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利用重叠延伸PCR和同源重组克隆技术制备HLA-A＊ 0201/HBc18-27单链三分子复合体

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东南大学学报（医学版） 2016年第6期35卷 825-831页

作者：许涛李晓娥吴优 khawar ali shahzad 王伟张雷沈传来东南大学医学院病原生物学与免疫学系江苏南京210009

目的：构建和表达HLA-A＊0201/HBc18-27单链三分子复合体。方法：利用重叠延伸PCR将HBc18-27、（Gly4Ser）3、β2m、（Gly4Ser）4和HLA-A＊0201胞外区依次串联为单链三聚体（single-chain trimer,SCT）融合基因,通过同源重组克隆技术插... 详细信息

目的：构建和表达HLA-A＊0201/HBc18-27单链三分子复合体。方法：利用重叠延伸PCR将HBc18-27、（Gly4Ser）3、β2m、（Gly4Ser）4和HLA-A＊0201胞外区依次串联为单链三聚体（single-chain trimer,SCT）融合基因,通过同源重组克隆技术插入到pET28a原核表达载体,用异丙基β-D-硫代吡喃半乳糖苷（IPTG）在大肠杆菌BL21（DE3）中诱导表达,采用金属螯合亲和层析法纯化目的蛋白,通过稀释复性法折叠成HLA-A＊0201/HBc18-27的复合单体,并生物素化,将单体包被到直径4.5μm的磁性微球,用构象特异性单抗（W6/32）进行荧光染色,流式细胞术分析其空间构象。结果：pET28a-HLA-A＊0201/HBc18-27基因序列和蛋白大小与预测一致;纯化后融合蛋白纯度约93%;流式细胞术分析显示pET28a-HLA-A＊0201/HBc18-27单体具有正确的空间构象。结论：利用重叠延伸PCR和同源重组克隆技术成功制备了HLA-A＊0201/HBc18-27的单链复合体,无需限制性内切酶和连接酶,发展了主要组织相容性单链复合体技术,有效简化了pMHCⅠ类分子的制备过程。

关键词：重叠延伸聚合酶链式反应同源重组主要组织相容性复合体

人类白细胞抗原-B等位基因多态性与江苏地区汉族人群2型糖尿病的相关性研究

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中国现代医学杂志 2016年第6期26卷 37-44页

作者：夏玲芝曹鹏李晓娥许涛 khawar ali shahzad 毛源张厚智沈传来南京金域医学检验所实验诊断部江苏南京210002 东南大学医学院病原生物学与免疫学系江苏南京210009

目的探讨人类白细胞抗原（HLA-B）等位基因多态性与江苏地区汉族人群2型糖尿病的相关性。方法采用聚合酶链反应直接测序分型法（PCR-SBT）对118例江苏地区汉族2型糖尿病（T2DM）个体进行HLA-B等位基因分型,计算各等位基因频率和血清型频... 详细信息

目的探讨人类白细胞抗原（HLA-B）等位基因多态性与江苏地区汉族人群2型糖尿病的相关性。方法采用聚合酶链反应直接测序分型法（PCR-SBT）对118例江苏地区汉族2型糖尿病（T2DM）个体进行HLA-B等位基因分型,计算各等位基因频率和血清型频率,并与中华骨髓库江苏分库汉族志愿者人群（644例和20 248例）进行统计学比较。结果 HLA-B＊1501的等位基因频率在江苏地区汉族T2DM人群中明显升高（8.90%vs 3.03%,Pc=0.000,^OR=3.13,95%CI=1.80～5.42,Power=98.23%）,而HLA-B＊5801的等位基因频率则明显下降（1.69%vs 8.62%,Pc=0.000,^OR=0.18,95%CI=0.07～0.50,Power=91.07%）;相对应地,HLA-B＊15的血清型频率在江苏地区汉族T2DM人群中明显升高（21.18%vs 13.03%,Pc=0.022,^OR=1.79,95%CI=1.26～2.55,Power=90.15%）,而HLA-B＊58的血清型频率则明显下降（1.69%vs 8.62%,Pc=0.000,^OR=0.18,95%CI=0.07～0.50,Power=91.09%）。结论 HLA-B＊1501等位基因可能与江苏地区汉族人群2型糖尿病的易感性有关,而HLA-B＊5801则可能有保护作用。这些结果首次提示HLA-I类基因多态性可能与中国人群2型糖尿病有关。

关键词： 2型糖尿病 HLA-B 等位基因多态性 PCR-SBT

携载avasimibe的PLGA微粒式人工抗原提呈细胞抗肿瘤作用研究

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携载avasimibe的PLGA微粒式人工抗原提呈细胞抗肿瘤作用研究

共载11种免疫分子的PLGA微粒式人工抗原提呈细胞的制备及其抗肿瘤作用研究

第十三届全国免疫学学术大会

作者：张雷 khawar ali shahzad 张立平刘健刚荣令东南大学医学院附属中大医院溧水分院东南大学医学院病原生物学与免疫学系

背景:以聚乳酸-羟基乙酸共聚物(PLGA)微粒为载体的人工抗原提呈细胞(aAPC)可包裹缓释第三信号分子,有效活化和扩增抗原特异性T细胞,是特异性免疫疗法的有效策略。Avasimibe是胆固醇酯化酶ACAT1的小分子抑制剂,能够有效增强CD8+T细胞的... 详细信息

背景:以聚乳酸-羟基乙酸共聚物(PLGA)微粒为载体的人工抗原提呈细胞(aAPC)可包裹缓释第三信号分子,有效活化和扩增抗原特异性T细胞,是特异性免疫疗法的有效策略。Avasimibe是胆固醇酯化酶ACAT1的小分子抑制剂,能够有效增强CD8+T细胞的抗肿瘤活性。目的 :以PLGA微粒为载体,制备可缓释avasimibe的人工抗原提呈细胞(aAPCavasimibe),探讨其在体外扩增肿瘤抗原特异性T细胞和在体内抑制肿瘤生长的能力。方法 :复乳溶剂挥发法制备内部包裹avasimibe的PLGA微粒,EDC/NHS法使其表面功能化后联合共价吸附H-2Kb/TRP2180-188二聚体及共刺激分子anti-CD28,制备aAPCavasimibe;在体外,与C57BL/6鼠脾细胞共培养,流式检测TRP2抗原特异性CTL细胞比例及特异性杀伤肿瘤细胞能力;尾静脉注射aAPCavasimibe至黑色素皮下载瘤鼠和肺转移鼠体内,流式检测模型鼠外周血、脾脏和肿瘤组织中抗原特异性CTL的频率变化,观察皮下瘤生长进度,计数肺转移病灶。结果:PLGA微粒对avasimibe的包封率在80%以上,且可缓慢释放,28天累积释放率大于85%。在体外,aAPCavasimibe与C57BL/6鼠脾细胞共培养7天,TRP2特异性CTL的比例可提高至66%,扩增后脾细胞特异性杀瘤细胞能力较传统两信号aAPC对照组提高359.6%;在体内,MaAPC输注可有效提升载瘤鼠外周血和脾脏中TRP2抗原特异性CTL的频率,显著提高肿瘤组织中抗原特异性CTL的浸润程度,并明显抑制皮下瘤的生长速度,减少肺转移鼠的转移病灶数量。结论 :可缓释avasimibe的aAPCavasimibe能更有效的促进肿瘤抗原特异性CTL扩增,提高机体抗肿瘤能力,在肿瘤免疫治疗中具有潜在应用价值。

关键词： PLGA avasimibe 人工抗原提呈细胞抗原特异性T细胞肿瘤免疫治疗

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共载11种免疫分子的PLGA微粒式人工抗原提呈细胞的制备及其抗肿瘤...

小鼠EAE模型中病理和免疫指标的动态监测及杀伤性人工抗原提呈细胞的初步治疗

第十一届全国免疫学学术大会

作者：张雷 khawar ali shahzad 许涛万昕汪礼敏裴伟亚沈传来东南大学医学院病原生物学与免疫学系

目的 :在磁珠或胶乳微球表面共展现p MHC分子和共刺激分子的非细胞性人工抗原提呈细胞(a APC)是特异性免疫疗法的新策略。但其不可生物降解性阻碍了体内应用。聚乳酸-羟基乙酸共聚物(PLGA)具有生物降解和相容性,是被FDA批准的药物递送... 详细信息

目的 :在磁珠或胶乳微球表面共展现p MHC分子和共刺激分子的非细胞性人工抗原提呈细胞(a APC)是特异性免疫疗法的新策略。但其不可生物降解性阻碍了体内应用。聚乳酸-羟基乙酸共聚物(PLGA)具有生物降解和相容性,是被FDA批准的药物递送常用材料。本研究以PLGA微粒为载体共载11种免疫分子,制备多功能式人工抗原提呈细胞(Ma APC),验证其表面共吸附能力和包裹缓释特征;探讨其体外扩增肿瘤抗原特异性T细胞和体内抑瘤生长的能力。方法:复乳溶剂挥发法制备内部包裹IL-2、IL-15、CCL21、anti-CTLA-4和anti-PD-1的PLGA微粒,EDC/NHS法使其表面功能化后联合共价吸附H-2Kb/TRP2180-188二聚体、H-2Db/gp10025-33二聚体、antiCD28、anti-4-1BB、anti-CD2以及抗吞噬分子CD47-Ig,制备Ma APC;在体外,与C57BL/6鼠脾细胞共培养,流式检测TRP2和gp100抗原特异性CTL比例;尾静脉注射Ma APC至黑色素瘤皮下载瘤鼠体内,流式监测外周血,脾脏和肿瘤组织中抗原特异性CTL的频率变化,观察肿瘤生长进度。结果 :PLGA微粒对5种免疫分子的包裹率均在65%以上,均可缓慢释放,28天累积释放率大于80%。其表面对其他6种免疫分子也可有效共吸附,各分子间吸附干扰效应较小;在体外,Ma APC与C57BL/6鼠脾细胞共培养7天后使TRP2和gp100特异性CTL的比例分别提高至71%和68%;在体内,Ma APC输注可有效提升载瘤鼠外周血和脾脏中TRP2和gp100抗原特异性CTL的频率,显著提高肿瘤组织中抗原特异性CTL的浸润,明显抑制皮下瘤生长速度。结论 :PLGA微粒既可表面展现又可包裹缓释常见免疫分子,是理想的a APC载体。共载11种免疫分子的Ma APC是一种新的、可生物降解的特异性主动免疫生物制剂。

关键词： PLGA 人工抗原提呈细胞抗原特异性T细胞抗肿瘤主动免疫

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小鼠EAE模型中病理和免疫指标的动态监测及杀伤性人工抗原提呈细...

Elastic-viscoplastic constitutive model for capturing the mechanical response of polymer composite at various strain rates

第十一届全国免疫学学术大会

作者：万昕裴伟亚张一鸣张雷 khawar ali shahzad 许涛王伟沈传来东南大学医学院病原生物学与免疫学系

目的 :观察小鼠EAE病程中临床、病理及自身反应性CD4和CD8 T细胞等指标的动态变化规律;探索以聚乳酸-羟基乙酸微球(PLGA-MPs)为载体的杀伤性人工抗原提呈细胞(Ka APCs)对EAE的特异性免疫疗法。方法 :以MOG35-55多肽诱导构建C57BL/6鼠的... 详细信息

目的 :观察小鼠EAE病程中临床、病理及自身反应性CD4和CD8 T细胞等指标的动态变化规律;探索以聚乳酸-羟基乙酸微球(PLGA-MPs)为载体的杀伤性人工抗原提呈细胞(Ka APCs)对EAE的特异性免疫疗法。方法 :以MOG35-55多肽诱导构建C57BL/6鼠的EAE模型;临床神经功能评分评估病程;HE和IHC染色观察急性初发期、高峰期和慢性恢复期EAE小鼠脑和脊髓组织中炎性细胞浸润以及病理相关蛋白MBP和GFAP的表达;ELISPOT法和p MHC多聚体荧光染色及流式分析法检测不同病期中脾脏及淋巴结中MOG40-54表位特异性CD8 T细胞和MOG35-55表位特异性CD4 T细胞的反应活性和频率;在PLGA-MPs表面共包被2种靶抗原(H-2Db/MOG40-54二聚体,H-2I-Ab/MOG35-55多聚体)、3种负调节分子(anti-Fas m Ab,PD-L1-Ig,TGF-β1)以及抗吞噬分子CD47-Ig,制备MOG抗原特异性的多功能Ka APCs;经尾静脉注射Ka APCs治疗EAE,通过临床神经功能评分初步评价其疗效。结果:EAE模型鼠中,脑和脊髓组织中炎性细胞浸润程度随病情加重而增多,MBP随病情加重而表达降低,GFAP随病情加重而表达上升;MOG特异性CD4和CD8 T细胞在抗原肽刺激后分泌IFN-γ的反应活性在发病初期最高,随后逐渐下降,而其频率却在发病高峰期最高,以后缓慢下降;Ka APCs的输注治疗能显著减轻小鼠EAE的疾病程度,也明显减少脑和脊髓组织中炎性浸润,作用机制正在研究。结论 :中枢神经组织中炎性细胞的浸润程度和病理相关蛋白的表达水平与小鼠EAE的病程发展相一致,但自身反应性T细胞的数量与其对抗原刺激的反应活性有不同的动态变化规律。可生物降解的PLGA微球为载体的多动能Ka APCs有治疗EAE的潜能。

关键词：实验性自身免疫性脑脊髓炎杀伤性人工抗原提呈细胞自身反应性T细胞

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Journal of Materials Science & Technology 2020年第22期54卷 12-17页

作者： shahzad Fateh ali Jitang Fan State Key Laboratory of Explosion Science and Technology Beijing Institute of TechnologyBeijing700087China

Recently, polymer composites, consisting of hard segments and soft segments blend, attract a broad research interest because of the high flow stress and large deformation under a high rate loading. They display an outstanding energy absorption capability for impact resistant engineering. In this paper,a physics-based elastic-viscoplastic constitutive model is developed which accounts the activation of molecular chains motion at different segments upon an applied stress as well as the rate dependency of mechanical properties. Accordingly, finite element simulations are conducted to derive the mechanical performance at various strain rates. It involves the linearly elastic deformation to yielding, softening behavior with a slight decrease of stress, plastic flow at a plateau stress accompanied by a considerable straining, densification with a fast increase of stress. The results achieved from model simulations match well with these obtained by experiments and the mechanical response in the strain rate range from 0.0001 s^(-1) to 8000 s^(-1) can be captured by this model. Finally, energy absorption capability of the polymer composite is discussed, and the physical mechanisms in material science are addressed by post-loading analysis. This work is full of interest to develop the constitutive model for designing the impact-resistant polymers and illustrating the dynamic mechanical response, which will promote the application of light-weight transparent protection structure against a high-speed impact.

关键词： Polymer composite Constitutive model Strain rate Mechanical behavior

Clinical applications of stem cell-derived exosomes

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Signal Transduction and Targeted Therapy 2024年第2期9卷 621-651页

作者： Fei Tan Xuran Li Zhao Wang Jiaojiao Li khawar shahzad Jialin Zheng Department of ORL-HNS Shanghai Fourth People’s Hospitaland School of MedicineTongji UniversityShanghaiChina Plasma Medicine and Surgical Implants Center Tongji UniversityShanghaiChina The Royal College of Surgeons in Ireland DublinIreland The Royal College of Surgeons of England LondonUK Center for Translational Neurodegeneration and Regenerative Therapy Tongji Hospital affiliated to Tongji University School of MedicineShanghaiChina Shanghai Frontiers Science Center of Nanocatalytic Medicine Tongji UniversityShanghaiChina

Although stem cell-based therapy has demonstrated considerable potential to manage certain diseases more successfully than conventional surgery,it nevertheless comes with inescapable drawbacks that might limit its clinical *** to stem cells,stem cell-derived exosomes possess numerous advantages,such as non-immunogenicity,non-infusion toxicity,easy access,effortless preservation,and freedom from tumorigenic potential and ethical *** can inherit similar therapeutic effects from their parental cells such as embryonic stem cells and adult stem cells through vertical delivery of their pluripotency or *** a thorough search and meticulous dissection of relevant literature from the last five years,we present this comprehensive,up-to-date,specialty-specific and disease-oriented review to highlight the surgical application and potential of stem cell-derived *** derived from stem cells(e.g.,embryonic,induced pluripotent,hematopoietic,mesenchymal,neural,and endothelial stem cells)are capable of treating numerous diseases encountered in orthopedic surgery,neurosurgery,plastic surgery,general surgery,cardiothoracic surgery,urology,head and neck surgery,ophthalmology,and obstetrics and *** diverse therapeutic effects of stem cells-derived exosomes are a hierarchical translation through tissue-specific responses,and cell-specific molecular signaling *** this review,we highlight stem cell-derived exosomes as a viable and potent alternative to stem cell-based therapy in managing various surgical *** recommend that future research combines wisdoms from surgeons,nanomedicine practitioners,and stem cell researchers in this relevant and intriguing research area.

关键词： viable stem capable

On the Exact Solution for Axisymmetric Flow and Heat Transfer over a Nonlinear Radially Stretching Sheet

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Chinese Physics Letters 2012年第8期29卷 168-171页

作者： Azeem shahzad Ramzan ali Masood Khan Department of Mathematics Quaid-i-Azam UniversityIslamabad 44000Pakistan

We investigate the boundary-layer flow and heat transfer of a magnetohydrodynamic viscous fluid over a nonlinear radially porous stretching sheet within a porous *** flow is generated due to a nonlinear stretching sheet and influenced by a continuous suction/blowing of the fluid through the porous *** governing momentum and thermal boundary layer equations are converted into ordinary differential equations by appropriate similarity *** exact solution for the velocity and the temperature fields are derived in the form of an incomplete Gamma *** analytic solutions are found by the homotopy analysis *** graphical results for velocity and temperature fields are presented and ***,the numerical values of the skin friction coefficient and the Nusselt number are calculated and discussed.

关键词： function stretching porous

Computing of LQR Technique for Nonlinear System Using Local Approximation

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Computer Systems Science & Engineering 2023年第7期46卷 853-871页

作者： Aamir shahzad ali Altalbe Mechanical Engineering Department The University of LahoreLahorePakistan Faculty of Computing and Information Technology King Abdulaziz UniversityJeddah80258Saudi Arabia

The main idea behind the present research is to design a state-feedback controller for an underactuated nonlinear rotary inverted pendulum module by employing the linear quadratic regulator(LQR)technique using local *** LQR is an excellent method for developing a controller for nonlinear *** provides optimal feedback to make the closed-loop system robust and stable,rejecting external ***-based optimal controller for a nonlinear system such as a rotatory inverted pendulum has not been designed and implemented using Newton-Euler,Lagrange method,and local ***,implementing LQR to an underactuated nonlinear system was vital to design a stable controller.A mathematical model has been developed for the controller design by utilizing the Newton-Euler,Lagrange *** nonlinear model has been linearized around an equilibrium *** and nonlinear models have been compared to find the range in which linear and nonlinear models’behaviour is *** LQR function and system dynamics have been used to estimate the controller *** the performance evaluation of the designed controller,Simulink has been *** and nonlinear models have been simulated along with the designed *** have been performed for the designed controller over the linear and nonlinear system under different conditions through varying system *** results show that the system is stable and robust enough to act against external *** controller maintains the rotary inverted pendulum in an upright position and rejects disruptions like falling under gravitational force or any external disturbance by adjusting the rotation of the horizontal link in both linear and nonlinear environments in a specific *** controller has been practically designed and *** is vivid from the results that the controller is robust enough to reject the disturbances in milliseconds and keeps the pendulum

关键词： Computing rotary inverted pendulum(RIP) modeling and simulation linear quadratic regulator(LQR) nonlinear system