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Visualization of perforin/gasdermin/complement-formed pores in real cell membranes using atomic force microscopy

Cellular & Molecular Immunology 2019年第6期16卷 611-620页

作者： Yuying Liu Tianzhen Zhang Yabo Zhou Jiping Li Xiaoyu Liang Nannan Zhou Jiadi Lv Jing Xie Feiran Cheng Yiliang Fang Yunfeng Gao Ning Wang Bo Huang Department of Immunology&National Key Laboratory of Medical Molecular Biology Institute of Basic Medical SciencesChinese Academy of Medical SciencesSchool of Basic Medicine Peking Union Medical CollegeBeijing 100005China Clinical Immunology Center Chinese Academy of Medical SciencesBeijing 100005China Beijing SOJO Electric Company Limited BeijingChina Department of Mechanical Science and Engineering College of EngineeringUniversity of Illinois at Urbana-ChampaignUrbanaIllinois 61801USA Laboratory for Cellular Biomechanics and Regenerative Medicine College of Life Science and TechnologyHuazhong University of Science and TechnologyWuhanHubei 430074China Department of Biochemistry&Molecular Biology Tongji Medical CollegeHuazhong University of Science&TechnologyWuhan 430030China

Different types of pores ubiquitously form in cell membranes,leading to various types of cell death that profoundly influence the fate of inflammation and the disease ***,these pores have never truly been visualized to *** force microscopy(AFM),which is emerging as a powerful tool to analyze the mechanical properties of biomolecules and cells,is actually an excellent imaging platform that allows biological samples to be visualized by probing surface roughness at the level of atomic ***,membrane pore structures were clearly visualized using *** visualization not only describes the aperture and depth of the pore complexes but also highlights differences among the pores formed by perforin and gasdermins in tumor cell membranes and by complement in immune cell ***,this type of visualization also reveals the dynamic process of pore formation,fusion,and repair.

关键词： membrane pore formation AFM perforin GSDMD/GSDME complement

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Distinct patterns of mucosal apoptosis in H pylori-associated gastric ulcer are associated with altered FasL and perforin cytotoxic pathways

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World Journal of Gastroenterology 2006年第38期12卷 6133-6141页

作者： Heitor SP Souza Marcelo S Neves Celeste CS Elia Claudio JA Tortori Ilana Dines Cesonia A Martinusso Kalil Madi Leonardo Andrade Morgana TL Castelo-Branco Departamento de Clínica Médica Instituto de Ciências Biomédicas Universidade Federal do Rio de Janeiro 21941-590 Brazil Laboratório Multidisciplinar de Pesquisa e Departamento de Anatomia Patologica Instituto de Ciências Biomédicas Universidade Federal do Rio de Janeiro 21941-590 Brazil Hospital Universitário Clementino Fraga Filho Departamento de Histologia e Embriologia Instituto de Ciências Biomédicas Universidade Federal do Rio de Janeiro 21941-590 Brazil Laboratório de Imunologia Celular Instituto de Ciências Biomédicas Universidade Federal do Rio de Janeiro 21941-590 Brazil

AIM: To analyze the level of apoptosis in different mucosal compartments and the differential expression of Fas/Fas-ligand and perforin in H pylori-associated gastric ulcer. METHODS: Antral specimens from patients with H pylori-related active gastric ulcer (GU), H pylori-related gastritis, and non-infected controls were analysed for densities and distribution of apoptotic cells determined by the TdT-mediated dUDP-biotin nick-end-labelling method. GU patients were submitted to eradication therapy with follow-up biopsy after 60 d. Fas, FasL, and perforin-expressing cells were assessed by immunoper- oxidase, and with anti-CD3, anti-CD20 and anti-CD68 by double immunofluorescence and confocal microscopy. Quantitative analysis was performed using a computer- assisted image analyser. RESULTS: H pylori-infected antrum showed greater surface epithelial apoptosis which decreased after eradi- cation therapy. In the lamina propria, higher rates of mononuclear cell apoptosis were observed in H pylori- gastritis. Co-expression of Fas with T-cell and macro- phage markers was reduced in GU. FasL- and perforin-expressing cells were increased in H pylori-infection and correlated with epithelial apoptosis. perforin-expressing cells were also increased in GU compared with H pylori- gastritis. CONCLUSION: Epithelial apoptosis is increased in H pylori-infection and correlates to FasL- and perforin- expression by T cells. Expression of perforin is correlated with the tissue damage, and may represent the enhance- ment of a distinct cytotoxic pathway in GU. Increased expression of FasL not paralleled by Fas on T-cells and macrophages may indicate a reduced susceptibility to the Fas/FasL-mediated apoptosis of lymphoid cells in H pylori-infection.

关键词： Gastric ulcer H pylori Apoptosis Fas receptor Fas ligand perforin

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Mice lacking perforin have improved regeneration of the injured femoral nerve

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Neural Regeneration Research 2022年第8期17卷 1802-1808页

作者： Igor Jakovčevski Monika von Düring David Lutz Maja Vulović Mohammad Hamad Gebhard Reiss Eckart Förster Melitta Schachner Institut für Anatomie und Klinische Morphologie Universität Witten/HerdeckeWittenGermany Department of Neuroanatomy and Molecular Brain Research Ruhr University BochumBochumGermany Department of Anatomy Faculty of Medical SciencesUniversity of KragujevacKragujevacSerbia Keck Center for Collaborative Neuroscience and Department of Cell Biology and Neuroscience Rutgers UniversityPiscatawayNJUSA

The role that the immune system plays after injury of the peripheral nervous system is still not completely ***,a natural killer cell-and T-lymphocyte-derived enzyme that mediates cytotoxicity,plays important roles in autoimmune diseases,infections and central nervous system trauma,such as spinal cord *** dissect the roles of this single component of the immune response to injury,we tested regeneration after femoral nerve injury in perforin-deficient(Pfp^(-/-))and wild-type control *** frame motion analysis showed better motor recovery in Pfp^(-/-)mice compared with control mice at 4 and 8 weeks after *** tracing of the motoneuron axons regrown into the motor nerve branch demonstrated more correctly projecting motoneurons in the spinal cord of Pfp^(-/-)mice compared with *** of regrown axons measured by g-ratio was more extensive in Pfp^(-/-)than in wild-type mice in the motor branch of the femoral ***^(-/-)mice displayed more cholinergic synaptic terminals around cell bodies of spinal motoneurons after injury than the injured *** histologically analyzed lymphocyte infiltration 10 days after surgery and found that in Pfp^(-/-)mice the number of lymphocytes in the regenerating nerves was lower than in wild-types,suggesting a closed blood-nerve barrier in Pfp^(-/-)*** conclude that perforin restricts motor recovery after femoral nerve injury owing to decreased survival of motoneurons and reduced myelination.

关键词： blood-nerve barrier femoral nerve injury locomotor recovery lymphocytes myelination NK-cells perforin reinnervation

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The Effect of Anti-Human CD134 Monoclonal Antibody on Phytohemagglutinin-Induced mRNA Expression of perforin in Peripheral Blood Mononuclear Cells

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Cellular & Molecular Immunology 2005年第6期2卷 467-471页

作者： Ming Li~1 Yuanchao Zhang~(1,2) ~1Department of Rheumatology and Immunology,Shandong Provincial Hospital,Shandong University,Jinan 250021,China ~2Department of Rheumatology and Immunology,Shandong Provincial Hospital,Shandong University,Jinan 250021,Shandong,China. Department of Rheumatology and Immunology Shandong Provincial Hospital Shandong University Jinan 250021 China.

CD134, a member of the tumor necrosis factor receptor superfamily, plays a crucial role in T cell survival. In this study, peripheral blood mononuclear cells （PBMCs） stimulated with phytohemagglutinin （PHA, 50 μg/ml） were treated by CD134 mAb （1μg/ml, 5 μg/ml, 10 μg/ml） for 6 h, 12 h, 24 h and 48 h. The level of perforin mRNA was measured by reverse transcription-polymerase chain reaction （RT-PCR） technique. Our data showed that the expression of perforin mRNA in PBMCs was down-regulated by CD134 mAb in a dose-dependent manner in range of 1 μg/ml to 5 μg/ml and dropped down to its minimum on 24 h （p 〈 0.05）. The level of perforin mRNA reached a plateau when the concentration of CD134 mAb exceeded 5 μg/ml. In conclusion, CD134 mAb can inhibit perforin expression, which may enhance the ability of T cells for survival. Cellular ＆ Molecular Immunology. 2005;2（6）： 467-471.

关键词： CD134 perforin phytohemagglutinin

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In situ detection of tumor infiltrating lymphocytes expressing perforin and fas-ligand genes in human HCC

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World Journal of Gastroenterology 1999年第1期5卷 17-19页

作者： QIAN Qi Jun, XUE Hui Bin, QU Zeng Qiang, FANG Shi Gang, CAO Hui Fang and WU Meng Chao

AIM To investigate the expression of perforin and fas ligand (fas L) of tumor infiltrating lymphocytes (TILs) in human hepatocellular carcinoma (HCC). METHODS By in situ hybridization and immunohistochemistry, the perforin and fas L gene expression of TILs was studied in 20 HCC cases. RESULTS Positive expression of perforin and fas L genes was detected in 16 HCC cases. One patient had expression of perforin and fas L genes in the majority of TILs and survived 1 5 years after tumor resection without HCC relapse. This seems that the presence of a large number of activated T cells might be beneficial for the antitumor immunity. In other cases, less than 10% of TILs were able to express perforin and fas L genes. CONCLUSION Although there were a number of T cells in HCC, only few of them were immunoactive and able to kill tumor cells. It seems important to promote further proliferation of these activated T cells in vitro or in vivo .

关键词： carcinoma, hepatocellular lymphocytes fas ligand genes perforin in situ hybridization liver neoplasms

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perforin Expression in <i>Theileria parva</i>Specific Cytotoxic T Cells Correlates with Cytotoxicity

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Open Journal of Immunology 2014年第3期4卷 117-127页

作者： Jerome Wendoh Rebecca Waihenya Rosemary Saya Elias Awino Vishvannath Nene Lucilla Steinaa International Livestock Research Institute Nairobi Kenya Jomo Kenyatta University of Agriculture and Technology Nairobi Kenya Kenya Medical Research Institute Nairobi Kenya

Theileria parva is a protozoan parasite that causes the disease East Coast fever in cattle which results in major economic losses in Eastern, Central and Southern Africa. Efforts to generate vaccines involve measurements of cytotoxic activity since CD8 cells are believed to be responsible for protection of the animals. CTL assays are time consuming, and may require use of radioactive material and they also impose a considerable amount of in vitro work, which may skew the response compared to ex vivo assays. Hence it would be beneficial to identify other markers that correlate with the cytotoxicity. In this in vitro study we examined if the number of tetramer positive CD8 cells and the staining intensity of these correlated with lysis of the target cells. Furthermore, we investigated if the expression of the activation marker perforin correlated with the cytotoxicity. perforin is involved in permeabilization of the cell membrane of the target cell. Bulk CTL lines and purified CD8 cell lines generated from cattle of the A18 BoLA (MHC) type were analysed for the Theileria parva specific immune responses using a peptide-MHC tetramer and antibodies for perforinin FACS analysis. Thelysis of target cells was determined by a chromium release assay. Results demonstrate that the percentage of tetramer positive cells, in six cell lines generated against the whole parasite, correlate with killing of PBMC pulsed with the peptide. The product of the percentages of perforin and tetramer double positive cells and the net MFI of perforin showed a perfect correlation with the cytotoxicity of the peptide pulsed PBMC. Likewise, the product of percentage perforin positive cells and the staining intensity had the best significant correlation with killing of the pulsed PBMC. The present results suggest that perforin could be a possible biomarker for the cytotoxicity to Theileria parva infections/immunizations.

关键词： Cytotoxic T Cells Biomarkers perforin Theileria parva

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Lymphocytes and perforin expression in endometrium during the menstrual cycle

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Chinese Medical Journal 2000年第10期113卷 66-69页

作者：田秀珠张丽珠张小为 Department of Obstetricsand Gynecology Third Clinical Medical SchoolBeijing Medical niversity

Obejctive To study the lymphocytes, perforin protein and mRNA expression in endometrium of different phases in the menstrual cycle and to explore the regularity and role of the lymphocytes and perforin in the reproductive system Methods Immunohistochemical and in situ hybridization techniques were employed to demonstrate the population, phenotype and mediator (perforin) of lymphocytes in the proliferative and secretory endometrium Results In different phases of the endometrium, there were very few CD 3 + typical T lymphocytes, CD 4 + helper lymphocytes and CD 8 + suppressor/cytotoxic T lymphocytes, while CD 56 + lymphocyte (also named NK like cell) were abundant CD 56 + lymphocytes were absent in postmenopausal endometrium A double immunohistochemical labelling method demonstrated the co expression of perforin protein and CD 56 antigen of cells in different phases of the endometrium Quantitative analysis of CD 56 + lymphocytes and perforin positive cells increased in number from the proliferative to the secretory endometrium ( P perforin mRNA positive cells formed islands in proliferative endometrial stroma, but were scattered in secretory endometrium There was difference between perforin gene expression and protein expression in the proliferative endometrium Conclusions Lymphocytes and perforin expression in the endometrium at different phases of the menstrual cycle are specific and may play an important role in the reproductive system They may exert a positive influence on embryo implantation and be involved in endometrial stroma breakdown during menstruation

关键词： lymphocyte perforin endometrium menstrual cycle

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perforin ameliorates high fat diet-induced non-alcoholic fatty liver disease

Perforin ameliorates high fat diet-induced non-alcoholic fat...

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第十二届全国免疫学学术大会

作者： Li Dehai jinan university

Non-alcoholic fatty liver disease(NAFLD) is a metabolic disease that seriously threatens human health,with a much higher prevalence among obese ***’s reported that intrahepatic immune cells haved important roles in regulating the *** studies showed perforin was a immune regulator in obesity-related metabolic *** perforin in regulating the process of NAFLD is not *** using high fat diet-induced non-alcoholic fatty liver disease model,We found that the presence of perforin could reduce the extent of nonalcoholic fatty liver *** with HFD-fed WT mice,perforin deficient mice showed increased body weight,liver weight,glucose tolerance,insulin *** Oil red and HE staining liver structure,we found significant accumulation of fat droplets phenomenon of Prf *** high fat diet-induced non-alcoholic fatty liver disease model,the livers of Prfmice showed upregulated expression of lipogenic genes in comparison with WT ***-fed Prf mice showed the higher ratio of IFN gamma in CD4 T ***,the proportion of NK cells,CD4 Tcells,CD8 Tcells,NKT cells were not *** conclusion,the findings support the function of perforin in non-alcoholic fatty liver disease and suggest that IFN gamma in inflamed liver is one possible avenue for therapeutic intervention.

关键词： perforin Non-alcoholic fatty liver disease Lipogenic IFN gamma

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perforin gene mutations in 77 Chinese patients with lymphomas

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World Journal of Emergency Medicine 2013年第2期4卷 128-132页

作者： Qi Ding Li-yun Yang Department of Hematology Peking University First Hospital Department of Clinical Laboratory General Hospital of Armed Police

BACKGROUND:perforin gene(PRF1) mutations have been reported in patients with lymphoma,but the prevalence and characteristics of PRF1 mutation have not been identified in Chinese patients with ***:Seventy-seven patients with lymphoma,including 6 patients with Hodgkin lymphoma and 71 patients with non-Hodgkin lymphoma,were *** samples from peripheral blood were used for PRF1 mutation detection by the PCR-sequencing ***:Eleven novel PRF1 mutations were found in 8 of the 77 patients with *** or compound monoallelic missense mutations in 3 patients indicated the severe impairment of perforin function,monoallelic missense mutations in 3 patients possibly contributed a genetic predisposition to malignancies,and synonymous mutations in 2 patients showed unknown ***:The frequency of EBV infection was similar in lymphoma patients with PRF1 mutations and those without the *** same PRF1 mutations were also found in DNA samples from nails or hair follicles from 4 patients with PRF1 mutations,suggesting that these mutations may be of germ-line origin.

关键词： perforin Mutation Lymphoma

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前列腺癌患者外周血淋巴细胞中perforin和granzyme-B mRNA的表达及意义

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中南大学学报（医学版） 2015年第4期40卷 387-391页

作者：何景良李鎏勋危文素魏竞超龙智张一川何乐业中南大学湘雅三医院泌尿外科长沙410013

目的:探讨前列腺癌(prostate cancer,PCa)患者外周血淋巴细胞(peripheral blood lymphocyte,PBL)中perforin和granzyme-B的表达水平及意义。方法:运用荧光定量RT-PCR检测60例PCa患者PBL中perforin和granzyme-B的m RNA的表达水平,同期检测40例良性前列腺增生(benign prostatic hyperplasia,BPH)患者及20例健康者作为对照。结果:PCa组PBL中perforin和granzyme-B m RNA的表达水平较BPH组及健康对照组均明显下降(Pperforin和granzyme-B m RNA表达水平较对照组下降,且与病理分级和临床分期负相关,提示PCa的发生发展与患者免疫状态下降有关。

关键词：前列腺癌淋巴细胞 perforin granzyme-B

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