Cartilage protective and anti-edema effects of JTF in osteoarthritis via inhibiting NCOA4-HMGB1-driven ferroptosis and aquaporin dysregulation.

BACKGROUND: Preventing joint edema is crucial in halting osteoarthritis (OA) progression. Growing clinical evidence indicate that Jianpi-Tongluo Formula (JTF) may have a promising anti-edema effect. However, the therapeutic properties of JTF and the underlying mechanisms remains unclear. MATERIALS AND METHODS: An OA rat model was established and employed to evaluate pharmacological effects of JTF in vivo based on dynamic histopathologic assessments and micro-CT observations. Then, OA-related genes and potential targets of JTF were identified through clinical transcriptomic data analysis and "disease gene-drug target" network analysis, which were verified by a series of in vivo experiments. RESULTS: JTF administration effectively reduced pain and joint edema, inhibited matrix degradation, chondrocyte apoptosis, and aquaporin expression in OA rats. Notably, JTF dose-dependently reversed damage-associated molecular patterns and inflammatory factor upregulation. Mechanically, our "disease gene-drug target" network analysis indicated that the NCOA4-HMGB1-GSK3B-AQPs axis, implicated in ferroptosis and aquaporin dysregulation, may be potentially served as a target of JTF against OA. Accordingly, JTF mitigated NCOA4, HMGB1, and GSK3B expression, oxidative stress, and iron metabolism aberrations in OA rats. Furthermore, JTF treatment significantly attenuated the aberrant upregulation of AQP1, AQP3, and AQP4 proteins observed in cartilage tissues of OA rats. CONCLUSION: Our data reveal for the first time that JTF may exert cartilage protective and anti-edema effects in osteoarthritis therapy by inhibiting NCOA4-HMGB1-driven ferroptosis and aquaporin dysregulation.

Shen Qi Wan ameliorates nephritis in chronic kidney disease via AQP1 and DEFB1 regulation.

Shen Qi Wan (SQW) has been proven to exert anti-inflammatory effects in the kidneys of CKD models accompanied by unclear therapeutic mechanisms. This study aims to evaluate the kidney-protective and anti-inflammatory effects of SQW and to elucidate its fundamental mechanisms for CKD treatment. Firstly, the main active components of SQW were identified by UPLC-Q-TOF/MS technique. Subsequently, we evaluated inflammatory factors, renal function and renal pathology changes following SQW treatment utilizing adenine-induced CKD mice and aquaporin 1 knockout (AQP1-/-) mice. Additionally, we conducted RNA-seq analysis and bioinformatics analysis to predict the SQW potential therapeutic targets and anti-nephritis pathways. Simultaneously, WGCNA analysis method and machine learning algorithms were used to perform a clinical prognostic analysis of potential biomarkers in CKD patients from the GEO database and validated through clinical samples. Lipopolysaccharide-induced HK-2 cells were further used to explore the mechanism. We found that renal collagen deposition was reduced, serum inflammatory cytokine levels decreased, and renal function was improved after SQW intervention. It can be inferred that ß-defensin 1 (DEFB1) may be a pivotal target, as confirmed by serum and renal tissue samples from CKD patients. Furthermore, SQW assuages inflammatory responses by fostering AQP1-mediated DEFB1 expression was confirmed in in vitro and in vivo studies. Significantly, the renal-protective effect of SQW is to some extent attenuated after AQP1 gene knockout. SQW could reduce inflammatory responses by modulating AQP1 and DEFB1. These findings underscore the potential of SQW as a promising contender for novel prevention and treatment strategies within the ambit of CKD management.

Shen Qi Wan attenuates renal interstitial fibrosis through upregulating AQP1.

Renal interstitial fibrosis (RIF) is the crucial pathway in chronic kidney disease (CKD) leading to the end-stage renal failure. However, the underlying mechanism of Shen Qi Wan (SQW) on RIF is not fully understood. In the current study, we investigated the role of Aquaporin 1 (AQP1) in SQW on tubular epithelial-to-mesenchymal transition (EMT). A RIF mouse model induced by adenine and a TGF-ß1-stimulated HK-2 cell model were etablished to explore the involvement of AQP 1 in the protective effect of SQW on EMT in vitro and in vivo. Subsequently, the molecular mechanism of SQW on EMT was explored in HK-2 cells with AQP1 knockdown. The results indicated that SQW alleviated kidney injury and renal collagen deposition in the kidneys of mice induced by adenine, increased the protein expression of E-cadherin and AQP1 expression, and decreased the expression of vimentin and α-smooth muscle actin (α-SMA). Similarly, treatmement with SQW-containing serum significantly halted EMT process in TGF-ß1 stimulated HK-2 cells. The expression of snail and slug was significantly upregulated in HK-2 cells after knockdown of AQP1. AQP1 knockdown also increased the mRNA expression of vimentin and α-SMA, and decreased the expression of E-cadherin. The protein expression of vimentin increased, while the expression of E-cadherin and CK-18 significantly decreased after AQP1 knockdown in HK-2 cells. These results revealed that AQP1 knockdown promoted EMT. Furthermore, AQP1 knockdown abolished the protective effect of SQW-containing serum on EMT in HK-2 cells. In sum, SQW attentuates EMT process in RIF through upregulation of the expression of AQP1.

XinLi formula, a traditional Chinese decoction, alleviates chronic heart failure via regulating the interaction of AGTR1 and AQP1.

BACKGROUND: XinLi formula (XLF) is a traditional Chinese medicine used in clinical practice to treat chronic heart failure (CHF) in humans, with remarkable curative effect. However, the mechanism remains unknown. PURPOSE: The goal of the current investigation was to determine how XLF affected CHF in a rat model of the condition brought on by ligation of the left anterior descending coronary artery, and to investigate the underlying mechanism. STUDY DESIGN AND METHODS: Cardiac function was detected by echocardiography. The contents of myocardial enzymes, Ang II, ALD, TGF-ß1, and inflammatory factors were measured by ELISA. Myocardial injury and myocardial fibrosis were evaluated by HE and Masson staining. Myocardial edema was assessed by cardiac mass index and transmission electron microscopy. Using Western blot and immunohistochemistry to examining the protein expression of inflammasome, TGF-ß1, AGTR1, and AQP1 in the left ventricle. Furthermore, the interaction of AGTR1 and AQP1 was evaluated by co-immunoprecipitation. RESULTS: XLF attenuated myocardial enzymes and myocardial injury, and improved cardiac function in rats with CHF after myocardial infarction. It also reduced Ang II and ALD levels in CHF rats, and suppressed the expression of AGTR1 and TGF-ß1, finally alleviated myocardial fibrosis. By mechanism, XLF inhibited the expression of NLRP3 inflammasome proteins, reduced the plasma contents of IL-1ß, IL-18, IL-6 and TNF-α. Additionally, XLF inhibited the expression of AQP1 and the interaction of AGTR1 and AQP1, alleviating myocardial edema. The common structure of the main chemical constituents of XLF were glycoside compounds with glycosyl. CONCLUSION: XLF ameliorated CHF, which was evidenced by the alleviation of myocardial fibrosis by inhibiting AGTR1/NLRP3 signal, as well as the attenuation of myocardial edema by suppressing the interaction of AGTR1 and AQP1.

Shen Qi Wan attenuates renal interstitial fibrosis through upregulating AQP1 / 中国天然药物

Renal interstitial fibrosis (RIF) is the crucial pathway in chronic kidney disease (CKD) leading to the end-stage renal failure. However, the underlying mechanism of Shen Qi Wan (SQW) on RIF is not fully understood. In the current study, we investigated the role of Aquaporin 1 (AQP1) in SQW on tubular epithelial-to-mesenchymal transition (EMT). A RIF mouse model induced by adenine and a TGF-β1-stimulated HK-2 cell model were etablished to explore the involvement of AQP 1 in the protective effect of SQW on EMT in vitro and in vivo. Subsequently, the molecular mechanism of SQW on EMT was explored in HK-2 cells with AQP1 knockdown. The results indicated that SQW alleviated kidney injury and renal collagen deposition in the kidneys of mice induced by adenine, increased the protein expression of E-cadherin and AQP1 expression, and decreased the expression of vimentin and α-smooth muscle actin (α-SMA). Similarly, treatmement with SQW-containing serum significantly halted EMT process in TGF-β1 stimulated HK-2 cells. The expression of snail and slug was significantly upregulated in HK-2 cells after knockdown of AQP1. AQP1 knockdown also increased the mRNA expression of vimentin and α-SMA, and decreased the expression of E-cadherin. The protein expression of vimentin increased, while the expression of E-cadherin and CK-18 significantly decreased after AQP1 knockdown in HK-2 cells. These results revealed that AQP1 knockdown promoted EMT. Furthermore, AQP1 knockdown abolished the protective effect of SQW-containing serum on EMT in HK-2 cells. In sum, SQW attentuates EMT process in RIF through upregulation of the expression of AQP1.

[Anti-ascites effect of total saponins of Phytolaccae Radix on mice with ascites and mechanism].

This study investigated the anti-ascites effect of the total saponins of Phytolaccae Radix(PRTS) and the mechanism.H22 cell suspension was used(ip) to induce ascites in ICR male mice, and the model mice were randomized into model group, positive drug group(furosemide, 6 mg·kg~(-1)), total extract of Phytolaccae Radix(PRTE) group, and PRTS(1.29 g·kg~(-1)).Another 10 male mice were selected as the blank group.Mice in the blank group and model group were given(ig) normal saline containing 0.5% CMC-Na, and those in the positive drug group, PRTE group, and PRTS group received(ig) corresponding doses of drugs, once a day, for 8 consecutive days.The ascites volume, urine volume, and fecal water content in mice with ascites, serum levels of antidiure-tic hormone(ADH), renin in renin-angiotensin-aldosterone system(RAAS), angiotensin Ⅱ(AngⅡ), and aldosterone(ALD), expression of aquaporin(AQP)1-AQP4 in kidney, expression of AQP1, AQP3 in colon, and expression of phosphatidylinositol 3-kinase/protein kinase B(PI3 K/Akt) pathway-related proteins were detected to explore the anti-ascites mechanism of PRTS.The results showed that the PRTS can increase the urine volume and fecal water content and decrease the ascites volume of ascites mice.Moreover, PRTS significantly reduced the expression of AQP1-AQP4 in kidney and AQP1, AQP3 in colon, serum levels of renin, AngⅡ, ALD, and ADH, and the expression of p-PI3 K and p-Akt in the kidney of ascites mice.PRTS exerts anti-ascites effect by promoting urination and defecation.The mechanism is that it inhibits the activities of RAAS and ADH and suppresses the phosphorylation of PI3 K/Akt signaling pathway, thereby restricting the expression of AQPs in the kidney and colon.

Qi Sui Zhu Shui Plaster Inhibits AQP1 and MAPK Signaling Reduces Liver Damage Induced by Cirrhotic Ascites.

Objective: At present, there is no special treatment for cirrhotic ascites in modern medicine. Qi Sui Zhu Shui plaster (QSZSP) has been used in ascites. The purpose of this study was to investigate the mechanism of action of QSZSP in the treatment of cirrhotic ascites and its relationship with aquaporin 1 (AQP1). Methods: Twenty-four rats were divided into four groups, six rats in each group. Carbon tetrachloride-olive oil is injected into modeling. The control and model groups are treated with blank gel plaster (2 cm × 2 cm), QSZSP low-dose group is treated with Qi Sui Zhu Shui plaster (1 cm × 1 cm), and QSZSP high-dose group is treated with Qi Sui Zhu Shui plaster (2 cm × 2 cm). The changes in body weight and abdominal circumference were measured, the histopathological changes in liver, kidney, and peritoneum were observed in HE staining, the biochemical indexes related to liver function were detected, and the changes in AQP1 expression and the activation of MAPK pathway in the liver, kidney, and peritoneal tissues were evaluated in IHC staining and Western blot. Results: After one week of injection of carbon tetrachloride-olive oil, the rats in the model group increased their body weight slowly, the abdominal circumference of the model rats continued to increase with time. After 16 weeks of construction of the cirrhotic ascites model, the liver, kidney, and peritoneum were significantly damaged, and the serum levels of TBiL, AST, ALT, Cr, BUN, K, Na, and Ca in the rats were significantly higher (P < 0.001) and ALB levels were significantly lower (P < 0.001) than those in the control group. After 4 weeks of treatment, the liver, kidney, and peritoneal injury were improved. TBiL, AST, ALT, Cr, BUN, K, Na, and Ca levels were significantly lower (P < 0.001) and ALB levels were significantly higher (P < 0.001) than those in the model group. The protein expression of AQP1, p-ERK, p-JNK, and p-p38 was found to be inhibited in the liver, kidney, and peritoneum. Conclusion: QSZSP inhibits the protein expression of AQP1 and MAPK signaling pathway in the liver, peritoneum, and kidney to alleviate liver, kidney, and peritoneal injury caused by cirrhotic ascites, thus reducing the abnormal growth of abdominal circumference.

Xinshuitong Capsule extract attenuates doxorubicin-induced myocardial edema via regulation of cardiac aquaporins in the chronic heart failure rats.

Doxorubicin (Dox), an effective antineoplastic drug, was limited use for cardiotoxicity. Xinshuitong Capsule (XST), a patented herbal formula, showed desirable beneficial effects in the treatment of chronic heart failure (CHF) patients. However, the drug on Dox-induced cardiotoxicity remains unclear. Ninety male Sprague-Dawley rats were randomized into two groups: 15 rats were selected as the normal group and 75 rats were injected intraperitoneally with Dox to establish CHF rat models, the success ones were randomly divided into five groups: low XST (LXST), medium XST (MXST) or high XST (HXST) (4.9, 9.8, or 19.6 g/kg d) administrated intragastrically twice a day for 4 weeks, with the captopril-treated group and the model group as comparison. The model group showed the cardiac functions generally impaired, and CHF mortality rate higher (47%) than those in the XST-treated groups (averaged 24%, P < 0.05). Compared with XST-treated groups, myocardial remodeling, inflammation and desarcomerization, and higher water content more severe in the cardiac tissue in the model group (P < 0.05), which was associated with higher expressions of mRNA or protein levels of AQP1, 4 and 7. Dox-impaired cardiac functions, cardiac remodeling and myocardial edema could be dose-dependently reverted by XST treatment. XST could inhibit AQP1, 4 and 7 at mRNA levels or at protein levels, which was associated with the attenuation of myocardial edema and cardiac remodeling, decreasing the ventricular stiffness and improving the cardiac functions and rats' survival. AQPs is involved in cardiac edema composed one of the mechanisms of Dox-induced cardiotoxicity, XSTvia inhibition of AQPs relieved the Dox-induced side effects.

Amelioration of airway inflammation and pulmonary edema by Teucrium stocksianum via attenuation of pro-inflammatory cytokines and up-regulation of AQP1 and AQP5.

Current study investigates the immunomodulatory effects of T. stocksianum using mouse model of ovalbumin (OVA)-induced allergic asthma. The mice were treated with methanolic extract, n-hexane, and ethyl acetate fractions for consecutive 7 days along with intranasal challenge. The mRNA expression levels of interleukin-4 (IL-4), IL-5, Aquaporin-1 (AQP1) and Aquaporin-5 (AQP5) were evaluated using reverse transcription polymerase chain reaction. The data showed that T. stocksianum significantly reduced airway inflammation as indicated by reduced inflammatory cell infiltration in lungs, and attenuated total and differential leukocyte counts both in blood and BALF. Expression levels of pro-inflammatory IL-4 and IL-5 in lungs were also found significantly reduced. T. stocksianum significantly reduced pulmonary edema as indicated by reduced lung wet/dry ratio and goblet cell hyperplasia. AQP1 and AQP5 expression levels were also found elevated in treatment groups. In conclusion, T. stocksianum possesses anti-asthmatic activity which may be attributed to reduction in IL-4 and IL-5 expression levels, and elevation in AQP1 and AQP5 expression levels.

The mechanism underlying hypaconitine-mediated alleviation of pancreatitis-associated lung injury through up-regulating aquaporin-1/TNF-α.

BACKGROUND/AIMS: Acute pancreatitis-associated lung injury (APALI) is one of the most common and most dangerous form of extra-pancreatic organ damage in severe acute pancreatitis (SAP). The treatment options for SAP were limited thus far; as a result, approximately 60%-80% of patients with SAP would die within a week. Hypaconitine (HC), one of the most important active ingredients in a Mongolian traditional medicine Radix Aconiti Kusnezoffii has an excellent anti-inflammatory effect. MATERIALS AND METHODS: To ascertain whether HC has a protective effect against APALI, we investigated the therapeutic effects and the underlying mechanisms in vivo and in vitro and attempted to elucidate the mechanism in detail. In this study, APALI rats and human pulmonary microvascular endothelial cells were treated with therapeutic doses of HC after establishing a model with sodium taurocholate and lipopolysaccharide, respectively. RESULTS: Serum amylase and lipase activity, lung wet/dry weight ratio, lung myeloperoxidase activity, and pancreatic and lung histopathological changes showed that HC alleviated APALI in a dose-dependent way, which can be abolished by an aquaporin-1 (AQP-1) knockdown. The results of the reverse transcriptase polymerase chain reaction, Western blot, and immunohistochemical staining confirmed the expression of AQP-1, a kind of transmembrane protein that mainly distributed in the membranes of pulmonary cells and contributed to maintain water balance in the body by interacting with tumor necrosis factor-alpha (TNF-α), is negatively associated with APALI. On the contrary, HC treatment up-regulated AQP-1 expression and down-regulated the TNF-α expression as a consequence in APALI. CONCLUSION: These results suggest that HC has a good anti-inflammatory therapeutic effect on APALI with a possible underlying mechanism that affects the AQP-1/TNF-α pathway.

The Effect of Shen Qi Wan Medicated Serum on NRK-52E Cells Proliferation and Migration by Targeting Aquaporin 1 (AQP1).

BACKGROUND Shen Qi Wan (SQW) as a well-known formula for the amelioration of kidney yang deficiency syndrome (KYDS), and it has been widely employed in traditional Chinese medicine (TCM). This study aimed to investigate the effect and underlying mechanism of SQW medicated serum on proliferation and migration in NRK-52E cells. MATERIAL AND METHODS We employed the real-time cell analysis (RTCA) system to investigate the effect of SQW medicated serum on proliferation and migration in NRK-52E cells. In addition, the migration was further investigated by using a wound-healing assay. The mRNA and protein expression level of aquaporin 1 (AQP1) of NRK-52E cells with SQW medicated serum-treated were quantified by real-time quantitative polymerase chain reaction (q-PCR) and western blot assay, respectively. Furthermore, NRK-52E cells were transfected with lentivirus AQP1-RNAi to assess migratory cell abilities in vitro. RESULTS The migratory abilities of NRK-52E cells were significantly increased after SQW medicated serum treatment (P<0.05), and no significant difference in cell proliferation. In addition, SQW medicated serum was significantly upregulated the mRNA and protein expression level of AQP1 in NRK-52E cells (P<0.05). Additionally, the in vitro metastasis test proved that knockdown of AQP1 suppressed migratory abilities according to RTCA and wound healing test while was reversed by SQW medicated serum (P<0.05). CONCLUSIONS Our study demonstrates that SQW medicated serum effectively promotes the migration of NRK-52E cells by increasing AQP1 expression, and AQP1 may be as a therapeutic target of SQW for renal injury treatment under KYDS.

Immunomodulatory and Anti-Inflammatory Potential of Curcumin for the Treatment of Allergic Asthma: Effects on Expression Levels of Pro-inflammatory Cytokines and Aquaporins.

Curcumin is well known for possessing anti-inflammatory properties and for its beneficial effects in the treatment of asthma. Current study investigates the immunomodulatory and anti-inflammatory effects of curcumin using mouse model of ovalbumin-induced allergic asthma. BALB/c mice were immunized with ovalbumin on day 0 and 14 to induce allergic asthma. Animals were treated with two different doses of curcumin (20 mg/kg and 100 mg/kg) and methylprednisolone from day 21 to 28. Mice were also daily challenged intranasally with ovalbumin during treatment period, and all groups were sacrificed at day 28. Histopathological examination showed amelioration of allergic asthma in treated groups as evident by the attenuation of infiltration of inflammatory cells, goblet cell hyperplasia, alveolar thickening, and edema and vascular congestion. Curcumin significantly reduced total and differential leukocyte counts in both bronchoalveolar lavage fluid and blood. Reverse transcription polymerase chain reaction analysis showed significantly suppressed mRNA expression levels of IL-4 and IL-5 (pro-inflammatory cytokines), TNF-α, TGF-ß (pro-fibrotic cytokines), eotaxin (chemokine), and heat shock protein 70 (marker of airway obstruction) in treated groups. Attenuation of these pro-inflammatory markers might have led to the suppression of airway inflammation. The expression levels of aquaporin-1 (AQP) and AQP-5 were found significantly elevated in experimental groups which might be responsible for reduction of pulmonary edema. In conclusion, curcumin significantly ameliorated allergic asthma. The anti-asthmatic effect might be attributed to the suppression of pro-inflammatory cytokines, and elevation of aquaporin expression levels, suggesting further studies and clinical trials to establish its candidature in the treatment of allergic asthma.

Regulation effects of naringin on diesel particulate matter-induced abnormal airway surface liquid secretion.

BACKGROUND: PM2.5 is closely related to the incidence and mortality of respiratory diseases. Diesel particulate matter (DPM) is the main component of particulate air pollution and an important source of PM2.5. HYPOTHESIS/PURPOSE: This study mainly explored the effect of DPM on airway surface liquid (ASL) secretion and the regulation of naringin in this process, to evaluate therapeutic potentials of naringin for the treatment of abnormal secretion of the respiratory tract caused by PM2.5. METHODS: The concentration of lysozyme was measured by Lysozyme Assay Kit. Total protein content was determined by the BCA Protein Assay Kit. The concentration of cAMP and MUC5AC, expressions of CFTR, AQP1, and AQP5 proteins were measured by ELISA. Expressions of CFTR, AQP1 and AQP5 mRNA were determined by qPCR. Amount of CFTR on the cell membrane was determined by immunofluorescence. RESULTS: The in vitro and in vivo studies had indicated that DPM could inhibit ASL secretion and increased the viscosity of the liquid. Naringin had the functions to attenuate DPM-induced injury, reduce liquid viscosity by reducing MUC5AC and total protein secretion, increase DPM-induced CFTR, AQP1, and AQP5 mRNA and protein expression, positively regulate apical CFTR insertion and promote CFTR activation by increasing intracellular cAMP. CONCLUSION: These results demonstrated that naringin had regulating effects on the DPM-induced abnormal secretion of the respiratory tract.

Vitamin D Regulates the Expressions of AQP-1 and AQP-4 in Mice Kidneys.

AIM: Vitamin D plays an important role in water and salt homeostasis. The aim of our study was to investigate the underlying relationship of Vitamin D and Aquaporins (AQP). METHODS: The behaviors of 1α (OH)-ase knockout mice and wild type mice were observed before analysis. The ICR mice were treated with vehicle or paricalcitol, a vitamin D analogue, followed by animals receiving a standard diet and free access to drinking water either with aliskiren (renin blocker; 37.5 mg aliskiren in 100 ml water), or telmisartan (a angiotensin II type I receptor blocker; 40 mg telmisartan in 100 ml water) a week before study. The expressions of AQP-1, AQP-4, and renin in mice kidneys were detected by western bolting, immunohistochemistry, and immunofluorescence. RESULTS: Diuresis and polydipsia were observed in 1α (OH)-ase knockout mice, and a decreased water intake and urine output in ICR mice was observed after paricalcitol treatment. Compared with wild type, the AQP-1 expressions were increased in renal papilla and AQP-4 expressions were decreased in renal proximal tubule of 1α(OH) ase knockout mice. In addition, AQP-1 was decreased in renal papilla and AQP-4 expressions were increased in proximal tubule by suppressing renin activity or supplement of Vitamin D analogue. After injecting renin into the lateral ventricle of the 1α(OH)ase knockout mice, the renin expression level was decreased in the kidney, followed by the decrease of AQP-1 in renal papilla and increase of AQP-4 in proximal tubule. CONCLUSIONS: Overall, Vitamin D and renin inhibitors have synergistic effects in regulating water channels in mice kidneys.

Therapeutic efficacy of zingerone against vancomycin-induced oxidative stress, inflammation, apoptosis and aquaporin 1 permeability in rat kidney.

Vancomycin (VCM) is a glycopeptidic broad-spectrum antibiotic against methicillin-resistant Staphylococcus aureus, though it has some adverse effects, including nephrotoxicity, that limit its usefulness. Zingerone (ZO), a component of dry ginger root, has several pharmacological activities due to its antioxidant, anti-inflammatory and antiapoptotic properties. The aim of this study was to determine the therapeutic efficacy of ZO against VCM-induced oxidative stress, inflammation, apoptosis and kidney aquaporin 1 (AQP1) levels in rats. Intraperitoneal administration of VCM (200 mg/kg body weight) for seven days increased kidney lipid peroxidation and decreased antioxidant enzyme activities, including kidney superoxide dismutase (SOD), catalase (CAT) and glutathione peroxidase (GPx). VCM increased serum creatinine and urea levels and induced histopathological changes while causing a decrease in AQP1 protein level. VCM also increased the levels of the inflammatory markers nuclear factor kappa B (NF-κB), B-cell lymphoma-3(Bcl-3), interleukin-1ß (IL-1ß), interleukin-33 (IL-33), tumor necrosis factor-α (TNF-α), prostaglandin E2 (PGE2), inducible nitric oxide synthase (iNOS), myeloperoxidase (MPO) and cyclooxygenase-2 (COX-2). Moreover, it activated the apoptotic pathway by increasing the expression levels of p53, Bcl-2 associated X protein (Bax), cysteine aspartate specific protease-3 (caspase-3) and 8-hydroxy-2'-deoxyguanosine (8-OHdG), which is a marker of oxidative DNA damage. Treatment with ZO (25 and 50 mg/kg body weight) at both doses prevented nephrotoxicity by ameliorating the histopathological alterations, oxidative stress, inflammation, apoptosis, oxidative DNA damage and renal AQP1 levels. The findings of the present study suggested that ZO attenuates VCM-induced nephrotoxicity.

[Effects of toxic fractions of Euphorbiae Ebracteolatae Radix on toxicity of mice intestinal tract and colon aquaporins expression level before and after vinegar processing].

To investigate the toxicity changes of Euphorbiae Ebracteolatae Radix (EER) before and after vinegar processing, toxic diterpenoids were concentrated with chloroform as extraction solvent from EER. Then the residue was extracted for non-chloroform extract with 95% ethanol and water after extraction with chloroform. The chloroform extraction of vinegar processed EER was prepared with the same method. The mice received the drug by oral administration. Moisture content in mice feces, duodenum and colon tissue, aquaporin AQP1, AQP3, AQP4 protein expression levels were assayed as the indexes to investigate the toxicity variation of chloroform fraction, non-chloroform fraction, as well as intestinal tract toxicity before and after vinegar processing of EER. The results showed that the chloroform fraction extracted from EER could significantly increase the moisture content in mice feces, duodenum and colon, and decrease AQP1 protein expression level, increase AQP3 and AQP4 protein expression levels in the colon. The intestinal toxicity of the chloroform extract was significantly higher than that of non-chloroform extract. The moisture content in mice feces, duodenum and colon was significantly decreased, and the AQPs protein expression tended to be normal in the colon after vinegar processing. The results showed that the chloroform fraction extracted from EER could lead to diarrhea, intestinal edema, and the intestinal toxicity action was associated with interfering AQPs protein expression and promoting intestinal fluid transport disorder in mice. Vinegar-processing could reduce intestinal toxicity of EER, so vinegar processing was considered to be the scientific processing method of EER.

The Aquaporin 1 Inhibitor Bacopaside II Reduces Endothelial Cell Migration and Tubulogenesis and Induces Apoptosis.

Expression of aquaporin-1 (AQP1) in endothelial cells is critical for their migration and angiogenesis in cancer. We tested the AQP1 inhibitor, bacopaside II, derived from medicinal plant Bacopa monnieri, on endothelial cell migration and tube-formation in vitro using mouse endothelial cell lines (2H11 and 3B11) and human umbilical vein endothelial cells (HUVEC). The effect of bacopaside II on viability, apoptosis, migration and tubulogenesis was assessed by a proliferation assay, annexin-V/propidium iodide flow cytometry, the scratch wound assay and endothelial tube-formation, respectively. Cell viability was reduced significantly for 2H11 at 15 µM (p = 0.037), 3B11 at 12.5 µM (p = 0.017) and HUVEC at 10 µM (p < 0.0001). At 15 µM, the reduced viability was accompanied by an increase in apoptosis of 38%, 50% and 32% for 2H11, 3B11 and HUVEC, respectively. Bacopaside II at ≥10 µM significantly reduced migration of 2H11 (p = 0.0002) and 3B11 (p = 0.034). HUVECs were most sensitive with a significant reduction at ≥7.5 µM (p = 0.037). Tube-formation was reduced with a 15 µM dose for all cell lines and 10 µM for 3B11 (p < 0.0001). These results suggest that bacopaside II is a potential anti-angiogenic agent.

[Effects of electroacupuncture preconditioning on expression of aquaporin-1 and activity of protein kinase C in myocardium of rats with acute myocardial ischemia-reperfusion injury].

OBJECTIVE: To explore the partial action mechanism and the myocardial protective effect differences between electroacupuncture (EA) preconditioning at "Neiguan"(PC 6) and "Taiyuan"(LU 9) in rats with acute myocardial ischemia-reperfusion injury. METHODS: Ninety-six Wistar rats were randomly assigned into a sham-operation group, a model group, a Neiguan group and a Taiyuan group, 24 rats in each one. The rats in the Neiguan group and Taiyuan group were treated with EA (2 Hz in frequency, 1 mA in intensity) at "Neiguan" (PC 6) and "Taiyuan" (LU 9) respectively, 20 min per treatment, once a day for consecutive 7 days. The rats in the sham-operation group and model group were treated with immobilization for the same time, and no EA was given. The model of myocardial ischemia-reperfusion injury was established in the model group, Neiguan group and Taiyuan group 24 h after the end of EA, while the rats in the sham-operation group were treated with sham operation (no ligation was made during surgery). The myocardial ischemic size, infarction size, activity of protein kinase C (PKC) and expression of aquaporin1 (AQP1) in each group were detected. RESULTS: Compared with sham-operation group, the myocardial ischemic size, infarction size, AQP1 expression and PKC activity in the model group were significantly increased (all P<0.01); compared with the model group and Taiyuan group, the myocardial ischemic size, infarction size, PKC activity and AQP1 expression were significantly decreased in the Neiguan group (P<0.01, P<0.05). By Pearson correlation analysis, the changes of AQP1 expression were positively correlated with those of PKC activity after EA preconditioning. CONCLUSIONS: EA preconditioning at "Neiguan" (PC 6) could significantly decrease myocardial AQP1 expression and PKC activity in rats with acute myocardial ischemia-reperfusion injuing, but the effect of EA preconditioning at "Taiyuan"(LU 9) is not obvious; its protective effect is likely to be achieved by inhibiting PKC activity and AQP1 expression.

Constitutively-stressed yeast strains are high-yielding for recombinant Fps1: implications for the translational regulation of an aquaporin.

BACKGROUND: We previously selected four strains of Saccharomyces cerevisiae for their ability to produce the aquaporin Fps1 in sufficient yield for further study. Yields from the yeast strains spt3Δ, srb5Δ, gcn5Δ and yTHCBMS1 (supplemented with 0.5 µg/mL doxycycline) that had been transformed with an expression plasmid containing 249 base pairs of 5' untranslated region (UTR) in addition to the primary FPS1 open reading frame (ORF) were 10-80 times higher than yields from wild-type cells expressing the same plasmid. One of the strains increased recombinant yields of the G protein-coupled receptor adenosine receptor 2a (A2aR) and soluble green fluorescent protein (GFP). The specific molecular mechanisms underpinning a high-yielding Fps1 phenotype remained incompletely described. RESULTS: Polysome profiling experiments were used to analyze the translational state of spt3Δ, srb5Δ, gcn5Δ and yTHCBMS1 (supplemented with 0.5 µg/mL doxycycline); all but gcn5Δ were found to exhibit a clear block in translation initiation. Four additional strains with known initiation blocks (rpl31aΔ, rpl22aΔ, ssf1Δ and nop1Δ) also improved the yield of recombinant Fps1 compared to wild-type. Expression of the eukaryotic transcriptional activator GCN4 was increased in spt3Δ, srb5Δ, gcn5Δ and yTHCBMS1 (supplemented with 0.5 µg/mL doxycycline); these four strains also exhibited constitutive phosphorylation of the eukaryotic initiation factor, eIF2α. Both responses are indicative of a constitutively-stressed phenotype. Investigation of the 5'UTR of FPS1 in the expression construct revealed two untranslated ORFs (uORF1 and uORF2) upstream of the primary ORF. Deletion of either uORF1 or uORF1 and uORF2 further improved recombinant yields in our four strains; the highest yields of the uORF deletions were obtained from wild-type cells. Frame-shifting the stop codon of the native uORF (uORF2) so that it extended into the FPS1 ORF did not substantially alter Fps1 yields in spt3Δ or wild-type cells, suggesting that high-yielding strains are able to bypass 5'uORFs in the FPS1 gene via leaky scanning, which is a known stress-response mechanism. Yields of recombinant A2aR, GFP and horseradish peroxidase could be improved in one or more of the yeast strains suggesting that a stressed phenotype may also be important in high-yielding cell factories. CONCLUSIONS: Regulation of Fps1 levels in yeast by translational control may be functionally important; the presence of a native uORF (uORF2) may be required to maintain low levels of Fps1 under normal conditions, but higher levels as part of a stress response. Constitutively-stressed yeast strains may be useful high-yielding microbial cell factories for recombinant protein production.

Protective effects of puerarin on acute lung and cerebrum injury induced by hypobaric hypoxia via the regulation of aquaporin (AQP) via NF-κB signaling pathway.

OBJECTIVE: Hypobaric hypoxia, frequently encountered at high altitude, may lead to lung and cerebrum injury. Our study aimed to investigate whether puerarin could exert ameliorative effects on rats exposed to hypobaric hypoxia via regulation of aquaporin (AQP) and NF-κB signaling pathway in lung and cerebrum. MATERIALS AND METHODS: 40 Sprague Dawley rats were divided into four groups (normal control group, hypobaric hypoxia group, puerarin group and dexamethasone group). Wet/dry ratio, blood gas, pathological changes of lung and cerebrum and spatial memory were observed in each group. Inflammatory cytokines in bronchoalveolar lavage fluid (BALF) were determined with ELISA and expression of AQP1, AQP4, NF-κB signaling pathway in lung and cerebrum with western blot RESULTS: Puerarin showed significant preventative effects on tissue injury and behavioral changes, as evidenced by histopathological findings and Morris water maze. In addition, levels of inflammatory cytokines in BALF decreased in the two preventative groups compared with those of hypobaric hypoxia group. AQP in lung and cerebrum increased under the condition of hypobaric hypoxia while was down regulated in both two preventative groups. NF-κB and IκB was also inhibited by puerarin. CONCLUSION: Our study suggested that lung and cerebrum injury, increased inflammatory cytokines in BALF and increased AQP1, AQP4 and NF-κB signaling pathway occurred under the condition of hypobaric hypoxia. Moreover, puerarin could prevent lung and cerebrum injury of rats exposed to hypobaric hypoxia via down-regulation of inflammatory cytokines, AQP1 and AQP4 expression and NF-κB signaling pathway.