Type 1 diabetes mellitus(T1DM) lacks insulin secretion due to autoimmune deficiency of pancreaticβ-cells.Protecting pancreatic islets and enhancing insulin secretion has been therapeutic approaches.Mannogalactoglucan...Type 1 diabetes mellitus(T1DM) lacks insulin secretion due to autoimmune deficiency of pancreaticβ-cells.Protecting pancreatic islets and enhancing insulin secretion has been therapeutic approaches.Mannogalactoglucan is the main type of polysaccharide from natural mushroom,which has potential medicinal prospects.Nevertheless,the antidiabetic property of mannogalactoglucan in T1DM has not been fully elucidated.In this study,we obtained the neutral fraction of alkali-soluble Armillaria mellea polysaccharide(AAMP-N) with the structure of mannogalactoglucan from the fruiting body of A.mellea and investigated the potential therapeutic value of AAMP-N in T1DM.We demonstrated that AAMP-N lowered blood glucose and improved diabetes symptoms in T1DM mice.AAMP-N activated unfolded protein response(UPR) signaling pathway to maintain ER protein folding homeostasis and promote insulin secretion in vivo.Besides that,AAMP-N promoted insulin synthesis via upregulating the expression of transcription factors,increased Ca^(2+) signals to stimulate intracellular insulin secretory vesicle transport via activating calcium/calmodulin-dependent kinase Ⅱ(CamkⅡ) and cAMP/PKA signals,and enhanced insulin secretory vesicle fusion with the plasma membrane via vesicle-associated membrane protein 2(VAMP2).Collectively,these studies demonstrated that the therapeutic potential of AAMP-N on pancreatic islets function,indicating that mannogalactoglucan could be natural nutraceutical used for the treatment of T1DM.展开更多
Objective Pseudomonas aeruginosa is a ubiquitous and opportunistic pathogen that uses the type Ⅲ secretion system （TTSS） to inject effector proteins directly into the cytosol of target cells to subvert the host cel...Objective Pseudomonas aeruginosa is a ubiquitous and opportunistic pathogen that uses the type Ⅲ secretion system （TTSS） to inject effector proteins directly into the cytosol of target cells to subvert the host cell＇s functions. Specialized bacterial chaperones are required for effective secretion of some effectors. To identify the chaperone of ExoS, the representative effector secreted by the TTSS of P aeruginosa, we analyzed the role of a postulated chaperone termed Orfl. Methods By allelic exchange, we constructed the mutant with the deletion of gene Orfl. Analysis of secreted and cell-associated fractions was performed by SDS-PAGE and Western blotting. Using strain expressing in trans Orfl, tagged by V5 polypeptide and histidine, protein-protein interaction was determined by affinity resin pull-down assay in combination with MALDI-TOF The role of Orfl in the expression of exoS was evaluated by gene reporter analysis. Results Pull-down assay showed that Orfl binds to ExoS and ExoT. Secretion profile analysis showed that Orfl was necessary for the optimal secretion of ExoS and ExoT. However, Orfl had no effect on the expression of exoS. Conclusion Orfl is important for the secretion of ExoS probably by maintaining ExoS in a secretion-competent conformation. We propose to name Orfl as SpcS for ＂specific Pseudomonas chaperone for ExoS＂.展开更多
Neuregulin-1 type Ⅲ is a key regulator in Schwann cell proliferation, committing to a myelinat- ing fate and regulating myelin sheath thickness. However, the expression pattern of neuregulin- 1 type III in the periph...Neuregulin-1 type Ⅲ is a key regulator in Schwann cell proliferation, committing to a myelinat- ing fate and regulating myelin sheath thickness. However, the expression pattern of neuregulin- 1 type III in the peripheral nervous system during developmental periods （such as the premyelin- ating stage, myelinating stage and postmyelinating stage） has rarely been studied. In this study, dorsal root ganglia were isolated from rats between postnatal day 1 and postnatal day 56. The expression pattern of neuregulin-1 type III in dorsal root ganglia neurons at various develop- mental stages were compared by quantitative real-time polymerase chain reaction, western blot assay and immunofluorescent staining. The expression of neuregulin-I type Ⅲ mRNA reached its peak at postnatal day 3 and then stabilized at a relative high expression level from postnatal day 3 to postnatal day 56. The expression of neuregulin-1 type III protein increased gradually from postnatal day 1, reached a peak at postnatal day 28, and then decreased at postnatal day 56. Immunofluorescent staining results showed a similar tendency to western blot assay results. Experimental findings indicate that the expression of neuregulin-1 type III in rat dorsal root ganglion was increased during the premyelinating （from postnatal day 2 to postnatal day 5） and myelinating stage （from postnatal day 5 to postnatal day 10）, but remained at a high level in the postmyelinating stage （after postnatal day 10）.展开更多
Objective To investigate the effects of TGF- β1 on proliferation and type N collagen mRNAexpression of rat renal interstitial libroblasts in vitro for elucidating the role of fibroblasts in renal interstitialfibrosis...Objective To investigate the effects of TGF- β1 on proliferation and type N collagen mRNAexpression of rat renal interstitial libroblasts in vitro for elucidating the role of fibroblasts in renal interstitialfibrosis. Methods The cells were cultured in media containing various concentrations of TGF- β1. Theproliferation and the type Ⅲ collagen mRNA expression of the cells were assayed with MTT method andRT- PCR respectively. Results TGF- β1, has a stimulating proliferation effect with dose- dependence and aincreasing expression of type Ⅲ collagen mRNA with both dose- and time- dependence to the renal fibroblasts. Conclusion It is suggested that TGF- β, is involved in proliferation of renal fibroblasts and their type Ⅲcollagen mRNA expression, and may play an important role in renal interstitial fibrosis.展开更多
目的探讨细胞外信号调节激酶1/2(ERK1/2)在转化生长因子-β1(TGF-β1)诱导的肺成纤维细胞合成Ⅰ、Ⅲ型胶原蛋白中的作用,及新型过氧化物酶Peroxiredoxin-1(Prx-1)对该作用的影响。方法体外培养肺成纤维细胞随机分为4组:对照组(0.4%血清)...目的探讨细胞外信号调节激酶1/2(ERK1/2)在转化生长因子-β1(TGF-β1)诱导的肺成纤维细胞合成Ⅰ、Ⅲ型胶原蛋白中的作用,及新型过氧化物酶Peroxiredoxin-1(Prx-1)对该作用的影响。方法体外培养肺成纤维细胞随机分为4组:对照组(0.4%血清)、TGF-β1组(5μg/L)、阴性转染组(TGF-β1+阴性对照si RNA)和Prx-1 si RNA转染组(TGF-β1+Prx-1 si RNA)。采用脂质体转染法转染si RNA,实时定量逆转录-聚合酶链反应(RT-PCR)检测转染后Prx-1 m RNA表达;Western blot检测Ⅰ和Ⅲ型胶原蛋白、ERK1/2及Prx-1表达;2,7-二氯荧光素二乙酸(DCFH-DA)检测活性氧(ROS)水平。结果 Prx-1 si RNA转染肺成纤维细胞后,Prx-1 m RNA表达明显降低,最大抑制率为92%。与对照组比较,TGF-β1组的Ⅰ和Ⅲ型胶原蛋白、ROS、磷酸化ERK1/2(p-ERK1/2)及Prx-1蛋白的表达水平均明显提高。与TGF-β1组比较,阴性转染组中的上述观察指标无明显变化,但Prx-1转染组的Ⅰ和Ⅲ型胶原蛋白、ROS、p-ERK1/2水平进一步提高,而Prx-1蛋白的表达被抑制。结论 TGF-β1能够诱导肺成纤维细胞生成ROS,并促进ERK1/2通路的激活,导致Ⅰ、Ⅲ型胶原蛋白合成增加,而Prx-1 si RNA可通过提高ROS水平进一步促进TGF-β1该作用。展开更多
The innate immune system is the first line of defense against invading pathogens. Innate immune cells recognize molecular patterns from the pathogen and mount a response to resolve the infection. The production of pro...The innate immune system is the first line of defense against invading pathogens. Innate immune cells recognize molecular patterns from the pathogen and mount a response to resolve the infection. The production of proinflammatory cytokines and reactive oxygen species, phagocytosis, and induced programmed cell death are processes initiated by innate immune cells in order to combat invading pathogens. However, pathogens have evolved various virulence mechanisms to subvert these responses. One strategy utilized by Gram-negative bacterial pathogens is the deployment of a complex machine termed the type Ⅲ secretion system(T3SS). The T3SS is composed of a syringe-like needle structure and the effector proteins that are injected directly into a target host cell to disrupt a cellular response. The three human pathogenic Yersinia spp.(Y. pestis, Y. enterocolitica, and Y. pseudotuberculosis) are Gramnegative bacteria that share in common a 70 kb virulence plasmid which encodes the T3 SS. Translocation of the Yersinia effector proteins(YopE, YopH, YopT, YopM, YpkA/YopO, and YopP/J) into the target host cell results in disruption of the actin cytoskeleton to inhibit phagocytosis, downregulation of proinflammatory cytokine/chemokine production, and induction of cellular apoptosis of the target cell. Over the past 25 years, studies on the Yersinia effector proteins have unveiled tremendous knowledge of how the effectors enhance Yersinia virulence. Recently, the long awaited crystal structure of YpkA has been solved providing further insights into the activation of the YpkA kinase domain. Multisite autophosphorylation by YpkA to activate its kinase domain was also shown and postulated to serve as a mechanism to bypass regulation by host phosphatases. In addition, novel Yersinia effector protein targets, such as caspase-1, and signaling pathways including activation of the inflammasome were identified. In this review, we summarize the recent discoveries made on Yersinia effector proteins and their contribution to Yersinia pathogenesis.展开更多
通过构建嗜水气单胞菌AH-1 Quorum Sensing(QS)2个关键调节基因ahyI,ahyR的突变菌株,来系统分析嗜水气单胞菌AH-1Ⅲ型分泌系统基因,揭示它们由QS系统调控.在ahyI突变菌中,TTSS分泌效应因子(effector)aexT量显著提高.通过构建LacZ-TTSS...通过构建嗜水气单胞菌AH-1 Quorum Sensing(QS)2个关键调节基因ahyI,ahyR的突变菌株,来系统分析嗜水气单胞菌AH-1Ⅲ型分泌系统基因,揭示它们由QS系统调控.在ahyI突变菌中,TTSS分泌效应因子(effector)aexT量显著提高.通过构建LacZ-TTSS基因启动子融合表达,进一步表明QS系统负调控编码TTSS组分的基因.展开更多
基金funded by the National Natural Science Foundation of China (32371341,31872674)the Scientific and Technologic Foundation of Jilin Province (20230202050NC)the Fundamental Research Funds for the Central Universities (CGZH202206)。
文摘Type 1 diabetes mellitus(T1DM) lacks insulin secretion due to autoimmune deficiency of pancreaticβ-cells.Protecting pancreatic islets and enhancing insulin secretion has been therapeutic approaches.Mannogalactoglucan is the main type of polysaccharide from natural mushroom,which has potential medicinal prospects.Nevertheless,the antidiabetic property of mannogalactoglucan in T1DM has not been fully elucidated.In this study,we obtained the neutral fraction of alkali-soluble Armillaria mellea polysaccharide(AAMP-N) with the structure of mannogalactoglucan from the fruiting body of A.mellea and investigated the potential therapeutic value of AAMP-N in T1DM.We demonstrated that AAMP-N lowered blood glucose and improved diabetes symptoms in T1DM mice.AAMP-N activated unfolded protein response(UPR) signaling pathway to maintain ER protein folding homeostasis and promote insulin secretion in vivo.Besides that,AAMP-N promoted insulin synthesis via upregulating the expression of transcription factors,increased Ca^(2+) signals to stimulate intracellular insulin secretory vesicle transport via activating calcium/calmodulin-dependent kinase Ⅱ(CamkⅡ) and cAMP/PKA signals,and enhanced insulin secretory vesicle fusion with the plasma membrane via vesicle-associated membrane protein 2(VAMP2).Collectively,these studies demonstrated that the therapeutic potential of AAMP-N on pancreatic islets function,indicating that mannogalactoglucan could be natural nutraceutical used for the treatment of T1DM.
基金This research was supported by the association "Vaincre la Mucoviscidose" of France
文摘Objective Pseudomonas aeruginosa is a ubiquitous and opportunistic pathogen that uses the type Ⅲ secretion system （TTSS） to inject effector proteins directly into the cytosol of target cells to subvert the host cell＇s functions. Specialized bacterial chaperones are required for effective secretion of some effectors. To identify the chaperone of ExoS, the representative effector secreted by the TTSS of P aeruginosa, we analyzed the role of a postulated chaperone termed Orfl. Methods By allelic exchange, we constructed the mutant with the deletion of gene Orfl. Analysis of secreted and cell-associated fractions was performed by SDS-PAGE and Western blotting. Using strain expressing in trans Orfl, tagged by V5 polypeptide and histidine, protein-protein interaction was determined by affinity resin pull-down assay in combination with MALDI-TOF The role of Orfl in the expression of exoS was evaluated by gene reporter analysis. Results Pull-down assay showed that Orfl binds to ExoS and ExoT. Secretion profile analysis showed that Orfl was necessary for the optimal secretion of ExoS and ExoT. However, Orfl had no effect on the expression of exoS. Conclusion Orfl is important for the secretion of ExoS probably by maintaining ExoS in a secretion-competent conformation. We propose to name Orfl as SpcS for ＂specific Pseudomonas chaperone for ExoS＂.
基金supported by grants from the National Program on Key Basic Research Project of China(973 Program),No.2014CB542206the National Natural Science Foundation of China,No.81201389,30973052Program for Changjiang Scholars and Innovative Research Team in University of Ministry of Education of China,No.IRT13051
文摘Neuregulin-1 type Ⅲ is a key regulator in Schwann cell proliferation, committing to a myelinat- ing fate and regulating myelin sheath thickness. However, the expression pattern of neuregulin- 1 type III in the peripheral nervous system during developmental periods （such as the premyelin- ating stage, myelinating stage and postmyelinating stage） has rarely been studied. In this study, dorsal root ganglia were isolated from rats between postnatal day 1 and postnatal day 56. The expression pattern of neuregulin-1 type III in dorsal root ganglia neurons at various develop- mental stages were compared by quantitative real-time polymerase chain reaction, western blot assay and immunofluorescent staining. The expression of neuregulin-I type Ⅲ mRNA reached its peak at postnatal day 3 and then stabilized at a relative high expression level from postnatal day 3 to postnatal day 56. The expression of neuregulin-1 type III protein increased gradually from postnatal day 1, reached a peak at postnatal day 28, and then decreased at postnatal day 56. Immunofluorescent staining results showed a similar tendency to western blot assay results. Experimental findings indicate that the expression of neuregulin-1 type III in rat dorsal root ganglion was increased during the premyelinating （from postnatal day 2 to postnatal day 5） and myelinating stage （from postnatal day 5 to postnatal day 10）, but remained at a high level in the postmyelinating stage （after postnatal day 10）.
文摘Objective To investigate the effects of TGF- β1 on proliferation and type N collagen mRNAexpression of rat renal interstitial libroblasts in vitro for elucidating the role of fibroblasts in renal interstitialfibrosis. Methods The cells were cultured in media containing various concentrations of TGF- β1. Theproliferation and the type Ⅲ collagen mRNA expression of the cells were assayed with MTT method andRT- PCR respectively. Results TGF- β1, has a stimulating proliferation effect with dose- dependence and aincreasing expression of type Ⅲ collagen mRNA with both dose- and time- dependence to the renal fibroblasts. Conclusion It is suggested that TGF- β, is involved in proliferation of renal fibroblasts and their type Ⅲcollagen mRNA expression, and may play an important role in renal interstitial fibrosis.
文摘目的探讨细胞外信号调节激酶1/2(ERK1/2)在转化生长因子-β1(TGF-β1)诱导的肺成纤维细胞合成Ⅰ、Ⅲ型胶原蛋白中的作用,及新型过氧化物酶Peroxiredoxin-1(Prx-1)对该作用的影响。方法体外培养肺成纤维细胞随机分为4组:对照组(0.4%血清)、TGF-β1组(5μg/L)、阴性转染组(TGF-β1+阴性对照si RNA)和Prx-1 si RNA转染组(TGF-β1+Prx-1 si RNA)。采用脂质体转染法转染si RNA,实时定量逆转录-聚合酶链反应(RT-PCR)检测转染后Prx-1 m RNA表达;Western blot检测Ⅰ和Ⅲ型胶原蛋白、ERK1/2及Prx-1表达;2,7-二氯荧光素二乙酸(DCFH-DA)检测活性氧(ROS)水平。结果 Prx-1 si RNA转染肺成纤维细胞后,Prx-1 m RNA表达明显降低,最大抑制率为92%。与对照组比较,TGF-β1组的Ⅰ和Ⅲ型胶原蛋白、ROS、磷酸化ERK1/2(p-ERK1/2)及Prx-1蛋白的表达水平均明显提高。与TGF-β1组比较,阴性转染组中的上述观察指标无明显变化,但Prx-1转染组的Ⅰ和Ⅲ型胶原蛋白、ROS、p-ERK1/2水平进一步提高,而Prx-1蛋白的表达被抑制。结论 TGF-β1能够诱导肺成纤维细胞生成ROS,并促进ERK1/2通路的激活,导致Ⅰ、Ⅲ型胶原蛋白合成增加,而Prx-1 si RNA可通过提高ROS水平进一步促进TGF-β1该作用。
基金Supported by the ASM Robert D Watkins Graduate FellowshipUC Davis Hellman Fellowship
文摘The innate immune system is the first line of defense against invading pathogens. Innate immune cells recognize molecular patterns from the pathogen and mount a response to resolve the infection. The production of proinflammatory cytokines and reactive oxygen species, phagocytosis, and induced programmed cell death are processes initiated by innate immune cells in order to combat invading pathogens. However, pathogens have evolved various virulence mechanisms to subvert these responses. One strategy utilized by Gram-negative bacterial pathogens is the deployment of a complex machine termed the type Ⅲ secretion system(T3SS). The T3SS is composed of a syringe-like needle structure and the effector proteins that are injected directly into a target host cell to disrupt a cellular response. The three human pathogenic Yersinia spp.(Y. pestis, Y. enterocolitica, and Y. pseudotuberculosis) are Gramnegative bacteria that share in common a 70 kb virulence plasmid which encodes the T3 SS. Translocation of the Yersinia effector proteins(YopE, YopH, YopT, YopM, YpkA/YopO, and YopP/J) into the target host cell results in disruption of the actin cytoskeleton to inhibit phagocytosis, downregulation of proinflammatory cytokine/chemokine production, and induction of cellular apoptosis of the target cell. Over the past 25 years, studies on the Yersinia effector proteins have unveiled tremendous knowledge of how the effectors enhance Yersinia virulence. Recently, the long awaited crystal structure of YpkA has been solved providing further insights into the activation of the YpkA kinase domain. Multisite autophosphorylation by YpkA to activate its kinase domain was also shown and postulated to serve as a mechanism to bypass regulation by host phosphatases. In addition, novel Yersinia effector protein targets, such as caspase-1, and signaling pathways including activation of the inflammasome were identified. In this review, we summarize the recent discoveries made on Yersinia effector proteins and their contribution to Yersinia pathogenesis.
文摘通过构建嗜水气单胞菌AH-1 Quorum Sensing(QS)2个关键调节基因ahyI,ahyR的突变菌株,来系统分析嗜水气单胞菌AH-1Ⅲ型分泌系统基因,揭示它们由QS系统调控.在ahyI突变菌中,TTSS分泌效应因子(effector)aexT量显著提高.通过构建LacZ-TTSS基因启动子融合表达,进一步表明QS系统负调控编码TTSS组分的基因.
