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中华蜜蜂肠粘蛋白AcMucin5AC-1的鉴定和定位分析
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作者 李小青 郭悦 +2 位作者 张洁 周泽扬 党晓群 《昆虫学报》 CAS CSCD 北大核心 2023年第9期1150-1160,共11页
【目的】本研究旨在鉴定中华蜜蜂Apis cerana cerana肠粘蛋白AcMucin5AC-1的结构、分布及对中肠的影响,为解析蜜蜂中肠的生理功能提供理论依据。【方法】利用生物信息学比较分析中华蜜蜂AcMucin5AC蛋白的序列特征;利用RT-qPCR检测AcMuci... 【目的】本研究旨在鉴定中华蜜蜂Apis cerana cerana肠粘蛋白AcMucin5AC-1的结构、分布及对中肠的影响,为解析蜜蜂中肠的生理功能提供理论依据。【方法】利用生物信息学比较分析中华蜜蜂AcMucin5AC蛋白的序列特征;利用RT-qPCR检测AcMucin 5 AC-1在中华蜜蜂4日龄幼虫中肠和表皮以及2日龄幼虫感染中华蜜蜂囊状幼虫病毒(Chinese sacbrood virus,CSBV)后0,12,24,48和72 h时中肠中的表达量。通过原核表达系统对中华蜜蜂AcMucin5AC-1进行表达,亲和层析柱纯化重组蛋白并制备相应多克隆抗体;Western blot检测AcMucin5AC-1在中华蜜蜂健康3-6日龄幼虫中以及4日龄幼虫中肠、表皮和围食膜中的表达,应用间接免疫荧光试验分析AcMucin5AC-1在中华蜜蜂4日龄幼虫中的定位。通过饲喂法利用RNAi沉默中华蜜蜂2日龄幼虫AcMucin 5 AC-1后12,24,48和72 h时分析RNAi干扰效率,利用苏木精-伊红(hematoxylin-eosin,HE)染色法探究RNAi干扰2日幼虫AcMucin 5 AC-1后24,48和72 h时对中华蜜蜂幼虫整体组织形态的影响。【结果】中华蜜蜂基因组中有8个AcMucin5AC基因,其氨基酸序列均含有粘蛋白结构域。RT-qPCR检测结果表明,AcMucin 5 AC-1在中华蜜蜂4日龄幼虫中肠中表达量较表皮中的高,且在感染CSBV病毒的12和72 h时的2日龄幼虫中肠中比对照组显著下调。成功获得约50 kD的重组蛋白AcMucin5AC-1和多克隆抗体,Western blot结果显示在中华蜜蜂3-6日龄幼虫以及4日龄幼虫中肠、表皮和围食膜的总蛋白中均能检测到AcMucin5AC-1。间接免疫荧光试验结果表明AcMucin5AC-1主要定位在4日龄幼虫中肠和围食膜。RNAi效率检测结果表明,2.0μg/头ds AcMucin 5 AC-1干扰24 h时中华蜜蜂AcMucin 5 AC-1较ds EGFP对照组表达量下调92%。HE染色检测结果表明,在AcMucin 5 AC-1的RNAi后72 h时中华蜜蜂幼虫整个肠腔的细胞间的致密程度减弱,形态结构紊乱。【结论】中华蜜蜂AcMucin5AC-1是位于幼虫的中肠和围食膜的粘蛋白,AcMucin 5 AC-1的下调影响中华蜜蜂幼虫中肠的形态,暗示该基因在幼虫中肠发育过程中可能发挥重要作用。 展开更多
关键词 中华蜜蜂 Mucin5ac 原核表达 RNAI 组织定位
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黏液-纤毛标志物Mucin 5AC-acetylated alpha-tubulin在过敏性鼻炎患者鼻黏膜中的表达 被引量:1
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作者 郭沐涛 周穗子 邱前辉 《中国临床新医学》 2023年第1期31-36,共6页
目的探究过敏性鼻炎(AR)患者鼻腔黏液-纤毛清除功能在鼻黏膜分子水平上的损伤表现和严重程度。方法选取2021年3月至2022年3月期间在我院进行鼻中隔偏曲手术的40例患者的下鼻甲黏膜组织,其中AR患者21例(AR组),非AR患者(对照组)19例。通... 目的探究过敏性鼻炎(AR)患者鼻腔黏液-纤毛清除功能在鼻黏膜分子水平上的损伤表现和严重程度。方法选取2021年3月至2022年3月期间在我院进行鼻中隔偏曲手术的40例患者的下鼻甲黏膜组织,其中AR患者21例(AR组),非AR患者(对照组)19例。通过免疫荧光染色方法在蛋白水平上观察及评估黏液分泌标志物Mucin 5AC(MUC5AC)和纤毛标志物acetylated alpha-tubulin(acet.α-tubulin)表达情况。结果与对照组相比,AR组黏液分泌的总免疫荧光强度增加32.365%,纤毛长度缩短将近1/2,纤毛脱落评分上升将近3倍,纤毛总免疫荧光强度下降24.555%。结论AR患者黏液分泌量显著增加,纤毛排列不齐、变短、稀疏以及脱落增加可能是导致AR患者鼻腔黏膜黏液-纤毛清除功能减弱,继而加重AR症状严重程度的分子依据。 展开更多
关键词 过敏性鼻炎 黏液-纤毛清除功能 Mucin 5ac Acetylated alpha-tubulin
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Gastric mucin expression in Helicobacter pylori-related, nonsteroidal anti-inflammatory drug-related and idiopathic ulcers 被引量:5
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作者 Doron Boltin Marisa Halpern +4 位作者 Zohar Levi Alex Vilkin Sara Morgenstern Samuel B Ho Yaron Niv 《World Journal of Gastroenterology》 SCIE CAS CSCD 2012年第33期4597-4603,共7页
AIM: To determine the pattern of secreted mucin expression in Helicobacter pylori (H. pylori)-related, nonsteroidal anti-inflammatory drug (NSAID)-related and idiopathic gastric ulcers. METHODS: We randomly selected 9... AIM: To determine the pattern of secreted mucin expression in Helicobacter pylori (H. pylori)-related, nonsteroidal anti-inflammatory drug (NSAID)-related and idiopathic gastric ulcers. METHODS: We randomly selected 92 patients with H. pylori-associated (n = 30), NSAID-associated (n = 18), combined H. pylori and NSAID-associated gastric ulcers (n = 24), and patients with idiopathic gastric ulcers (n = 20). Immunohistochemistry for T-cell CD4/CD8, andfor mucin 5AC (MUC5AC) and mucin 6 (MUC6), was performed on sections of the mucosa from the ulcer margin. Inflammation score was assessed according to the Sydney system. RESULTS: MUC5AC was expressed on the surface epithelium (98.9%) and neck glands (98.9%) with minimal expression in the deep glands (6.5%). MUC6 was strongly expressed in the deep glands (97.8%), variable in the neck glands (19.6%) and absent in the surface epithelium (0%). The pattern of mucin expression in idiopathic ulcer margins was not different from the expression in ulcers associated with H. pylori, NSAIDs, or combined H. pylori and NSAIDs. CD4/CD8 ratio was higher in H. pylori-positive patients (P = 0.009). Idiopathic ulcers are associated with hospitalized patients and have higher bleeding and mortality rates. CONCLUSION: Idiopathic ulcers have a unique clinical profile. Gastric mucin expression in idiopathic gastric ulcers is unchanged compared with H. pylori and/or NSAID-associated ulcers. 展开更多
关键词 Idiopathic ulcer Mucin Mucin 5ac Mucin 6 Helicobacter pylori
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Qingfei oral liquid downregulates TRPV1 expression to reduce airway inflammation and mucus hypersecretion injury caused by respiratory syncytial virus infection and asthma in mice 被引量:1
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作者 Xiao-Ping Jing Wu-Ning Yan +1 位作者 Wei-Wei Cheng Hai-Rong Zeng 《Traditional Medicine Research》 2020年第4期229-237,共9页
Objective:Qingfei oral liquid(QF),an experimental Chinese medicine prescription developed from the ancient priscription of traditional Chinese medicines Ma Xin Shi Gan decoction and Tingli Dazao Xie Fei decoction,has ... Objective:Qingfei oral liquid(QF),an experimental Chinese medicine prescription developed from the ancient priscription of traditional Chinese medicines Ma Xin Shi Gan decoction and Tingli Dazao Xie Fei decoction,has been effectively used since decades to treat patients with viral pneumonia and asthma.In our previous study,we had demonstrated that QF can significantly reduce airway hyperresponsiveness,hyperemia,lung tissue edema,inflammatory lung tissue infiltration in mice,airway mucus secretion,and peripheral airway collagen hyperplasia;however,its mechanism of action is unknown.Methods:Fifty 6–8-week-old male BALB/c mice were equally and randomly divided into five groups:the control,ovalbumin(OVA),OVA+respiratory syncytial virus(RSV),QF,and dexamethasone(Dxms)groups.The QF group was administered QF at 1.17 g·kg−1·d−1,the Dxms group received dexamethasone injections at 0.2 mg·kg−1·d−1,and the remaining groups were administered PBS.Inflammation in the lung tissue was assessed by hematoxylin and eosin(HE),periodic acid–Schiff(PAS),and Van Gieson staining.ELISA was used to evaluate the IL-13,IL-25,and IL-33 in the mice.Western blotting was used to examine changes in the proteins levels of transient receptor potential vanilloid-1(TRPV1)and mucin 5AC(MUC5AC)in the lung tissues of mice.Results:Histopathological evaluation revealed that the OVA and OVA+RSV groups exhibited lung tissue edema and inflammatory lung tissue infiltration in the HE staining and airway secretions in the PAS staining;collagen hyperplasia around the airway was increased in these two groups compared with the control group.The QF group exhibited significantly reduced lung tissue edema,inflammatory lung tissue infiltration,airway secretions,and collagen hyperplasia around the airway compared with the OVA+RSV group.We analyzed the serum levels of IL-13,IL-25,and IL-33 in the mice and found that these levels were higher in the OVA and OVA+RSV groups than in the control group(P<0.05 in the OVA group,P<0.01 in the OVA+RSV group).The QF group exhibited significantly decreased serum levels of IL-13,IL-25,and IL-33 compared with the OVA+RSV group(all P<0.05).The Dxms group also exhibited significant decreases in the serum levels of IL-13 and IL-33(all P<0.05)but no significant decrease in the serum levels of IL-25 compared with the RSV+OVA group.Finally,we examined the protein levels of TRPV1 and MUC5AC in the lung tissues of mice using Western blotting.After identifying RSV infection in the mice with asthma,the protein levels of TRPV1 and MUC5AC in the lung tissues of mice were significantly higher than those in the control group(P<0.05,P<0.01).We found that compared with RSV+OVA,QF can significantly downregulate the protein level of TRPV1;further,the protein level of MUC5AC was also significantly reduced(all P<0.001).Conclusion:QF can inhibit RSV replication and reduce airway inflammation and mucus hypersecretion injury caused by RSV infection and asthma,and its mechanism of action may be associated with the downregulation of TRPV1 expression and a decrease in airway mucus hypersecretion injury. 展开更多
关键词 Qingfei oral liquid Viral pneumonia ASTHMA Respiratory syncytial virus infection Transient receptor potential vanilloid-1 Mucin 5ac
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Effect of electroacupuncture on expressions of acetylcholine and mucin 5AC in the lungs of rats with chronic obstructive pulmonary disease 被引量:6
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作者 Cong Wen-juan Li Jing +6 位作者 Liao Yu-jing Zhang Xin-fang Jiang Chuan-wei Xiang Shui-ying Huang Wen-biao Liu Xiao-yun Liu Zi-bing 《Journal of Acupuncture and Tuina Science》 CSCD 2018年第3期133-139,共7页
Objective: To observe the effect of electroacupuncture (EA) on the expressions of acetylcholine (ACh) and mucin 5AC (MUC5AC) in the lungs of rats with chronic obstructive pulmonary disease (COPD), and explore... Objective: To observe the effect of electroacupuncture (EA) on the expressions of acetylcholine (ACh) and mucin 5AC (MUC5AC) in the lungs of rats with chronic obstructive pulmonary disease (COPD), and explore the mechanism of EA in treating COPD. Methods: Thirty Sprague-Dawley (SD) rats were randomly divided into a control group, a COPD group, and an EA group, with 10 rats in each group. The control group was a group of normal rats. The COPD rat model was induced by cigarette smoke combined with lipopolysaccharide (LPS). The COPD rats were treated with EA at bilateral Feishu (BL 13) and Zusanli (ST 36) in the EA group, 30 rain each time, once a day, successively for 14 d. The lung function was tested. The contents of ACh and MUC5AC in lungs and bronchoalveolar lavage fluid (BALF) were detected by enzyme-linked immunosorbent assay (ELISA). Pearson method was used to analyze the correlation between pulmonary function and the content of MUC5AC in lungs. The mRNA and protein expressions of MUC5AC in lung tissues were detected by real-time polymerase chain reaction (RT-PCR) and Western blot (WB), respectively. The immune response of MUC5AC was observed by immunohistochemistry. Results: Eight rats were left in each group, and the other two died. Compared with the control group, the total airway resistance (Raw) increased significantly and dynamic compliance (Cdyn) decreased significantly in the COPD group (P〈0.01); compared with the COPD group, the Raw level declined significantly and Cdyn increased significantly in the EA group (P〈0.01). The contents of ACh and MUC5AC in the lungs and BALF were remarkably higher in the COPD group compared with those in the control group (P〈0.01, P〈0.001); compared with the COPD group, the contents of ACh and MUC5AC were significantly lower in the EA group (P〈0.05, P〈0.001). There was a negative correlation between MUC5AC content and lung function (P〈0.001). The mRNA and protein expressions of MUC5AC in the lungs were significantly higher in the COPD group than in the control group (P〈0.001); compared with the COPD group, the expressions were significantly lower in the EA group (P〈0.01). Compared with the control group, the immune response of MUC5AC in the airway epithelium significantly increased in the COPD group (P〈0.001); the immune response of MUC5AC was significantly lower in the EA group compared with that in the COPD group (P〈0.001). Conclusion: EA treatment can improve the lung function of COPD rats, which may be related to its effect in the down-regulation of ACh and MUC5AC contents in the lungs as well as the inhibition of mucus hypersecretion. 展开更多
关键词 Acupuncture Therapy ELECTROACUPUNCTURE Pulmonary Disease Chronic Obstructive POINT Feishu (BL 13) POINT Zusanli (ST 36) ACETYLCHOLINE Mucin 5ac
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Curcumin Inhibits Lipopolysaccharide-lnduced Mucin 5AC Hypersecretion and Airway Inflammation via Nuclear Factor Erythroid 2-Related Factor 2 被引量:6
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作者 Xiao-Ping Lin Cheng Xue +3 位作者 Jia-Min Zhang Wei-Jing Wu Xiao-Yang Chen Yi-Ming Zeng 《Chinese Medical Journal》 SCIE CAS CSCD 2018年第14期1686-1693,共8页
Background: Excess mucus production is an important pathophysiological feature of chronic inflammatory airway diseases. Effective therapies are currently lacking. The aim of the study was to evaluate the effects ofcu... Background: Excess mucus production is an important pathophysiological feature of chronic inflammatory airway diseases. Effective therapies are currently lacking. The aim of the study was to evaluate the effects ofcurcumin (CUR) on lipopolysaccharide (LPS)-induced mucus secretion and inflammation, and explored the underlying mechanism in vivo and in vitro. Methods: For the in vitro study, human bronchial epithelial (NCI-H292) cells were pretreated with CUR or vehicle for 30 min, and then exposed to LPS for 24 h. Next, nuclear factor erythroid 2-related factor 2 (Nrf2) was knocked down with Nrf2 small interfering RNA (siRNA) to confirm the specific role of Nrf2 in mucin regulation of CUR in NCI-H292 cells. In vivo, C57BL/6 mice were randomly assigned to three groups (n = 7 for each group): control group, LPS group, and LPS + CUR group. Mice in LPS and LPS + CUR group were injected with saline or CUR (50 mg/kg) intraperitoneally 2 h before intratracheal instillation with LPS ( 100 μg/ml) for 7 days. Cell lysate and lung tissue were obtained to measured Mucin 5AC (MUC5AC) and Nrf2 mRNA and protein expression by a real-time polymerase chain reaction and Western blotting. Bronchoalveolar lavage fluid (BALF) was collected to enumerate total cells and neutrophils. HistopathologicaI changes of the lung were observed. Data were analyzed by one-way analysis of variance. Student's t-test was used when two groups were compared. Results: CUR significantly decreased the expression ofMUC5AC mRNA and protein in NCI-H292 cells exposed to LPS. This effect was dose dependent (2.424 ± 0.318 vs. 7.169 ± 1.785, t = 4.534, and 1.060 ± 0.197 vs. 2.340 ± 0.209, t = 7.716; both P 〈 0.05, respectively) and accompanied by increased mRNA and protein expression of Nrf2 (1.952 ± 0.340 vs. 1.142 ± 0.176, t = -3.661, and 2.010 ± 0.209 vs. 1.089 ±0. 132, t = -6.453; both P 〈 0.05, respectively). Furthermore, knockdown of Nrf2 with siRNA increased MUC5A C mRN A expression by 47.7%, compared with levels observed in the siRNA-negative group (6.845 ± 1.478 vs. 3.391 ± 0.517, t = -3.821, P 〈 0.05). Knockdown of Nrf2 with siRNA also markedly increased MUC5A C protein expression in NCI-H292 cells. CUR also significantly decreased LPS-induced mRNA and protein expression of MUC5A C in mouse lung ( 1.672 ± 0.721 vs. 5.961 ± 2.452, t = 2.906, and 0.480 ± 0.191 vs. 2.290 ± 0.834, t = 3.665, respectively; both P 〈 0.05). Alcian blue/periodic acid-Schiff staining also showed that CUR suppressed mucin production. Compared with the LPS group, the numbers of inflammatory cells (247 ± 30 vs. 334 ± 24, t = 3.901, P 〈 0.05) and neutrophils (185 ± 22 vs. 246 ± 20, t = 3.566, P 〈 0.05) in BALF decreased in the LPS + CUR group, as well as reduced inflammatory cell infiltration in lung tissue. Conclusion: CUR inhibits LPS-induced airway mucus hypersecretion and inflammation through activation of Nrf2 possibly. 展开更多
关键词 Airway Mucin CURCUMIN Mucin 5ac Nuclear Factor Erythroid 2-Related Factor 2
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Increased expression of human calcium-activated chloride channel 1 is correlated with mucus overproduction in the airways of Chinese patients with chronic obstructive pulmonary disease 被引量:8
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作者 WANG Ke FENG Yu-ling WEN Fu-qiang CHEN Xue-rong OU Xue-mei XU Dan YANG Jie DENG Zhi-pin 《Chinese Medical Journal》 SCIE CAS CSCD 2007年第12期1051-1057,共7页
Background Chronic obstructive pulmonary disease (COPD) is usually complicated with mucus overproduction in airway. Recently the increased expression of the human calcium-activated chloride channel 1 (CaCC1) was f... Background Chronic obstructive pulmonary disease (COPD) is usually complicated with mucus overproduction in airway. Recently the increased expression of the human calcium-activated chloride channel 1 (CaCC1) was found to play an important role in mucus overproduction in the asthmatic airways. To investigate the relationship of CaCC1 and mucus overproduction in the airway of Chinese patients with COPD, the expressions of CaCC1, MUC5AC and mucus in bronchial tissues were examined. Methods Bronchial tissues were obtained from fiberoptic bronchoscopy and bronchial biopsy in West China Hospital from April to July in 2004. Twenty-five patients were diagnosed as the patients with COPD overproduction, and other 20 were the control subjects. The expressions of CaCC1, MUC5AC and mucin in bronchial tissues were detected by reverse transcriptase-polymerase chain reaction (RT-PCR), in situ hybridization with digoxigenin (DIG)-Iabeled RNA probe, immunohistochemical and alcian blue-periodic acid Schiff (AB-PAS) staining, respectively. Results Compared with the control group, the stronger expressions of CaCC1 were further detected throughout the bronchial tissues from patients with COPD (P〈0.01). Furthermore, the stronger expressions of the CaCC1 mRNA were related to the severity of airflow obstruction. Samples from COPD showed a stronger staining for MUC5AC than those in control subjects (P〈0.01) and AB-PAS staining revealed more mucins in COPD patients' submucosal gland comparing with that in control subjects (P〈0.01). Expression levels of the CaCC1 mRNA were respectively negatively correlated with the patients' forced expiratory volume in one second (FEV~) / forced vital capacity (FVC) data, FEV1% predicted data, V50% predicted data, V25% predicted data (r=-0.43, r=-0.43, r=-0.35, r=-0.36, P〈0.01, P〈0.01, P〈0.05, P〈0.05). While the expression levels of the CaCC1 mRNA were well correlated with the expression levels of the MUC5AC mRNA of airway epithelium and the PAS-AB stained area of submucosal glands (r=0.39, r=0.46, P〈0.05, P〈0.01). Expression levels of the MUC5AC mRNA were negatively correlated with the patients' FEV1/FVC data (P=0.01), FEV1% pred data (P=-0.01), V50% predicted data, V25% predicted data(r=-0.53, r=-0.53, r=-0.48, r=-0.43, P〈0.01, P〈0.01, P〈0.01, P〈0.01). While the expression levels of the MUC5AC mRNA were well correlated with the positively PAS-AB stained area of submucosal gland (P〈0.05), and the correlation coefficients were 0.43. Conclusion These results suggest that the stronger gene expression of CaCC1 exists, complicated with mucus overproduction in the airwav of Chinese patients with COPD. 展开更多
关键词 calcium-activated chloride channel 1 mucin 5ac MUCIN mucus overproduction chronic obstructive pulmonary disease
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