pcDNA3.1/hIL-18真核表达载体的构建及表达

目的 :构建重组真核表达质粒pcDNA3.1/IL 18,并在哺乳动物细胞COS 7和Rlc310中进行瞬时和稳定性表达。方法 :从含hIL 18基因的中介载体 pGEM TEasy( pGEM T/hIL 18)中 ,以限制性内切酶酶切方法获得目的片段 ,克隆入真核表达质粒 pcDNA3.1( )中。以脂质体法转染COS 7和Rlc310细胞 ,用RT PCR检测IL 18mRNA的水平 ,免疫组化染色法检测蛋白表达。结果 :构建了hIL 18基因的重组真核表达质粒pcDNA3.1/IL 18,并可在哺乳动物细胞中瞬时、稳定表达 ,获得了可稳定表达hIL 18基因的Rlc310细胞株。结论 :pcDNA3.1/IL 18的构建及表达 ,为IL 18抗肿瘤作用的研究奠定了基础     

Th1型细胞因子基因对结核分枝杆菌基因疫苗诱导BALB/c小鼠产生抗CFP10抗体水平的影响

目的：研究分别表达含IL—12和IL-18基因的质粒，对结核分枝杆菌(Mycobacterium tuberculosis，MTB)H37R1株CFP1O基因疫苗诱导免疫应答的影响。方法：从正常人外周血单个核细胞(PMBCs)中提取RNA，用RT—PCR扩增IL-18 cDNA，并克隆人载体pGEM—Teasy中。测序证实后，亚克隆至真核表达载体pcDNA3．1的BamH Ⅰ和EcoR Ⅰ酶切位点。将分别表达小鼠IL—12和人IL—18基因的真核表达质粒pcmlL12和pclL18，与MTB CFP10基因疫苗联合肌注免疫BALB／c小鼠，共免疫3次，每次间隔2wk。每次免疫后2wk采血、分离血清，用ELISA检测小鼠血清抗CFP10抗体的滴度。结果：用RT—PCR成功地从人PMBC的RNA中扩增出IL—18 cDNA，测序结果正确，用BamH Ⅰ和EcoR Ⅰ酶切鉴定证实，目的基因已插入载体pcDNA3．1中，阳性克隆命名为pcIL18。pcCFP10组第1次免疫后，血清抗CFP10抗体的平均滴度为1：600，末次免疫后的滴度为1：4000。pcIL18 pcCFP10组联合免疫后，血清抗CFP10抗体的滴度高于pcCFP10组，最终达1：8000。而pcmIL12 pcCFP10组联合免疫后滴度仅为1：200。结论：pcIL18与CFP10基因疫苗联合免疫，可增强CFP10抗原的特异性体液免疫应答；pcmIL12则可使CFP10基因疫苗产生的抗体水平降低。pcIL18 pcCFP10基因联合免疫是否具有增强CFP10抗原特异性细胞免疫的作用有待进一步研究。     

Increased expression levels of E-cadherin,cytokeratin 18 and 19 observed in preeclampsia were not correlated with disease severity

Introduction

Preeclampsia is a pregnancy-specific disorder and placental factor(s) contribute to the pathogenesis of preeclampsia. Turnover of villous trophoblast is affected by impaired placental perfusion in preeclampsia. Expression and localisation of cadherins and cytokeratins are involved in the pathogenesis of preeclampsia. However, studies describing the associations between cadherins and cytokeratins in preeclampsia are limited. The aim of this study was to investigate the expression of E-cadherin, N-cadherin, cytokeratin 18 and cytokeratin 19 in placentae from women with preeclampsia in order to determine whether their expression differs with disease severity.

Methods

29 preeclamptic placentae and 25 normotensive placentae were included in this study. The expression of E-cadherin, cytokeratin 18, cytokeratin 19 andN-cadherin was quantified by immunohistochemistry and western blotting.

Results

E-cadherin, cytokeratin 18 and cytokeratin 19 were expressed predominantly in the syncytiotrophoblast of the placenta and the expression of E-cadherin, cytokeratin 18 and cytokeratin 19 was significantly increased in preeclampsia compared to normotensive pregnancies. However, there was no significant difference in expression between severe preeclampsia and mild preeclampsia. In addition, there was no difference in the expression of N-cadherin between preeclampsic and normotensive pregnancies.

Discussion

Our data demonstrated increased expression of E-cadherin, cytokeratin 18 and cytokeratin 19 in the syncytiotrophoblast of preeclamptic placentae, but this increase was not correlated with disease severity.

Conclusion

Our data suggests that E-cadherin and cytokeratins are involved in the pathogenesis of preeclampsia.     

反复种植失败的相关对策新进展

辅助生殖技术的迅速发展使众多不孕症患者借助体外受精-胚胎移植(IVF-ET)及其衍生技术获得了后代。然而很大一部分妇女经历多次优质胚胎移植亦不能获得妊娠,反复种植失败(RIF)已经成为阻碍妊娠率进一步提高的瓶颈问题,且日益受到生殖医学界的广泛关注。就目前的条件而言,对RIF患者给予药物或者机械操作以提高子宫内膜容受性,行宫腹腔镜检查排除宫腔及盆腔病变以改善胚胎种植环境,通过辅助孵化、选择性囊胚移植、植入前胚胎遗传学筛查、共培养等技术提高胚胎着床能力都有可能改善和提高其种植率及妊娠率。RIF成为了我们亟待解决的问题,现综述近年有关反复种植失败的相关对策新进展。     

Sarcocystis Species Lethal for Domestic Pigeons

A large number of Sarcocystis spp. infect birds as intermediate hosts, but pigeons are rarely affected. We identified a novel Sarcocystis sp. that causes lethal neurologic disease in domestic pigeons in Germany. Experimental infections indicated transmission by northern goshawks, and sequence analyses indicated transnational distribution. Worldwide spread is possible.     

Comparison of different compressed sensing algorithms for low SNR 19F MRI applications—Imaging of transplanted pancreatic islets and cells labeled with perfluorocarbons

Transplantation of pancreatic islets is a possible treatment option for patients suffering from Type I diabetes. In vivo imaging of transplanted islets is important for assessment of the transplantation site and islet distribution. Thanks to its high specificity, the absence of intrinsic background signal in tissue and its potential for quantification, ¹⁹F MRI is a promising technique for monitoring the fate of transplanted islets in vivo. In order to overcome the inherent low sensitivity of ¹⁹F MRI, leading to long acquisition times with low signal‐to‐noise ratio (SNR), compressed sensing (CS) techniques are a valuable option. We have validated and compared different CS algorithms for acceleration of ¹⁹F MRI acquisition in a low SNR regime using pancreatic islets labeled with perfluorocarbons both in vitro and in vivo. Using offline simulation on both in vitro and in vivo low SNR fully sampled ¹⁹F MRI datasets of labeled islets, we have shown that CS is effective in reducing the image acquisition time by a factor of three to four without seriously affecting SNR, regardless of the particular algorithms used in this study, with the exception of CoSaMP. Using CS, signals can be detected that might have been missed by conventional ¹⁹F MRI. Among different algorithms (SPARSEMRI, OMMP, IRWL1, Two‐level and CoSAMP), the two‐level l₁ method has shown the best performance if computational time is taken into account. We have demonstrated in this study that different existing CS algorithms can be used effectively for low SNR ¹⁹F MRI. An up to fourfold gain in SNR/scan time could be used either to reduce the scan time, which is beneficial for clinical and translational applications, or to increase the number of averages, to potentially detect otherwise undetected signal when compared with conventional ¹⁹F MRI acquisitions. Potential applications in the field of cell therapy have been demonstrated.     

基于PET/CT形变配准技术对放疗前后靶区变化的评估

PET图像提供的新陈代谢信息可用于判断放疗后肿瘤的复发区域,对于制订精确的放疗计划具有重要的临床意义。研究采用多分辨率形变配准的方法提取放疗前后CT图像的形变域,并将其作用于放疗前PET图像,与放疗后的PET图像相比较,通过设定图像中SUV 的阈值,判断勾画轮廓之间的重叠率,以获得图像中的高摄取区域,回顾性指导精确放疗。研究针对22例肺癌病例,实验结果显示放疗后残留的高代谢区域和放疗前GTV重叠较好:当阈值设定为SUV_max的70%、80%和90%时,对应的重叠率分别为(95.2±0.6)%、(96.6±3.4)%和100%;当阈值设定为SUV2.5和SUV5.0时,对应的重叠率为(86.0±6.6)%和(97.0±3.0)%。对氟代脱氧葡萄糖(FDG)高摄取区域的高重叠率表明病变区域在放疗前后的位置相对稳定,放疗后的残余肿瘤基本上位于放疗前靶区对FDG的摄取区域。初步实验结果证明,研究可用于判断靶区区域对放疗的反应,回顾性指导在放疗计划中,针对放疗后残余的靶区加大照射剂量,保护危及器官和组织,精确放疗。     

多发性硬化的PET显像

多发性硬化属神经内科罕见病,其发病机制与病理基础尚未完全阐明.目前临床上缺乏疾病特异性影像诊断方法,结构影像与疾病临床表现、进展与转归的相关性差.基于核素示踪原理的PET显像为该病的机制研究带来了契机.本文主要叙述了目前对多发性硬化的PET正电子示踪剂研究现状与进展.     

乙醇性胃粘膜损伤大鼠胃粘膜血液动力学变化及丹参提取物-F的作用

目的 :探讨不同浓度乙醇以及在事先给予丹参提取物 F ( DSE-F)和消炎痛的情况下大鼠胃粘膜血液动力学变化。方法 :用组织反射光谱分析仪 ( TS-2 0 0 )测定胃粘膜表层血液量 ( ΔEr)和 Hb氧饱和度 ( F)。结果 :( 1)胃粘膜损伤程度随乙醇浓度增高而加大 ;( 2 )低浓度乙醇引起胃粘膜充血 ,而高浓度则导致静脉淤血 ;( 3 )乙醇诱导的胃粘膜血液动力学变化因预先给予 DSE-F或消炎痛而减轻。结论 :( 1)乙醇性胃粘膜损伤的致病因素是高浓度乙醇所致的胃粘膜淤血缺氧 ;( 2 ) DSE-F对乙醇性胃粘膜损伤的保护作用在于减轻乙醇所致的胃粘膜血液动力学变化 ,后者可能与内源性前列腺素有关。     

Polymorphism of interleukin-18 promoter influences the onset of kidney graft function after transplantation

It has been well recognized that the promoter polymorphisms of interleukin-18 (IL-18) influence the level of cytokine expression. In our previously published data, we showed constitutive IL-18 expression in the epithelium of renal distal tubules in patients after kidney transplantation and significantly elevated IL-18 expression during acute rejection. In this study, we evaluated the clinical significance of two functional promoter polymorphisms of the IL-18 gene at positions -607 A/C (rs1946518) and -137 C/G (rs187238) in patients after kidney transplantation and looked for associations with the onset of graft function and the incidence of rejection episodes. Promoter polymorphisms in 124 patients and 103 unrelated controls were evaluated by sequence-specific primer polymerase chain reaction and the allele, genotype and haplotype frequencies were statistically correlated. We found a statistically different distribution of the allele frequency of -607 A/C polymorphism between patients with immediate or delayed onset of kidney graft function. Data showed that the C allele, which contributes to higher IL-18 expression, is more frequent in patients with delayed onset of function (P = 0.03, odds ratio = 1.93; 95% confidence interval = 1.15-3.25). A/C single nucleotide polymorphisms of the IL-18 promoter at position -607 may influence the onset of early kidney allograft function.