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Objective To study the effects of estrogen on bicarbonate secretion of duodenal mucosal , and to observe estrogen receptor (ER) subtypes of estrogen.Methods Sixteen 4-week-old male C57 mice were divided into control group and estrogen group , with eight mice in each group .The mice serum level of estrogen was detected by chemiluminescence .The expression of cystic fibrosis transmembrane conductance regulator (CFTR), solute carrier family 26 (SLC26) A3 and SLC26A6 in the duodenum tissues were determined by real-time polymerase chain reaction ( RT-PCR).After SCBN cells treated with estrogen , ERαand ERβblocking agent, and transfected with silenced ER αand ERβfor 24 and 48 hours, the expression levels of CFTR, SLC26A3 and SLC26A6 mRNA in cells were detected by RT-PCR.The effects of estrogen before and after silenced ER αand ERβon bicarbonate secretion of SCBN cells were observed by high-speed ion imaging system.T test and rank sum test were used for statistical analysis .Results Compared with that of control group , the serum estrogen level of estrogen group was significantly high ((4 874 ±942) pmol/L vs.(657 ±187) pmol/L,t＝-11.579, P?0.01). The expression levels of CFTR, SLC26A3 and SLC26A6 mRNAs in duodenum tissues of estrogen group were higher than those of control group (0.856 ±0.302 vs.0.452 ±0.246, 2.910 ±1.680 vs.1.120 ±0.540, 1.272 ± 0.667 vs.0.319 ±0.114), and the differences were statistically significant ( t ＝-2.317,-2.483 and-3.721, all P?0.05).Compared with those treated with estrogen for 24 and 48 hours, the levels of CFTR mRNA and SLC26A6 mRNA were lower after the ERβblocking agent were added into estrogen for 24 and 48 hours (CFTR mRNA: 0.171 ±0.059 vs.0.522 ±0.260 and 0.111 ±0.014 vs.0.578 ±0.297; SLC26A6 mRNA:0.486 ±0.289 vs.1.118 ±0.571 and 0.492 ±0.231 vs.1.551 ±0.605), and the differences were statistically significant (tCFTR mRNA＝2.974 and 2.655, tSLC26A6 mRNA＝2.393 and 3.272; all P?0.05).Compared with those of silenced ERαgroup, the levels of CFTR mRNA, SLC26A3 mRNA and SLC26A6 mRNA were higher after ERα silenced and then added estrogen for 24 and 48 hours (24 h: 5.073 ±2.270 vs.1.185 ±0.494, 1.796 ±1.168 vs.0.468 ±0.108 and 3.085 ±1.357 vs.0.706 ±0.347; 48 h: 5.025 ±1.998 vs.1.185 ±0.494, 1.557 ± 0.653 vs.0.468 ±0.108 and 3.290 ±1.750 vs.0.706 ±0.347), and the differences were statistically significant (t24 h＝-4.122,-2.773 and -4.162, t48 h ＝-4.604,-4.034 and -3.250; all P?0.05).Compared with that of silenced ERαgroup, the bicarbonate secretion increased after ER αsilenced and then added estrogen for 24 and 48 hours (0.72 ±0.17 and 1.15 ±0.25 vs.0.35 ±0.17), and pH also elevated, and the differences were statistically significant (t＝-6.516 and -12.387, both P?0.01).Conclusion Estrogen mainly up-regulates the expression of bicarbonate transporter protein in duodenal mucosal epithelial cells through ER β, and promotes the bicarbonate secretion of duodenal mucosa .
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Objective@#To study the effects of estrogen on bicarbonate secretion of duodenal mucosal, and to observe estrogen receptor (ER) subtypes of estrogen.@*Methods@#Sixteen 4-week-old male C57 mice were divided into control group and estrogen group, with eight mice in each group. The mice serum level of estrogen was detected by chemiluminescence. The expression of cystic fibrosis transmembrane conductance regulator (CFTR), solute carrier family 26 (SLC26) A3 and SLC26A6 in the duodenum tissues were determined by real-time polymerase chain reaction (RT-PCR). After SCBN cells treated with estrogen, ERα and ERβ blocking agent, and transfected with silenced ERα and ERβ for 24 and 48 hours, the expression levels of CFTR, SLC26A3 and SLC26A6 mRNA in cells were detected by RT-PCR. The effects of estrogen before and after silenced ERα and ERβ on bicarbonate secretion of SCBN cells were observed by high-speed ion imaging system. T test and rank sum test were used for statistical analysis.@*Results@#Compared with that of control group, the serum estrogen level of estrogen group was significantly high ((4 874±942) pmol/L vs. (657±187) pmol/L, t=-11.579, P<0.01). The expression levels of CFTR, SLC26A3 and SLC26A6 mRNAs in duodenum tissues of estrogen group were higher than those of control group (0.856±0.302 vs. 0.452±0.246, 2.910±1.680 vs. 1.120±0.540, 1.272±0.667 vs. 0.319±0.114), and the differences were statistically significant (t=-2.317, -2.483 and -3.721, all P<0.05). Compared with those treated with estrogen for 24 and 48 hours, the levels of CFTR mRNA and SLC26A6 mRNA were lower after the ERβ blocking agent were added into estrogen for 24 and 48 hours (CFTR mRNA: 0.171±0.059 vs. 0.522±0.260 and 0.111±0.014 vs. 0.578±0.297; SLC26A6 mRNA: 0.486±0.289 vs. 1.118±0.571 and 0.492±0.231 vs. 1.551±0.605), and the differences were statistically significant (tCFTR mRNA=2.974 and 2.655, tSLC26A6 mRNA=2.393 and 3.272; all P<0.05). Compared with those of silenced ERα group, the levels of CFTR mRNA, SLC26A3 mRNA and SLC26A6 mRNA were higher after ERα silenced and then added estrogen for 24 and 48 hours (24 h: 5.073±2.270 vs. 1.185±0.494, 1.796±1.168 vs. 0.468±0.108 and 3.085±1.357 vs. 0.706±0.347; 48 h: 5.025±1.998 vs. 1.185±0.494, 1.557±0.653 vs. 0.468±0.108 and 3.290±1.750 vs. 0.706±0.347), and the differences were statistically significant (t24 h=-4.122, -2.773 and -4.162, t48 h=-4.604, -4.034 and -3.250; all P<0.05). Compared with that of silenced ERα group, the bicarbonate secretion increased after ERα silenced and then added estrogen for 24 and 48 hours (0.72±0.17 and 1.15±0.25 vs. 0.35±0.17), and pH also elevated, and the differences were statistically significant (t=-6.516 and -12.387, both P<0.01).@*Conclusion@#Estrogen mainly up-regulates the expression of bicarbonate transporter protein in duodenal mucosal epithelial cells through ERβ, and promotes the bicarbonate secretion of duodenal mucosa.
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Objective To explore the relation between genetic polymorphisms and the expression in colonic tissues of solute-linked carrier family 26 member A3 (SLC26A3) and susceptibility of Crohn's disease (CD) in Han population of Zhejiang Province.Methods A total of 265 CD patients and 566 gender-and age-matched healthy individuals were enrolled.Alleles and genotypes of SLC26A3 (rs17154444,rs7810937,rs7785539,rs2108225,rs6951457) were examined by SNaPshot.The linkage disequilibrium (LD) and haplotype were also analyzed.Eight patients with colonic CD and eight genderand age-matched patients with benign colonic polyps (control group) were selected.The expression level of SLC26A3 protein in the colonic tissue was detected by immunohistochemistry.T test and rank-sum test were performed for statistical analysis.Unconditional Logistic regression analysis was used to analyze the distributions of SLC26A3 polymorphisms and their effects on the clinicopathological features of CD patients.Results The frequencies of mutant allele of rs2108225,rs7785539 and rs6951457 of the CD group were 53.77％ (285/530),4.72％ (25/530) and 2.83％ (15/530),and the frequencies of mutant genotype were 76.23 ％ (202/265),9.43 ％ (25/265) and 5.66 ％ (15/265),which were lower than those of the control group (60.95％,690/1 132;8.13％,92/1 132;6.10％,69/1 132;83.92％,475/566;15.37％,87/566 and 11.84％,67/566),and the differences were statistically significant (all P＜0.05).The frequencies of mutant allele of rs17154444 and rs7810937 of the CD group were 10.19％ (54/530) and 34.91 ％ (185/530),and the frequencies of mutant genotype were 18.49 ％ (49/265) and 56.23 ％ (149/ 265),compared with those of the control group (8.30％,94/1 132;30.92％,350/1 132;15.55％,88/566 and 51.77％,293/566),the differences were not statistically significant (all P＞0.05).The frequency of mutant allele G of rs2108225 in patients with ileal CD was 47.89 ％ (91/190),and the frequency of mutant genotypeAG+GG was 65.26％(62/95),which were both lower than those of colonic CD (61.62％,122/198 and 85.86％,85/99),and the differences were statistically significant (both P＜0.012 5).rs7810937,rs7785539 and rs2108225 were in a strong linkage disequilibrium.The frequencies of haplotypes AGG and ACA of the CD group were 53.96％ (286/530) and 4.34％ (23/530),which were lower than those of the control group (60.07％,680/1 132 and 7.51％,85/1 132),and the differences were statistically significant (52 =5.534,P=0.019;x2 =5.967,P=0.015).And the frequency of haplotype AGA of the CD group was 8.30％ (44/530),which was higher than that of the control group (1.15％,13/1 132),and the difference was statistically significant (x2 =7.793,P＜0.01).Furthermore,the expression level of SLC26A3 protein in colonic tissues of eight colonic CD patients was 0.19±0.07,which was lower than that of patients with benign colonic polyps (0.26 ±-0.03),and the difference was statistically significant (t=2.55,P=0.023).In addition,the expression levels of SLC26A3 protein in patients carrying genotype GG or AG of rs2108225 were 0.19±0.03 and 0.10±0.01,respectively,which were lower than that of patients carrying genotype AA (0.26± 0.02),and the differences were statistically significant (t=3.19,P=0.033;t=9.06,P=0.003).Conclusions The genetic polymorphismns and their haplotypes of SLC26A3 (rs7785539,rs2108225 and rs6951457) are associated with the susceptibility of CD,and SLC26A3 (rs2108225) polymorphism may affect the expression level of SLC26A3 protein in the colonic tissues.
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Congenital chloride diarrhea (CCD) is a rare autosomal recessive disease, which is characterized by electrolyte absorption defect due to impaired function of the Cl-/HCO3 - exchanger in the ileum and the colon. Its main features are profuse watery diarrhea, high fecal chloride concentration, and failure to thrive. Profuse watery diarrhea characterized by a high concentration of chloride in stools results in hypochloremia, hyponatremia, and dehydration with metabolic alkalosis. Early detection and therapeutic intervention can prevent life-threatening symptoms of CCD and growth failure. Recently, several therapies, such as proton pump inhibitors and butyrate, have been suggested for amelioration of diarrhea. Here, we report a case of CCD in a preterm male infant who was successfully treated with an oral proton pump inhibitor.
Subject(s)
Humans , Infant , Male , Absorption , Alkalosis , Butyrates , Colon , Dehydration , Diarrhea , Failure to Thrive , Hyponatremia , Ileum , Omeprazole , Proton Pump Inhibitors , Proton Pumps , ProtonsABSTRACT
Objective To analyze the clinical characteristics and mutation of SLC26A3 gene of a patient with congenital chloride diarrhea in order to deepen the understanding of the disease.Methods The clinical data of the patient who was admitted in Affiliated Hospital of Capital Pediatric Institute in June 2014 were collected.Venous blood of the proband and his parents (2 mL for each) had been extracted for genomic DNA isolation.The 21 exons of SLC26A3 gene were amplified with polymerase chain reaction and screened for mutations by sequencing.Results The main clinical features of the patient included polyhydramnios,preterm,normal birth weight,watery diarrhea,low weight and severe electrolyte disturbances with hypochloremia,hypokalemia,hyponatremia and metabolic alkalosis.Renin angiotensin and aldosterone were high.His urine chloride concentration was low and fecal chloride concentration was high (＞ 90mmol/L).After oral salt substitution therapy with KCl and NaCl [3 mmol/(kg · d),4 mmol/(kg · d)],the electrolyte was better,alkalosis was alleviated,and growth and development were improved.The gene analysis revealed that the patient carried nt1631T ＞ A homozygous mutation on exon 15 which lead to Ile544Asn mutation in the predicted SLC26A3 transmembrane protein sequence,which was considered to be responsible for the functional abnormality of the Cl-/HCO3-protein.His parents were carriers of SLC26A3 gene and their clinical phenotype was normal.Conclusions Congenital chloride diarrhea is a rare autosomal recessive disorder and easily misdiagnosed.The patient of early postnatal diarrhea with persistent hypochloremia,hypokalemia,hyponatremia and metabolic alkalosis should be thought about this disease.Genetic analysis can help make the diagnosis.The prognosis is good if a patient has an early diagnosis and appropriate management.
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ObjectiveTo discuss the clinical diagnosis, treatment and genetic diagnosis of congenital chloride diarrhea (CCD), a rare autosomal recessive disease.Methods One month old boy with persistent diarrhea, hypochloremia, hyponatremia, hypokalemia and metabolic alkalosis, his stool electrolyte testing, clinical treatment and follow-up, as well as his and his parents’ SLC26A3 gene mutation analysis were retrospectively analyzed.Results The fecal electrolyte testing showed that the levels of Cl- and K+ were increased and the level of Cl- was much higher than the sum of Na+ and K+. After replacement therapy with NaCl and KCl, the blood electrolyte recovered to normal. Follow-up 4 years, the boy had a normal growth and development. Mutation analysis onSLC26A3 gene showed there was a homozygous mutation of 239G>A and both his father and mother carried the same heterozygous mutation. This mutation was ifrst discovered in China.Conclusions The sequencing analysis ofSLC26A3 mutation may help to diagnosis CCD.
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Objective The expression and impaired function of ion channels might be one of the pathophysiological mecha -nisms responsible for diarrhea in inflammatory bowel disease ( IBD) .Proper animal model is the key to explore detailed pathophysiolog-ical process.The purpose of this study was to build a rat model of acute colitis induced by dextran sodium sulfate (DSS) in C57BL/6 mice and evaluate diarrhea-associated clinical , histological , pathological parameters and expressions of ion channel protein . Methods C57BL/6J mice of model group were treated with 4%DSS solution for 7 days to induce acute colitis.Mice body weight, stool moisture, stool consistency and the degree of hematochezia were recorded .The histopathological changes of mice colon specimens were observed visually and microcosmically, and the ion channel SLC26A3 protein was detected by Western Blot . Results All experimental mice survived.In the experiment, compared with control group , bloody diarrhea and weight lose occurred in model group , along with increased stool moisture ([73.30 ±8.31]% after experiment vs [44.32 ±6.42]% before experiment, P=0.004), and rapidly in-creased disease activity index (DAI) of acute colitis ([3.50 ±0.87] after experiment vs [1.0 ±0.00] before experiment, P=0.000).At the end of this experiment , compared with control group , the model group resulted in higher colonic damage score and pathological inflammation score (P=0.00, P=0.002), significantly shortened co-lon (P=0.00) and decreased expression of SLC26A3. Conclusion The intestinal mucosal injury and phenotypic features of 4%DSS-induced acute colitis are very similar to those of human ulcerative colitis .Impaired expression of intestinal ion transporter SLC26 A3 coexists with diarrhea in model group mice , and this model can support the research on mechanism of functional changes of ion channels in inflammatory diarrhea .
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Congenital chloride diarrhea (CLD) is an autosomal recessive disorder with the hallmark of persistent watery Cl(-)-rich diarrhea from birth. Mutations in the solute carrier family 26, member 3 (SLC26A3) gene, which encodes a coupled Cl-/HCO3- exchanger in the ileum and colon, are known to cause CLD. Although there are a few reports of CLD patients in Korea, none of these had been confirmed by genetic analysis. Here, we describe the case of a Korean infant with clinical features of CLD. Using direct sequencing analysis, we identified 2 sequence variants: a missense variant of unknown significance (c.525G>C; p.Arg175 Ser) and a splicing mutation (c.2063-1G>T) in the SLC26A3 gene; these had been inherited from the father and mother, respectively. Whilst CLD is rare, its main symptom, diarrhea, is very common in infants. Hence, the diagnosis of CLD can prove difficult. Mutational analysis of the SLC26A3 gene should be considered as a viable method to confirm a diagnosis of CLD in Korean infants with persistent diarrhea.