Publications by authors named "Wen-Hui Wang"

176 Publications

Perianesthetic mortality in English Bulldogs: a retrospective analysis in 2010 - 2017.

BMC Vet Res 2022 May 25;18(1):198. Epub 2022 May 25.

Department of Molecular and Biomedical Sciences, College of Veterinary Medicine, North Carolina State University, Raleigh, NC, USA.

Background: Many veterinarians consider English Bulldogs to have a greater perianesthetic mortality risk. The aims of this study were to 1) determine total and anesthesia-related, perianesthetic mortality (PAM) rates in English Bulldogs (EB), 2) identify potential risk factors associated with mortality in EB, and 3) determine the difference in the perianesthetic mortality rates between EB, other-brachycephalic breeds (OB), and non-brachycephalic breeds (NB). Records from EB that were anesthetized between 2010 and 2017, were investigated. OB and NB were enrolled to match with each EB based on a procedure and age from the study period. Data collected in EB included: age, ASA status, weight, procedure types, anesthetic and analgesic management, anesthetic duration, anesthetic recovery location, and cause of death. Age and cause of death were determined from OB and NB. Fisher's exact test was used to compare PAM rate and age in EB, OB, and NB. Mann-Whitney U test was used to compare EB survivor and EB non-survivor. Logistic regression models were used to identify factors and odds ratio (OR) associated with PAM in EB.

Result: Two hundred twenty nine EB, 218 OB, and 229 NB were identified. The total and anesthesia-related PAM rates in EB were 6.6 and 3.9%, respectively. EB had a greater total PAM rate compared with OB (p = 0.007). ASA status was different between survivors and non-survivors in EB (p < 0.01). Risk factors identified regardless of the cause of death were premedication with full μ opioids (OR = 0.333, p = 0.114), continuous infusion of ketamine post-operatively (OR = 13.775, p = 0.013), and acepromazine administration post-operatively (OR = 7.274, p = 0.004). The most common cause of death in EB was postoperative respiratory dysfunction (87.5%).

Conclusion: Total and anesthesia-related mortality in EB is considerable. Most deaths in EB occurred during the postoperative period secondary to respiratory complications.
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http://dx.doi.org/10.1186/s12917-022-03301-9DOI Listing
May 2022

Infection Restrain IgA, IgG, and IgM Cells Residence in Sheep () Small Intestine.

Front Vet Sci 2022 28;9:878467. Epub 2022 Apr 28.

College of Veterinary Medicine, Gansu Agricultural University, Lanzhou, China.

Secreted immunoglobulin A (SIgA), IgG, and IgM play a crucial role in forming the intestinal mucosal immune barrier, and parasites could disturb the host's immune response by releasing various immunomodulatory molecules. is an important pathogen parasitizing in the sheep small intestine. It is aimed to explore the residence characteristics of IgA, IgG, and IgM cells in the sheep small intestine, and the influence of infection on them. Control group ( = 6) and infected group ( = 6) were selected, respectively, and the three subtype cells residing in the small intestine were systematically observed and analyzed. The results showed that in the Control group, the three types of positive cells were all distributed diffusely, and the total densities in jejunum, duodenum and ileum was gradually declined in turn. Notably, the change trend of IgA and IgG cells densities were both congruent with the total densities, and the differences among them were significant, respectively ( < 0.05); the IgM cells density was the highest in duodenum, followed by jejunum and ileum, there was no significant difference between duodenum and jejunum ( > 0.05), but both significantly higher than in ileum ( < 0.05). In the Infected group, their total densities in duodenum, jejunum and ileum were gradually declined in turn. Notably, the IgA and IgM cells densities change trend was the same as the total densities, and the differences among them were significant, respectively ( < 0.05). The IgG cells density in duodenum was the highest, followed by ileum and jejunum and there was significantly difference among them ( < 0.05). The comparison results between Control and Infected groups showed that from the duodenum, jejunum to ileum, IgA, IgG, and IgM cells were all reduced significantly, respectively. The results suggest that the three types of positive cells were resided heterogeneously in the small intestinal mucosa, that is, significant region-specificity; infection could not change their diffuse distribution characteristics, but strikingly, reduce their resident densities, and the forming mucosal immune barrier were significantly inhibited. It provided powerful evidence for studying on the molecular mechanism of evasion from immune surveillance by strongly inhibiting the host's mucosal immune barrier.
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http://dx.doi.org/10.3389/fvets.2022.878467DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9096708PMC
April 2022

Mineralocorticoid Receptor Antagonists Cause Natriuresis in the Absence of Aldosterone.

Hypertension 2022 May 4:101161HYPERTENSIONAHA12219159. Epub 2022 May 4.

Division of Nephrology and Hypertension, Department of Medicine, Oregon Health and Science University, Portland (Y.M., X.-T.S., A.S., J.A.M., D.H.E.).

Background: MR (mineralocorticoid receptor) antagonists are recommended for patients with resistant hypertension even when circulating aldosterone levels are not high. Although aldosterone activates MR to increase epithelial sodium channel (ENaC) activity, glucocorticoids also activate MR but are metabolized by 11βHSD2 (11β-hydroxysteroid dehydrogenase type 2). 11βHSD2 is expressed at increasing levels from distal convoluted tubule (DCT) through collecting duct. Here, we hypothesized that MR maintains ENaC activity in the DCT2 and early connecting tubule in the absence of aldosterone.

Methods: We studied AS (aldosterone synthase)-deficient (AS) mice, which were backcrossed onto the same C57Bl6/J strain as kidney-specific MR knockout (KS-MR) mice. KS-MR mice were used to compare MR expression and ENaC localization and cleavage with AS mice.

Results: MR was highly expressed along DCT2 through the cortical collecting duct (CCD), whereas no 11βHSD2 expression was observed along DCT2. MR signal and apical ENaC localization were clearly reduced along both DCT2 and CCD in KS-MR mice but were fully preserved along DCT2 and were partially reduced along CCD in AS mice. Apical ENaC localization and ENaC currents were fully preserved along DCT2 in AS mice and were not increased along CCD after low salt. AS mice exhibited transient Na wasting under low-salt diet, but administration of the MR antagonist eplerenone to AS mice led to hyperkalemia and decreased body weight with higher Na excretion, mimicking the phenotype of MR mice.

Conclusions: Our results provide evidence that MR is activated in the absence of aldosterone along DCT2 and partially CCD, suggesting glucocorticoid binding to MR preserves sodium homeostasis along DCT2 in AS mice.
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http://dx.doi.org/10.1161/HYPERTENSIONAHA.122.19159DOI Listing
May 2022

The long β2,3-sheets encoded by redundant sequences play an integral role in the channel function of P2X7 receptors.

J Biol Chem 2022 Apr 30:102002. Epub 2022 Apr 30.

School of Basic Medicine and Clinical Pharmacy and State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009, China. Electronic address:

P2X receptors are a class of non-selective cation channels widely distributed in the immune and nervous systems, and their dysfunction is a significant cause of tumors, inflammation, leukemia, and immune diseases. P2X7 is a unique member of the P2X receptor family with many properties that differ from other subtypes in terms of primary sequence, the architecture of N- and C-terminals, and channel function. Here, we suggest that the observed lengthened β2- and β3-sheets and their linker (loop β2,3), encoded by redundant sequences, play an indispensable role in the activation of the P2X7 receptor. We show that deletion of this longer structural element leads to the loss of P2X7 function. Furthermore, by combining mutagenesis, chimera construction, surface expression, and protein stability analysis, we found that the deletion of the longer β2,3-loop affects P2X7 surface expression, but more importantly, that this loop affects channel gating of P2X7. We propose that the longer β2,3-sheets may have a negative regulatory effect on a loop on the head domain and on the structural element formed by E171 and its surrounding regions. Understanding the role of the unique structure of the P2X7 receptor in the gating process will aid in the development of selective drugs targeting this subtype.
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http://dx.doi.org/10.1016/j.jbc.2022.102002DOI Listing
April 2022

P2X3-selective mechanism of Gefapixant, a drug candidate for the treatment of refractory chronic cough.

Comput Struct Biotechnol J 2022 31;20:1642-1653. Epub 2022 Mar 31.

School of Basic Medicine and Clinical Pharmacy and State Key Laboratory of Natural Medicines, China Pharmaceutical University, Nanjing 210009, China.

Gefapixant/AF-219, a selective inhibitor of the P2X3 receptor, is the first new drug other than dextromethorphan to be approved for the treatment of refractory chronic cough (RCC) in nearly 60 years. To date, seven P2X subtypes (P2X1-7) activated by extracellular ATP have been cloned, and subtype selectivity of P2X inhibitors is a prerequisite for reducing side effects. We previously identified the site and mechanism of action of Gefapixant/AF-219 on the P2X3 receptor, which occupies a pocket consisting of the left flipper (LF) and lower body (LB) domains. However, the mechanism by which AF-219 selectively acts on the P2X3 receptor is unknown. Here, we combined mutagenesis, chimera construction, molecular simulations, covalent occupation and chemical synthesis, and find that the negative allosteric site of AF-219 at P2X3 is also present in other P2X subtypes, at least for P2X1, P2X2 and P2X4. By constructing each chimera of AF-219 sensitive P2X3 and insensitive P2X2 subtypes, the insensitive P2X2 subtype was made to acquire the inhibitory properties of AF-219 and AF-353, an analog of AF-219 with higher affinity. Our results suggest that the selectivity of AF-219/AF-353 for P2X3 over the other P2X subtypes is determined by a combination of the accessibility of P2X3 binding site and the internal shape of this pocket, a finding that could provide new perspectives for drug design against P2X3-mediated diseases such as RCC, idiopathic pulmonary fibrosis, hypertension and overactive bladder disorder.
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http://dx.doi.org/10.1016/j.csbj.2022.03.030DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9014320PMC
March 2022

Moniezia benedeni infection enhances neuromedin U (NMU) expression in sheep (Ovis aries) small intestine.

BMC Vet Res 2022 Apr 19;18(1):143. Epub 2022 Apr 19.

College of Veterinary Medicine, Gansu Agricultural University, Lanzhou, 730070, Gansu, China.

Background: Neuromedin U (NMU) plays an important role in activating the group 2 innate lymphoid cells (ILC2s) and initiating the host's anti-parasitic immune responses. It is aimed to explore the distribution characteristics of NMU in the sheep small intestine and the influence of Moniezia benedeni infection on them. In the present study, the pET-28a-NMU recombinant plasmids were constructed, and Escherichia coli. BL21 (DE3) were induced to express the recombinant protein. And then, the rabbit anti-sheep NMU polyclonal antibody was prepared and immunofluorescence staining was performed with it. The expression levels of NMU in the intestine of normal and Moniezia benedeni-infected sheep were detected by ELISA.

Results: The results showed that the molecular weight of the obtained NMU recombinant protein was consistent with the expected molecular (13 kDa) and it was expressed in the form of inclusion body. The titer and specificity of obtained rabbit anti-sheep NMU polyclonal antibody were good. The results of immunofluorescence analysis showed that the nerve fibers which specifically expressed NMU mainly extended from the ganglion in the submucosal to lamina propria (LP) in the sheep small intestine, and the expression level was relatively high; especially on the nerve fibers of LP around the intestinal glands. The expression levels were gradually increased from the duodenum to the ileum, and the levels in the jejunum and ileum were significantly higher than that in the duodenum (P < 0.05). In addition, scattered NMU positive cells were distributed in the epithelium of the jejunal crypts. Moniezia benedeni infection increased the expression of NMU in each intestinal segment, especially in the jejunum and ileum there were significant increase (P < 0.05).

Conclusions: It was suggested that Moniezia benedeni infection could be detected by the high expression of NMU in sheep enteric nervous, and which laid the foundation for further studies on whether NMU exerts anti-parasitic immunity by activating ILC2s. In addition, NMU was expressed in some intestinal gland epitheliums, which also provided a basis for studying its roles in regulation of the immune homeostasis. The present study laid the foundation for further revealing the molecular mechanism of sheep's neural-immune interaction network perceiving the colacobiosis of parasites.
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http://dx.doi.org/10.1186/s12917-022-03243-2DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC9016964PMC
April 2022

Expression characteristics of polymeric immunoglobulin receptor in Bactrian camel (Camelus bactrianus) lungs.

PLoS One 2022 4;17(3):e0264815. Epub 2022 Mar 4.

College of Veterinary Medicine, Gansu Agricultural University, Lanzhou, Gansu, China.

Polymeric immunoglobulin receptor (pIgR), the transmembrane transporter of polymeric immunoglobulin A and M, has multiple immune functions. To explore the characteristics of pIgR expression in Bactrian camel lungs, twelve healthy adult (2-7 years old) Bactrian camels were systematically studied. The results showed that pIgR was mainly expressed in the cytoplasm and membrane of ciliated cells, as well as in the cytoplasm and membrane of basal cells, serous cells of bronchial glands, club cells and alveolar type 2 cells in Bactrian camel lungs. Specially, as the bronchial branches extended, the pIgR expression level in ciliated cells significantly declined (p<0.05), and the corresponding bronchial luminal areas obviously decreased (p<0.05). However, pIgR was not expressed in goblet cells, endocrine cells, alveolar type 1 cells and mucous cells of bronchial glands. The results demonstrated that ciliated cells continuously distributed throughout the whole bronchial tree mucosa were the major expression sites of pIgR, and pIgR was also expressed in basal cells, serous cells of bronchial glands, club cells and alveolar type 2 cells, which would facilitate secretory immunoglobulin A (SIgA) transmembrane transport by pIgR and form an intact protective barrier. Moreover, the pIgR expression level in ciliated cells was positively correlated with the bronchial luminal areas; but negatively correlated with the cleanliness of airflow through the bronchial cross-sections, showing that the pIgR expression level in the bronchial epithelium was inhomogeneous. Our study provided a foundation for further exploring the regulatory functions of immunoglobulins (i.e., SIgA) after transport across the membrane by pIgR in Bactrian camel lungs.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0264815PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8896721PMC
April 2022

Thymopentin-Mediated Inhibition of Cancer Stem Cell Stemness Enhances the Cytotoxic Effect of Oxaliplatin on Colon Cancer Cells.

Front Pharmacol 2022 15;13:779715. Epub 2022 Feb 15.

Interventional Cancer Institute of Chinese Integrative Medicine, Putuo Hospital, Shanghai University of Traditional Chinese Medicine, Shanghai, China.

Thymopentin (TP5) is an immunomodulatory pentapeptide that has been widely used in malignancy patients with immunodeficiency due to radiotherapy and chemotherapy. Here, we propose that TP5 directly inhibits the stemness of colon cancer cells HCT116 and therefore enhances the cytotoxicity of oxaliplatin (OXA) in HCT116 cells. In the absence of serum, TP5 was able to induce cancer stemness reduction in cultured HCT116 cells and significantly reduced stemness-related signals, such as the expression of surface molecular markers (CD133, CD44 and CD24) and stemness-related genes (ALDH1, SOX2, Oct-4 and Nanog), and resulted in altered Wnt/β-catenin signaling. Acetylcholine receptors (AchRs) are implicated in this process. OXA is a common chemotherapeutic agent with therapeutic effects in various cancers. Although TP5 had no direct effect on the proliferation of HCT116, this pentapeptide significantly increased the sensitivity of HCT116 to OXA, where the effect of TP5 on the stemness of colon cancer cells through stimulation of AchRs may contribute to this process. Our results provide a promising strategy for increasing the sensitivity of colon cancer cells to chemotherapeutic agents by incorporating immunomodulatory peptides.
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http://dx.doi.org/10.3389/fphar.2022.779715DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8886222PMC
February 2022

A novel effect of PDLIM5 in α7 nicotinic acetylcholine receptor upregulation and surface expression.

Cell Mol Life Sci 2022 Jan 10;79(1):64. Epub 2022 Jan 10.

Neurobiology Research Center, School of Medicine, Shenzhen Campus of Sun Yat-Sen University, No. 66, Gongchang Road, Guangming District, Shenzhen, 518107, Guangdong, People's Republic of China.

Nicotinic acetylcholine receptors (nAChRs) are widespread throughout the central nervous system. Signaling through nAChRs contributes to numerous higher-order functions, including memory and cognition, as well as abnormalities such as nicotine addiction and neurodegenerative disorders. Although recent studies indicate that the PDZ-containing proteins comprising PSD-95 family co-localize with nicotinic acetylcholine receptors and mediate downstream signaling in the neurons, the mechanisms by which α7nAChRs are regulated remain unclear. Here, we show that the PDZ-LIM domain family protein PDLIM5 binds to α7nAChRs and plays a role in nicotine-induced α7nAChRs upregulation and surface expression. We find that chronic exposure to 1 μM nicotine upregulated α7, β2-contained nAChRs and PDLIM5 in cultured hippocampal neurons, and the upregulation of α7nAChRs and PDLIM5 is increased more on the cell membrane than the cytoplasm. Interestingly, in primary hippocampal neurons, α7nAChRs and β2nAChRs display distinct patterns of expression, with α7nAChRs colocalized more with PDLIM5. Furthermore, PDLIM5 interacts with α7nAChRs, but not β2nAChRs in native brain neurons. Knocking down of PDLIM5 in SH-SY5Y abolishes nicotine-induced upregulation of α7nAChRs. In primary hippocampal neurons, using shRNA against PDLIM5 decreased both surface clustering of α7nAChRs and α7nAChRs-mediated currents. Proteomics analysis and isothermal titration calorimetry (ITC) results show that PDLIM5 interacts with α7nAChRs through the PDZ domain, and the interaction between PDLIM5 and α7nAChRs can be promoted by nicotine. Collectively, our data suggest a novel cellular role of PDLIM5 in the regulation of α7nAChRs, which may be relevant to plastic changes in the nervous system.
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http://dx.doi.org/10.1007/s00018-021-04115-yDOI Listing
January 2022

[Hydrochemical Characteristics and Controlling Factors of Surface Water and Groundwater in Wuding River Basin].

Huan Jing Ke Xue 2022 Jan;43(1):220-229

College of Natural Resources and Environment, Northwest A&F University, Yangling 712100, China.

Taking Wuding River as the research object, the study explored the hydrochemical characteristics and discussed the source of solute and control factors of groundwater and surface water in the basin, in order to provide a reference for water quality management. Considering the seasonal differences, water samples were collected during the dry season and the flood season. By comprehensively using graphic methods, correlation analysis, and forward deduction models, we analyzed the temporal and spatial evolution characteristics of water chemistry, explored the formation mechanism of water chemistry, and quantified the contribution rates of different sources to solutes. The results showed that the overall water quality was weakly alkaline in Wuding River basin. HCO and Na were the main anions and cations in the water, respectively, and the main water chemistry type was HCO·SO-Na·Ca. The water quality gradually deteriorated along the river course from west to east, and the sampling points that exceeded the level Ⅲ water were mainly distributed in tributaries during the dry season and downstream during the flood season. The cation exchange effect increased the Na and K in the water, and NO and HCO differed significantly in different seasons, which may be affected by seasonal precipitation leaching soil and land use types. Evaporite weathering and silicate weathering were the main sources of solute contribution in the Wuding River basin, which were 35.0% and 46.5% in the dry season and 46.7% and 42.3% in the flood season,respectively.
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http://dx.doi.org/10.13227/j.hjkx.202103039DOI Listing
January 2022

[Global centennial research and the internationalization of acupuncture and moxibustion: in the perspective of SCI database (1921-2020)].

Zhongguo Zhen Jiu 2021 Dec;41(12):1410-4

School of Acupuncture-Moxibustion and Tuina, Changchun University of CM, Changchun 130117, Jilin Province, China.

The paper retrieves and analyzes SCI articles on acupuncture-moxibustion published in the world from 1921 to 2020. It is found that the overall growth of SCI articles on acupuncture-moxibustion in both China and global countries is increasing, and the proportion of publication amount in China is increased gradually. It is believed that the articles on acupuncture-moxibustion researches from 1921 to 2020 in the world collected in SCI database indicate three stages, i.e. scattered publication, internationalization and great contribution on acupuncture-moxibustion in TCM. The paper investigates the first SCI article on acupuncture-moxibustion in the world and in China respectively and analyzes the main disciplines, research institutions and journal distribution, as well as the highly cited articles in the global countries. It is proposed that acupuncture-moxibustion research in China should reflect the academic ideological characteristics of acupuncture-moxibustion in TCM, develop the interdisciplinary research and deepen the cooperation with high-level scientific institutions so as to improve the international academic influence of acupuncture-moxibustion in TCM.
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http://dx.doi.org/10.13703/j.0255-2930.20210506-0003DOI Listing
December 2021

ROMK channels are inhibited in the aldosterone-sensitive distal nephron of renal tubule Nedd4-2-deficient mice.

Am J Physiol Renal Physiol 2022 01 29;322(1):F55-F67. Epub 2021 Nov 29.

Department of Pharmacology, New York Medical College, Valhalla, New York.

We used whole cell recording to examine the renal outer medullary K channel (ROMK or Kir1.1) and epithelial Na channel (ENaC) in the late distal convoluted tubule (DCT2)/initial connecting tubule (CNT) and in the cortical collecting duct (CCD) of kidney tubule-specific neural precursor cell-expressed developmentally downregulated protein 4-2 (Nedd4-2) knockout mice (Ks-Nedd4-2 KO) and floxed neural precursor cell-expressed developmentally downregulated 4-like () mice (control). Tertiapin Q (TPNQ)-sensitive K currents (ROMK) were smaller in both the DCT2/CNT and CCD of Ks-Nedd4-2 KO mice on a normal diet than in control mice. Neither high dietary salt intake nor low dietary salt intake had a significant effect on ROMK activity in the DCT2/CNT and CCD of control and Ks-Nedd4-2 KO mice. In contrast, high dietary K intake (HK) increased, whereas low dietary K intake (LK) decreased TPNQ-sensitive K currents in floxed mice. However, the effects of dietary K intake on ROMK channel activity were absent in Ks-Nedd4-2 KO mice since neither HK nor LK significantly affected TPNQ-sensitive K currents in the DCT2/CNT and CCD. Moreover, TPNQ-sensitive K currents in the DCT2/CNT and CCD of Ks-Nedd4-2 KO mice on HK were similar to those of control mice on LK. Amiloride-sensitive Na currents in the DCT2/CNT and CCD were significantly higher in Ks-Nedd4-2 KO mice than in floxed mice on a normal K diet. HK increased ENaC activity of the DCT2/CNT only in control mice, but HK stimulated ENaC of the CCD in both control and Ks-Nedd4-2 KO mice. Moreover, the HK-induced increase in amiloride-sensitive Na currents was larger in Ks-Nedd4-2 KO mice than in control mice. Deletion of Nedd4-2 increased with no lysine kinase 1 expression and abolished HK-induced inhibition of with no lysine kinase 1. We conclude that deletion of Nedd4-2 increases ENaC activity but decreases ROMK activity in the aldosterone-sensitive distal nephron and that HK fails to stimulate ROMK, but robustly increases ENaC activity in the CCD of Nedd4-2-deficient mice. We demonstrate that renal outer medullary K (ROMK) channel activity is inhibited in the late distal convoluted tubule/initial connecting tubule and cortical collecting duct of neural precursor cell-expressed developmentally downregulated protein 4-2 (Nedd4-2)-deficient mice. Also, deletion of Nedd4-2 abolishes the stimulatory effect of dietary K intake on ROMK. The lack of high K-induced stimulation of ROMK is associated with the absence of high K-induced inhibition of with no lysine kinase 1.
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http://dx.doi.org/10.1152/ajprenal.00306.2021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8714254PMC
January 2022

Biosynthesis and evaluation of a novel highly water-soluble quercetin glycoside derivative.

J Asian Nat Prod Res 2021 Oct 14:1-7. Epub 2021 Oct 14.

School of Food Science and Pharmaceutical Engineering, Nanjing Normal University, Nanjing 210023, China.

Quercetin () was converted into quercetin 7--succinyl glucoside () by used FJ18 as a solvent-resistant whole-cell biocatalyst. The structure of the new compound was confirmed by LC-MS analysis and NMR spectroscopy. The water-solubility of this novel quercetin 7--succinyl glucoside () was approximately 1000 times higher than that of native quercetin (). Quercetin () and quercetin 7--succinyl glucoside () exhibited significant DPPH scavenging capacity with IC values of 23.55 and 36.05 µM, respectively. Both compounds showed moderate cytotoxic effects against the two human cancer cell lines (MCF-7 and HepG2) with IC values ranging from 39.45-63.38 µM.
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http://dx.doi.org/10.1080/10286020.2021.1981875DOI Listing
October 2021

GSK1702934A and M085 directly activate TRPC6 via a mechanism of stimulating the extracellular cavity formed by the pore helix and transmembrane helix S6.

J Biol Chem 2021 10 28;297(4):101125. Epub 2021 Aug 28.

Department of Basic Medicine and Clinical Pharmacy, China Pharmaceutical University, Nanjing, China. Electronic address:

Transient receptor potential canonical (TRPC) channels, as important membrane proteins regulating intracellular calcium (Ca) signaling, are involved in a variety of physiological and pathological processes. Activation and regulation of TRPC are more dependent on membrane or intracellular signals. However, how extracellular signals regulate TRPC6 function remains to be further investigated. Here, we suggest that two distinct small molecules, M085 and GSK1702934A, directly activate TRPC6, both through a mechanism of stimulation of extracellular sites formed by the pore helix (PH) and transmembrane (TM) helix S6. In silico docking scanning of TRPC6 identified three extracellular sites that can bind small molecules, of which only mutations on residues of PH and S6 helix significantly reduced the apparent affinity of M085 and GSK1702934A and attenuated the maximal response of TRPC6 to these two chemicals by altering channel gating of TRPC6. Combing metadynamics, molecular dynamics simulations, and mutagenesis, we revealed that W679, E671, E672, and K675 in the PH and N701 and Y704 in the S6 helix constitute an orthosteric site for the recognition of these two agonists. The importance of this site was further confirmed by covalent modification of amino acid residing at the interface of the PH and S6 helix. Given that three structurally distinct agonists M085, GSK1702934A, and AM-0883, act at this site, as well as the occupancy of lipid molecules at this position found in other TRP subfamilies, it is suggested that the cavity formed by the PH and S6 has an important role in the regulation of TRP channel function by extracellular signals.
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http://dx.doi.org/10.1016/j.jbc.2021.101125DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8458982PMC
October 2021

Deletion of renal Nedd4-2 abolishes the effect of high K intake on Kir4.1/Kir5.1 and NCC activity in the distal convoluted tubule.

Am J Physiol Renal Physiol 2021 07 24;321(1):F1-F11. Epub 2021 May 24.

Department of Pharmacology, New York Medical College, Valhalla, New York.

High-dietary K (HK) intake inhibits basolateral Kir4.1/Kir5.1 activity in the distal convoluted tubule (DCT), and HK-induced inhibition of Kir4.1/Kir5.1 is essential for HK-induced inhibition of NaCl cotransporter (NCC). Here, we examined whether neural precursor cell expressed developmentally downregulated 4-2 (Nedd4-2) deletion compromises the effect of HK on basolateral Kir4.1/Kir5.1 and NCC in the DCT. Single-channel recording and whole cell recording showed that neither HK decreased nor low-dietary K (LK) increased basolateral Kir4.1/Kir5.1 activity of the DCT in kidney tubule-specific Nedd4-2 knockout (Ks-Nedd4-2 KO) mice. In contrast, HK inhibited and LK increased Kir4.1/Kir5.1 activity in control mice [neural precursor cell expressed developmentally downregulated 4-like ]. Also, HK intake decreased the negativity of K current reversal potential in the DCT (depolarization) only in control mice but not in Ks-Nedd4-2 KO mice. Renal clearance experiments showed that HK intake decreased, whereas LK intake increased, hydrochlorothiazide-induced renal Na excretion only in control mice, but this effect was absent in Ks-Nedd4-2 KO mice. Western blot analysis also demonstrated that HK-induced inhibition of phosphorylated NCC (Thr) and total NCC was observed only in control mice but not in Ks-Nedd4-2 KO mice. Furthermore, expression of all three subunits of the epithelial Na channel in Ks-Nedd4-2 KO mice on HK was higher than in control mice. Thus, plasma K concentrations were similar between and Ks-Nedd4-2 KO mice on HK for 7 days despite high NCC expression. We conclude that Nedd4-2 plays a role in regulating HK-induced inhibition of Kir4.1/Kir5.1 and NCC in the DCT. Basolateral Kir4.1/Kir5.1 in the distal convoluted tubule plays an important role as a "K sensor" in the regulation of renal K excretion after high K intake. We found that neural precursor cell expressed developmentally downregulated 4-2 (Nedd4-2) a role in mediating the effect of K diet on Kir4.1/Kir5.1 and NaCl cotransporter because high K intake failed to inhibit basolateral Kir4.1/Kir5.1 and NaCl cotransporter in kidney tubule-specific Nedd4-2 knockout mice.
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http://dx.doi.org/10.1152/ajprenal.00072.2021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8321800PMC
July 2021

Deletion of Kir5.1 abolishes the effect of high Na intake on Kir4.1 and Na-Cl cotransporter.

Am J Physiol Renal Physiol 2021 06 26;320(6):F1045-F1058. Epub 2021 Apr 26.

Department of Pharmacology, New York Medical College, Valhalla, New York.

High sodium (HS) intake inhibited epithelial Na channel (ENaC) in the aldosterone-sensitive distal nephron and Na-Cl cotransporter (NCC) by suppressing basolateral Kir4.1/Kir5.1 in the distal convoluted tubule (DCT), thereby increasing renal Na excretion but not affecting K excretion. The aim of the present study was to explore whether deletion of Kir5.1 compromises the inhibitory effect of HS on NCC expression/activity and renal K excretion. Patch-clamp experiments demonstrated that HS failed to inhibit DCT basolateral K channels and did not depolarize K current reversal potential of the DCT in Kir5.1 knockout (KO) mice. Moreover, deletion of Kir5.1 not only increased the expression of Kir4.1, phospho-NCC, and total NCC but also abolished the inhibitory effect of HS on the expression of Kir4.1, phospho-NCC, and total NCC and thiazide-induced natriuresis. Also, low sodium-induced stimulation of NCC expression/activity and basolateral K channels in the DCT were absent in Kir5.1 KO mice. Deletion of Kir5.1 decreased ENaC currents in the late DCT, and HS further inhibited ENaC activity in Kir5.1 KO mice. Finally, measurement of the basal renal K excretion rate with the modified renal clearance method demonstrated that long-term HS inhibited the renal K excretion rate and steadily increased plasma K levels in Kir5.1 KO mice but not in wild-type mice. We conclude that Kir5.1 plays an important role in mediating the effect of HS intake on basolateral K channels in the DCT and NCC activity/expression. Kir5.1 is involved in maintaining renal ability of K excretion during HS intake. Kir5.1 plays an important role in mediating the effect of high sodium intake on basolateral K channels in the distal convoluted tubule and Na-Cl cotransporter activity/expression.
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http://dx.doi.org/10.1152/ajprenal.00004.2021DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8285650PMC
June 2021

Protective Efficacy of Inactivated Vaccine against SARS-CoV-2 Infection in Mice and Non-Human Primates.

Virol Sin 2021 Oct 9;36(5):879-889. Epub 2021 Apr 9.

Center for Biosafety Mega-Science, Wuhan Institute of Virology, Chinese Academy of Sciences, Wuhan, 430071, China.

The ongoing coronavirus disease 2019 (COVID-19) pandemic caused more than 96 million infections and over 2 million deaths worldwide so far. However, there is no approved vaccine available for severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the disease causative agent. Vaccine is the most effective approach to eradicate a pathogen. The tests of safety and efficacy in animals are pivotal for developing a vaccine and before the vaccine is applied to human populations. Here we evaluated the safety, immunogenicity, and efficacy of an inactivated vaccine based on the whole viral particles in human ACE2 transgenic mouse and in non-human primates. Our data showed that the inactivated vaccine successfully induced SARS-CoV-2-specific neutralizing antibodies in mice and non-human primates, and subsequently provided partial (in low dose) or full (in high dose) protection of challenge in the tested animals. In addition, passive serum transferred from vaccine-immunized mice could also provide full protection from SARS-CoV-2 infection in mice. These results warranted positive outcomes in future clinical trials in humans.
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http://dx.doi.org/10.1007/s12250-021-00376-wDOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8034048PMC
October 2021

Deletion of renal Nedd4-2 abolishes the effect of high sodium intake (HS) on Kir4.1, ENaC, and NCC and causes hypokalemia during high HS.

Am J Physiol Renal Physiol 2021 05 5;320(5):F883-F896. Epub 2021 Apr 5.

Department of Pharmacology, New York Medical College, Valhalla, New York.

Neural precursor cell expressed developmentally downregulated protein 4-2 (Nedd4-2) regulates the expression of Kir4.1, thiazide-sensitive NaCl cotransporter (NCC), and epithelial Na channel (ENaC) in the aldosterone-sensitive distal nephron (ASDN), and Nedd4-2 deletion causes salt-sensitive hypertension. We now examined whether Nedd4-2 deletion compromises the effect of high-salt (HS) diet on Kir4.1, NCC, ENaC, and renal K excretion. Immunoblot analysis showed that HS diet decreased the expression of Kir4.1, Ca-activated large-conductance K channel subunit-α (BKα), ENaCβ, ENaCγ, total NCC, and phospho-NCC (at Thr) in floxed neural precursor cell expressed developmentally downregulated gene 4-like () mice, whereas these effects were absent in kidney-specific Nedd4-2 knockout (Ks-Nedd4-2 KO) mice. Renal clearance experiments also demonstrated that Nedd4-2 deletion abolished the inhibitory effect of HS diet on hydrochlorothiazide-induced natriuresis. Patch-clamp experiments showed that neither HS diet nor low-salt diet had an effect on Kir4.1/Kir5.1 currents of the distal convoluted tubule in Nedd4-2-deficient mice, whereas we confirmed that HS diet inhibited and low-salt diet increased Kir4.1/Kir5.1 activity in mice. Nedd4-2 deletion increased ENaC currents in the ASDN, and this increase was more robust in the cortical collecting duct than in the distal convoluted tubule. Also, HS-induced inhibition of ENaC currents in the ASDN was absent in Nedd4-2-deficient mice. Renal clearance experiments showed that HS intake for 2 wk increased the basal level of renal K excretion and caused hypokalemia in Ks-Nedd4-2-KO mice but not in mice. In contrast, plasma Na concentrations were similar in and Ks-Nedd4-2 KO mice on HS diet. We conclude that Nedd4-2 plays an important role in mediating the inhibitory effect of HS diet on Kir4.1, ENaC, and NCC and is essential for maintaining normal renal K excretion and plasma K ranges during long-term HS diet. The present study suggests that Nedd4-2 is involved in mediating the inhibitory effect of high salt (HS) diet on Kir4.1/kir5.1 in the distal convoluted tubule, NaCl cotransporter function, and epithelial Na channel activity and that Nedd4-2 plays an essential role in maintaining K homeostasis in response to a long-term HS diet. This suggests the possibility that HS intake could lead to hypokalemia in subjects lacking proper Nedd4-2 E3 ubiquitin ligase activity in aldosterone-sensitive distal nephron.
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http://dx.doi.org/10.1152/ajprenal.00555.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC8174810PMC
May 2021

Time to relapse after treatment withdrawal for different biologics used to treat plaque psoriasis.

Chin Med J (Engl) 2020 Nov 19;133(24):2998-3000. Epub 2020 Nov 19.

Department of Dermatology, Peking University Third Hospital, Beijing 100191, China.

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http://dx.doi.org/10.1097/CM9.0000000000001232DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7752671PMC
November 2020

Low toxicity and high immunogenicity of an inactivated vaccine candidate against COVID-19 in different animal models.

Emerg Microbes Infect 2020 Dec;9(1):2606-2618

National Institutes for Food and Drug Control, Beijing, People's Republic of China.

The ongoing COVID-19 pandemic is causing huge impact on health, life, and global economy, which is characterized by rapid spreading of SARS-CoV-2, high number of confirmed cases and a fatality/case rate worldwide reported by WHO. The most effective intervention measure will be to develop safe and effective vaccines to protect the population from the disease and limit the spread of the virus. An inactivated, whole virus vaccine candidate of SARS-CoV-2 has been developed by Wuhan Institute of Biological Products and Wuhan Institute of Virology. The low toxicity, immunogenicity, and immune persistence were investigated in preclinical studies using seven different species of animals. The results showed that the vaccine candidate was well tolerated and stimulated high levels of specific IgG and neutralizing antibodies. Low or no toxicity in three species of animals was also demonstrated in preclinical study of the vaccine candidate. Biochemical analysis of structural proteins and purity analysis were performed. The inactivated, whole virion vaccine was characterized with safe double-inactivation, no use of DNases and high purity. Dosages, boosting times, adjuvants, and immunization schedules were shown to be important for stimulating a strong humoral immune response in animals tested. Preliminary observation in ongoing phase I and II clinical trials of the vaccine candidate in Wuzhi County, Henan Province, showed that the vaccine is well tolerant. The results were characterized by very low proportion and low degree of side effects, high levels of neutralizing antibodies, and seroconversion. These results consistent with the results obtained from preclinical data on the safety.
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http://dx.doi.org/10.1080/22221751.2020.1852059DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7733911PMC
December 2020

Biochemical and antigenic characterization of the structural proteins and their post-translational modifications in purified SARS-CoV-2 virions of an inactivated vaccine candidate.

Emerg Microbes Infect 2020 Dec;9(1):2653-2662

Wuhan Institute of Biological Products, Co. Ltd, Wuhan, People's Republic of China.

In the face of COVID-19 pandemic caused by the newly emerged SARS-CoV-2, an inactivated, Vero cell-based, whole virion vaccine candidate has been developed and entered into phase III clinical trials within six months. Biochemical and immunogenic characterization of structural proteins and their post-translational modifications in virions, the end-products of the vaccine candidate, would be essential for the quality control and process development of vaccine products and for studying the immunogenicity and pathogenesis of SARS-CoV-2. By using a panel of rabbit antisera against virions and five structural proteins together with a convalescent serum, the spike (S) glycoprotein was shown to be N-linked glycosylated, PNGase F-sensitive, endoglycosidase H-resistant and cleaved by Furin-like proteases into S1 and S2 subunits. The full-length S and S1/S2 subunits could form homodimers/trimers. The membrane (M) protein was partially N-linked glycosylated; the accessory protein 3a existed in three different forms, indicative of cleavage and dimerization. Furthermore, analysis of the antigenicity of these proteins and their post-translationally modified forms demonstrated that S protein induced the strongest antibody response in both convalescent and immunized animal sera. Interestingly, immunization with the inactivated vaccine did not elicit antibody response against the S2 subunit, whereas strong antibody response against both S1 and S2 subunits was detected in the convalescent serum. Moreover, vaccination stimulated stronger antibody response against S multimers than did the natural infection. This study revealed that the native S glycoprotein stimulated neutralizing antibodies, while bacterially-expressed S fragments did not. The study on S modifications would facilitate design of S-based anti-SARS-CoV-2 vaccines.
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http://dx.doi.org/10.1080/22221751.2020.1855945DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7738289PMC
December 2020

Predictive value of serum VEGF, IL-1 and TNF-α in the treatment of thromboangiitis obliterans by revascularization.

Exp Ther Med 2020 Dec 16;20(6):232. Epub 2020 Oct 16.

The Department of Interventional Radiology of The First Hospital of Lanzhou University, Lanzhou, Gansu 730000, P.R. China.

Effect of revascularization in the treatment of thromboangiitis obliterans (TAO) and the predictive value of serum vascular endothelial growth factor (VEGF), interleukin-1 (IL-1) and tumor necrosis factor-α (TNF-α) of risk factors of amputation were investigated. From April 2012 to August 2015, a total of 117 patients with TAO admitted to the First Hospital of Lanzhou University were selected. Patients treated with revascularization combined with prostaglandin sodium and cilostazol were enrolled in group A (67 patients), and patients treated with sodium and cilostazol were enrolled in group B (50 patients). The clinical efficacy was evaluated by calculating the intermittent claudication distance and the ankle brachial index (ABI) of patients. The occurrence probability of nausea and vomiting, skin pruritus, abdominal pain, coagulation abnormalities and amputation were recorded. The concentration of serum VEGF, IL-1 and TNF-α were measured using enzyme-linked immunosorbent assay (ELISA). After treatment, the intermittent claudication distance, ABI and efficiency of group A was markedly higher than that of group B (P<0.05). After treatment, serum VEGF concentration in group A was clearly higher than that in group B (P<0.05), and IL-1 and TNF-α levels were much lower than those in group B (P<0.05). The amputation rate in group A was significantly lower than that in group B (P<0.05). Patients with amputation in both groups were enrolled in the study group (24 cases), and those without amputation were included in the control group (93 cases). The serum VEGF concentration in the study group before treatment was significantly lower than that in the control group (P<0.05), while IL-1 and TNF-α levels were significantly higher than those of the control group (P<0.05). In conclusion, pretreatment serum VEGF, IL-1 and TNF-α had a positive diagnostic value for poor prognosis of patients with amputation, and low concentration of VEGF and higher concentration of IL-1 and TNF-α are the risk factors for amputations in patients with TAO.
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http://dx.doi.org/10.3892/etm.2020.9362DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7604734PMC
December 2020

An angiosarcoma in the breast associated with Kasabach-Merritt syndrome: Breast Images.

Breast J 2021 01 13;27(1):62-64. Epub 2020 Oct 13.

Weifang People's Hospital, Oncology, Weifang, China.

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http://dx.doi.org/10.1111/tbj.14079DOI Listing
January 2021

Bacterial community analysis on the different mucosal immune inductive sites of gastrointestinal tract in Bactrian camels.

PLoS One 2020 8;15(10):e0239987. Epub 2020 Oct 8.

College of Veterinary Medicine, Gansu Agricultural University, Lanzhou, Gansu, China.

The microbial communities colonize the mucosal immune inductive sites could be captured by hosts, which could initiate the mucosal immune responses. The aggregated lymphoid nodule area (ALNA) and the ileal Payer's patches (PPs) in Bactrian camels are both the mucosal immune inductive sites of the gastrointestinal tract. Here, the bacteria community associated with the ALNA and ileal PPs were analyzed using of 16S rDNA-Illumina Miseq sequencing. The mutual dominant bacterial phyla at the two sites were the Bacteroidetes, Firmicutes, Verrucomicrobia and Proteobacteria, and the mutual dominant genus in both sits was Prevotella. The abundances of the Fibrobacter, Campylobacter and RFP12 were all higher in ALNA than in ileal PPs. While, the abundances of the 5-7N15, Clostridium, and Escherichia were all higher in ileal PPs than in ALNA. The results suggested that the host's intestinal microenvironment is selective for the symbiotic bacteria colonizing the corresponding sites, on the contrary, the symbiotic bacteria could impact on the physiological functions of this local site. In ALNA and ileal PPs of Bactrian camel, the bacteria which colonized different immune inductive sites have the potential to stimulate different immune responses, which is the result of the mutual selection and adaptation between microbial communities and their host.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0239987PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7544057PMC
December 2020

Efficacy and safety of wait and see strategy versus radical surgery and local excision for rectal cancer with cCR response after neoadjuvant chemoradiotherapy: a meta-analysis.

World J Surg Oncol 2020 Aug 31;18(1):232. Epub 2020 Aug 31.

Department of Medical Imaging, Cancer Hospital of China Medical University, Liaoning Cancer Hospital & Institute, No 44 Xiaoheyan Road, Dadong District, Shenyang, 110042, Liaoning Province, PR China.

Background: Neoadjuvant therapy can shrink tumors, increase anus preservation rate, and protect anal function. Radical surgery need cut off the diseased bowel, clean up the lymph nodes, and then restore bowel function. It could bring traumatic effect and poor postoperative quality of life to the patient. Local resection requires removal of the diseased bowel with circular negative margin. The surgical trauma is small, and the postoperative quality of life is good. In this meta-analysis, we aimed to evaluate the efficacy and safety between wait and see strategy (WS), radical surgery (RS), and local excision (LE) of rectal cancer patients with clinical complete response (cCR) response after neoadjuvant chemoradiotherapy.

Methods: We searched PubMed, Cochrane Library, CNKI (China National Knowledge Infrastructure), and Wanfang databases to compare wait and see strategy with radical surgery and local excision for rectal cancer with cCR response after neoadjuvant chemoradiotherapy up to March 2020. We collected the data of local recurrence, distant metastasis, cancer-related death, overall survival, and disease-free survival and used RevMan 5.0 to carry out the meta-analysis. Continuous data were evaluated by the standardized mean differences (SMD) with 95% confidence intervals (95% CIs), and dichotomous data were evaluated by relative risks (ORs or RRs) with 95% CIs. We aimed to compare the advantages and disadvantages of the three groups.

Results: Eleven English studies with 1131 patients were included. There were 412 patients in WS group, 678 patients in RS group, and 41 patients in LE group. WS group had a higher local recurrence rate than RS group (OR 7.32, 95% CI 3.58 to 14.95, P < 0.001). There was no significant difference in the other data between the three groups.

Conclusion: Compared with the RS group, the WS group had an increased risk of local recurrence. However, the WS group had a similar DFS and OS compared with the RS group and the local excision group. Hence, we speculated that the WS group would have similar results as the surgery group for patients with cCR status.
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http://dx.doi.org/10.1186/s12957-020-02003-6DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7457353PMC
August 2020

Distal convoluted tubule Cl concentration is modulated via K channels and transporters.

Am J Physiol Renal Physiol 2020 09 27;319(3):F534-F540. Epub 2020 Jul 27.

Division of Nephrology and Hypertension, School of Medicine, Oregon Health and Science University, Portland, Oregon.

Cl-sensitive with-no-lysine kinase (WNK) plays a key role in regulating the thiazide-sensitive Na-Cl cotransporter (NCC) in the distal convoluted tubule (DCT). Cl enters DCT cells through NCC and leaves the cell across the basolateral membrane via the Cl channel ClC-K2 or K-Cl cotransporter (KCC). While KCC is electroneutral, Cl exit via ClC-K2 is electrogenic. Therefore, an alteration in DCT basolateral K channel activity is expected to influence Cl movement across the basolateral membrane. Although a role for intracellular Cl in the regulation of WNK and NCC has been established, intracellular Cl concentrations ([Cl]) have not been directly measured in the mammalian DCT. Therefore, to measure [Cl] in DCT cells, we generated a transgenic mouse model expressing an optogenetic kidney-specific Cl-Sensor and measured Cl fluorescent imaging in the isolated DCT. Basal measurements indicated that the mean [Cl] was ~7 mM. Stimulation of Cl exit with low-Cl hypotonic solutions decreased [Cl], whereas inhibition of KCC by DIOA or inhibition of ClC-K2 by NPPB increased [Cl], suggesting roles for both KCC and ClC-K2 in the modulation of [Cl] . Blockade of basolateral K channels (Kir4.1/5.1) with barium significantly increased [Cl]. Finally, a decrease in extracellular K concentration transiently decreased [Cl], whereas raising extracellular K transiently increased [Cl], further suggesting a role for Kir4.1/5.1 in the regulation of [Cl]. We conclude that the alteration in ClC-K2, KCC, and Kir4.1/5.1 activity influences [Cl] in the DCT.
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http://dx.doi.org/10.1152/ajprenal.00284.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7509289PMC
September 2020

Epoxyeicosatrienoic acid metabolites inhibit Kir4.1/Kir5.1 in the distal convoluted tubule.

Am J Physiol Renal Physiol 2020 06 20;318(6):F1369-F1376. Epub 2020 Apr 20.

Department of Pharmacology, New York Medical College, Valhalla, New York.

Cytochrome -450 (Cyp) epoxygenase-dependent metabolites of arachidonic acid (AA) have been shown to inhibit renal Na transport, and inhibition of Cyp-epoxygenase is associated with salt-sensitive hypertension. We used the patch-clamp technique to examine whether Cyp-epoxygenase-dependent AA metabolites inhibited the basolateral 40-pS K channel (Kir4.1/Kir5.1) in the distal convoluted tubule (DCT). Application of AA inhibited the basolateral 40-pS K channel in the DCT. The inhibitory effect of AA on the 40-pS K channel was specific because neither linoleic nor oleic acid was able to mimic the effect of AA on the K channel. Inhibition of Cyp-monooxygenase with -methylsulfonyl-12,12-dibromododec-11-enamide or inhibition of cyclooxygenase with indomethacin failed to abolish the inhibitory effect of AA on the 40-pS K channel. However, the inhibition of Cyp-epoxygenase with -methylsulfonyl-6-(propargyloxyphenyl)hexanamide abolished the effect of AA on the 40-pS K channel in the DCT. Moreover, addition of either 11,12-epoxyeicosatrienoic acid (EET) or 14,15-EET also inhibited the 40-pS K channel in the DCT. Whole cell recording demonstrated that application of AA decreased, whereas -methylsulfonyl-6-(propargyloxyphenyl)hexanamide treatment increased, Ba-sensitive K currents in the DCT. Finally, application of 14,15-EET but not AA was able to inhibit the basolateral 40-pS K channel in the DCT of mice. We conclude that Cyp-epoxygenase-dependent AA metabolites inhibit the basolateral Kir4.1/Kir5.1 in the DCT and that Cyp2c44-epoxygenase plays a role in the regulation of the basolateral K channel in the mouse DCT.
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http://dx.doi.org/10.1152/ajprenal.00018.2020DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7311705PMC
June 2020

Renal Tubule Nedd4-2 Deficiency Stimulates Kir4.1/Kir5.1 and Thiazide-Sensitive NaCl Cotransporter in Distal Convoluted Tubule.

J Am Soc Nephrol 2020 06 15;31(6):1226-1242. Epub 2020 Apr 15.

Department of Pharmacology, New York Medical College, Valhalla, New York

Background: The potassium channel Kir4.1 forms the Kir4.1/Kir5.1 heterotetramer in the basolateral membrane of the distal convoluted tubule (DCT) and plays an important role in the regulation of the thiazide-sensitive NaCl cotransporter (NCC). Kidney-specific deletion of the ubiquitin ligase Nedd4-2 increases expression of NCC, and coexpression of Nedd4-2 inhibits Kir4.1/Kir5.1 . Whether Nedd4-2 regulates NCC expression in part by regulating Kir4.1/Kir5.1 channel activity in the DCT is unknown.

Methods: We used electrophysiology studies, immunoblotting, immunostaining, and renal clearance to examine Kir4.1/Kir5.1 activity in the DCT and NCC expression/activity in wild-type mice and mice with kidney-specific knockout of Nedd4-2, Kir4.1, or both.

Results: Deletion of Nedd4-2 increased the activity/expression of Kir4.1 in the DCT and also, hyperpolarized the DCT membrane. Expression of phosphorylated NCC/total NCC and thiazide-induced natriuresis were significantly increased in the Nedd4-2 knockout mice, but these mice were normokalemic. Double-knockout mice lacking both Kir4.1/Kir5.1 and Nedd4-2 in the kidney exhibited increased expression of the epithelial sodium channel -subunit, largely abolished basolateral potassium ion conductance (to a degree similar to that of kidney-specific Kir4.1 knockout mice), and depolarization of the DCT membrane. Compared with wild-type mice, the double-knockout mice displayed inhibited expression of phosphorylated NCC and total NCC and had significantly blunted thiazide-induced natriuresis as well as renal potassium wasting and hypokalemia. However, NCC expression/activity was higher in the double-knockout mice than in Kir4.1 knockout mice.

Conclusions: Nedd4-2 regulates Kir4.1/Kir5.1 expression/activity in the DCT and modulates NCC expression by Kir4.1-dependent and Kir4.1-independent mechanisms. Basolateral Kir4.1/Kir5.1 activity in the DCT partially accounts for the stimulation of NCC activity/expression induced by deletion of Nedd4-2.
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http://dx.doi.org/10.1681/ASN.2019090923DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7269345PMC
June 2020

Effect of Angiotensin II on ENaC in the Distal Convoluted Tubule and in the Cortical Collecting Duct of Mineralocorticoid Receptor Deficient Mice.

J Am Heart Assoc 2020 04 25;9(7):e014996. Epub 2020 Mar 25.

Department of Pharmacology New York Medical College Valhalla NY.

Background Angiotensin II stimulates epithelial Na channel (ENaC) by aldosterone-independent mechanism. We now test the effect of angiotensin II on ENaC in the distal convoluted tubule (DCT) and cortical collecting duct (CCD) of wild-type (WT) and kidney-specific mineralocorticoid receptor knockout mice (KS-MR-KO). Methods and Results We used electrophysiological, immunoblotting and renal-clearance methods to examine the effect of angiotensin II on ENaC in KS-MR-KO and wild-type mice. High K intake stimulated ENaC in the late DCT/early connecting tubule (DCT2/CNT) and in the CCD whereas low sodium intake stimulated ENaC in the CCD but not in the DCT2/CNT. The deletion of MR abolished the stimulatory effect of high K and low sodium intake on ENaC, partially inhibited ENaC in DCT2/CNT but almost abolished ENaC activity in the CCD. Application of losartan inhibited ENaC only in DCT2/CNT of both wild-type and KS-MR-KO mice but not in the CCD. Angiotensin II infusion for 3 days has a larger stimulatory effect on ENaC in the DCT2/CNT than in the CCD. Three lines of evidence indicate that angiotensin II can stimulate ENaC by MR-independent mechanism: (1) angiotensin II perfusion augmented ENaC expression in KS-MR-KO mice; (2) angiotensin II stimulated ENaC in the DCT2/CNT but to a lesser degree in the CCD in KS-MR-KO mice; (3) angiotensin II infusion augmented benzamil-induced natriuresis, increased the renal K excretion and corrected hyperkalemia of KS-MR-KO mice. Conclusions Angiotensin II-induced stimulation of ENaC occurs mainly in the DCT2/CNT and to a lesser degree in the CCD and MR plays a dominant role in determining ENaC activity in the CCD but to a lesser degree in the DCT2/CNT.
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http://dx.doi.org/10.1161/JAHA.119.014996DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7428622PMC
April 2020
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