目的探索酮还原酶家族1成员C3(aldo-keto reductase family 1 member C3,AKR1C3)对乳腺癌恶性细胞生物学行为的干预作用及对程序性细胞死亡蛋白/程序性死亡-配体1(programmed cell death protein1/programmed death-ligand1,PD-1/PD-L)...目的探索酮还原酶家族1成员C3(aldo-keto reductase family 1 member C3,AKR1C3)对乳腺癌恶性细胞生物学行为的干预作用及对程序性细胞死亡蛋白/程序性死亡-配体1(programmed cell death protein1/programmed death-ligand1,PD-1/PD-L)通路的影响。方法把MCF-7人乳腺癌细胞中NC组和AKR1C3组分别转染空质粒和AKR1C3质粒,采用MTT法检测转染后24 h、48 h、72 h细胞活力;采用流式细胞技术测定各组细胞的存活率以及早期、晚期凋亡比例;通过Transwell实验对各组细胞的迁移和侵袭能力进行检测;通过Western blot检测各组细胞PD-1、PD-L1、蛋白激酶B(protein kinase b,AKT)蛋白表达水平。使用C57BL/6小鼠构建荷瘤模型,将采用人乳腺癌MCF-7细胞转染NC质粒和AKR1C3质粒进行细胞荷瘤,每3 d测量瘤体积,持续21 d,绘制两组小鼠肿瘤生长曲线,并于实验终点测量肿瘤质量。结果相较于NC组,AKR1C3组细胞活力降低(P<0.05),并且具有时间依赖效应(P<0.05),迁移和侵袭能力降低(P<0.05),早期凋亡和晚期凋亡比例升高(P<0.05),PD-1、PD-L1、AKT蛋白表达水平降低(P<0.05)。动物实验表明,AKR1C3组小鼠肿瘤体积降低,肿瘤质量下降(P<0.05)。结论AKR1C3可以抑制人乳腺癌细胞恶性生物学行为,抑制PD-1/PDL1信号通路蛋白表达。展开更多
构建使用了PD-1抑制剂的肿瘤患者出现甲状腺功能障碍的风险预测模型,分析使用PD-1肿瘤抑制剂导致的甲状腺功能障碍的相关风险因素,设计监测预警系统。选取2020年—2023年广西医科大学附属肿瘤医院1225例使用PD-1抑制剂肿瘤患者的临床资...构建使用了PD-1抑制剂的肿瘤患者出现甲状腺功能障碍的风险预测模型,分析使用PD-1肿瘤抑制剂导致的甲状腺功能障碍的相关风险因素,设计监测预警系统。选取2020年—2023年广西医科大学附属肿瘤医院1225例使用PD-1抑制剂肿瘤患者的临床资料,包括人口学特征、既往史、实验室检测等63个变量。本文选取相关性前10/20/30/40/50/60个变量的4种传统机器学习模型进行性能比较。通过F1分数、灵敏度、准确率、精确率、特异性曲线下面积(Area Under the Curve,AUC)评估以上预测模型的性能,并利用Shapley加性解释(Shapley Additive Explanation,SHAP)可视化解释本文的机器学习模型。与促甲状腺激素相关性排名前10的变量依次为:羟丁酸脱氢酶、乳酸脱氢酶、淋巴细胞绝对值、天门冬氨酸转移酶、钙离子、碱性磷酸酶、谷氨酰转肽酶、单核细胞绝对值、红细胞分布宽度SD、胆碱酯酶。建立了使用PD-1抑制剂的肿瘤患者出现甲状腺功能障碍的风险预测模型,并在全局解释和局部解释的层面上分别作出模型预测结果影响的解释。展开更多
目的:探究卵巢癌(ovarian cancer,OV)浸润性CD4^(+)调节性T细胞(regulatory T cell,Treg)的脂质摄取与积累情况,及其与CD4^(+)Treg中程序性细胞死亡蛋白1(programmed cell death protein 1,PD-1)和细胞毒性T淋巴细胞相关蛋白-4(cytotoxi...目的:探究卵巢癌(ovarian cancer,OV)浸润性CD4^(+)调节性T细胞(regulatory T cell,Treg)的脂质摄取与积累情况,及其与CD4^(+)Treg中程序性细胞死亡蛋白1(programmed cell death protein 1,PD-1)和细胞毒性T淋巴细胞相关蛋白-4(cytotoxic Tlymphocyte associated protein 4,CTLA-4)表达的相关性。方法:采用亲脂性荧光染料BODIPY^(TM)493/503和荧光脂肪酸探针BODIPY^(TM)500/510 C1 C12分别检测OV组织中的或与不同OV细胞系(ES-2、SKOV3、CAOV3)上清液共培养的人源CD4^(+)Treg的细胞内脂质含量和脂质摄取能力;使用脂肪酸氧化抑制剂(Etomoxir)、脂肪酸合成抑制剂(C75)和脂肪酸摄取抑制剂磺基-N-琥珀酰亚胺油酸酯(sulfo-N-succinimidyl oleate,SSO)干预脂质代谢;通过流式细胞术分析CD4^(+)Treg上免疫抑制分子PD-1和CTLA-4的表达。结果:OV组织CD4^(+)Treg相比传统CD4^(+)T细胞表现出更高的脂质含量和脂质摄取能力(P均<0.01)。在体外实验中,与基础培养基相比,OV细胞培养上清可显著提升CD4^(+)Treg的胞内脂质含量与脂质摄取能力(P均<0.05),其中CAOV3来源的上清作用最为显著。此外,CAOV3上清液还能提升CD4^(+)Treg中PD-1与CTLA-4的表达(P<0.05)。CD4^(+)Treg的脂质积累随CAOV3上清浓度增加呈剂量依赖性上升(P<0.05),且其对胞外荧光脂肪酸类似物的摄取能力具有浓度依赖性(P<0.05)。脂肪酸摄取抑制剂SSO可有效逆转CAOV3上清液诱导的CD4^(+)Treg脂质积累及PD-1、CTLA-4的高表达(P<0.05);而脂肪酸氧化抑制剂Etomoxir与合成抑制剂C75则无显著影响。结论:OV微环境通过促进CD4^(+)Treg的脂质摄取,提高细胞内脂质含量,促进其免疫抑制分子PD-1和CTLA-4表达。靶向脂肪酸摄取途径可能是逆转OV中Treg介导的免疫抑制的潜在策略。展开更多
Leaves and glumes act as lateral organs and have essential effects on photosynthesis and seed morphology,thus affecting yield.However,the molecular mechanisms controlling their polarity development in rice still need ...Leaves and glumes act as lateral organs and have essential effects on photosynthesis and seed morphology,thus affecting yield.However,the molecular mechanisms controlling their polarity development in rice still need further study.Here,we isolated a polarity defect of lateral organs 1(pdl1)mutant in rice,which exhibits twisted/filamentous-shaped leaves and cracked/filamentous-shaped lemmas caused by defects in polarity development.PDL1 encodes a SUPPRESSOR OF GENE SILENCING 3 protein localized in the cytoplasmic granules.PDL1 is expressed in the shoot apical meristem,inflorescence meristem,floral meristem,and lateral organs including leaves and floral organs.PDL1 is involved in the synthesis of tasiR-ARF,which may subsequently modulate the expression of OsARFs.Meanwhile,the expression levels of abaxial miR165/166 and the adaxial identity genes OSHBs were respectively increased and reduced significantly.The results of this study clarify the molecular mechanism by which PDL1-mediated tasiR-ARF synthesis regulates the lateral organ polarity development in rice.展开更多
Rice blast,caused by Magnaporthe oryzae,is a fungal disease that causes devastating damage to rice production worldwide.During infection,pathogens secrete effector proteins that modulate plant immunity.Disulfide bond ...Rice blast,caused by Magnaporthe oryzae,is a fungal disease that causes devastating damage to rice production worldwide.During infection,pathogens secrete effector proteins that modulate plant immunity.Disulfide bond formation catalyzed by protein disulfide isomerases(PDI)is essential for protein folding and maturation.However,the biological function of Pdi1 in M.oryzae has not yet been characterized.In this study,we identified the endoplasmic reticulum(ER)-located protein,MoPdi1,in M.oryzae.MoPdi1 regulates conidiation,cell wall stress,and pathogenicity of M.oryzae.Furthermore,the CGHC active sites in the a and a'redox domain of MoPdi1 were essential for the biological function of MoPDI1.Further tests demonstrated that MoPdi1 was involved in the regulation of ER stress and positively regulated ER phagy.We also found that MoPdi1 interacted with MoHut1.Deletion of MoPDI1 led to the bereft of MoHut1 dimerization,which depends on the formation of disulfide bonds.In addition,MoPdi1 affected the normal secretion of the cytoplasmic effector AVR-Pia.We provided evidence that MoHut1 is important for the vegetative growth,conidiation,and pathogenicity in M.oryzae.Therefore,our findings could provide a suitable target point for designing antifungal agrochemicals against rice blast fungus.展开更多
The mechanistic target of rapamycin(m TOR) is a serine/threonine kinase that plays a pivotal role in cellular growth, proliferation, survival, and metabolism. In the central nervous system(CNS), the mTOR pathway regul...The mechanistic target of rapamycin(m TOR) is a serine/threonine kinase that plays a pivotal role in cellular growth, proliferation, survival, and metabolism. In the central nervous system(CNS), the mTOR pathway regulates diverse aspects of neural development and function. Genetic mutations within the m TOR pathway lead to severe neurodevelopmental disorders, collectively known as “mTORopathies”(Crino, 2020). Dysfunctions of m TOR, including both its hyperactivation and hypoactivation, have also been implicated in a wide spectrum of other neurodevelopmental and neurodegenerative conditions, highlighting its importance in CNS health.展开更多
旨在制备猫PD-L1抗原并筛选单抗,探讨其在犬、猫肿瘤检测和诊断中的应用,同时制备猫源化嵌合单链抗体,为猫肿瘤的治疗提供基础。本研究通过PCR扩增猫PD-L1基因,构建原核表达重组质粒PET-MPD-L1,通过在宿主BL21(DE3)中表达制备PD-L1抗原...旨在制备猫PD-L1抗原并筛选单抗,探讨其在犬、猫肿瘤检测和诊断中的应用,同时制备猫源化嵌合单链抗体,为猫肿瘤的治疗提供基础。本研究通过PCR扩增猫PD-L1基因,构建原核表达重组质粒PET-MPD-L1,通过在宿主BL21(DE3)中表达制备PD-L1抗原,免疫小鼠后细胞融合,ELISA方法进行杂交瘤细胞筛选,3次亚克隆后建立MPD-L1杂交瘤细胞系。测定杂交瘤细胞上清、腹水效价、抗体亚类、Western blot和免疫组织化学(IHC)特性。扩增轻、重链可变区和猫IgG1 Fc基因,降落PCR将重链可变区(Variable region of the Heavy chain,VH)和轻链可变区(Variable region of the Light chain,VL)拼接成scFv,进一步拼接出scFv-fFc,构建pFast-MPD-L1-scFv和pFast-MPD-L1-scFv-fFc重组质粒,在昆虫杆状病毒表达系统中表达。结果表明,猫PD-L1在上清中大量表达且纯度较好,杂交瘤细胞2A5A2F4D7分泌的抗体属于IgG 2a,轻链为κ亚型,细胞上清、腹水抗体效价分别为1∶3200和1∶102400,Western blot结果表明,该单抗能与表达的猫PD-L1反应,IHC结果显示,该单抗可以识别犬猫肿瘤组织细胞表达的PD-L1,可以反映PD-L1在肿瘤组织中的表达情况。VH和VL与IMGT数据库中相应基因序列相符,具有4个框架区和3个高变区,猫源化嵌合单链抗体在昆虫杆状病毒表达系统中成功表达。综上,本研究制备的猫PD-L1单克隆抗体在犬猫肿瘤检测和预后中具有一定的价值,猫源化单链抗体的成功制备为其在犬猫肿瘤病治疗中的应用打下了基础。展开更多
文摘构建使用了PD-1抑制剂的肿瘤患者出现甲状腺功能障碍的风险预测模型,分析使用PD-1肿瘤抑制剂导致的甲状腺功能障碍的相关风险因素,设计监测预警系统。选取2020年—2023年广西医科大学附属肿瘤医院1225例使用PD-1抑制剂肿瘤患者的临床资料,包括人口学特征、既往史、实验室检测等63个变量。本文选取相关性前10/20/30/40/50/60个变量的4种传统机器学习模型进行性能比较。通过F1分数、灵敏度、准确率、精确率、特异性曲线下面积(Area Under the Curve,AUC)评估以上预测模型的性能,并利用Shapley加性解释(Shapley Additive Explanation,SHAP)可视化解释本文的机器学习模型。与促甲状腺激素相关性排名前10的变量依次为:羟丁酸脱氢酶、乳酸脱氢酶、淋巴细胞绝对值、天门冬氨酸转移酶、钙离子、碱性磷酸酶、谷氨酰转肽酶、单核细胞绝对值、红细胞分布宽度SD、胆碱酯酶。建立了使用PD-1抑制剂的肿瘤患者出现甲状腺功能障碍的风险预测模型,并在全局解释和局部解释的层面上分别作出模型预测结果影响的解释。
文摘目的:探究卵巢癌(ovarian cancer,OV)浸润性CD4^(+)调节性T细胞(regulatory T cell,Treg)的脂质摄取与积累情况,及其与CD4^(+)Treg中程序性细胞死亡蛋白1(programmed cell death protein 1,PD-1)和细胞毒性T淋巴细胞相关蛋白-4(cytotoxic Tlymphocyte associated protein 4,CTLA-4)表达的相关性。方法:采用亲脂性荧光染料BODIPY^(TM)493/503和荧光脂肪酸探针BODIPY^(TM)500/510 C1 C12分别检测OV组织中的或与不同OV细胞系(ES-2、SKOV3、CAOV3)上清液共培养的人源CD4^(+)Treg的细胞内脂质含量和脂质摄取能力;使用脂肪酸氧化抑制剂(Etomoxir)、脂肪酸合成抑制剂(C75)和脂肪酸摄取抑制剂磺基-N-琥珀酰亚胺油酸酯(sulfo-N-succinimidyl oleate,SSO)干预脂质代谢;通过流式细胞术分析CD4^(+)Treg上免疫抑制分子PD-1和CTLA-4的表达。结果:OV组织CD4^(+)Treg相比传统CD4^(+)T细胞表现出更高的脂质含量和脂质摄取能力(P均<0.01)。在体外实验中,与基础培养基相比,OV细胞培养上清可显著提升CD4^(+)Treg的胞内脂质含量与脂质摄取能力(P均<0.05),其中CAOV3来源的上清作用最为显著。此外,CAOV3上清液还能提升CD4^(+)Treg中PD-1与CTLA-4的表达(P<0.05)。CD4^(+)Treg的脂质积累随CAOV3上清浓度增加呈剂量依赖性上升(P<0.05),且其对胞外荧光脂肪酸类似物的摄取能力具有浓度依赖性(P<0.05)。脂肪酸摄取抑制剂SSO可有效逆转CAOV3上清液诱导的CD4^(+)Treg脂质积累及PD-1、CTLA-4的高表达(P<0.05);而脂肪酸氧化抑制剂Etomoxir与合成抑制剂C75则无显著影响。结论:OV微环境通过促进CD4^(+)Treg的脂质摄取,提高细胞内脂质含量,促进其免疫抑制分子PD-1和CTLA-4表达。靶向脂肪酸摄取途径可能是逆转OV中Treg介导的免疫抑制的潜在策略。
基金supported by the National Natural Science Foundation of China(32470354,31900612,and 31971919)the Natural Science Foundation of Chongqing,China(cstc2020jcyj-jqX0020)+1 种基金the Foundation for Innovative Research Groups of the Natural Science Foundation of Chongqing,China(cstc2021jcyj-cxttX0004)the Chongqing Talent Program Foundation,China(cstc2024ycjhbgzxm0063)。
文摘Leaves and glumes act as lateral organs and have essential effects on photosynthesis and seed morphology,thus affecting yield.However,the molecular mechanisms controlling their polarity development in rice still need further study.Here,we isolated a polarity defect of lateral organs 1(pdl1)mutant in rice,which exhibits twisted/filamentous-shaped leaves and cracked/filamentous-shaped lemmas caused by defects in polarity development.PDL1 encodes a SUPPRESSOR OF GENE SILENCING 3 protein localized in the cytoplasmic granules.PDL1 is expressed in the shoot apical meristem,inflorescence meristem,floral meristem,and lateral organs including leaves and floral organs.PDL1 is involved in the synthesis of tasiR-ARF,which may subsequently modulate the expression of OsARFs.Meanwhile,the expression levels of abaxial miR165/166 and the adaxial identity genes OSHBs were respectively increased and reduced significantly.The results of this study clarify the molecular mechanism by which PDL1-mediated tasiR-ARF synthesis regulates the lateral organ polarity development in rice.
基金supported by the National Natural Science Foundation of China(32202253)the Natural Science Foundation of Anhui Higher Education Institutions,China(KJ2020A0102)the Talent Research Project of Anhui Agricultural University,China(rc342001)。
文摘Rice blast,caused by Magnaporthe oryzae,is a fungal disease that causes devastating damage to rice production worldwide.During infection,pathogens secrete effector proteins that modulate plant immunity.Disulfide bond formation catalyzed by protein disulfide isomerases(PDI)is essential for protein folding and maturation.However,the biological function of Pdi1 in M.oryzae has not yet been characterized.In this study,we identified the endoplasmic reticulum(ER)-located protein,MoPdi1,in M.oryzae.MoPdi1 regulates conidiation,cell wall stress,and pathogenicity of M.oryzae.Furthermore,the CGHC active sites in the a and a'redox domain of MoPdi1 were essential for the biological function of MoPDI1.Further tests demonstrated that MoPdi1 was involved in the regulation of ER stress and positively regulated ER phagy.We also found that MoPdi1 interacted with MoHut1.Deletion of MoPDI1 led to the bereft of MoHut1 dimerization,which depends on the formation of disulfide bonds.In addition,MoPdi1 affected the normal secretion of the cytoplasmic effector AVR-Pia.We provided evidence that MoHut1 is important for the vegetative growth,conidiation,and pathogenicity in M.oryzae.Therefore,our findings could provide a suitable target point for designing antifungal agrochemicals against rice blast fungus.
基金supported by grants from Simons Foundation (SFARI 479754),CIHR (PJT-180565)the Scottish Rite Charitable Foundation of Canada (to YL)funding from the Canada Research Chairs program。
文摘The mechanistic target of rapamycin(m TOR) is a serine/threonine kinase that plays a pivotal role in cellular growth, proliferation, survival, and metabolism. In the central nervous system(CNS), the mTOR pathway regulates diverse aspects of neural development and function. Genetic mutations within the m TOR pathway lead to severe neurodevelopmental disorders, collectively known as “mTORopathies”(Crino, 2020). Dysfunctions of m TOR, including both its hyperactivation and hypoactivation, have also been implicated in a wide spectrum of other neurodevelopmental and neurodegenerative conditions, highlighting its importance in CNS health.
文摘旨在制备猫PD-L1抗原并筛选单抗,探讨其在犬、猫肿瘤检测和诊断中的应用,同时制备猫源化嵌合单链抗体,为猫肿瘤的治疗提供基础。本研究通过PCR扩增猫PD-L1基因,构建原核表达重组质粒PET-MPD-L1,通过在宿主BL21(DE3)中表达制备PD-L1抗原,免疫小鼠后细胞融合,ELISA方法进行杂交瘤细胞筛选,3次亚克隆后建立MPD-L1杂交瘤细胞系。测定杂交瘤细胞上清、腹水效价、抗体亚类、Western blot和免疫组织化学(IHC)特性。扩增轻、重链可变区和猫IgG1 Fc基因,降落PCR将重链可变区(Variable region of the Heavy chain,VH)和轻链可变区(Variable region of the Light chain,VL)拼接成scFv,进一步拼接出scFv-fFc,构建pFast-MPD-L1-scFv和pFast-MPD-L1-scFv-fFc重组质粒,在昆虫杆状病毒表达系统中表达。结果表明,猫PD-L1在上清中大量表达且纯度较好,杂交瘤细胞2A5A2F4D7分泌的抗体属于IgG 2a,轻链为κ亚型,细胞上清、腹水抗体效价分别为1∶3200和1∶102400,Western blot结果表明,该单抗能与表达的猫PD-L1反应,IHC结果显示,该单抗可以识别犬猫肿瘤组织细胞表达的PD-L1,可以反映PD-L1在肿瘤组织中的表达情况。VH和VL与IMGT数据库中相应基因序列相符,具有4个框架区和3个高变区,猫源化嵌合单链抗体在昆虫杆状病毒表达系统中成功表达。综上,本研究制备的猫PD-L1单克隆抗体在犬猫肿瘤检测和预后中具有一定的价值,猫源化单链抗体的成功制备为其在犬猫肿瘤病治疗中的应用打下了基础。
