Diderm bacteria,characterized by an additional lipid membrane layer known as the outer membrane,fold their outer membrane proteins(OMPs)via theβ-barrel assembly machinery(BAM)complex.Understanding how the BAM complex...Diderm bacteria,characterized by an additional lipid membrane layer known as the outer membrane,fold their outer membrane proteins(OMPs)via theβ-barrel assembly machinery(BAM)complex.Understanding how the BAM complex,particularly its key component BamA,assists in OMP folding remains crucial in bacterial cell biology.Recent research has focused primarily on the structural and functional characteristics of BamA within the Gracilicutes clade,such as in Escherichia coli(E.coli).However,another major evolutionary branch,Terrabacteria,has received comparatively less attention.An example of a Terrabacteria is Deinococcus radiodurans(D.radiodurans),a Gram-positive bacterium that possesses a distinctive outer membrane structure.In this study,we first demonstrated that theβ-barrel domains of BamA are not interchangeable between D.radiodurans and E.coli.The structure of D.radiodurans BamA was subsequently determined at 3.8Åresolution using cryo-electron microscopy,revealing obviously distinct arrangements of extracellular loop 4(ECL4)and ECL6 after structural comparison with their counterparts in gracilicutes.Despite the overall similarity in the topology of theβ-barrel domain,our results indicate that certain ECLs have evolved into distinct structures between the Terrabacteria and Gracilicutes clades.While BamA and its function are generally conserved across diderm bacterial species,our findings underscore the evolutionary diversity of this core OMP folder among bacteria,offering new insights into bacterial physiology and evolutionary biology.展开更多
The aerobic granular sludge was cultivated in a pilot-scale sequencing batch reactor （SBR）, and some of the granules were stored at 8 ℃ for 150 d. Extracellular polymeric substances （EPS） of sludge samples were e...The aerobic granular sludge was cultivated in a pilot-scale sequencing batch reactor （SBR）, and some of the granules were stored at 8 ℃ for 150 d. Extracellular polymeric substances （EPS） of sludge samples were extracted and analyzed during the granulation and storage process. The results show that the contents of protein and EPS increase along with the granulation process, while polysaccharides remain almost unchanged. The content of protein in EPS is almost two-fold larger than that of polysaccharides in granular sludge cultivated with municipal wastewater. Moreover, some of the granules disintegrate during storage, corresponding to the decrease of protein contents in EPS. Three peaks are identified in three-dimensional excitation emission matrix （EEM） fluorescence spectra of the EPS in the aerobic granules. Two peaks （A and B） are attributed to the protein-like fluorophores, and the third （peak C） is related to visible fulvic-like substances. Peak A gradually disappears during storage, while a new peak related to ultraviolet fulvic acid （peak D） is formed. The formation and the stability of aerobic granules are closely dependent on the quantity and composition of EPS proteins. Peak C has no obvious changes during granulation, while the fulvic-like substances present an increase in fluorescence intensities during storage, accompanied with an increase in structural complexity. The fulvie-like substances are also associated with the disintegration of the aerobic granules.展开更多
In this paper,Sulfobacillus thermosulfidooxidans ST was selected for use in bioleaching of pyrite and chalcopyrite.The adsorption experiments revealed that more cells were adsorbed on the surface of pyrite than on the...In this paper,Sulfobacillus thermosulfidooxidans ST was selected for use in bioleaching of pyrite and chalcopyrite.The adsorption experiments revealed that more cells were adsorbed on the surface of pyrite than on the surface of chalcopyrite.The role of extracellular DNA(eDNA)in the bioleaching process was investigated by depletion of eDNA using DNase I.The number of cells attached on the chalcopyrite and pyrite surfaces decreased on a large scale,and the lag phase of cell growth increased,causing the leaching percentages of pyrite and chalcopyrite to decrease by approximately 11.6%and 20.5%,respectively.The formation and distribution of eDNA secreted during bioleaching was assessed by a fluorescent dye-based method and visualized by confocal laser scanning microscopy(CLSM).The content of eDNA increased with bioleaching time.Furthermore,ST showed a stronger capacity to produce eDNA on the surface of pyrite than on the surface of chalcopyrite.These results showed that the removal of eDNA has a more significant effect on the bioleaching of chalcopyrite than on pyrite.展开更多
The most common age-related neurodegenerative disease is Alzheimer disease(AD),the imbalance between Aβ generation and clearance accumulated in extracellular plaques(ECS). And aggregated hyperphosphorylated tau prote...The most common age-related neurodegenerative disease is Alzheimer disease(AD),the imbalance between Aβ generation and clearance accumulated in extracellular plaques(ECS). And aggregated hyperphosphorylated tau protein in intraneuronal neurofibrillary tangles, together with loss of cholinergic neurons, synaptic alterations, and chronic inflammation within the brain. These lead to progressive impairment of cognitive function. The sporadic form of AD is characterized by an overall impairment in Aβ clearance.(1) Immunotherapy for AD: Immunotherapy targeting Aβ clearance is believed to be a promising approach and is under active clinical investigation. Autophagy is a conserved pathway for degrading abnormal protein aggregates and is crucial for Aβ clearance. We found that oral vaccination with a recombinant AAV/Aβ vaccine increased the clearance of Aβ from the brain and improved cognitive ability in AD animal models, while the underlying mechanisms were not well understood. In this study, we first demonstrated that oral vaccination with r AAV/Aβ decreased the p62 level and up-regulated the LC3 B-Ⅱ/LC3 B-Ⅰ ratio in APP/PS1 mouse brain, suggesting enhanced autophagy. Further, inhibition of the Akt/m TOR pathway may account for autophagy enhancement. We also found increased anti-Aβ antibodies in the sera of APP/PS1 mice with oral vaccination, accompanied by elevation of complement factors C1 q and C3 levels in the brain. Our results indicate that autophagy is closely involved in oral vaccination-induced Aβ clearance and modulating the autophagy pathway may be an important strategy for pre-clinical stage of AD(PCAD) prevention and intervention.(2)ECS in AD: Strategies that promote local growth of lymphatic vessels have the potential to improve clearance Aβ by meningeal lymphatics. whether enhancing clearance at the blood–brain barrier can improve lymphatic drainage function, remains to be addressed. Understanding substance transportation in brain ECS, especially in deep brain is essential for the complete answer to the question of how brain functions in the absence of a lymphatic drainage pathway. The brain interstitial fluid(ISF) drainage in deep brain was recently studied using a tracer-based MRI method, and a non-uniform ISF drainage was demonstrated with various distribution territories and movement speeds in different regions. In addition, Aβ deposition in the ECS and rescued memory in an APP/PS1 transgenic mouse of AD model,our findings are expected to have a potentially significant influence on brain-inspired and artificial intelligence, the future of which is very promising. Immunotherapy and local brain drug delivery via the brain ECS could circumvent the BBB and reduce systemic toxicity. Updated knowledge of the whole-brain ISF drainage system(ISS) provides a beneficial reference for improving the immunotherapeutic efficacy via ISS. New insights into how behave our and genetics modify ECS-ISS function should lead to the development of new preventive tools for PCAD and novel immunotherapy targeting Aβ clearance therapeutic targets.展开更多
The interaction between the cell membrane and the extracellular matrix is crucial for many cellular functions by modulating mechanosensitive signaling pathways.Physical properties of the extracellular matrix such as s...The interaction between the cell membrane and the extracellular matrix is crucial for many cellular functions by modulating mechanosensitive signaling pathways.Physical properties of the extracellular matrix such as stiffness and topography affect such interactions.Our recent work reveals that surface topography of tens to hundreds of nanometer scale modulates cell signaling by activating intracellular curvature-sensitive proteins.We use vertical nanostructures protruding from a flat surface as a platform to induce precise curvatures on the cell membrane and to probe biological processes in live cells.Vertical nanopillars deform the plasma membrane inwards and induce membrane curvature when the cell engulfs them,leading to a reduction of the membrane-substrate gap distance.We found that the high membrane curvature induced by vertical nanopillars significantly affects the distribution of curvature-sensitive proteins and stimulates several cellular processes in live cells including cellular endocytosis and cytoskeleton dynamics.Our studies show a strong interplay between biological cells and nano-featured surfaces,which is an essential consideration for future development of interfacing devices.展开更多
When the loop-mediated isothermal amplification(LAMP)assay is used for detecting target genes,DNA extraction is unnecessary in many cases.Simple pretreatment(e.g.heating)is enough to obtain rather sensitive responses....When the loop-mediated isothermal amplification(LAMP)assay is used for detecting target genes,DNA extraction is unnecessary in many cases.Simple pretreatment(e.g.heating)is enough to obtain rather sensitive responses.Even test samples without any pretreatment can be used as template.This feature suggests that LAMP is superior to PCR in developing point-of-care test strategies.In this study,using Stx1 gene from E.coli as model,we verified that viable cells,dead cells and extracellular DNA could function as template in the LAMP assay.In the incubation at 63℃,viable bacteria in the LAMP reaction mixture lysed completely within 2 min,providing DNA template for nucleic acid amplification.The Stx1 gene in diluted culture medium,spiked tap water,spiked seawater and real seawater all could be detected,with or without the step of DNA extraction.We found that the complex substances in real sample(e.g.natural seawater)exhibited considerable inhibitory effect on the sensitivity of the LAMP assay.These outcomes are meaningful for building a point-of-care strategy by employing the LAMP assay for environmental monitoring,bio-resource surveys,food safety,etc.in particular those based on environmental DNA.展开更多
Background:XIAP-associated factor 1(XAF1)negatively regulates the function of the X-linked inhibitor of apoptosis protein(XIAP),a member of the IAP family that exerts antiapoptotic effects.The extracellular signal-reg...Background:XIAP-associated factor 1(XAF1)negatively regulates the function of the X-linked inhibitor of apoptosis protein(XIAP),a member of the IAP family that exerts antiapoptotic effects.The extracellular signal-regulated kinase(ERK)pathway is thought to increase cell proliferation and to protect cells from apoptosis.The aim of the study was to investigate the correlation between the ERK1/2 signaling pathway and XAF1 in colon cancer.Methods:Four human colon cancer cell lines,HCT1116 and Lovo(wildtype p53),DLD1 and SW1116(mutant p53),were used.Lovo stable transfectants with XAF1 sense and antisense were established.The effects of dominant-negative MEK1(DN-MEK1)and MEK-specific inhibitor U0126 on the ERK signaling pathway and expression of XAF1 and XIAP proteins were determined.The transcription activity of core XAF1 promoter was assessed by dual luciferase reporter assay.Cell proliferation was measured by MTT assay.Apoptosis was determined by Hoechst 33258 staining.Results:U0126 increased the expression of XAF1 in a time-and dose-dependent manner.A similar result was obtained in cells transfected with DN-MEK1 treatment.Conversely,the expression of XIAP was down-regulated.Activity of the putative promoter of the XAF1 gene was significantly increased by U0126 treatment and DN-MEK1 transient transfection.rhEGF-stimulated phosphorylation of ERK appeared to have little or no effect on XAF1 expression.Overexpression of XAF1 was more sensitive to U0126-induced apoptosis,whereas down-regulation of XAF1 by antisense reversed U0126-induced inhibition of cell proliferation.Conclusions:XAF1 expression was up-regulated by inhibition of the ERK1/2 pathway through transcriptional regulation,which required de novo protein synthesis.The results suggest that XAF1 mediates apoptosis induced by the ERK1/2 pathway in colon cancer.展开更多
Extracellular polysaccharide from Tremella aurantialba (TEP) significantly inhibited H2O2-induced haemolysis of RBCs, significantly decreased the amount of malondialdehyde (MDA) produced in mice liver homogenates as a...Extracellular polysaccharide from Tremella aurantialba (TEP) significantly inhibited H2O2-induced haemolysis of RBCs, significantly decreased the amount of malondialdehyde (MDA) produced in mice liver homogenates as a result of autoxidation, and inhibited VitC-Fe2+-induced swelling of liver mitochondria.展开更多
目的:探究尿液细胞外囊泡中关键蛋白细胞骨架相关蛋白4(cytoskeleton⁃associated protein 4,CKAP4)作为生物标志物预测糖尿病肾病(diabetic kidney disease,DKD)进展的能力。方法:纳入南京医科大学第一附属医院143例肾活检证实DKD的2型...目的:探究尿液细胞外囊泡中关键蛋白细胞骨架相关蛋白4(cytoskeleton⁃associated protein 4,CKAP4)作为生物标志物预测糖尿病肾病(diabetic kidney disease,DKD)进展的能力。方法:纳入南京医科大学第一附属医院143例肾活检证实DKD的2型糖尿病患者以及肾脏恶性肿瘤患者10例。免疫组化法检测肾组织中CKAP4的表达水平。采用Spearman相关分析CKAP4表达水平与临床指标的相关性。计算受试者工作特征(receiver operating characteristic,ROC)曲线下面积(area under the curve,AUC),以评估CKAP4表达是否能有效区分肾脏预后不良的患者。采用ROC曲线分析确定预测肾脏事件的CKAP4评分的最佳临界值(最高约登指数)。此外,进行时间依赖性AUC分析,以表征肾活检后6个月以上CKAP4的预测准确性。通过Cox比例风险模型随访CKAP4和DKD进展之间的风险比。将单变量分析中有统计学意义(P<0.05)的临床病理因素作为协变量纳入多变量Cox比例风险模型分析。采用Kaplan⁃Meier分析评价CKAP4高表达组和CKAP4低表达组肾活检后6个月以上生存的差异。结果:与肿瘤患者癌旁正常肾脏组织相比,CKAP4在DKD患者肾脏组织中表达增高,差异有统计学意义(P<0.001)。CKAP4在不同分期的DKD患者肾组织中表达有差异,其中Ⅱ期与Ⅲ期、Ⅱ期与Ⅳ期、Ⅲ期与Ⅳ期之间的差异均有统计学意义(P均<0.05)。DKD患者CKAP4表达与血清肌酐、尿素氮、24 h尿蛋白呈正相关,与肾小球滤过率估计值、血红蛋白呈负相关。DKD患者中位随访期为2.22年,63例(44.06%)患者出现DKD进展。Pearson相关分析显示,CKAP4表达水平随DKD病理分级的升高而升高(r=0.808,P<0.001)。多因素Cox回归分析显示,CKAP4高表达与DKD进展的风险增加相关(HR=4.120,95%CI:1.730~9.811,P=0.001)。此外,在Kaplan⁃Meier生存分析中,CKAP4高表达组患者的肾脏终点事件的发生率显著高于CKAP4低表达组(P<0.001)。绘制了包括CKAP4分类在内的列线图来预测DKD进展风险(C⁃index:0.689)。结论:来源于尿液细胞外囊泡的CKAP4是DKD患者肾活检后6个月以上疾病进展的独立危险因素。展开更多
目的探讨细胞外水分比率(ratio of extracellular water to total body water,ETR)对急性缺血性卒中(acute ischemic stroke,AIS)患者出院结局的影响。方法回顾性分析2018年7月—2024年6月在北京市房山区良乡医院神经内科住院的AIS患者...目的探讨细胞外水分比率(ratio of extracellular water to total body water,ETR)对急性缺血性卒中(acute ischemic stroke,AIS)患者出院结局的影响。方法回顾性分析2018年7月—2024年6月在北京市房山区良乡医院神经内科住院的AIS患者,根据出院结局将患者分为结局良好(mRS评分<3分)组和结局不良(mRS评分≥3分)组。比较两组患者的临床资料和ETR等指标的差异,采用多因素logistic回归分析探讨ETR对患者出院结局的影响,并在不同性别、年龄、BMI、骨骼肌质量指数及入院NIHSS评分亚组中分析ETR对出院结局的影响。结果共纳入AIS患者147例,平均年龄(68.9±12.2)岁,其中男性80例(54.4%),结局良好组107例(72.8%),结局不良组40例(27.2%)。多因素l ogi sti c回归分析显示,ETR升高(OR 2.09,95%CI 1.16~3.76,P=0.014)和ETR≥40.0%(OR 6.24,95%CI 1.33~29.32,P=0.020)是AI S患者出院结局不良的危险因素。亚组分析结果稳健,无明显交互作用(P交互>0.05)。在年龄<71岁、BMI≥24 kg/m2以及入院NIHSS评分≥3分亚组中,ETR对出院结局不良的影响具有显著性。结论ETR是AIS患者出院结局的独立影响因素,高ETR可显著增加出院结局不良的风险。展开更多
基金supported by the Fundamental Research Funds for the Central Universities(WK9100000063)the Fundamental Research Funds for the Central Universities(WK9100000031)+3 种基金the National Natural Science Foundation of China(32270035,32271241)the Anhui Provincial Natural Science Foundation(2208085MC40,2008085QC98)the Talent Fund Project of Biomedical Sciences and Health Laboratory of Anhui Province,University of Science and Technology of China(BJ9100000003)the start-up funding from the University of Science and Technology of China(KY9100000034,KJ2070000082).
文摘Diderm bacteria,characterized by an additional lipid membrane layer known as the outer membrane,fold their outer membrane proteins(OMPs)via theβ-barrel assembly machinery(BAM)complex.Understanding how the BAM complex,particularly its key component BamA,assists in OMP folding remains crucial in bacterial cell biology.Recent research has focused primarily on the structural and functional characteristics of BamA within the Gracilicutes clade,such as in Escherichia coli(E.coli).However,another major evolutionary branch,Terrabacteria,has received comparatively less attention.An example of a Terrabacteria is Deinococcus radiodurans(D.radiodurans),a Gram-positive bacterium that possesses a distinctive outer membrane structure.In this study,we first demonstrated that theβ-barrel domains of BamA are not interchangeable between D.radiodurans and E.coli.The structure of D.radiodurans BamA was subsequently determined at 3.8Åresolution using cryo-electron microscopy,revealing obviously distinct arrangements of extracellular loop 4(ECL4)and ECL6 after structural comparison with their counterparts in gracilicutes.Despite the overall similarity in the topology of theβ-barrel domain,our results indicate that certain ECLs have evolved into distinct structures between the Terrabacteria and Gracilicutes clades.While BamA and its function are generally conserved across diderm bacterial species,our findings underscore the evolutionary diversity of this core OMP folder among bacteria,offering new insights into bacterial physiology and evolutionary biology.
基金Project(2006AA06Z318) supported by the National High-Tech Research and Development Program of China
文摘The aerobic granular sludge was cultivated in a pilot-scale sequencing batch reactor （SBR）, and some of the granules were stored at 8 ℃ for 150 d. Extracellular polymeric substances （EPS） of sludge samples were extracted and analyzed during the granulation and storage process. The results show that the contents of protein and EPS increase along with the granulation process, while polysaccharides remain almost unchanged. The content of protein in EPS is almost two-fold larger than that of polysaccharides in granular sludge cultivated with municipal wastewater. Moreover, some of the granules disintegrate during storage, corresponding to the decrease of protein contents in EPS. Three peaks are identified in three-dimensional excitation emission matrix （EEM） fluorescence spectra of the EPS in the aerobic granules. Two peaks （A and B） are attributed to the protein-like fluorophores, and the third （peak C） is related to visible fulvic-like substances. Peak A gradually disappears during storage, while a new peak related to ultraviolet fulvic acid （peak D） is formed. The formation and the stability of aerobic granules are closely dependent on the quantity and composition of EPS proteins. Peak C has no obvious changes during granulation, while the fulvic-like substances present an increase in fluorescence intensities during storage, accompanied with an increase in structural complexity. The fulvie-like substances are also associated with the disintegration of the aerobic granules.
基金Projects(31470230,51320105006,51604308)supported by the National Natural Science Foundation of ChinaProject(2017RS3003)supported by the Youth Talent Foundation of Hunan Province of China+1 种基金Project(2018JJ2486)supported by the Natural Science Foundation of Hunan Province of ChinaProject(2018WK2012)supported by the Key Research and Development Projects in Hunan Province,China。
文摘In this paper,Sulfobacillus thermosulfidooxidans ST was selected for use in bioleaching of pyrite and chalcopyrite.The adsorption experiments revealed that more cells were adsorbed on the surface of pyrite than on the surface of chalcopyrite.The role of extracellular DNA(eDNA)in the bioleaching process was investigated by depletion of eDNA using DNase I.The number of cells attached on the chalcopyrite and pyrite surfaces decreased on a large scale,and the lag phase of cell growth increased,causing the leaching percentages of pyrite and chalcopyrite to decrease by approximately 11.6%and 20.5%,respectively.The formation and distribution of eDNA secreted during bioleaching was assessed by a fluorescent dye-based method and visualized by confocal laser scanning microscopy(CLSM).The content of eDNA increased with bioleaching time.Furthermore,ST showed a stronger capacity to produce eDNA on the surface of pyrite than on the surface of chalcopyrite.These results showed that the removal of eDNA has a more significant effect on the bioleaching of chalcopyrite than on pyrite.
基金NBRD Program of China(2016YFC1306302 2016YFC1305903)+3 种基金National Natural Science Foundation of China(81571044 81471633 61450004 and 81171015)
文摘The most common age-related neurodegenerative disease is Alzheimer disease(AD),the imbalance between Aβ generation and clearance accumulated in extracellular plaques(ECS). And aggregated hyperphosphorylated tau protein in intraneuronal neurofibrillary tangles, together with loss of cholinergic neurons, synaptic alterations, and chronic inflammation within the brain. These lead to progressive impairment of cognitive function. The sporadic form of AD is characterized by an overall impairment in Aβ clearance.(1) Immunotherapy for AD: Immunotherapy targeting Aβ clearance is believed to be a promising approach and is under active clinical investigation. Autophagy is a conserved pathway for degrading abnormal protein aggregates and is crucial for Aβ clearance. We found that oral vaccination with a recombinant AAV/Aβ vaccine increased the clearance of Aβ from the brain and improved cognitive ability in AD animal models, while the underlying mechanisms were not well understood. In this study, we first demonstrated that oral vaccination with r AAV/Aβ decreased the p62 level and up-regulated the LC3 B-Ⅱ/LC3 B-Ⅰ ratio in APP/PS1 mouse brain, suggesting enhanced autophagy. Further, inhibition of the Akt/m TOR pathway may account for autophagy enhancement. We also found increased anti-Aβ antibodies in the sera of APP/PS1 mice with oral vaccination, accompanied by elevation of complement factors C1 q and C3 levels in the brain. Our results indicate that autophagy is closely involved in oral vaccination-induced Aβ clearance and modulating the autophagy pathway may be an important strategy for pre-clinical stage of AD(PCAD) prevention and intervention.(2)ECS in AD: Strategies that promote local growth of lymphatic vessels have the potential to improve clearance Aβ by meningeal lymphatics. whether enhancing clearance at the blood–brain barrier can improve lymphatic drainage function, remains to be addressed. Understanding substance transportation in brain ECS, especially in deep brain is essential for the complete answer to the question of how brain functions in the absence of a lymphatic drainage pathway. The brain interstitial fluid(ISF) drainage in deep brain was recently studied using a tracer-based MRI method, and a non-uniform ISF drainage was demonstrated with various distribution territories and movement speeds in different regions. In addition, Aβ deposition in the ECS and rescued memory in an APP/PS1 transgenic mouse of AD model,our findings are expected to have a potentially significant influence on brain-inspired and artificial intelligence, the future of which is very promising. Immunotherapy and local brain drug delivery via the brain ECS could circumvent the BBB and reduce systemic toxicity. Updated knowledge of the whole-brain ISF drainage system(ISS) provides a beneficial reference for improving the immunotherapeutic efficacy via ISS. New insights into how behave our and genetics modify ECS-ISS function should lead to the development of new preventive tools for PCAD and novel immunotherapy targeting Aβ clearance therapeutic targets.
基金supported by two NIH grants 1R01GM125737 and 1R01GM117263 to BC
文摘The interaction between the cell membrane and the extracellular matrix is crucial for many cellular functions by modulating mechanosensitive signaling pathways.Physical properties of the extracellular matrix such as stiffness and topography affect such interactions.Our recent work reveals that surface topography of tens to hundreds of nanometer scale modulates cell signaling by activating intracellular curvature-sensitive proteins.We use vertical nanostructures protruding from a flat surface as a platform to induce precise curvatures on the cell membrane and to probe biological processes in live cells.Vertical nanopillars deform the plasma membrane inwards and induce membrane curvature when the cell engulfs them,leading to a reduction of the membrane-substrate gap distance.We found that the high membrane curvature induced by vertical nanopillars significantly affects the distribution of curvature-sensitive proteins and stimulates several cellular processes in live cells including cellular endocytosis and cytoskeleton dynamics.Our studies show a strong interplay between biological cells and nano-featured surfaces,which is an essential consideration for future development of interfacing devices.
文摘When the loop-mediated isothermal amplification(LAMP)assay is used for detecting target genes,DNA extraction is unnecessary in many cases.Simple pretreatment(e.g.heating)is enough to obtain rather sensitive responses.Even test samples without any pretreatment can be used as template.This feature suggests that LAMP is superior to PCR in developing point-of-care test strategies.In this study,using Stx1 gene from E.coli as model,we verified that viable cells,dead cells and extracellular DNA could function as template in the LAMP assay.In the incubation at 63℃,viable bacteria in the LAMP reaction mixture lysed completely within 2 min,providing DNA template for nucleic acid amplification.The Stx1 gene in diluted culture medium,spiked tap water,spiked seawater and real seawater all could be detected,with or without the step of DNA extraction.We found that the complex substances in real sample(e.g.natural seawater)exhibited considerable inhibitory effect on the sensitivity of the LAMP assay.These outcomes are meaningful for building a point-of-care strategy by employing the LAMP assay for environmental monitoring,bio-resource surveys,food safety,etc.in particular those based on environmental DNA.
基金Shanghai Medical Key Discipline Construction Foundation(05-Ⅲ-005-017).
文摘Background:XIAP-associated factor 1(XAF1)negatively regulates the function of the X-linked inhibitor of apoptosis protein(XIAP),a member of the IAP family that exerts antiapoptotic effects.The extracellular signal-regulated kinase(ERK)pathway is thought to increase cell proliferation and to protect cells from apoptosis.The aim of the study was to investigate the correlation between the ERK1/2 signaling pathway and XAF1 in colon cancer.Methods:Four human colon cancer cell lines,HCT1116 and Lovo(wildtype p53),DLD1 and SW1116(mutant p53),were used.Lovo stable transfectants with XAF1 sense and antisense were established.The effects of dominant-negative MEK1(DN-MEK1)and MEK-specific inhibitor U0126 on the ERK signaling pathway and expression of XAF1 and XIAP proteins were determined.The transcription activity of core XAF1 promoter was assessed by dual luciferase reporter assay.Cell proliferation was measured by MTT assay.Apoptosis was determined by Hoechst 33258 staining.Results:U0126 increased the expression of XAF1 in a time-and dose-dependent manner.A similar result was obtained in cells transfected with DN-MEK1 treatment.Conversely,the expression of XIAP was down-regulated.Activity of the putative promoter of the XAF1 gene was significantly increased by U0126 treatment and DN-MEK1 transient transfection.rhEGF-stimulated phosphorylation of ERK appeared to have little or no effect on XAF1 expression.Overexpression of XAF1 was more sensitive to U0126-induced apoptosis,whereas down-regulation of XAF1 by antisense reversed U0126-induced inhibition of cell proliferation.Conclusions:XAF1 expression was up-regulated by inhibition of the ERK1/2 pathway through transcriptional regulation,which required de novo protein synthesis.The results suggest that XAF1 mediates apoptosis induced by the ERK1/2 pathway in colon cancer.
文摘Extracellular polysaccharide from Tremella aurantialba (TEP) significantly inhibited H2O2-induced haemolysis of RBCs, significantly decreased the amount of malondialdehyde (MDA) produced in mice liver homogenates as a result of autoxidation, and inhibited VitC-Fe2+-induced swelling of liver mitochondria.
文摘目的:探究尿液细胞外囊泡中关键蛋白细胞骨架相关蛋白4(cytoskeleton⁃associated protein 4,CKAP4)作为生物标志物预测糖尿病肾病(diabetic kidney disease,DKD)进展的能力。方法:纳入南京医科大学第一附属医院143例肾活检证实DKD的2型糖尿病患者以及肾脏恶性肿瘤患者10例。免疫组化法检测肾组织中CKAP4的表达水平。采用Spearman相关分析CKAP4表达水平与临床指标的相关性。计算受试者工作特征(receiver operating characteristic,ROC)曲线下面积(area under the curve,AUC),以评估CKAP4表达是否能有效区分肾脏预后不良的患者。采用ROC曲线分析确定预测肾脏事件的CKAP4评分的最佳临界值(最高约登指数)。此外,进行时间依赖性AUC分析,以表征肾活检后6个月以上CKAP4的预测准确性。通过Cox比例风险模型随访CKAP4和DKD进展之间的风险比。将单变量分析中有统计学意义(P<0.05)的临床病理因素作为协变量纳入多变量Cox比例风险模型分析。采用Kaplan⁃Meier分析评价CKAP4高表达组和CKAP4低表达组肾活检后6个月以上生存的差异。结果:与肿瘤患者癌旁正常肾脏组织相比,CKAP4在DKD患者肾脏组织中表达增高,差异有统计学意义(P<0.001)。CKAP4在不同分期的DKD患者肾组织中表达有差异,其中Ⅱ期与Ⅲ期、Ⅱ期与Ⅳ期、Ⅲ期与Ⅳ期之间的差异均有统计学意义(P均<0.05)。DKD患者CKAP4表达与血清肌酐、尿素氮、24 h尿蛋白呈正相关,与肾小球滤过率估计值、血红蛋白呈负相关。DKD患者中位随访期为2.22年,63例(44.06%)患者出现DKD进展。Pearson相关分析显示,CKAP4表达水平随DKD病理分级的升高而升高(r=0.808,P<0.001)。多因素Cox回归分析显示,CKAP4高表达与DKD进展的风险增加相关(HR=4.120,95%CI:1.730~9.811,P=0.001)。此外,在Kaplan⁃Meier生存分析中,CKAP4高表达组患者的肾脏终点事件的发生率显著高于CKAP4低表达组(P<0.001)。绘制了包括CKAP4分类在内的列线图来预测DKD进展风险(C⁃index:0.689)。结论:来源于尿液细胞外囊泡的CKAP4是DKD患者肾活检后6个月以上疾病进展的独立危险因素。
文摘目的探讨细胞外水分比率(ratio of extracellular water to total body water,ETR)对急性缺血性卒中(acute ischemic stroke,AIS)患者出院结局的影响。方法回顾性分析2018年7月—2024年6月在北京市房山区良乡医院神经内科住院的AIS患者,根据出院结局将患者分为结局良好(mRS评分<3分)组和结局不良(mRS评分≥3分)组。比较两组患者的临床资料和ETR等指标的差异,采用多因素logistic回归分析探讨ETR对患者出院结局的影响,并在不同性别、年龄、BMI、骨骼肌质量指数及入院NIHSS评分亚组中分析ETR对出院结局的影响。结果共纳入AIS患者147例,平均年龄(68.9±12.2)岁,其中男性80例(54.4%),结局良好组107例(72.8%),结局不良组40例(27.2%)。多因素l ogi sti c回归分析显示,ETR升高(OR 2.09,95%CI 1.16~3.76,P=0.014)和ETR≥40.0%(OR 6.24,95%CI 1.33~29.32,P=0.020)是AI S患者出院结局不良的危险因素。亚组分析结果稳健,无明显交互作用(P交互>0.05)。在年龄<71岁、BMI≥24 kg/m2以及入院NIHSS评分≥3分亚组中,ETR对出院结局不良的影响具有显著性。结论ETR是AIS患者出院结局的独立影响因素,高ETR可显著增加出院结局不良的风险。
