Vasopressin Expressed in Hypothalamic CRF Neurons Causes Impaired Water Diuresis in Secondary Adrenal Insufficiency.

Patients with secondary adrenal insufficiency can present with impaired free water excretion and hyponatremia, which is due to the enhanced secretion of vasopressin (AVP) despite increased total body water. AVP is produced in magnocellular neurons in the paraventricular nucleus of the hypothalamus (PVH) and supraoptic nucleus and in parvocellular corticotropin-releasing factor (CRF) neurons in the PVH. This study aimed to elucidate whether magnocellular AVP neurons or parvocellular CRF neurons coexpressing AVP are responsible for the pathogenesis of hyponatremia in secondary adrenal insufficiency. The number of CRF neurons expressing copeptin, an AVP gene product, was significantly higher in adrenalectomized AVP-floxed mice (AVPfl/fl) than in sham-operated controls. Adrenalectomized AVPfl/fl mice supplemented with aldosterone showed impaired water diuresis under ad libitum access to water or after acute water loading. They became hyponatremic after acute water loading, and it was revealed under such conditions that aquaporin-2 (AQP2) protein levels were increased in the kidney. Furthermore, translocation of AQP2 to the apical membrane was markedly enhanced in renal collecting duct epithelial cells. Remarkably, all these abnormalities observed in the mouse model for secondary adrenal insufficiency were ameliorated in CRF-AVP-/- mice that lacked AVP in CRF neurons. Our study demonstrates that CRF neurons in the PVH are responsible for the pathogenesis of impaired water excretion in secondary adrenal insufficiency.

Mineralocorticoids induce polyuria by reducing apical aquaporin-2 expression of the kidney in partial vasopressin deficiency.

The symptoms of diabetes insipidus may be masked by the concurrence of adrenal insufficiency and emerge after the administration of hydrocortisone, occasionally at high doses. To elucidate the mechanism underlying polyuria induced by the administration of high-dose corticosteroids in the deficiency of arginine vasopressin (AVP), we first examined the secretion of AVP in three patients in whom polyuria was observed only after the administration of high-dose corticosteroids. Next, we examined the effects of dexamethasone or aldosterone on water balance in wild-type and familial neurohypophyseal diabetes insipidus (FNDI) model mice. A hypertonic saline test showed that AVP secretion was partially impaired in all patients. In one patient, there were no apparent changes in AVP secretion before and after the administration of high-dose corticosteroids. In FNDI mice, unlike dexamethasone, the administration of aldosterone increased urine volumes and decreased urine osmolality. Immunohistochemical analyses showed that, after the administration of aldosterone in FNDI mice, aquaporin-2 expression was decreased in the apical membrane and increased in the basolateral membrane in the collecting duct. These changes were not observed in wild-type mice. The present data suggest that treatment with mineralocorticoids induces polyuria by reducing aquaporin-2 expression in the apical membrane of the kidney in partial AVP deficiency.

Immunohistochemical Expression of AQP2 and HSP70 in Broiler Kidney Tissue Treated with Salix tetrasperma Roxb. Extract under Heat Exposure.

The administration of plant extracts to broilers may be a way to mitigate the effects of heat stress. The importance of AQP2 and HSP70 compounds in maintaining the homeostasis of the chicken body when it is subjected to heat stress is well established. This study aims to determine the effect of giving the ethanolic extract of the leaves of Salix tetrasperma Roxb. on the immunohistochemical expression of AQP2 and HSP70 in exposed and unexposed broiler kidney tissue. This study used 36 samples of 28-day-old chicken kidneys. Chickens were kept in individual cages, provided with feed and drinking water ad libitum. The design used was a completely randomized design with 6 treatments and 6 replications: (a) chickens were reared in conditions exposed to heat (HS + 0); (b) chickens were reared in conditions exposed to heat and given Salix extract at a dose of 50 mg/L drinking water (HS + 50); (c) chickens were reared under heat-exposed conditions and given Salix extract at a dose of 100 mg/L drinking water (HS + 100); (d) chickens were reared in conditions without exposure to heat (n-HS + 0); (e) chickens were reared in conditions without exposure to heat and given Salix extract at a dose of 50 mg/L drinking water (nHS + 50); and (f) chickens were reared in conditions exposed without exposure to heat and given 100 mg/L drinking water (nHS + 100) of Salix extract. Salix extract was given for 24 hours and was renewed every 6 hours. The results showed that giving Salix extract 100 mg/L in drinking water to chickens exposed to heat (HS + 100) reduced the value of the H/L ratio. Giving Salix extract 50-100 mg/L in drinking water caused an upregulated AQP2 expression; on the other hand, it downregulated HSP-70 expression, in chicken kidney tubules both exposed to heat stress and nonexposed to heat stress. In conclusion, exposure to heat stress in broiler chickens and giving Salix extract can increase the formation of aquaporin 2 compounds and suppress the formation of HSP70.

Bilobetin induces kidney injury by influencing cGMP-mediated AQP-2 trafficking and podocyte cell cycle arrest.

BACKGROUND: Ginkgo biloba (Gb) extracts have been used as a traditional Chinese medicine. Gb contains flavonoids, which are considered to be its active ingredients and have been used in the treatment of a variety of diseases. However, few scientific research studies on the side effects of flavonoid in Gb have been reported. PURPOSE: The present study aimed to investigate the effect of bilobetin on the kidney of Sprague-Dawley (SD) rats. STUDY DESIGN AND RESULT: In this study, rats were injected with 50 mg/kg of bilobetin, a biflavone isolated from Gb, for 7 days and aristolochic acid was used as positive controls. The results showed that the body weight and urine output of the rats were dramatically decreased, and urinary protein increased after the intraperitoneal injection of bilobetin compared with the control group. Bilobetin treatment showed vacuolar degeneration in the renal tubular epithelium, glomerular atrophy by histostaining, and podocyte fusion by electron microscopy. This study further showed that bilobetin promoted the trafficking of aquaporin 2 (AQP-2) onto the plasma membrane to achieve the function of urine concentration by in vivo study in rats and in vitro study in IMCD-3 cells. The redistribution of AQP-2 is due to increased expression of cGMP in IMCD-3 cells, which in turn promoted the phosphorylation of AQP-2 at site Ser-256. The proteinuria caused by bilobetin may be attributed to podocyte cell cycle arrest at G2/M transition, which is may associated with AKT and MAPK signaling. CONCLUSIONS: The current study showed that bilobetin has some side effects on kidneys at a dose of 50 mg/kg in SD rats and provides insight into the potential detrimental effects of monomeric ingredients in Gb.

Short-Term High-Salt Diet Increases Corin Level to Regulate the Salt-Water Balance in Humans and Rodents.

BACKGROUND: Dietary sodium and potassium affect the fluctuation in blood pressure (BP) and renal function. Corin, with its enzymatic activity to convert pro-atrial natriuretic peptide (pro-ANP) to biologically active ANP, regulates BP, cardiac, and renal functions. We investigated whether corin expression responds to a high-salt (HS) diet to regulate salt and water balance. METHODS: Forty-two volunteers followed 3 sequential diets for 7 days each: a low-salt (LS) diet (3.0 g/day NaCl), a HS diet (18.0 g/day NaCl), followed by an HS diet with K+ supplementation (HS + K+) (18.0 g/day NaCl and 4.5 g/day KCl). RESULTS: Corin level was higher with the HS diet than the LS and HS + K+ diets and was positively correlated with systolic BP (SBP) and 24-hour urinary Na+ and microalbumin (U-mALB) excretion. In rodents, serum and renal levels of corin were transiently increased with the HS diet and were decreased if the HS diet was continued for up to 7 days. HS loading increased SBP, 24-hour urinary Na+, U-mALB excretion, and the expression of proprotein convertase subtilisin/kexin-6 (PCSK6), a corin activator. Knockdown of PCSK6 or corin in high salt-treated M1-cortical collecting duct (M1-CCD) cells increased the expression of aquaporin 2 (AQP2) and ß-epithelial Na+ channel (ß-ENaC). CONCLUSIONS: Short-term HS may induce the PCSK6-corin-ANP-AQP2/ß-ENaC pathway in the kidney. Enhanced serum corin level in humans and rodents is positively correlated with HS-induced SBP and 24-hour urinary Na+ and U-mALB excretion, which suggests that corin is involved in the salt-water balance in response to HS intake. CLINICAL TRIALS REGISTRATION: Public Trials Registry Number NCT02915315.

Quantitative evaluation of Oryeongsan and its action on water regulation in renal inner medullary collecting duct cells.

ETHNOPHARMACOLOGICAL RELEVANCE: Oryeongsan (ORS, Wulingsan) has been reported to possess renal protective effects from renal diseases such as diabetes-induced renal damage, and nephrocalcinosis. AIM OF THE STUDY: This study was conducted to evaluate the quantitative analysis and the inhibitory effect of ORS on hypertonic stress-induced water channel and apoptosis in murine inner medullary collecting duct cell line (mIMCD-3). MATERIALS AND METHODS: Chromatographic and NMR spectroscopic analysis were performed and water balance regulation was determined by Western blot, RT-PCR, and immunofluorescnece. RESULTS: Seven active principles (5-hydroxymethylfurfural, alismoxide, methyl(-)trans-cinnamate, adenine, guanosine, adenosine, and ferulic acid) in ORS were isolated and the structures were identified mainly by NMR spectroscopic analysis. In addition, contents of these metabolites in ORS were evaluated by HPLC analysis. Pretreatment with ORS significantly attenuated the hypertonic stress (175mM NaCl)-induced increase in protein levels of AQP2 and apical membrane insertion. ORS also attenuated osmolyte sodium-myo-inositol transporter (SMIT) expression and tonicity-responsive enhancer binding protein (TonEBP) mRNA under hypertonic stress. Those actions of ORS presented the similar effect of PKA inhibitor which AQP2 expression throughout the inhibition of vasopressin-mediated cAMP/PKA signal pathway. On the other hand, pretreatment with ORS attenuated hypertonic stress-induced cell death. Hypertonic stress-induced Bax or caspase-3 expression was decreased by ORS, resulting in anti-apoptotic effect. CONCLUSIONS: The present data suggest that the beneficial effect of ORS in water balance and apoptosis against hypertonic stress of renal collecting ducts.

Influence of Sanao Tang on urine volume and expression of aquaporin 2 in rats with respiratory function impairment induced by passive smoking and lipopolysaccharide.

OBJECTIVE: To investigate the effect of Sanao Tang (SAT) on urine volume and the expression of aquaporin- 2 (AQP2) in rats with lung dysfunction induced by passive smoking and lipopolysaccharide. METHODS: Totally 45 healthy Specific pathogen Free Wistar Rats were randomized into 3 groups: normal control group, model group and SAT group. A rat model of respiratory dysfunction induced by exposure to cigarette smoking and lipopolysaccharide (LPS). Lavage of decoction of the Chinese medicine was performed for rats in the SAT group. Anires 2005 System was used to analyze the pulmonary function. Urine of rats was collected through metabolism cage method. Enzyme-linked immunosorbent assay (ELISA) was used to determine content of antidiuretic hormone (ADH), angiotensin Ⅱ (AngⅡ), atrial natriuretic factor (ANP), endothelin 1 (ET-1), nitric oxide (NO) and prostaglandin E2 (PGE2) in serum, lung and kidney. The expression of AQP2 in rat renal tissue was determined with real-time fluorescence quantitative PCR (RT-PCR). RESULTS: (a) In comparison with the normal control, It was found that enforced vital capacity (FVC), 1-second forced expiratory volume/forced vital capacity% (FEV(1)/FVC%), 24 h urine volume content of NO and PGE2 were decreased, while AQP2mRNA level and content of ADH, Agn Ⅱ, ANP and ET-1 were increased in the model group with statistical significance (P < 0.05). (b) In comparison with the model group, It was found that FVC, FEV(1), FEV(1)/FVC%, 24 h urine volume, content of PGE2 and NO decreased, while AQP2mRNA level, content of ANP, ADH and Ang Ⅱ decreased in the SAT group with statistical significance (P < 0.05). CONCLUSION: SAT might effectively regulate the urine volume in the modeled rats; ADH, Ang Ⅱ, ANP, ET-1, NO and PGE2 might play an important role in the regulation on urine volume by lungs. This might be the mechanisms underpinning the function of lung governing water passage in terms of the theory of Traditional Chinese Medicine.

[Effect of Yixintai Granule on mRNA and protein expression levels of AQP, in renal medulla of chronic heart failure rabbits].

OBJECTIVE: To explore the effect of Yixintai Granule (YG) on mRNA and protein expression levels of AQP2 in renal medulla of chronic heart failure (CHF) rabbits. METHODS: CHF rat model was established by ear marginal vein injection of adriamycin. Successfully modeled rabbits were divided into the model group, the high (8.4 g/kg), middle (4.2 g/kg), and low dose (2.1 g/kg) YG group, and the Furosemide group (2 mg/kg). Besides, a normal control group was set up. Equal volume of physiological saline was administered to rabbits of the model group and the normal control group by gastrogavage. YG at different doses was administered to rabbits of the 3 YG groups by gastrogavage. The intervention lasted for 4 weeks, once per day. After treatment the urine volume and pathomorphological changes of renal medulla tissue were observed. mRNA and its protein expression levels of AQP2 were detected. RESULTS: Compared with the normal control group, the urine volume decreased significantly, mRNA and protein expression levels of renal medulla AQP2 increased significantly in the model group (all P < 0.01). Compared with the model group, the urine volume increased significantly, and mRNA and protein expression levels of renal medulla AQP2 decreased significantly in all medicated groups (all P < 0.01). Compared with the low dose YG group, the urine volume significantly increased and the mRNA expression level of renal medulla AQP2 significantly decreased in the middle and high dose YG groups (all P < 0.01). The expression level of AQP2 protein significantly decreased in the high dose YG group (P < 0.01). Pathological changes of the renal medulla was the most obviously seen in the model group. But they were alleviated to various degrees in all medicated groups. They were more obviously attenuated in the middle and high dose YG groups. CONCLUSION: YG could improve CHF possibly through down-regulating mRNA and protein expression levels of AQP2 in renal medulla, and elevating the urine volume.

[Relationship between regulation effect of salvia miltiorrhiza on AQP2 in kidney and promoting blood circulation and diuresis].

Partial nature of "promoting blood circulation and dieresis" of Salvia Miltiorrhizain was initially demonstrated by investigating the regulation effect of AQP2 expression in kidney of trauma blood stasis model rats with the Salvia Miltiorrhizain so as to provide guidance for its clinical deployment of administration. Random allocation was taken to averagely divide 30 SD rats into two groups: 10 rats in normal group and 20 rats in blood stasis syndrome group. Trauma blood stasis rat model was established by quantitatively beating. Then the rat model group was divided into model group and salvia group. After 7 days of treatment, the rat kidney AQP2 expression was detected, the content of urine AQP2 was compared and the damaged local muscle and kidney pathological changes were observed by immunohistochemical method and western blot method. Compared with that of the normal group, rats in model group had inflammatory cells infiltration, blood stasis and edema of the injured local muscles and up-regulated AQP2 expression, decreasing urinary output, and kidney tissues blood stasis and edema (P < 0.05). On the other hand, compared with that of the model group, those parameters of rats in salvia group were all decreasing except urine output (P < 0.05). Such result indicated that Salvia Miltiorrhiza can reduce trauma blood stasis rat content of urine AQP2 and down-regulated AQP2 expression in kidney tissue, so as to reduce the reabsorption of water by renal tubular and increase urine output. The promoting blood circulation effect of Salvia Miltiorrhizain can alleviate the degree of the damaged tissue edema and encourage urine drainage. This therapy is closely related to the effect of regulating AQP2 in kidney by salvia, so the purpose of this study by verifying "promoting blood circulation and diuresis" as the mechanism for the regulation effect of the salvia on AQP2 expression.

Rhubarb tannins extract inhibits the expression of aquaporins 2 and 3 in magnesium sulphate-induced diarrhoea model.

Tannins, a group of major active components of Chinese rhubarb and widely distributed in nature, have a significant antidiarrhoeal activity. Aquaporins (AQPs) 2 and 3 play important roles in regulating water transfer during diarrhoea. The present study aims to determine the effect of the total tannins extract of rhubarb on aquaporins (AQPs) 2 and 3 in diarrhoea mice and HT-29 cells both induced by magnesium sulphate (MgSO4). Our results showed that rhubarb tannins extract (RTE) significantly decreased the faecal water content in colon and evaluation index of defecation of diarrhoea mice. Interestingly, RTE could markedly reduce the mRNA and protein expression levels of AQPs 2 and 3 in apical and lateral mucosal epithelial cells in the colons of diarrhoea mice and HT-29 cells both induced by MgSO4 in a dose-dependent manner. Furthermore, RTE suppressed the production of cyclic monophosphate- (cAMP-) dependent protein kinase A catalytic subunits α (PKA C-α) and phosphorylated cAMP response element-binding protein (p-CREB, Ser133) in MgSO4-induced HT-29 cells. Our data showed for the first time that RTE inhibit AQPs 2 and 3 expression in vivo and in vitro via downregulating PKA/p-CREB signal pathway, which accounts for the antidiarrhoeal effect of RTE.

Sclederma of Poria cocos exerts its diuretic effect via suppression of renal aquaporin-2 expression in rats with chronic heart failure.

ETHNOPHARMACOLOGICAL RELEVANCE: Sclederma of Poria cocos (Hoelen) has been used as a diuretic in traditional Asian medicine. However, the underlying mechanism by which Sclederma of Poria cocos (hoelen) exerts its diuretic effect has not been well identified. The aim of the present study was to evaluate the effects of Sclederma of Poria cocos (hoelen) in rats with chronic heart failure (CHF) induced by acute myocardial infarction and to investigate the underlying mechanisms. MATERIALS AND METHODS: An aqueous extract of Sclederma of Poria cocos (hoelen) (2.4 g/kg/d, 1.2 g/kg/d or 0.6 g/kg/d) or furosemide (20 mg/kg/d) was administered orally to male Sprague-Dawley rats starting on the day of coronary ligation. The urine output of all rats was quantified and collected every day for 1 or 4 weeks. The expression of aquaporin-2 (AQP2) was examined after treatment for 1 or 4 weeks. RESULTS: Urinary output increased significantly and urinary osmolality decreased after oral administration of Sclederma of Poria cocos (hoelen) for both 1 and 4 weeks. Sclederma of Poria cocos (hoelen) caused less electrolyte disorder than furosemide. Furthermore, Sclederma of Poria cocos (hoelen) reduced the levels of plasma BNP in CHF rats, whereas furosemide had no effect. Importantly, both mRNA and protein expression of AQP2 were down-regulated and urinary excretion of AQP2 was decreased after administration of Sclederma of Poria cocos (hoelen) to CHF rats. Similarly, Sclederma of Poria cocos (hoelen) reduced plasma arginine vasopressin (AVP) level and down-regulated vasopressin type 2 receptor (V2R) mRNA expression. CONCLUSIONS: Sclederma of Poria cocos (hoelen) exerts its diuretic effect and improves cardiac function in CHF rats via the AVP-V2R-AQP2 axis.

[Effect of Euphorbia kansui on urination and kidney AQP2, IL-1beta and TNF-alpha mRNA expression of mice injected with normal saline].

OBJECTIVE: To observe the effect of Euphorbia kansui (E. KS) alcohol extracts on urination and kidney-related expressions of mice injected with normal saline and to discuss its impact on kidney. METHOD: Mice intraperitoneally injected with normal saline were observed for urination and changes in kidney-related histiocytic factors of after intragastrical administration of E. KS and compared with normal mice. RESULT: E. KS alcohol extracts can promote urination of mice injected with normal saline and enhance peripheral serum creatinine, with no obvious pathological change showed in tissue sections. It had a certain effect on reducing AQP2 expression and enhancing TNF-alpha expression. CONCLUSION: Euphorbia kansui in large dose has a remarkable effect on kidney but may be accompanied with pathological reactions to some extent, especially the dose of 1.2 g x kg(-1). The pathological reactions may be related with increased serum creatinine and TNF-alpha expression.

Effect of Poria cocos on hypertonic stress-induced water channel expression and apoptosis in renal collecting duct cells.

ETHNOPHARMACOLOGICAL RELEVANCE: A major physiological role of the kidney is to regulate body water and urine concentration. Aquaporin-2 (AQP2), a family of water channels, plays an important role in the urinary concentrating process and regulation of water balance in the kidney. The dried sclerotia of Poria cocos Wolf has been known to have a diuretic effect and used for the treatment of chronic edema and nephrosis. AIM OF THE STUDY: This study was conducted to evaluate the inhibitory effect of the sclerotia of Poria cocos (WPC) on hypertonic stress-induced AQP2 expression and apoptosis in inner medullary collecting duct cell lines (IMCD-3). MATERIALS AND METHODS: Hypertonic stress was induced by 175mM NaCl. Inhibitory effect of WPC on hypertonic stress-induced AQP2 expression and apoptosis were determined by western blot, RT-PCR, and immunofluorescence. RESULTS: Hypertonic stress (175mM NaCl) increased in the levels of AQP2 expression by hypertonicity in IMCD-3 cells. WPC attenuated the hypertonicity-induced increase in protein and mRNA levels of AQP2 in a concentration-dependent manner. Pretreatment with WPC attenuated hypertonicity-induced cell death. Hypertonicity increased serum- and glucocorticoid-inducible protein kinase (Sgk1) phosphorylation, however, WPC attenuated the hypertonicity-induced Sgk1 activation. Tonicity-responsive enhancer binding protein (TonEBP) mRNA was also recovered by WPC under hypertonic stress. Pretreatment with WPC presented the similar effect of PKA inhibitor which decreased hypertonic stress-induced AQP2 expression. Hypertonicity increased cAMP levels and the changes were blocked by WPC. On the other hand, hypertonic stress-induced Bax or caspase-3 expression was decreased by WPC, resulting in anti-apoptotic effect. CONCLUSIONS: These results provided evidence that the beneficial effect of WPC in water balance against in vitro hypertonic stress of renal collecting ducts. In addition, WPC exhibits anti-apoptotic property response to hypertonic stress. Thus, these data suggests that WPC has benefit for the therapeutic approach to the inhibition of renal disorder.

[Effect and mechanism of emodin for regulating aquaporin-2 expression in cultured NRK cells].

OBJECTIVE: To investigate the effect and mechanism of emodin for regulating aquapoin-2 (AQP2) in NRK cells cultured in vitro. METHODS: Experiments on NRK cells cultured with alpha-DMEM medium in vitro were conducted in two steps. (1) Cells were randomly divided into 4 groups: the control group, and the three emodin treated groups treated with different dosages of emodin (5, 10 and 20 mg/L) respectively. After 24 h treatment, the location of AQP2 was decided by indirect immunofluorescene, and the AQP2 protein and mRNA expression levels were detected by Western blot and semiquantive RT-PCR. (2) Cells were randomly divided into 4 groups, the control group, and the three treated groups treated respectively with 10 mg/L 8-Bromo-cAMP, 20 mg/L emodin, and 20 mg/L emodin +10 mg/L 8-Bromo-cAMP. The activity of protein kinase A (PKA) in NRK cells after 24 h treatment was determined with non-radioactive detecting method. RESULTS: AQP2 was located at the cell membrane of NRK cells. Western blot and semiquantitive RT-PCR found that AQP2 protein and mRNA expressions were significantly decreased in NRK cells of groups treated by 10 mg/L and 20 mg/L emodin (P < 0.05). PKA activity determination showed significantly decreased phosphorylation level of PKA in NRK cells of groups treated with 20 mg/L emodin group (P < 0.05). CONCLUSION: Emodin can inhibit the genetic transcription and the translation of AQP2 gene in NRK cells, which demonstrates that the change of AQP2 expression regulated by emodin may be correlated with the diuresis effect of rhubarb, and it is likely that the regulation is going through PKA signal pathway.

[Impacts of the formula of Suoquanwan(SQW) on expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency].

OBJECTIVE: To observe the impacts of the formula of Suoquanwan (SQW) on the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency. METHODS: The model rats were induced by adenine (250 mg/kg) for 4 weeks, then treated respectively with SQW or dDAVP. The expression of AQP-2 mRNA and AVPR-V2 mRNA in kidney of Yang-deficiency model by realtime fluorescence quantitative PCR method were investigated. RESULTS: In model rats, the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney decreased, dDAVP and SQW high dose could increased the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney. The others had no influence on the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney. CONCLUSION: SQW can increase the expression of AQP-2 mRNA and AVPR-V2 mRNA in the kidney of rat polyuria model of Yang-deficiency.

[Effect of total anthraquinone in rheum on aquaporin 2 expression in rat distal colon].

OBJECTIVE: To investigate effect of total anthraquinone in rheum on aquaporin 2 expression in rat distal colon. METHOD: SD rats were randomly divided into control group, low dose group, middle dose group and high dose group. Gavaged to control group, and treated group were administered saline and total anthraquinone in rheum with dosage of 0.14, 2.5, 4.5 g x kg(-1) x d(-1), respectively. All rats were put sacrificed after 5 days and stool in full length colon was gently collected to detect water content stool. Distal colon was removed to detect AQP2 expression with immunohistochemistry, western blot and RT-PCR. RESULT: No diarrhea was found in low dose group and control group, there were not significant difference water content of stool and AQP2 expression between low dose group and control group. However, soft feces and loose stools occurred in diarrheic dose group, loose stools and watery stool appeared in high dose group. Stool water content increased in diarrheic dose group and High dose group, expression of AQP2 decreased evidently in these two groups (P < 0.01). CONCLUSION: Total anthraquinone in rheum can reduce the transcription and translation of AQP2 in rats' distal colon, increase fecal water content, which probably is one of the mechanisms of diarrhea caused by total anthraquinone in rheum.

[Effect of rhubarb on expressions of aquaporin-2 and -4 in rat's kidney].

OBJECTIVE: To investigate the effect of rhubarb on expressions of aquaporin-2 and 4 (AQP2 and AQP4) in rat's kidney. METHODS: Thirty-two SD rats were randomly divided into 4 groups, the normal control group, and the three rhubarb groups medicated via gastrogavage with low, mid and high dose of rhubarb extract (total anthraquinone) respectively. The 6 h and 24 h urine volume were measured, and the protein and mRNA expressions of AQP2 and AQP4 in renal tissue were determined with immunohistochemistry, Western blot and RT-PCR. RESULTS: No significant difference between the control group and the low dose rhubarb treated group was found in urine volume, as well as in AQP2 and AQP4 protein and mRNA expressions. But the urine volume was obviously higher, the protein and mRNA expressions of AQP2 and AQP4 were markedly lower in rats after mid/high dose rhubarb medication respectively when compared with those in the normal controls (all P < 0.01). CONCLUSION: Rhubarb can inhibit the protein and mRNA expressions of AQP2 and AQP4 in rats' kidney, which probably is one of the mechanisms of rhubarb for diuresis.

Effects of water restriction on gene expression in mouse renal medulla: identification of 3betaHSD4 as a collecting duct protein.

To identify novel gene targets of vasopressin regulation in the renal medulla, we performed a cDNA microarray study on the inner medullary tissue of mice following a 48-h water restriction protocol. In this study, 4,625 genes of the possible approximately 12,000 genes on the array were included in the analysis, and of these 157 transcripts were increased and 63 transcripts were decreased by 1.5-fold or more. Quantitative, real-time PCR measurements confirmed the increases seen for 12 selected transcripts, and the decreases were confirmed for 7 transcripts. In addition, we measured transcript abundance for many renal collecting duct proteins that were not represented on the array; aquaporin-2 (AQP2), AQP3, Pax-8, and alpha- and beta-Na-K-ATPase subunits were all significantly increased in abundance; the beta- and gamma-subunits of ENaC and the vasopressin type 1A receptor were significantly decreased. To correlate changes in mRNA expression with changes in protein expression, we carried out quantitative immunoblotting. For most of the genes examined, changes in mRNA abundances were not associated with concomitant protein abundance changes; however, AQP2 transcript abundance and protein abundance did correlate. Surprisingly, aldolase B transcript abundance was increased but protein abundance was decreased following 48 h of water restriction. Several transcripts identified by microarray were novel with respect to their expression in mouse renal medullary tissues. The steroid hormone enzyme 3beta-hydroxysteroid dehydrogenase 4 (3betaHSD4) was identified as a novel target of vasopressin regulation, and via dual labeling immunofluorescence we colocalized the expression of this protein to AQP2-expressing collecting ducts of the kidney. These studies have identified several transcripts whose abundances are regulated in mouse inner medulla in response to an increase in endogenous vasopressin levels and could play roles in the regulation of salt and water excretion.