Effects of mycophenolic acid on IL-6 expression of human renal proximal and distal tubular cells in vitro.

BACKGROUND: Interleukin-6 (IL-6) is a multifunctional cytokine which regulates immune responses and host defence mechanisms. IL-6 has been found to be increased in certain inflammatory conditions of the kidney, in which tubular epithelial cells play a pivotal role. Human renal tubular cells express IL-6. Until now no data about the effect of the immunosuppressant drug mycophenolic acid (MPA) on IL-6 expression were available. METHODS: Proximal and distal tubular epithelial cells (PTC/DTC) have been isolated immunomagnetically. Confluent monolayers were stimulated with interleukin-1beta (IL-1beta; 25 U/ml), IL-1beta+ MPA (0.25-50 micro M) or MPA alone for 48 h. Release of IL-6 protein into the supernatant was evaluated with an enzyme immunoassay, IL-6 mRNA expression was evaluated using the Quantikine mRNA kit. RESULTS: After IL-1beta stimulation, a highly significant 2.6- (PTC) and 3.8-fold (DTC) upregulation of IL-6 expression was detectable. IL-6 mRNA was upregulated by IL-1beta [1.57- (PTC) and 2.03-fold (DTC)]. MPA inhibited this cytokine-induced IL-6 expression in a dose-dependent manner. Incubation with the lowest MPA concentration had no effect on the stimulated upregulation, whereas all higher doses significantly decreased IL-6 expression. Dexamethasone significantly inhibited the cytokine-induced IL-6 protein release in PTC, but not in DTC. CONCLUSIONS: In this study we demonstrated for the first time an inhibitory effect of MPA on the stimulated IL-6 expression of renal tubular epithelial cells. In contrast to older data, which showed a synergistic upregulation of the expression of a CC-chemokine by a combination of cytokines and MPA, in the present study we could demonstrate an immunosuppressive effect of MPA on the expression of an important cytokine.     

乳猪肝细胞的分离、培养及功能检测

目的 为构建生物人工肝进行肝细胞的准备。方法 采用胶原酶半原位灌流法分离单个乳猪肝细胞,并对其活力及单层和聚集培养后的白蛋白、尿素合成功能进行检测。结果 采用本方法从每头乳猪中分离到的单个肝细胞数为(3.1±1.5)×10~(10),活性超过95％。在加入激素和生长因子的培养基中单层培养时,肝细胞功能良好,可维持2周左右。而在未加入激素和生长因子的培养中肝细胞虽能存活1周,但功能于24h后即丧失。球形聚集培养可实现肝细胞的大量培养,且生物学活性较单层培养显著提高。结论 采用胶原酶半原位灌注法所得单个乳猪肝细胞基本能满足构建生物人工肝对肝细胞数量的要求。聚集培养接种密度大,细胞生物学活性高,可用于构建生物人工肝。     

Rosiglitazone down-regulates lipopolysaccharide-induced expression of CD40 and intercellular adhesion molecule 1 in rat peritoneal mesothelial cells through a NF-κB dependent mechanism

Objective To investigate the effect and mechanism by which PPARγ ligand, rosiglitasone, regulates the expression of CD40 and intercellular adhesion molecule 1 (ICAM-1) in the rat peritoneal mesothelial cells (RPMCs) induced by lipopolysaccharide (LPS). Methods RPMCs were harvested from Sprague-Dawley rat peritoneal cavity and maintained under defined in vitro conditions. The cells were randomly divided into groups as follows: medium, LPS (5 mg/L), LPS (5 mg/L)+BAY11-7085(5 μmol/L, NF-κB inhibitor), rosiglitazone (10 μmol/L or 20 μmol/L, peroxisome proliferator-activated receptor γ activator), LPS (5 mg/L)+rosiglitazone (10 μmol/L)+GW9662 (3 μmol/L, peroxisome proliferator-aetivatcd receptor γ antagonist), and LPS (5 mg/L)+vehicle (DMSO 0.2 ml/L). The expressions of CD40 and ICAM-1 RNA in RPMCs were examined by RT-PCR after 3 hour treatment, and the protein expressions of CD40, ICAM-1, p-NF-κB p65 and p-IκBα were examined by Western blot or immunofluorescence after 24 hour treatment. Results Following treatment with LPS, both the expressions of CD40 and ICAM-1 protein in RPMCs were up-regulated significantly (P<0.05), and the phosphoralation of p65 was increased greatly (1.10±0.17 vs 0.55±0.06, P<0.05). BAY11-7085 (5 μmol/L) significantly decreased the protein expression of p-p65 (0.22±0.11 vs 1.10±0.17, P<0.01), CD40 (0.34±0.02 vs 0.50±0.06, P<0.05) and ICAM-1 (0.35±0.16 vs 0.74±0.03, P<0.05). Pretreated with rosiglitazone for 3 h then added with LPS for 1 h, the levels of p-p65, CD40 and ICAM-1 in RPMCs were significantly decreased compared with those of LPS group (0.77±0.08 vs 0.90±0.10, P相似文献   

再生障碍性贫血造血细胞Fas抗原表达与T淋巴细胞功能的研究

目的 研究造血细胞凋亡与T淋巴细胞免疫在再生障碍性贫血(再障，Aplastic anemia，AA)发病机制中的作用及两者相关性。方法 采用流式细胞仪测定40例AA患者和15例非血液、免疫系统疾病对照者骨髓单个核细胞(BMMNC)的总CD34^ 、CD34^ Fas^ 、CD34^ Fas^-、CD3^ 、CD8^ 、CD3^ 、CD3^ CD25^ 标记值。结果 (1)与对照组比较，AA组总CD34^ 细胞％明显减少而其Fas表达率(以占总CD34^ 细胞％为计)明显增高。(2)AA组CD34^ 细胞％数与其Fas抗原表达率无明显负相关。(3)AA组T细胞％明显增多，且以CD8^ 细胞和CD3^ CD25^ 细胞增多为主。(4)AA组CD34^ 细胞％数与其T细胞活化状态无显著负相关。(5)AA组CD34^ 细胞Fas表达率与其T细胞活化状态无显著正相关。结论 AA骨髓存在着造血细胞数量减少和T淋巴细胞亚群数量、功能的异常，造血细胞数量减少还可能与Fas以外途径诱导的凋亡过度有关。骨髓造血细胞凋亡过度可能有活化T淋巴细胞免疫以外的途径诱导Fas途径或活化T淋巴细胞可以通过Fas之外的途径诱导造血细胞凋亡。     

膜表达热休克蛋白70真核表达载体的构建及其对肿瘤细胞的转染

目的构建普遍适合性的膜表达热休克蛋白70(HSP70)的真核表达载体,修饰并建立HSP 70表达并结合在细胞膜表面的肿瘤细胞.方法RT-PCR法从人胚肾中获取HSP 70的cDNA,扩增成带酶切位点的目的基因.选择载体pDisplay,将目的基因插入载体多克隆酶位点中.DNA测序证实后,以脂质体转染的方法,将载体转染到黑色素瘤细胞.用G418抗性筛选获得阳性克隆.结果酶切电泳和DNA测序证实载体构建正确;目的基因的上游带信号肽序列,下游带跨膜序列.RT-PCR产物电泳、Westem Blotting、激光共聚焦显微镜和流式细胞术证实大量HSP 70表达并结合在转染的黑色素瘤细胞膜表面.结论成功地构建了膜表达HSP 70的真核表达载体;转染到黑色素瘤细胞后,细胞膜表面表达丰富的HSP70.转染细胞可以进一步制备新型瘤苗.     

生长激素、肝细胞生长因子和烟酰胺对人胎胰岛细胞的增殖作用

目的研究生长激素(GH)，肝细胞生长因子(HGF)和烟酰胺(NIC)对体外培养的人胎胰岛细胞的增殖作用及其交互作用。方法采用La(2^7)正交设计法在体外培养的人胎胰岛细胞的各组中分别加入不同浓度及组合的GH，HGF和NIC，培养48h后，收集各孔细胞，DTZ染色，计数。结果GH，HGF和NIC均起主要作用，HGF和NIC的交互作用不可忽视，最佳的生长因子组合及适配浓度为GH(100ng／ml)HGF(25ng／ml)NIC(100mmol／L)。结论GH，HGF和NIC均能促进体外培养的胰岛细胞的增殖，且组合GH(100ng／ml)HGF(25ng／ml)NIC(100mmol／L)的作用最大。     

骨髓间质干细胞体外预构组织工程化肌腱的实验研究

目的探讨骨髓间质干细胞与胶原-聚羟基乙酸的细胞相容性,为构建组织工程化肌腱寻求理想方法.方法以贴壁法分离、培养骨髓间质干细胞,并检测CD44.在实验组中将骨髓间质干细胞置入含胶原-聚羟基乙酸的DMEM培基中培养:在对照组中将骨髓间质干细胞置入DMEM培基中培养.通过MTT方法比较两组的细胞活性和生长情况,并对实验组进行超微观察.以骨髓间质干细胞为种子细胞,以胶原-聚羟基乙酸为支架在体外预构组织工程化肌腱.结果以贴壁法原代培养骨髓间质干细胞,11天细胞即汇合成片,检测CD44示阳性.骨髓间质干细胞接种于胶原-聚羟基乙酸中混合培养后14天生长良好,始终保持89%以上的细胞活力,与对照组比较无显著差别;实验组细胞数未发生明显改变,而对照组从第4天开始即发生增殖.透射电镜示实验组细胞培养14天后仍保持旺盛的分泌功能.体外预构的组织工程化肌腱具有良好的形态,细胞伸展成梭形,沿聚羟基乙酸缝线大致平行排列.结论骨髓间质干细胞与胶原-聚羟基乙酸的细胞相容性好.以骨髓间质干细胞为种子细胞,以胶原-聚羟基乙酸为支架可在体外初步预构组织工程化肌腱.     

Bleeding from foveolar hyperplasia developing on a gastrointestinal stromal tumor of the stomach

A 52‐year‐old Japanese woman who presented with gastrointestinal (GI) bleeding underwent a proximal gastrectomy for a gastrointestinal stromal tumor (GIST) with a foveolar hyperplasia at the apex of the tumor, 4.5 cm in size, located in the upper body of the stomach. Although GIST are often asymptomatic and are found only incidentally, clinical symptoms such as bleeding, abdominal pain, or obstruction, occasionally lead to a premorbid diagnosis. When submucosal tumors present GI bleeding, the source of the bleeding usually is an ulceration of the mucosa over the tumor. However, in the present study, it was thought that the bleeding originated from the region of foveolar hyperplasia.     

中国仓鼠肺成纤维细胞和HeLa细胞DNA氧化损伤的自身修复能力与比较

目的:研究不同氧化相关因素对中国仓鼠肺成纤维细胞(CHL)和HeLa细胞DNA损伤的自身修复情况.方法:将CHL细胞和HeLa细胞用不同氧化相关因素处理一定时间[CHL细胞:过氧化氢(H2O2)25 min,重铬酸钾(K2Cr2O7)105 min,阿霉素(Dox)75 min;HeLa细胞:H2O2 25 min,K2Cr2O7 105 min],随后立即去毒培养0、0.5、1、2、3 h,以碱性单细胞凝胶电泳技术检测DNA链断裂情况.结果:①CHL细胞经H2O2、K2Cr2O7、Dox作用后引起DNA链断裂,去毒培养1 h链断裂修复明显(P<0.01);去毒培养2～3 h,前两毒剂的损伤组完全修复,而Dox组链断裂仍高于未损伤组;②HeLa细胞经H2O2、K2Cr2O7作用后引起DNA链断裂,去毒培养0.5 h链断裂明显修复(P<0.01),去毒培养1 h则完全修复;③CHL细胞和HeLa细胞损伤后修复的拖尾率与修复时间的回归系数显著不同(P<0.05).结论:两种细胞在氧化性DNA损伤后均迅速启动自身修复,但HeLa细胞比CHL细胞有更快的修复能力;同时这两种细胞由Dox所致损伤修复能力均较H2O2、K2Cr2O7所致的差.     

Why study transport of peptides and proteins at the neurovascular interface

The blood–brain barrier (BBB) is an immense neurovascular interface. In neurodegenerative, ischemic, and traumatic disorders of the central nervous system (CNS), the BBB may hinder the delivery of many therapeutic peptides and proteins to the brain and spinal cord. Fortunately, the mistaken dogma that peptides and proteins do not cross the BBB has been corrected during the past two decades by the accumulating evidence that peptides and proteins in the periphery exert potent effects in the CNS. Not only can peptides and proteins serve as carriers for selective therapeutic agents, but they themselves may directly cross the BBB after delivery into the bloodstream. Their passage may be mediated by simple diffusion or specific transport, both of which can be affected by interactions in the blood compartment (outside the BBB) and within the endothelial cells (at the BBB level). Although the majority of current delivery strategies focuses on modification of the molecule to be delivered, understanding the mechanisms of transport will eventually facilitate regulation of the BBB directly. We review the different aspects of interactions and discuss recent advances in the cell biology of peptide/protein transport across the BBB. Better understanding of the nature and regulation of the transport systems at the BBB will provide a new direction to enhance the interactions of peripheral peptides and proteins with the CNS.