In vitro responses of human pulp cells and 3T3 mouse fibroblasts to six contemporary dental restoratives

In vitro responses of human primary pulp cells (HPCs) and 3T3 mouse fibroblasts to six contempo-rary commercial dental restoratives were evaluated using the WST-1 assay. The results show that Fuji II is not cytotoxic to both cells. Fuji II LC is not cyto-toxic to HPCs but cytotoxic to 3T3 cells, indicating that 3T3 cells are more vulnerable to 2-hydroxyethyl methacrylate (HEMA) than HPCs. Vitremer is very cytotoxic probably due to having diphenyliodonium chloride and HEMA in it. Z100 is very cytotoxic probably due to having triethylene glycol dimethacry-late (TEGDMA) in it. P60 is cytotoxic but less cyto-toxic than Z100 probably due to no TEGDMA in it. Durelon is the most cytotoxic among the six materials studied probably due to the high cytotoxicity of zinc ions. Additionally, the cytotoxcity of the tested mate-rials was found to be dose-dependent.

Cytotoxicity of seven recent dentine bonding agents on mouse 3T3 fibroblast cells

Today it is generally accepted that most bonding agents are cytotoxic. In this study the relative cytotoxicity of seven recent dentine bonding agents on mouse 3T3 fibroblast cells were investigated. Materials and Methods. Near-confluent mouse 3T3 fibroblast cells were exposed to Dulbecco Modified Eagle’s Medium containing extractions from the seven different bonding agents. The cell survival rate was then determined using the standard MTT assay. Results. The cell survival rate ranking is: iBond (94%) < Gbond (78%) < Xeno V (71%) < Adper Easy Bond (63%) < Xeno V+ (61%) < Adper Scotchbond SE (33%) < XP Bond (32%). Part A of Adper Scotchbond SE had a survival rate of 35% and part B 38%. These two parts did not differ significantly. Adper Scotchbond SE and XP Bond do not differ significantly. While Xeno V+, Xeno V and Adper Easy Bond do not differ. (p < 5%;Tukey-Kramer Multiple-Comparison Test). Conclusion. All of the tested adhesive bonding agents were cytotoxic with survival rate of 3T3 cells between 94% to 31%. Of the 7 bonding agents tested iBond was found to be only slightly toxic and by far the least toxic. The two bonding agents (XP Bond and Adper Scotchbond SE) containing UDMA plus TEGDMA plus HEMA plus camphorquinone were found to be the most toxic.

Human ovarian neoplasm cell CD147 stimulates production and activation of matrix metalloproteinases in co-cultures with mouse fibroblasts

Objective: To investigate the expression of CD147 on human ovarian neoplasm cell lines and its influence on production and activation of matrix metallproteinases(MMPs). Methods: The expression of CD147 on different human ovarian neoplasm cell lines was studied by western blotting. Co-culture was carried out to investigate the stimulative effect of the positive expression CD147 cell HO-8910 on the production of MMPs of fibroblast cell in vitro. Zymography and immune blotting were used to study the production and activity of positive MMPs, at the time, to explore the relation between CD147 and MMPs. Results: CD147 was positively presented in 2 ovarian neoplasm cell lines(HO-8910,3-AO), but in SKOV3, TC-1,NIN3T3 cell was negative. MMP-2 and MMP-9 were detected by HO-8910 cell line, mouse fibroblast cell and co-culture cells; but the expression in co-culture cell is obviously higher than individual cultures of each type alone.CD147 stimulated MMPs in dose-dependent manner. Conclusion: CD147 causes increased production and activation of MMP-2, MMP-9.CD147 is probably a indirect marker of some ovarian cancer cells with invasion and metastasis.

Tryptophan 2,3-dioxygenase-positive matrix fibroblasts fuel breast cancer lung metastasis via kynurenine-mediated ferroptosis resistance of metastatic cells and T cell dysfunction

Background:Tumor metastasis is a major threat to cancer patient survival.The organ-specific niche plays a pivotal role in tumor organotropic metas-tasis.Fibroblasts serve as a vital component of the metastatic microenviron-ment,but how heterogeneous metastasis-associated fibroblasts(MAFs)promote organotropic metastasis is poorly characterized.Here,we aimed to decipher the heterogeneity of MAFs and elucidate the distinct roles of these fibroblasts in pulmonary metastasis formation in breast cancer.Methods:Mouse models of breast cancer pulmonary metastasis were estab-lished using an in vivo selection method of repeated injections of metastatic cells purified from the mouse lung.Single-cell RNA-sequencing(scRNA-seq)was employed to investigate the heterogeneity of MAFs.Transgenic mice were used to examine the contribution of tryptophan 2,3-dioxygenase-positive matrix fibroblasts(TDO2^(+)MFs)in lung metastasis.Results:We uncovered 3 subtypes of MAFs in the lung metastatic microenviron-ment,and their transcriptome profiles changed dynamically as lung metastasis evolved.As the predominant subtype,MFs were exclusively marked by platelet-derived growth factor receptor alpha(PDGFRA)and mainly located on the edge of the metastasis,and T cells were enriched around MFs.Notably,high MF sig-natures were significantly associated with poor survival in breast cancer patients.Lung metastases were markedly diminished,and the suppression of T cells was dramatically attenuated in MF-depleted experimental metastatic mouse mod-els.We found that TDO2^(+)MFs controlled pulmonary metastasis by producing kynurenine(KYN),which upregulated ferritin heavy chain 1(FTH1)level in dis-seminated tumor cells(DTCs),enabling DTCs to resist ferroptosis.Moreover,TDO2^(+)MF-secreted chemokines C-C motif chemokine ligand 8(CCL8)and C-C motif chemokine ligand 11(CCL11)recruited T cells.TDO2^(+)MF-derived KYN induced T cell dysfunction.Conditional knockout of Tdo2 in MFs diminished lung metastasis and enhanced immune activation.Conclusions:Our study reveals crucial roles of TDO2^(+)MFs in promoting lung metastasis and DTCs’immune evasion in the metastatic niche.It suggests that targeting the metabolism of lung-specific stromal cells may be an effective treatment strategy for breast cancer patients with lung metastasis.

Effects of salvianolic acid-A on NIH/3T3 fibroblast proliferation,collagen synthesis and gene expression

AIM To investigate the mechanisms ofsalvianolic acid A（SA-A）against liver fibrosisin vitro.METHODS NIH/3T3 fibroblasts were culturedroutinely,and incubated with 10-4mol/L-10-7mol/L SA-A for 22 h.The cell viability wasassayed by[3H]proline incorporation,cellproliferation by[3H]TdR incorporation,cellcollagen synthetic rate was measured with[3H]proline impulse and collagenase digestionmethod.The total RNA was prepared from thecontrol cells and the drug treated cellsrespectively,and α（1）I pro-collagen mRNAexpression was semi-quantitatively analyzedwith RT-PCR.RESULTS 10-4mol/L SA-A decreased cellviability and exerted some cytotoxiciy,while10-5mol/L-10-7mol/L SA-A did not affect cellviability,but inhibited cell proliferationsignificantly,and 10-6mol/L SA-A had the besteffect on cell viability among theseconcentrations of drugs.10-5mol/L-10-6mol/LSA-A inhibited intracellular collagen syntheticrate,but no significant influence on extracellularcollagen secretion.Both 10-5mol/L and10-6mol/L SA-A could decrease α（1）I pro-collagen mRNA expression remarkably.CONCLUSION SA-A had potent action againstliver fibrosis.It inhibited NIH/3T3 fibroblastproliferation,intracellular collagen syntheticrate and type I pro-collagen gene expression,which may be one of the main mechanisms of thedrug.

TCRγδ^+CD^(3+)CD^(4-)CD^(8-)双阴性T细胞在3型鼠肝炎病毒诱导的小鼠慢性病毒性肝炎发病中的作用

目的本研究拟探讨在MHV-3诱导的慢性病毒性肝炎模型小鼠外周血和脾脏TCRγδ+DNT细胞在T淋巴细胞中比例变化以及其主要表面分子标记的变化。方法取60只C3H/Hej小鼠,腹腔注射10pfu MHV-3;应用流式细胞仪三色或四色荧光分析法分别检测小鼠外周血和脾脏TCRγδ+DNT细胞在T淋巴细胞中的比例及其CD25、CD28、CD30和CD44表达的变化。结果 MHV-3诱导的慢性病毒性肝炎小鼠各时间点外周血和脾脏TCRγδ+DNT在T淋巴细胞中的比例较对照组显著升高(P<0.05);在感染后10天达到最高峰:外周血和脾脏TCRγδ+DNT细胞占T淋巴细胞的比例分别为4.8±1.5%和4.5±1.3%;模型小鼠外周血和脾脏TCRγδ+DNT细胞的表型均为TCRγδ+CD3+CD4-CD8-CD25-CD28-CD30-CD44+。结论感染MHV-3后C3H/Hej小鼠体内TCRγδ+DNT细胞的数量增加,提示TCRγδ+DNT细胞参与了MHV-3诱导的小鼠慢性病毒性肝炎的发生和发展过程。

基因扫描分析小鼠T细胞受体CDR3谱型检测T细胞克隆技术的建立

目的建立小鼠T细胞受体(TCR)互补决定区3(CDR3)谱型的基因扫描分析术,为分析T细胞克隆提供研究方法。方法应用RT-PCR方法分别扩增近交系小鼠C57BL/6H-2b,Balb/CH-2d和F1代小鼠(Balb/c×C57BL/6F1H-2d/b,CB6F1)脾脏T细胞的21个TCRVα家族和18个TCRVβ家族的CDR3,并用基因扫描分析扩增产物以确定T细胞的克隆性。结果3种小鼠脾脏T细胞均表达所有的TCRVα和VβCDR3基因,基因扫描显示全部TCRCDR3谱型呈高斯(钟型)分布,提示均为多克隆T细胞。各家族CDR3基因表达频率接近,但呈现不同的多态性和长度分布。结论基因扫描分析TCRαβCDR3基因谱型是检测小鼠T细胞克隆性的精确敏感方法。

Tim-3对B16F10细胞共培养的小鼠脾淋巴细胞增殖的影响

目的:明确T细胞免疫球蛋白黏蛋白-3(Tim-3)对B16F10细胞共培养的小鼠脾淋巴细胞增殖的影响。方法:构建Tim-3真核表达载体后,CCK-8法检测Tim-3融合蛋白对B16F10共培养的小鼠脾淋巴细胞的增殖。结果:Tim-3融合蛋白转染组淋巴细胞增殖活力低于未转染组和转染空载体组,差异有统计学意义(P<0.05)。结论:Tim-3融合蛋白抑制B16F10共培养的小鼠脾淋巴细胞的增殖。

哮喘小鼠肺组织和肺T细胞β1,3-N-乙酰糖基转移酶的表达

目的研究支气管哮喘模型小鼠肺组织及肺T细胞中三种β1,3-N-乙酰糖基转移酶(Fringe)RFNG、LFNG、MFNG的基因和蛋白表达的特点,探讨Fringe是否与哮喘发病有关。方法使用卵白蛋白腹腔注射后雾化吸入法制备BALB/c小鼠哮喘模型,并设置生理盐水对照组。经Perco1及尼龙毛法分离获取肺T细胞。采用半定量RT-PCR检测两组小鼠肺组织及肺T细胞中RFNG、LFNG、MFNGmRNA表达;Westernblot检测两组小鼠肺组织中RFNG、LFNG、MFNG蛋白表达。结果两组小鼠肺组织及肺T细胞中均存在三种Fringe表达。哮喘组肺组织及肺T细胞的RFNGmRNA表达较对照组增加(肺组织:0.92±0.35比0.51±0.13,P<0.01;肺T细胞:0.33±0.06比0.18±0.07,P<0.01);LFNGmRNA表达较对照组减少(肺组织:0.77±0.32比1.61±0.31,P<0.01;肺T细胞:0.49±0.19比0.71±0.03,P<0.01)。MFNG在肺组织中的表达无明显差异(肺组织:1.44±0.29比1.70±0.44,P>0.05),但MFNG在肺T细胞中的表达明显减少(0.11±0.03比0.52±0.18,P<0.01)。肺组织中三种Fringe蛋白表达与肺组织的mRNA结果相似。哮喘组的RFNG蛋白表达高于对照组(1.17±0.04比0.68±0.07,P<0.05),MFNG蛋白表达在两者之间没有明显的差异(8.10±0.60比9.12±0.07,P>0.05),而LFNG的蛋白表达则低于对照组(4.11±0.38比6.41±0.11,P<0.05)。结论三种Fringe的异常表达可能与支气管哮喘的发病有关。

骨髓内皮细胞产物对 CFU-F、CFU-GM和WEHI-3细胞生长的作用

为了研究骨髓内皮细胞的造血调控功能 ,采用串联超滤技术从小鼠骨髓内皮细胞无血清条件培养液 ( m BMEC- CM)中制备出不同相对分子质量 ( Mr)范围的超滤组分 ,进行细胞集落形成实验 ,检测了 m BMEC- CM及其超滤组分对骨髓成纤维祖细胞 ( CFU- F)、粒巨噬系造血祖细胞 ( CFU- GM)和小鼠粒单系白血病细胞系 WEHI- 3细胞生长的作用 .结果显示 :m BMEC- CM抑制 CFU- F的生长 ,对 CFU- GM和 WEHI- 3细胞的生长却未见明显影响 ;Mr>1 0 4 组分对 CFU- F的生长无明显作用 ,但促进 CFU- GM的生长和抑制 WEHI- 3细胞的生长 ;Mr<3× 1 0 3组分对这 3种细胞的集落形成均有抑制作用 .

In-Vitro Cellular Responses of Human Dental Primary Cells to Dental Filling Restoratives

In vitro cytotoxicity of six contemporary commercial dental filling restoratives on human dental primary cells, pulp cells (HPCs) and human gingival fibroblasts (HGFs), were tested using WST-1 assay. Continuous 3T3 mouse fibroblast cell lines were used for comparison. The results show that conventional glass-ionomer cement (GIC) Fuji II is not cytotoxic to all the cells. Resin-modified GIC (RMGIC) Fuji II LC is not cytotoxic to both HPCs and HGFs but cytotoxic to 3T3 cells. RMGIC Vitremer and resin composite Z100 are very cytotoxic to all the cells. Resin composite P60 is cytotoxic but much less cytotoxic than Z100. Polycarboxylate cement Durelon is the most cytotoxic among the six tested materials. It was found that continuous 3T3 cell lines were more vulnerable to leachable cytotoxic components than primary HPCs and HGFs. It was also found that the cytotoxcity of the tested materials was dose-dependent.

Fibroblasts upregulate expression of adhesion molecules and promote lymphocyte retention in 3D fibroin/gelatin scaffolds

Bioengineered scaffolds are crucial components in artificial tissue construction.In general,these scaffolds provide inert three-dimensional(3D)surfaces supporting cell growth.However,some scaffolds can affect the phenotype of cultured cells,especially,adherent stromal cells,such as fibroblasts.Here we report on unique properties of 3D fibroin/gelatin materials,which may rapidly induce expression of adhesion molecules,such as ICAM-1 and VCAM-1,in cultured primary murine embryonic fibroblasts(MEFs).In contrast,two-dimensional(2D)fibroin/gelatin films did not show significant effects on gene expression profiles in fibroblasts as compared to 3D culture conditions.Interestingly,TNF expression was induced in MEFs cultured in 3D fibroin/gelatin scaffolds,while genetic or pharmacological TNF ablation resulted in diminished ICAM-1 and VCAM-1 expression by these cells.Using selective MAPK inhibitors,we uncovered critical contribution of JNK to 3D-induced upregulation of these adhesion molecules.Moreover,we observed ICAM-1/VCAM-1-dependent adhesion of lymphocytes to fibroblasts cultured in 3D fibroin/gelatin scaffolds,but not on 2D fibroin/gelatin films,suggesting functional reprogramming in stromal cells,when exposed to 3D environment.Finally,we observed significant infiltration of lymphocytes into 3D fibroin/gelatin,but not into collagen scaffolds in vivo upon subcapsular kidney implantation in mice.Together our data highlight the important features of fibroin/gelatin scaffolds,when they are produced as 3D sponges rather than 2D films,which should be considered when using these materials for tissue engineering.

高表达IDO的树突状细胞抑制T细胞增殖的实验研究

目的:研究基因转染后高表达吲哚胺2,3-双加氧酶(IDO)的树突状细胞(DC)对T细胞增殖的抑制作用,以探讨其在器官移植中排斥反应的作用。方法:采用逆转录-聚合酶链式反应(RT-PCR)从腹膜炎小鼠的小肠组织中获取IDO全长cDNA片段,将其克隆至真核表达质粒pTracer中,构建真核表达质粒IDO-pTracer,随后用LipofectamineTM2000介导转染小鼠脾脏DC,分为未转染组、空载体转染组、IDO基因转染组和IDO基因转染并添加1-甲基色氨酸(1-MT)组,以混合淋巴细胞反应的方法检测转染后DC对同种异系T细胞增殖的作用。结果:克隆的小鼠IDO基因全长cDNA大小约1.3kb,无碱基突变,以构建的重组真核表达质粒IDO-pTracer转染DC后,混合淋巴细胞反应结果显示IDO基因转染组的刺激指数(SI)低于未转染组、空载体转染组及IDO基因转染并添加1-MT组,差异均有统计学意义(P<0.05)。结论:成功构建了IDO基因的真核表达载体IDO-pTracer,且IDO基因转染DC具有抑制异系T细胞增殖的作用。

多房棘球绦虫重组Bb—EmⅡ／3-Em14—3—3疫苗诱导BALB／c小鼠T淋巴细胞亚群变化的研究

目的探讨多房棘球绦虫（Em）重组Bb-EmⅡ／3-Eml4．3．3疫苗免疫和Em原头节攻击后小鼠脾CD4＋和CD8＋T淋巴细胞亚群的变化。方法用重组Bb-EmⅡ／3-Em14．3．3疫苗分别通过皮下注射、肌肉注射、鼻腔黏膜接种以及口服接种免疫BALB／c小鼠，双歧杆菌（Bb）液和磷酸盐缓冲液（PBS）为对照，12周后，用50个Em原头蚴腹腔注射进行攻击，攻击感染18周剖杀小鼠，检获泡球蚴组织，称取重量，计算减蚴率；分离脾细胞，流式细胞仪检测脾CD4＋和CD8＋T淋巴细胞亚群的百分比。结果疫苗免疫组（皮下注射、肌肉注射、鼻腔黏膜接种、口服接种）小鼠检获泡球蚴重量[（0．77±0．52）、（0．87±0．60）、（2．17±0．50）、（3．06±0．15）g]均明显低于PBS对照组[（3．54±0．32）g．P〈0．05或〈0．01]。疫苗免疫组（皮下注射、肌肉注射、鼻腔黏膜接种、口服接种）小鼠脾CD4＋T细胞亚群[（28．2±2．5）％、（25．0±2．7）％、（24．0±1.3）％、（23．0±1．8）％]显著高于PBS对照组[（16．1±2．2）％，P均〈0．01]；各疫苗免疫组小鼠CD8＋T细胞亚群水平均轻微升高，但组间比较差异无统计学意义（F=1．36，P〉0．05）。皮下注射组小鼠CD4＋T细胞亚群高于肌肉注射组、鼻腔黏膜接种组和口服接种组（P均〈0．05）。结论CD4＋T细胞亚群在Em重组Bb-EmⅡ／3-Em14-3-3疫苗诱导的小鼠抗Em原头节攻击感染的保护性免疫机制中起关键作用，疫苗皮下注射是一种较好的免疫途径。

TSA体外抑制Ⅱ型胶原诱导的抗原特异性T细胞增殖

目的:检测TSA对Ⅱ型胶原诱导的T细胞增殖的抑制作用。方法:采用CIA小鼠模型,收集引流淋巴结细胞。MTT法观察不同浓度TSA对T细胞的抑制作用,Annexin-v观察细胞凋亡情况。用Caspase 3试剂盒检测Caspase 3酶活性。结果:TSA呈时间和剂量依赖性抑制T细胞增殖,经20ng/ml TSA处理后,T细胞凋亡率增高,呈现时间依赖性效应关系。20ng/ml TSA作用T细胞后,实验组与对照组相比Caspase3活性明显升高。结论:TSA可能通过上调Caspase3活性,诱导细胞凋亡,从而抑制T细胞增殖。

Carcinogenic potential of duodenal reflux juice from patients with long-standing postgastrectomy

AIM: To determine whether study on the carcinogenic potential of reflux juice from patients with remote gastrectomy could clarify the inherent relationship between duodenal reflux and gastric stump cancer. METHODS: A total of 37 reflux juice samples (13 Billroth I, 24 Billroth II) were employed in the present study. A two-stage transformation assay using BALB/c 3T3 cells was carried out to test the initiating or promoting activity of these samples. RESULTS: Two of 18 (11.1%) reflux samples exerted initiating activities, whereas 9/19 (47.4%) samples enhanced the MNNG-initiating cell transformation, suggesting the duodenal reflux juice might more frequently possess the tumor-promoter activity (P = 0.029). In addition, there was no difference in initiating activities of the samples irrespective of surgical procedures (P = 0.488), while Billroth II samples exhibited stronger tumor-promoter activity than Billroth I samples (P = 0.027). Furthermore, the promoter activities were well correlated with the histological changes of the stomas (r(s) = 0.625, P = 0.004), but neither their cytotoxicities nor initiating activities had this correlation (Probabilities were 0.523 and 0.085, respectively). CONCLUSION: The duodenal reflux juice from patients with remote postgastrectomy did have carcinogenic potential, and suggested that tumor-promoting activity should principally account for the high incidence of gastric cancer in gastrectomy patients. In contrast, it is difficult to explain the high stump-cancer incidence with the N-nitroso compounds theory-a popular theory for the intact stomach carcinogenesis, and it seemed to be justified to focus chemo-prevention of this cancer on the tumor-promoting potential of reflux juice.

Novel nervous and multi-system regenerative therapeutic strategies for diabetes mellitus with mTOR

Throughout the globe,diabetes mellitus（DM） is increasing in incidence with limited therapies presently available to prevent or resolve the significant complications of this disorder.DM impacts multiple organs and affects all components of the central and peripheral nervous systems that can range from dementia to diabetic neuropathy.The mechanistic target of rapamycin（m TOR） is a promising agent for the development of novel regenerative strategies for the treatment of DM.m TOR and its related signaling pathways impact multiple metabolic parameters that include cellular metabolic homeostasis,insulin resistance,insulin secretion,stem cell proliferation and differentiation,pancreatic β-cell function,and programmed cell death with apoptosis and autophagy.m TOR is central element for the protein complexes m TOR Complex 1（m TORC1） and m TOR Complex 2（m TORC2） and is a critical component for a number of signaling pathways that involve phosphoinositide 3-kinase（PI 3-K）,protein kinase B（Akt）,AMP activated protein kinase（AMPK）,silent mating type information regulation 2 homolog 1（Saccharomyces cerevisiae）（SIRT1）,Wnt1 inducible signaling pathway protein 1（WISP1）,and growth factors.As a result,m TOR represents an exciting target to offer new clinical avenues for the treatment of DM and the complications of this disease.Future studies directed to elucidate the delicate balance m TOR holds over cellular metabolism and the impact of its broad signaling pathways should foster the translation of these targets into effective clinical regimens for DM.

Insights into chitosan hydrogels on dentine bond strength and cytotoxicity

Contemporary dental adhesives show favorable immediate results in terms of bonding effectiveness. However, the durability of resin-dentin bonds is their major problem. Materials and Methods: Preparation of 3 chitosan-antioxidant hydrogels was achieved using modified hydrogel preparation method. Their effect on the bond strength to dentine both short term (after 24 hours) and long term (after 6 months) were evaluated using shear bond strength measurements using Instron Universal Testing Mascine). The SEM was used to study the surface of the hydrogels. The cell survival rate (cytotoxicity) of the antioxidants resveratrol, β-carotene and propolis towards Balb/c 3T3 mouse fibroblast cells was also assessed using the standard MTT assay. Results: It was found that chitosan-H treated dentine gives significantly (p β-carotene (92%) > propolis (68%) > resveratrol (33%). Conclusion: the antioxidant-chitosan hydrogels significantly improved bonding to dentine with or without phosphoric acid treatment. The pH of the growth medium had a high influence on the cell survival rate of Balb/c mouse 3T3 fibroblast cells. The release of the antioxidant β-carotene would not have an influence on the pulp cells. These materials might address the current perspectives for improving bond durability.

调节性T细胞在3型鼠肝炎病毒诱导的小鼠暴发型肝炎模型中作用的初步探讨

目的检测调节性T细胞（Tr）在3型鼠肝炎病毒（MHV-3）诱导的小鼠暴发型肝炎模型中的比例变化及细胞因子表达，初步探讨Tr在该疾病模型中的作用。方法通过腹腔注射MHV-3感染BALB／cJ小鼠诱导暴发型肝炎，观察小鼠的生存时间，检测血清谷丙转氨酶（ALT）、谷草转氨酶（AST）水平，利用苏木精-伊红（HE）染色法检测肝脏病理学改变，分离感染不同时间点外周血、脾脏以及肝脏中的淋巴细胞，利用流式细胞术来检测Tr的比例以及细胞因子IL-10表达水平。结果BALB／cJ小鼠感染MHV-3后，全部在3—6d内死亡，血清ALT、AST水平随着感染时间延长逐渐升高，HE染色显示肝脏组织炎症及坏死程度逐渐加重，流式细胞术检测发现随着感染时间延长，小鼠肝脏中的rrr的比例明显升高。同时肝脏Tr分泌细胞因子IL-10的比例以及肝脏组织IL-10的mRNA水平逐渐升高。结论MHV-3诱导的小鼠暴发型肝炎模型中Tr在肝脏中的比例和功能显著升高，这种代偿性升高提示Tr可能发挥调节机体过度免疫反应的功能。

Tim-3在药物性急性肾损伤动物模型中的表达及作用机制

目的探索T细胞免疫球蛋白黏蛋白分子3(Tim-3)在急性肾损伤(AKI)中的作用,并研究其作用机制。方法将6~8周C57雄性小鼠随机分为对照组(CTRL组)、顺铂组(AKI组)及顺铂+阻断Tim-3组(RMT组),每组4只,AKI组使用顺铂(20 mg/kg)腹腔注射,构建AKI动物模型,检测肾脏损伤情况与Tim-3表达情况。RMT组使用抗体RMT3-23(200μg/只)阻断Tim-3分子,建立AKI模型,检测AKI组与RMT组肾脏损伤情况以及Tim-3、自噬相关分子的表达情况。结果肾脏近端小管出现损伤,Western blotting结果显示AKI组Tim-3表达水平较CTRL组增加(t=3.876,P=0.0082)。与AKI组相比,RMT组的NGAL和P62表达水平均增高,但差异无统计学意义(tNGAL=1.664,P=0.1570;tP62=1.991,P=0.1031),LC3II表达下降,差异有统计学意义(t=5.901,P=0.0020),自噬受到抑制且肾脏小管损伤加重(P=0.0101)。结论Tim-3可以通过调节肾小管上皮细胞自噬活性,减轻顺铂诱导的AKI肾损伤。