Granzyme B: A novel therapeutic target for treatment of atopic dermatitis

Granzyme B is a serine protease that can play multiple roles in intracellular and extracellular perforin-dependent or non-perforin-dependent mechanisms. Granzyme B has been found to be an important factor involved in the pathogenesis of atopic dermatitis and is increased in both skin lesions and peripheral blood of atopic dermatitis patients. In this article, we review the correlation between granzyme B and atopic dermatitis to provide a novel therapeutic targeting option for clinical treatment of the latter.

Preliminary study of the role of IL-18/IL-18BP in mediating cytotoxic ability of NK-92MI cells against endothelial cells from GTKO porcine models / 器官移植

Objective To evaluate the role and potential mechanism of interleukin (IL)-18/IL-18 binding protein (BP) in mediating the killing effect of natural killer (NK)-92MI cells upon endothelial cells from α-1, 3- galactosyltransferase gene-knockout (GTKO) porcine models. Methods NK-92MI cells were divided into the NK, NK+IL-18, NK+GTKO, IL-18+NK+GTKO and IL-18+IL-18BP+NK+GTKO groups. The messenger ribonucleic acid (mRNA) levels of inflammation-related genes in NK-92MI cells were detected by quantitative real-time reverse transcription polymerase chain reaction (qRT-PCR). The killing effect of NK-92MI cells on endothelial cells from GTKO porcine models was evaluated by lactate dehydrogenase (LDH) assay. The apoptosis of endothelial cells from GTKO porcine models was assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay. The expression levels of proteins with killing effect and apoptosis-related proteins were determined by Western blot. Results Compared with the NK, NK+IL-18 and NK+GTKO groups, the expression levels of interferon (IFN)-γ, tumor necrosis factor (TNF)-α, IL-8, IL-3, IL-6 and granulocyte-macrophage colony stimulating factor (GM-CSF) mRNA were up-regulated in NK-92MI cells in the IL-18+NK+GTKO group, and the differences were statistically significant (all P < 0.05). Compared with the IL-18+NK+GTKO group, the expression levels of IFN-γ, TNF-α, IL-8, IL-3, IL-6 and GM-CSF mRNA were down-regulated in NK-92MI cells in the IL-18+IL-18BP+NK+GTKO group, and the differences were statistically significant (all P < 0.05). Compared with the NK+GTKO group, the expression levels of perforin, granzyme B and IFN-γ proteins in NK-92MI cells were up-regulated, the killing rate of NK-92MI cells against endothelial cells from GTKO porcine models was enhanced, the apoptosis rate of endothelial cells from GTKO porcine models was increased, and the ratios of B cell lymphoma-2 (Bcl-2)-associated X protein (Bax)/Bcl-2 and cleaved Caspase-3/Caspase-3 in endothelial cells from GTKO porcine models were elevated in the IL-18+NK+GTKO group, and the differences were statistically significant (all P < 0.05). Compared with the IL-18+NK+GTKO group, the expression levels of perforin, granzyme B and IFN-γ proteins were down-regulated, the killing rate of NK-92MI cells against endothelial cells from GTKO porcine models was decreased, the apoptosis rate of endothelial cells from GTKO porcine models was decreased, and the ratios of Bax/Bcl-2 and cleaved Caspase-3/Caspase-3 in endothelial cells from GTKO porcine models were declined in the IL-18+IL-18BP+NK+GTKO group, and the differences were statistically significant (all P < 0.05). Conclusions IL-18BP may block the expression of inflammation-related genes in NK-92MI cells induced by IL-18 and the killing effect of NK-92MI cells on endothelial cells from GTKO porcine models.

The level of Tim-3+CD8+ T cells can serve as a potential marker for evaluating the severity of acute graft-versus-host disease after haplo-PBSCT

Early and accurate diagnosis of acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation is crucial for the prognosis of patients. This study identified a potential biomarker for the severity of aGVHD after human leukocyte antigen (HLA)-haploidentical peripheral blood hematopoietic stem cell transplantation (haplo-PBSCT). We included 20 healthy subjects and 57 patients who underwent haplo-PBSCT. Of these patients, 22 developed aGVHD after haplo-PBSCT. The results showed that patients with aGVHD had significantly increased levels of Tim-3+/Perforin+/Granzyme B+CD8+ T cells, but significantly decreased Galectin-9. The differences in Galectin-9 and Tim-3+/Granzyme B+CD8+ T cells between grade I-II aGVHD and III-IV aGVHD were also significant. In vitro, the apoptosis of CD8+ T cells from aGVHD patients was significantly increased after Tim-3/Galectin-9 pathway activation, which decreased Granzyme B secretion. As revealed by univariate analysis, the level of Tim-3+CD8+ T cells was a risk factor for severe aGVHD. ROC analysis demonstrated that high levels of Tim-3+CD8+ T cells had a significant diagnostic value for severe aGVHD, with an area under the curve of 0.854 and cut-off value of 14.155%. In conclusion, the binding of Tim-3 with exogenous Galectin-9 can promote apoptosis of CD8+ T cells and affect the secretion of Granzyme B. Tim-3+CD8+ T cells have the potential to serve as immunological markers for assessing the severity of aGVHD after haplo-PBSCT and identifying patients at a higher risk for severe aGVHD.

Antitumor impact of amygdalin on adaptive immune response in BALB/c mice with breast cancer

Amygdalin is a potential therapeutically target in cancer. The main purpose of this experimental study was to evaluate the therapeutic effect of amygdalin in the mice model of breast cancer. The percentages of CD4, CD8 T lymphocyte, intracellular IFN-?, and Granzyme B were assessed in spleen cells of tumorized mice treated with 50 and 150 mg/kg of amygdalin (AG50 and AG150). The expression of caspase 3 and p53, tumor size, and survival rate of Balb/c mice was determined in tumor tissue after amygdalin administration. No significant difference was observed in the frequency of CD4+ and CD8+ T cells in the three study groups. However, a significantly increased level of granzyme B in CD8+ T cells, as well as a significant decrease in the level of IL-10 in CD4+ T cells was detected in the AG50 group compared to the AG150. There was no significant difference in the expression of caspase 3 and P53 between the two groups. A significant change was seen in tumor size and survival rate of AG50 and AG150 groups compared to the controls. Our findings indicated that antitumor effect of amygdalin in vivo was probably due to stimulating the effective immune response, not apoptotic genes induction

Effect of IL-15 addition on asbestos-induced suppression of human cytotoxic T lymphocyte induction

BACKGROUND@#Asbestos fibers possess tumorigenicity and are thought to cause mesothelioma. We have previously reported that exposure to asbestos fibers causes a reduction in antitumor immunity. Asbestos exposure in the mixed lymphocyte reaction (MLR) showed suppressed induction of cytotoxic T lymphocytes (CTLs), accompanied by a decrease in proliferation of CD8@*METHODS@#For MLR, human peripheral blood mononuclear cells (PBMCs) were cultured with irradiated allogenic PBMCs upon exposure to chrysotile B asbestos at 5 μg/ml for 7 days. After 2 days of culture, IL-15 was added at 1 ng/ml. After 7 days of MLR, PBMCs were collected and analyzed for phenotypic and functional markers of CD8@*RESULTS@#IL-15 addition partially reversed the decrease in CD3@*CONCLUSION@#These findings indicate that CTLs induced upon exposure to asbestos possess dysfunctional machinery that can be partly compensated by IL-15 supplementation, and that IL-15 is more effective in the recovery of proliferation and granzyme B levels from asbestos-induced suppression of CTL induction compared with IL-2.

Effect of asbestos exposure on differentiation and function of cytotoxic T lymphocytes

Asbestos exposure is known to cause malignant mesothelioma, which is associated with poor prognosis. We focused on and examined the effect of asbestos exposure on the differentiation and function of cytotoxic T lymphocytes (CTLs). CTLs have the ability to specifically attack tumor cells after being differentiated from naïve CD8 T cells following antigen stimulation. Exposure to chrysotile B asbestos suppressed the differentiation of CTLs during the mixed lymphocyte reaction (MLR) and was associated with a decrease in proliferation of CD8 T cells. Additionally, in an effort to investigate the mechanism associated with suppressed CTL differentiation upon exposure to asbestos, we focused on IL-2, a cytokine involved in T cell proliferation. Our findings indicated that insufficient levels of IL-2 are not the main cause for the suppressed induction of CTLs by asbestos exposure, although they suggest potential improvement in the suppressed CTL function. Furthermore, the functional properties of peripheral blood CD8 lymphocytes from asbestos-exposed individuals with pleural plaque (PP) and patients with malignant mesothelioma (MM) were examined. MM patients showed lower perforin levels in CD8 lymphocytes following stimulation compared with PP-positive individuals. The production capacity of IFN-γ in the MM group tended to be lower compared with healthy volunteers or PP-positive individuals. In an effort to determine whether chronic and direct asbestos exposure affected the function of CD8 T cells, cultured human CD8 T cells were employed as an in vitro model and subjected to long-term exposure to chrysotile (CH) asbestos. This resulted in decreased levels of intracellular perforin and secreted IFN-γ. Those findings underlie the possibility that impaired CD8 lymphocyte function is caused by asbestos exposure, which fail to suppress the development of MM. Our studies therefore reveal novel effects of asbestos exposure on CTLs, which might contribute towards the development and implementation of an effective strategy for the prevention and cure of malignant mesothelioma.

Study on quantity and function of CD8+T cells in patients with repeated implantation failure / 实用医学杂志

Objective To investigate the quantity and function of CD8+T cells in peripheral blood of pa-tients with repeated implantation failure(RIF).Methods Thirty-seven patients with RIF and 19 healthy controls were enrolled in this study.The peripheral blood and endometrium were collected during the mid-luteal phase.The percentage of peripheral CD8+T subsets and the levels of perforin and granzyme B of peripheral CD8+T cells were determined by flow cytometry assay.The percentage of endometrial CD8+T cells was detected by IHC,the produc-tion of perforin and granzyme B of endometrial CD8+T cells was detected by IF. Results Compared with the con-trol group,the percentage of peripheral CD8+T cells in patients with RIF was not significantly changed(37.22% vs. 37.15%,P>0.05).However,the porportion of endometrial CD8+T cells in the RIF group was higher than that in the control group(1.99% vs.3.77%,P<0.001).The levels of perforin and granzyme B in peripheral blood and en-dometrial CD8+T cells in patients with RIF were similar with those in the control group.Conclusions Compared to the control group,the percentage of endometrial CD8+T was markedly upregulated in patients with RIF.However, the production of perforin and granzyme B were similar between the control group and the RIF group.

Regulatory mechanism of PESV on tumor-infiltrating natural killer cells in liver orthotopic transplantation tumor / 中国免疫学杂志

Objective:To investigate the regulatory mechanism of PESV on tumor-infiltrating natural killer ( NK) cells in a mice model with H22 orthotopic transplantation tumor .Methods:Suspensions of H22 cells were injected into the lobe of liver on C 57BL/6 mice for establishing liver orthotopic transplantation tumor model ,then the mice were randomly divided into four groups:normal group , control group ,PESV low dose group ( PESV-L ) and PESV high dose group ( PESV-H ) .Mice were either sacrificed for mechanistic studies or survival followed 14 days of therapy.The volume and weight of the tumor were measured .The proportion of infiltrating NK cells was measured by flow cytometry and the expression of NK 1.1(NK) cells was investigated by immunohistochemistry method .The expression of perforin and granzyme B were further investigated by real-time PCR.Results: In contrast to control group , the tumor inhibition rate was 15.38%and 30.77% in PESV-L group and PESV-H group respectively.The survival showed that PESV-H could significantly prolong the survival time of mice ,and life extension rate was 34.06%,(P<0.05).Histological analysis revealed significant pleomorphism of the neoplastic cells and invasive extendion in control group ,while there were more necrosis and less degree of atypia in PESV-L and PESV-H.The level of tumor-infiltrating NK cell was significantly higher in PESV-H than in tumor-bearing control group [(5.91±0.49)%vs.(3.69±0.50)%,P<0.05],and NK cells were infiltrating in peritumoral lesions.The mRNA of perforin and granzyme B in PESV-H were respectively 3.62 and 5.82 times than that of control group ( P<0.05 ) .Conclusion: These findings suggest that the treatment of PESV might increase the infiltration of natural killer cells in the orthotopic transplantation tumor and contribute to NK cells migration to the tumor , which induct and maintain the activities of natural killer cells against tumor cells by expressing perforin and granzyme B in vivo .

Effects of quercitrin on the proliferation and the cytotoxicity of human γδT cells / 中华微生物学和免疫学杂志

Objective To investigate the in vitro effects of quercitrin on the proliferation and the cytotoxicity of human γδT cells.Methods Peripheral blood mononuclear cells (PBMCs) were isolated from healthy subjects and cultured with isopentenyl pyrophosphate and IL -2 to induce human γδT cells.The hu-manγδT cells were cultured with quercitrin at various concentrations for 48 hours.CCK-8 kits were used to analyze the in vitro proliferation and cytotoxic activities of γδT cells.Flow cytometry was performed to meas-ure the expression of granzyme B and perforin in γδT cells.The expression of p-ERK, p-Akt and Bcl-2 at protein level were detected by Western blot .Results The percentage of human γδT cells in PBMCs was in-creased from (2.96±1.83)%to (88.94±2.36)%after 10 days of culture.The quercitrin at concentrations of 10 to 80 μg/ml could promote the growth of γδT cells and up-regulate the expression of granzyme B , per-forin, p-ERK, p-Akt and Bcl-2 in a dose dependent manner .The cytolytic activities of γδT cells against co-lonic carcinoma cells ( HCT116 ) were enhanced by quercitrin .Conclusion Quercitrin could promote the proliferation of γδT cells and enhance the expression of granzyme B and perforin at certain concentrations in vitro.ERK1/2 and Akt signal transduction systems might be involved in the process .

Distribution of QPY and RAH haplotypes of granzyme B gene in distinct Brazilian populations / Distribuição dos haplótipos QPY e RAH do gene da granzima B em grupos populacionais brasileiros

INTRODUCTION: The cytolysis mediated by granules is one of the most important effector functions of cytotoxic T lymphocytes and natural killer cells. Recently, three single nucleotide polymorphisms (SNPs) were identified at exons 2, 3, and 5 of the granzyme B gene, resulting in a haplotype in which three amino acids of mature protein Q48P88Y245 are changed to R48A88H245, which leads to loss of cytotoxic activity of the protein. In this study, we evaluated the frequency of these polymorphisms in Brazilian populations. METHODS: We evaluated the frequency of these polymorphisms in Brazilian ethnic groups (white, Afro-Brazilian, and Asian) by sequencing these regions. RESULTS: The allelic and genotypic frequencies of SNP 2364A/G at exon 2 in Afro-Brazilian individuals (42.3% and 17.3%) were significantly higher when compared with those in whites and Asians (p < 0.0001 and p = 0.0007, respectively). The polymorphisms 2933C/G and 4243C/T also were more frequent in Afro-Brazilians but without any significant difference regarding the other groups. The Afro-Brazilian group presented greater diversity of haplotypes, and the RAH haplotype seemed to be more frequent in this group (25%), followed by the whites (20.7%) and by the Asians (11.9%), similar to the frequency presented in the literature. CONCLUSIONS: There is a higher frequency of polymorphisms in Afro-Brazilians, and the RAH haplotype was more frequent in these individuals. We believe that further studies should aim to investigate the correlation of this haplotype with diseases related to immunity mediated by cytotoxic lymphocytes, and if this correlation is confirmed, novel treatment strategies might be elaborated.

INTRODUÇÃO: A citólise mediada por grânulos é uma das mais importantes funções efetoras de linfócitos T citotóxicos e células natural killer. Recentemente, três polimorfismos de nucleotídeo único foram identificados nos éxons 2, 3 e 5 do gene da granzima B, resultando em um haplótipo em que três aminoácidos da proteína madura Q48P88Y245 são alterados para R48A88H245, o que leva à perda da atividade citotóxica da proteína. No presente estudo, avaliamos a frequência desses polimorfismos em populações brasileiras. MÉTODOS:Avaliamos a frequência desses polimorfismos em grupos étnicos brasileiros (brancos, afro-brasileiros e asiáticos) por sequenciamento. RESULTADOS: As frequências alélica e genotípica do polimorfismo 2364A/G no éxon 2 em indivíduos afro-brasileiros (42,3% e 17,3%) foram significativamente maiores (p < 0,0001 e p = 0,0007) quando comparadas a brancos e asiáticos. Os polimorfismos 2933C/G e 4243C/T também foram mais frequentes em afro-brasileiros, mas sem diferença significativa. O grupo afro-brasileiro apresentou maior diversidade de haplótipos e o haplótipo RAH foi mais frequente nesse grupo (25%), seguidos pelos brancos (20,7%) e asiáticos (11,9%), semelhante à frequência apresentada na literatura. CONCLUSÕES: Há uma maior frequência de polimorfismos em afro-brasileiros e o haplótipo RAH foi mais frequente nesses indivíduos. Acreditamos que novos estudos devem ter como objetivo a investigação da correlação deste haplótipo com doenças relacionadas com a imunidade mediada por linfócitos citotóxicos, e se essa correlação for confirmada, novas estratégias de tratamento poderão ser elaboradas.

Granzyme B, B-cell lymphoma/leukemia-2 gene and BH3-interacting domain death agonist in cell apoptais / 国际外科学杂志

Granzyme B is an important effect factor of cytotoxic T lymphocytes in the immune killer function,can quickly induce cell apoptosis of the target cell.It is complicated for the reasons and the ways of the occurrence of the cell apoptosis,many genes are involved in gene regulation of cell apoptosis,and the B-cell lymphoma/leukemia-2 gene family members play a crucial role in the process of cell apoptosis.This paper mainly reviews the correlation of GrB and Bcl-2,Bid with cell apoptosis.

Relapsed extranodal NK/T-cell lymphoma in the mandible, Nasal Type: a case report and review of the literatures

Extranodal NK/T-cell lymphoma (NTCL), nasal type is rare and highly fatal malignant neoplasm. Early diagnosis and establishing treatment plan are very difficult. Furthermore, NTCL in the mandible is an extremely rare condition. The clinical significance of presented case is the very rare location of NTCL. To the best of author's knowledge, this is the first reported case of NTCL of the mandible in the literature.

The variety of the mRNA expression of perforin, granzyme B by expanded NK cells with different cytokines from human peripheral blood / 中华微生物学和免疫学杂志

Objective To study the relationship between the expression of perforin, granzyme B and the cytotoxicity of NK cells, after NK cells were purified and expanded with different cytokines. Meth-otis It was detected that the mRNA expressions of perforin, ganzyme B from 8 donors, and the cytotoxcity of NK cells to target I(562 cells by competitive qualitative RT-PCR. Results The mRNA expressions of perforin, granzyme B by purified and expanded NK cells with different cytokines were markedly enhanced, and IL-2 +IL-15 group, IL-2 + IL-12 + IL-15 group were significantly higher than others. It is consistent that the relationship between the mRNA expressions of NK cells and the cytotoxicity of NK cells to I(562 cells. Conclusion The mRNA expressions of perforin, granzyme B of NK cells with different cytokines were obviously.

n-3 PUFA can inhibit the apoptosis of intestinal epithelial cells of chronic rejection after small intestinal transplantation / 肠外与肠内营养

Objective: The aim of our work was to investigate the effects of n-3 polyunsaturated fatty acids on apoptosis, granzyme B and perforin expression of intestinal epithelial cells of chronic rejection after small intestinal transplantation. Methods: Small bowel transplantation was performed and rats were divided into three groups: Group 1, Lewis-to-Lewis, group 2, F344-to-Lewis, dietary corn oil, Group 3, F344-to-Lewis, dietary fish oil. All recipients were killed after 16 weeks of posttransplantation. The apoptosis rate of mucosal cells were evaluated by flow cytometry. The expressions of granzyme B and perforin were analyzed by reverse transcriptase RT-PCR. Results: A high apoptotic rate was observed when the allografts demonstrated one or more histological features of chronic rejection. N-3 polyunsaturated fatty acids decreased the rate of the apoptosis and inhibitted the expressions of granzyme B and perforin. Conclusion: N-3 polyunsaturated fatty acids can suppress the chronic rejection in small intestinal transplantation.

Research on expression of granzyme B in myocardium with experimental viral myocarditis / 中华微生物学和免疫学杂志

Objective To explore the role of granzyme B( GzmB) in infiltrated myocardial cells of mice with viral myocarditis( VMC) induced by coxsackievirus B3(CVB3) during the injury of myocardium and further to elucidate the pathogenesis of VMC. Methods Thirty-six BALB/c male mice, 5 to 6 weeks old, were randomly divided into two groups: the normal group, 10, and the model group, 26. On the eleventh day all mice were killed and the hearts were taken out for inspection whether there were pathological changes in the myocardium by macroscopy and microscopy. The expression of GzmB was detected in myocardium with immunohistochemistry and hemi-quantitive reverse transcript-polymerase chain reaction ( RT-PCR). Results GzmB positive cells in the mice myocardial cells infiltrated were detected by immunohistochemistry in the model group,and the number of GzmB positive cells is remarkable positive correlation to myocardial pathological score(r =0. 859, P =0.000) , however, no such cells were discovered in the normal murine hearts. Test of RT-PCR showed markedly high expression of GzmB mRNA in mice myocardium of the model group, however, no expression of GzmB mRNA in the normal group. Conclusion The expression of GzmB in the infiltrating myocardiocytes probably plays some role during the injury of myocardium, which mediated cytotoxicity effect is one of important mechanisms of myocardial injury in viral myocarditis. The expression quantity of GzmB in the infiltrating myocardiocytes is significant positive correlation to the severity of myocardial injury.

Analysis of granzyme B mRNA and perforin mRNA levels in urine for renal transplantation patients with de-layed graft function / 中华泌尿外科杂志

Objective To explore the clinical value of the level of granzyme B and perforin mRNA in urine for the diagnosis of renal transplantation patients with delayed graft function (DGF). Methods Twenty-four cases of renal transplantation patients with DGF were included in this study. Seventy-three u-rine specimens were obtained from these patients who received graft biopsies. Among the 24 cases, ureteral obstruction occurred in 2 cases, vascular thrombosis in 1 case, acute CsA intoxication in 3 cases, acute tubu-lar necrosis (ATN) in 7 cases, ATN complicating borderline change in 2 cases, ATN complicating acute re-jection (AR) in 3 cases, AR in 6 cases. Total RNA was isolated from the urinary cells. Messenger RNA (mRNA) encoding the cytotoxic proteins perforin and granzyme B gene were measured with the quantitative polymerase-chain-reaction assay-(RT-PCR). SPSS13.0 software was used for data analysis. Levels of mRNA were log-transformed before analysis. Results The levels of perforin and granzyme B mRNA in u-rine among the ureteral obstruction group, vascular thrombosis group, acute CsA intoxication group and ATN group were very low. There was no significant difference among these groups (P>0.05). However,among the ATN complicating borderline change group 1.22, 0. 97 fg/μg, ATN complicating AR group (1.20±0.39), (1.07±0.30)fg/μg, and AR group(11.13±0. 33), (1.01±0.19)fg/μg, the levels were increased significantly(P<0.001). Conclusion Measurement of mRNA encoding the cytotoxic proteins perforin and granzyme B gene in urinary cells in renal transplantation patients with DGF could be helpful to etiological diagnosis of DGF, and might be used as an index for the appropriate management of the borderline change.

Analysis of perforin and granzyme B gene single nucleotide polymorphisms in Han population of Chongqing / 重庆医科大学学报

T of PRF1 gene was found.There was a strong linkage disequilibrium among the SNPs of GZMB.Conclusion:7 SNPs were detected in PRF1 and GZMB genes all together,two of which were first identified. There is apparent linkage disequilibrium among SNPs of PRF1and GZMB gene in Han population of Chongqing.

Effect of perforin and granzyme B in peripheral blood lymphocytes on diagnosis of acute rejection in renal transplantation / 中国医师进修杂志

Objective To investigate the clinical value of perforin and granzyme B expression in peripheral blood lymphocytes and diagnosis of acute rejection in renal transplantation. Methods Sixty-seven recipients of renal allograft were included in the study. The expression of perforin and granzyme B of peripheral blood lymphocytes was studied by quantitative reverse transcription polymerase chain reaction (RT-PCR). The recipients were divided into four groups, including 7 cases of acute rejection as group 1, 8 cases of delayed graft function as group 2, 27 cases of stable function as group 3, 25 cases of long-term survival as group 4. Results The expression of perforin and granzyme B in group 1 was significantly higher than that of the other three groups (P

Effect of Perforin Antibody on the Expression of Granzyme B in Viral Myocarditis / 实用儿科临床杂志

Objective To explore the effect of perforin(PFP) antibody on the expression of Granzyme B (GzmB) in viral myocarditis.Methods Forty-five 4-week-old BALB/c male mice were randomly divided into 3 groups,including normal control group (n=15),viral control group (n=15) and PFP antibody therapy group (n=15).The mice in normal control group were inoculated with 0.15 mL Eagle reagent,and the mice in viral control group and PFP antibody therapy group were moculated with 0.15 mL TCID501012 L-1 coxsackievirus B3.The mice in PFP antibody therapy group were inoculated with PFP antibody (0.1 mg/kg) on 6 h and 3 d post inoculation.The mice in 3 groups were sacrificed on day 10 post inoculation.Extracted their hearts,then fixed,dehydrated,embeded and sliced the myocardial tissues.The expressions of GzmB proteins in myocardium were determined by immunohistochemistry.The areas of GzmB were measured by the Leica Qwin V3 system.SPSS 11.5 software was used to analyze the data.Results There were 4 mouse dead in viral control group,and 2 mouse dead in PFP autibody therapy group,no mice dead in normal contol group.The expressions of GzmB were not found in myocardial tissues of normal control group,but there were lots of expressions of GzmB in myocardial tissues of viral control group[(67.13?1.82)%].Athough the expressions of GzmB in PFP antibody therapy group[(6.98?1.34)%] were significantly decreased compared with those in myocardial tissues of viral control group(q=66.180 P

Detection of activated cytotoxic cells in lymphoma by tissue microarray / 解放军医学杂志

Objective To detect the expression and distribution of activated cytotoxic cells in types of lymphoma with tissue microarray, and provide evidences for clinical treatment and prognosis. Methods Immunohistochemical staining by S-P technique was used to detect the expression and distribution of perforin and granzyme B in the lymphoma tissue microarray, composed of 60 cases of lymphoma tissue. Ten cases of NK/T-cell lymphoma routine sections were used for relative research, and ten cases of reactive hyperplasia were used for comparison. Results In the tissue microarray, originated from intranode and extranode were 48 cases and 12 cases, respectively; consisting of 42 cases of B-cell lymphoma, 16 cases of T-cell lymphoma, two cases of Hodgkin's disease. 42 cases of B-cell lymphoma cells were negative in perforin and granzyme B. In Ten cases of peripheral T-cell lymphoma, perforin and granzyme B positive were eight cases and nine cases, respectively, but the positive cells were no tumor cells. In 12 cases of NK/T-cell lymphoma (two in the tissue microarray, ten routine sections), both perforin and granzyme B were strongly positive. B-cell lymphoma, T-cell lymphoma and NK/T-cell lymphoma differed significantly (P<0.01). Conclusion Perforin and granzyme B were immunity markers for the identification of activated cytotoxic cells, which also could be used as diagnostic markers of NK/T-cell lymphoma. Their expression in B-cell lymphoma reflected the anti-tumor immunologic reaction of host. Tissue microarray technology has the behavior of high-throughput, convenient, effective, small experiment error, good replication and so on, and can be used as a useful tool for research of lymphoma.