional Chinese medicine and its application in diseases related to water metabolism. However, there are still limitations that warrant further exploration and improvement. First, most current research on the regulation of AQP2 by TCM primarily involves in vitro cellular studies and animal experiments and lacks large-scale, high-quality randomized controlled clinical trial data. Due to differences between in vivo environments and actual clinical conditions, the extrapolation and reproducibility of experimental results in clinical settings still require further validation. Second, compound traditional Chinese medicines often contain multiple herbs, characterized by their multicomponent and multitarget synergistic effects. Although existing studies suggest that these effects may be mediated through the AVP-V2R-AQP2 signaling pathway or other AQP subtypes, the specific drug–receptor binding modes, downstream signaling networks, and the mechanisms of interaction between components lack systematic and in-depth elucidation. AQP2's regulation is distinguished into short-term and long-term mechanisms at the molecular level; for example, short-term regulation primarily involves the trafficking and localization of AQP2, whereas long-term regulation is associated with gene transcription and protein synthesis. Current research on the effects of TCM over different durations on AQP2 is insufficient, making it difficult to comprehensively assess the optimal timing for intervention and the sustainability of effects at various stages of disease. Although this article focuses on the pivotal role of AQP2 in water metabolism, other AQPs (such as AQP1, AQP3, and AQP4) also play significant roles in fluid balance across various tissues and organs. Whether interventions with TCM synergistically or adversely affect the expression or function of other AQPs has been minimally studied and warrants further exploration. The application of TCM emphasizes individualized treatment based on differential diagnosis; coupled with the fact that the quality of Chinese medicinal materials, preparation techniques, and methods of administration have not yet been completely standardized, this may lead to variations in clinical efficacy and safety across different studies or regions. Future efforts are required to continually improve the quality control of Chinese medicines and standardize clinical dosing. Although there are some exploratory studies suggesting potential synergistic or antagonistic interactions when TCM is used with diuretics and antihypertensive drugs, the current research is not systematic and lacks rigorously designed pharmacokinetic and pharmacodynamic trials and conclusive clinical evidence.