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1.
Gastroenterology ; 2021 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-34780722

RESUMO

BACKGROUND: Polygenic and environmental factors are underlying causes of inflammatory bowel disease (IBD). We hypothesized that integration of the genetic loci controlling a metabolite's abundance, with known IBD genetic susceptibility loci, may help resolve metabolic drivers of IBD. METHODS: We measured the levels of 1300 metabolites in the serum of 484 ulcerative colitis (UC) and 464 Crohn's disease (CD) patients and 365 controls. Differential metabolite abundance was determined for disease status, subtype, clinical and endoscopic disease activity as well as IBD phenotype including disease behavior, location and extent. To inform on the genetic basis underlying metabolic diversity, we integrated metabolite and genomic data. Genetic colocalization and Mendelian randomization (MR) analyses were performed using known IBD risk loci to explore whether any metabolite was causally associated with IBD. RESULTS: We found 173 genetically controlled metabolites (mQTL, 9 novel) within 63 non-overlapping loci (7 novel). Furthermore, several metabolites significantly associated with IBD disease status and activity as defined by clinical and endoscopic indexes. This constitutes a resource for biomarker discovery and IBD biology insights. Using this resource, we show that a novel mQTL for serum butyrate levels containing ACADS was not supported as causal for IBD; replicate the association of serum omega-6 containing lipids with the fatty acid desaturase 1/2 locus and identify these metabolites as causal for CD through MR; and validate a novel association of serum plasmalogen and TMEM229B, which was predicted causal for CD. CONCLUSION: An exploratory analysis combining genetics and unbiased serum metabolome surveys can reveal novel biomarkers of disease activity and potential mediators of pathology in IBD.

2.
J Asian Nat Prod Res ; : 1-7, 2021 Oct 14.
Artigo em Inglês | MEDLINE | ID: mdl-34647847

RESUMO

Quercetin (1) was converted into quercetin 7-O-succinyl glucoside (2) by used Bacillus amyloliquefaciens 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-O-succinyl glucoside (2) was approximately 1000 times higher than that of native quercetin (2). Quercetin (1) and quercetin 7-O-succinyl glucoside (2) exhibited significant DPPH scavenging capacity with IC50 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 IC50 values ranging from 39.45-63.38 µM.

3.
J Biol Chem ; 297(4): 101125, 2021 10.
Artigo em Inglês | MEDLINE | ID: mdl-34461094

RESUMO

Transient receptor potential canonical (TRPC) channels, as important membrane proteins regulating intracellular calcium (Ca2+i) 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.


Assuntos
Sinalização do Cálcio , Ativação do Canal Iônico/efeitos dos fármacos , Simulação de Dinâmica Molecular , Canal de Cátion TRPC6/química , Canal de Cátion TRPC6/metabolismo , Células HEK293 , Humanos , Estrutura Secundária de Proteína , Canal de Cátion TRPC6/genética
4.
Am J Physiol Renal Physiol ; 321(1): F1-F11, 2021 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-34029145

RESUMO

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 (Nedd4l)flox/flox]. 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 (Thr53) 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 Nedd4lflox/flox 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.NEW & NOTEWORTHY 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.


Assuntos
Túbulos Renais Distais/metabolismo , Ubiquitina-Proteína Ligases Nedd4/deficiência , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Membro 3 da Família 12 de Carreador de Soluto/metabolismo , Animais , Transporte Biológico/fisiologia , Transporte de Íons/fisiologia , Camundongos , Camundongos Knockout , Técnicas de Patch-Clamp/métodos , Canais de Potássio Corretores do Fluxo de Internalização/genética , Membro 3 da Família 12 de Carreador de Soluto/genética
5.
Am J Physiol Renal Physiol ; 320(5): F883-F896, 2021 05 01.
Artigo em Inglês | MEDLINE | ID: mdl-33818128

RESUMO

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, Ca2+-activated large-conductance K+ channel subunit-α (BKα), ENaCß, ENaCγ, total NCC, and phospho-NCC (at Thr53) in floxed neural precursor cell expressed developmentally downregulated gene 4-like (Nedd4lfl/fl) 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 Nedd4lflox/flox 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 Nedd4lflox/flox mice. In contrast, plasma Na+ concentrations were similar in Nedd4lflox/flox 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.NEW & NOTEWORTHY 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.


Assuntos
Canais Epiteliais de Sódio/metabolismo , Hipopotassemia/etiologia , Ubiquitina-Proteína Ligases Nedd4/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Sódio na Dieta/efeitos adversos , Animais , Antibacterianos/farmacologia , Transporte Biológico , Doxiciclina/farmacologia , Canais Epiteliais de Sódio/genética , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Hipopotassemia/induzido quimicamente , Hipopotassemia/genética , Transporte de Íons/fisiologia , Camundongos , Camundongos Knockout , Ubiquitina-Proteína Ligases Nedd4/genética , Néfrons/metabolismo , Técnicas de Patch-Clamp , Potássio/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/genética , Sódio/metabolismo , Sódio na Dieta/administração & dosagem , Membro 3 da Família 12 de Carreador de Soluto/genética , Membro 3 da Família 12 de Carreador de Soluto/metabolismo
6.
Am J Physiol Renal Physiol ; 320(6): F1045-F1058, 2021 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-33900854

RESUMO

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. NEW & NOTEWORTHY 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.


Assuntos
Canais de Potássio Corretores do Fluxo de Internalização/metabolismo , Simportadores de Cloreto de Sódio/metabolismo , Sódio na Dieta/administração & dosagem , Sódio na Dieta/farmacologia , Animais , Feminino , Regulação da Expressão Gênica/efeitos dos fármacos , Túbulos Renais Distais/efeitos dos fármacos , Túbulos Renais Distais/metabolismo , Masculino , Camundongos , Camundongos Knockout , Neurônios , Técnicas de Patch-Clamp , Canais de Potássio Corretores do Fluxo de Internalização/genética , Simportadores de Cloreto de Sódio/genética
7.
Virol Sin ; 36(5): 879-889, 2021 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-33835391

RESUMO

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.


Assuntos
Vacinas contra COVID-19/imunologia , COVID-19 , Animais , Anticorpos Neutralizantes/sangue , Anticorpos Antivirais/sangue , COVID-19/prevenção & controle , Camundongos , Camundongos Transgênicos , Primatas , SARS-CoV-2 , Glicoproteína da Espícula de Coronavírus , Vacinas de Produtos Inativados/imunologia
11.
Exp Ther Med ; 20(6): 232, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33149786

RESUMO

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.

12.
Emerg Microbes Infect ; 9(1): 2653-2662, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33232205

RESUMO

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.


Assuntos
Vacinas contra COVID-19 , Processamento de Proteína Pós-Traducional , SARS-CoV-2/isolamento & purificação , Proteínas Estruturais Virais , Vírion , Animais , Antígenos Virais/análise , Antígenos Virais/metabolismo , Vacinas contra COVID-19/química , Vacinas contra COVID-19/imunologia , Bovinos , Chlorocebus aethiops , Humanos , Coelhos , SARS-CoV-2/imunologia , Vacinas de Produtos Inativados/química , Vacinas de Produtos Inativados/imunologia , Células Vero , Proteínas Estruturais Virais/química , Proteínas Estruturais Virais/imunologia , Proteínas Estruturais Virais/isolamento & purificação , Vírion/química , Vírion/imunologia , Vírion/isolamento & purificação
13.
Emerg Microbes Infect ; 9(1): 2606-2618, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-33241728

RESUMO

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.


Assuntos
Vacinas contra COVID-19/imunologia , SARS-CoV-2 , Animais , Anticorpos Antivirais , Vacinas contra COVID-19/efeitos adversos , Feminino , Imunidade Humoral , Masculino , Vacinas de Produtos Inativados/efeitos adversos , Vacinas de Produtos Inativados/imunologia
14.
PLoS One ; 15(10): e0239987, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-33031424

RESUMO

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.


Assuntos
Trato Gastrointestinal/microbiologia , Imunidade nas Mucosas , Tecido Linfoide/microbiologia , Microbiota , Animais , Bactérias/genética , Bactérias/isolamento & purificação , Bacteroidetes/genética , Bacteroidetes/isolamento & purificação , Biodiversidade , Camelus , Fibrobacter/genética , Fibrobacter/isolamento & purificação , Sequenciamento de Nucleotídeos em Larga Escala , Tecido Linfoide/imunologia , Análise de Componente Principal , RNA Ribossômico 16S/química , RNA Ribossômico 16S/metabolismo , Simbiose
15.
World J Surg Oncol ; 18(1): 232, 2020 Aug 31.
Artigo em Inglês | MEDLINE | ID: mdl-32862826

RESUMO

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.


Assuntos
Terapia Neoadjuvante , Neoplasias Retais , Quimiorradioterapia , China , Humanos , Recidiva Local de Neoplasia/terapia , Prognóstico , Qualidade de Vida , Neoplasias Retais/terapia , Resultado do Tratamento , Conduta Expectante
16.
Am J Physiol Renal Physiol ; 319(3): F534-F540, 2020 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-32715757

RESUMO

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-]i) have not been directly measured in the mammalian DCT. Therefore, to measure [Cl-]i 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-]i was ~7 mM. Stimulation of Cl- exit with low-Cl- hypotonic solutions decreased [Cl-]i, whereas inhibition of KCC by DIOA or inhibition of ClC-K2 by NPPB increased [Cl-]i, suggesting roles for both KCC and ClC-K2 in the modulation of [Cl-]i . Blockade of basolateral K+ channels (Kir4.1/5.1) with barium significantly increased [Cl-]i. Finally, a decrease in extracellular K+ concentration transiently decreased [Cl-]i, whereas raising extracellular K+ transiently increased [Cl-]i, further suggesting a role for Kir4.1/5.1 in the regulation of [Cl-]i. We conclude that the alteration in ClC-K2, KCC, and Kir4.1/5.1 activity influences [Cl-]i in the DCT.


Assuntos
Cloretos/metabolismo , Túbulos Renais Distais/fisiologia , Canais de Potássio/metabolismo , Simportadores de Cloreto de Sódio/metabolismo , Animais , Cloretos/química , Fenômenos Eletrofisiológicos , Camundongos , Imagem Molecular , Simportadores de Cloreto de Sódio/genética
17.
Am J Physiol Renal Physiol ; 318(6): F1369-F1376, 2020 06 01.
Artigo em Inglês | MEDLINE | ID: mdl-32308018

RESUMO

Cytochrome P-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 N-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 N-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 N-methylsulfonyl-6-(propargyloxyphenyl)hexanamide treatment increased, Ba2+-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 Cyp2c44-/- 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.


Assuntos
Ácido 8,11,14-Eicosatrienoico/análogos & derivados , Ácido Araquidônico/farmacologia , Família 2 do Citocromo P450/metabolismo , Túbulos Renais Distais/efeitos dos fármacos , Bloqueadores dos Canais de Potássio/farmacologia , Canais de Potássio Corretores do Fluxo de Internalização/antagonistas & inibidores , Ácido 8,11,14-Eicosatrienoico/metabolismo , Ácido 8,11,14-Eicosatrienoico/farmacologia , Amidas/farmacologia , Animais , Ácido Araquidônico/metabolismo , Família 2 do Citocromo P450/antagonistas & inibidores , Família 2 do Citocromo P450/genética , Inibidores Enzimáticos/farmacologia , Túbulos Renais Distais/metabolismo , Masculino , Potenciais da Membrana , Camundongos da Linhagem 129 , Camundongos Knockout , Bloqueadores dos Canais de Potássio/metabolismo , Canais de Potássio Corretores do Fluxo de Internalização/metabolismo
18.
J Am Soc Nephrol ; 31(6): 1226-1242, 2020 06.
Artigo em Inglês | MEDLINE | ID: mdl-32295826

RESUMO

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 in vitro. 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 α-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.


Assuntos
Túbulos Renais Distais/metabolismo , Ubiquitina-Proteína Ligases Nedd4/fisiologia , Canais de Potássio Corretores do Fluxo de Internalização/fisiologia , Simportadores de Cloreto de Sódio/fisiologia , Tiazidas/farmacologia , Animais , Canais Epiteliais de Sódio/fisiologia , Camundongos , Camundongos Knockout
19.
J Am Heart Assoc ; 9(7): e014996, 2020 04 07.
Artigo em Inglês | MEDLINE | ID: mdl-32208832

RESUMO

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.


Assuntos
Angiotensina II/farmacologia , Canais Epiteliais de Sódio/metabolismo , Túbulos Renais Coletores/efeitos dos fármacos , Túbulos Renais Distais/efeitos dos fármacos , Receptor Tipo 1 de Angiotensina/agonistas , Receptores de Mineralocorticoides/deficiência , Animais , Hiperpotassemia/tratamento farmacológico , Hiperpotassemia/genética , Hiperpotassemia/metabolismo , Hiperpotassemia/fisiopatologia , Túbulos Renais Coletores/metabolismo , Túbulos Renais Coletores/fisiopatologia , Túbulos Renais Distais/metabolismo , Túbulos Renais Distais/fisiopatologia , Potenciais da Membrana , Camundongos Knockout , Natriurese/efeitos dos fármacos , Potássio/urina , Receptor Tipo 1 de Angiotensina/metabolismo , Receptores de Mineralocorticoides/genética , Eliminação Renal/efeitos dos fármacos
20.
Hypertension ; 75(2): 439-448, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31865783

RESUMO

The inhibition of Type II angiotensin II receptor (AT2R) or BK2R (bradykinin type II receptor) stimulates basolateral Kir4.1/Kir5.1 in the distal convoluted tubule (DCT) and activates thiazide-sensitive NCC (Na-Cl cotransporter). The aim of the present study is to examine the role of AT2R and BK2R in mediating the effect of HK (high dietary K+) intake on the basolateral K+ channels, NCC, and renal K+ excretion. Feeding mice (male and female) with HK diet for overnight significantly decreased the basolateral K+ conductance, depolarized the DCT membrane, diminished the expression of pNCC (phosphorylated NCC) and tNCC (total NCC), and decreased thiazide-sensitive natriuresis. Overnight HK intake also increased the expression of cleaved ENaC-α and -γ subunits but had no effect on NKCC2 expression. Pretreatment of the mice (male and female) with PD123319 and HOE140 stimulated the expression of tNCC and pNCC, augmented hydrochlorothiazide-induced natriuresis, and increased the negativity of the DCT membrane. The deletion of Kir4.1 not only decreased the NCC activity but also abolished the stimulatory effect of PD123319 and HOE140 perfusion on NCC activity. Moreover, the effect of overnight HK loading on Kir4.1/Kir5.1 in the DCT and NCC expression/activity was compromised in the mice treated with AT2R/BK2R antagonists. Renal clearance study showed that inhibition of AT2R and BK2R impairs renal K+ excretion in response to overnight HK loading, and the mice pretreated with PD123319 and HOE140 were hyperkalemic during HK intake. We conclude that synergistic activation of AT2R and BK2R is required for the effect of overnight HK diet on Kir4.1/Kir5.1 in the DCT and NCC activity.


Assuntos
Hiperpotassemia/metabolismo , Túbulos Renais Distais/metabolismo , Potássio/metabolismo , Receptor B2 da Bradicinina/metabolismo , Receptores de Angiotensina/metabolismo , Animais , Transporte Biológico , Modelos Animais de Doenças , Feminino , Hiperpotassemia/patologia , Immunoblotting , Túbulos Renais Distais/patologia , Masculino , Potenciais da Membrana , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptor B2 da Bradicinina/efeitos dos fármacos , Receptores de Angiotensina/efeitos dos fármacos
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