Cytotoxic CD4⁺ T cells (CD4⁺ CTLs) represent a novel subset of T cells with cytotoxic effects. They recognize target cells in an antigen-specific manner, relying on class II major histocompatibility complex (MHC-II) interactions. CD4⁺ CTLs exert cytotoxic effects on target cells by secreting cytotoxic molecules such as granzymes, perforin, and granulysin. Recent studies have revealed their significant roles in various autoimmune diseases. This review focuses on the differentiation, phenotypic characteristics, and roles of CD4⁺ CTLs in different types of autoimmune disorders, aiming to provide new insights for the prevention and treatment of these diseases.
Humans ; Autoimmune Diseases/immunology* ; CD4-Positive T-Lymphocytes/immunology* ; Animals ; T-Lymphocytes, Cytotoxic/immunology* ; Perforin/immunology* ; Granzymes/immunology*

Objectives To investigate interleukin 36γ (IL-36γ) expression, and analyze the influence of IL-36γ to CD8⁺ T cell activity in chronic obstructive pulmonary diseases (COPD) patients with secondary fungal pneumonia. Methods Peripheral blood was collected from 47 COPD patients, 39 COPD patients with secondary fungal pneumonia, and 20 controls. Bronchial alveolar lavage fluid (BALF) was isolated from 27 COPD patients with secondary fungal pneumonia. CD8⁺ T cells were purified. The levels of four IL-36 isoforms in plasma and BALF were measured by enzyme linked immunosorbent assay (ELISA). CD8⁺ T cells were stimulated with recombinant human IL-36γ. The levels of interferon γ(IFN-γ), tumor necrosis factor α(TNF-α), perforin and granzyme B in the cultured supernatants were measured by ELISA. Recombinant human IL-36γ-stimulated CD8⁺ T cells were co-cultured with NCI-H1882 cells in either direct cell-to-cell contact or Transwell^TM manner. The levels of IFN-γ, TNF-α, and lactate dehydrogenase in the cultured supernatants were assessed. The percentage of target cell death was calculated. Results Plasma IL-36α, IL-36β, and IL-36γ levels were significantly elevated in both COPD group and COPD with secondary fungal pneumonia group compared with those in control group. However, only plasma IL-36γ level was higher in COPD with secondary fungal pneumonia group than that in COPD group [(200.11±99.95)pg/mL vs (53.03±87.18)pg/mL, P=0.023]. There was no remarkable difference in plasma IL-36 receptor antagonist level among three groups. IL-36γ level in BALF from infectious site was higher than that from non-infectious site in COPD with secondary fungal pneumonia group [(305.82±59.60)pg/mL vs (251.93±76.01)pg/mL, P=0.011]. IL-36γ stimulation enhanced IFN-γ, TNF-α, perforin and granzyme B secreted by CD8⁺ T cells. When IL-36γ-stimulated CD8⁺ T cells were directly mixed with NCI-H1882 cells for co-culture, the percentage of cell death was increased [(16.06±3.67)% vs (11.47±2.36)%, P=0.002]. When using Transwell^TM plate for non-contact co-culture, IL-36γ-stimulated CD8⁺ T cell-mediated death of NCI-H1882 cells showed no significant difference compared to that without stimulation [(4.77±0.78)% vs (4.99±0.92)%, P=0.554]. Conclusion IL-36γ level in plasma and infectious site is elevated in COPD patients with secondary fungal pneumonia, which enhances the cytotoxicity of CD8⁺ T cells in peripheral blood and infectious microenviroment.
Humans ; Pulmonary Disease, Chronic Obstructive/complications* ; CD8-Positive T-Lymphocytes/metabolism* ; Male ; Female ; Aged ; Middle Aged ; Interferon-gamma/metabolism* ; Interleukin-1/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Lung Diseases, Fungal/complications* ; Bronchoalveolar Lavage Fluid/chemistry* ; Perforin/metabolism* ; Pneumonia/immunology* ; Granzymes/metabolism*

Objective To investigate the differences of type 2 innate lymphocytes (ILC2) and interlukin 9 (IL-9) between chronic lymphocytic leukemia (CLL) patients and healthy controls, and to understand the effects of ILC2 on the function of regulatory T cells (Tregs), CD8⁺ T cells and CLL cells through IL-9. Methods Flow cytometry was used to detect the levels of ILC2 and Tregs in the peripheral blood of 45 newly diagnosed CLL patients and 24 healthy controls, and the expressions of granzyme B and perforin in CD8⁺ T cells in the peripheral blood of 28 patients and 15 healthy controls; ELISA was used to detect the level of IL-9 in the serum. ILC2 of patients and healthy controls was sorted by immunomagnetic beads and cultured separately, and the level of IL-9 in the culture supernatant was measured by ELISA. ILC2 sorted from CLL patients and healthy control-derived peripheral blood mononuclear cells(PBMCs) were co-cultured with the B cell leukemia MEC-1 cells, one group was supplemented with IL-9 antibody and the other group was not. After 72 hours of culture, the ratio of Tregs, programmed death 1 (PD-1), T cell immunoglobulin and immunoreceptor tyrosine-based inhibitory motif domain (TIGIT), cytotoxic T lymphocyte antigen 4 (CTLA-4) on Tregs, granzyme B and perforin in CD8⁺ T cells were measured by flow cytometry, IL-9 level of the culture supernatant was measured by ELISA, the apoptosis of MEC-1 cells was measured by Annexin V-PI. Results Compared with the healthy control group, the levels of ILC2, Tregs and IL-9 in the CLL group increased significantly. The levels of granzyme B and perforin in CD8⁺ T cells were positively correlated in the peripheral blood of CLL patients. Compared with the healthy control group, IL-9 levels in the supernatant of sorted ILC2 from CLL patients increased. In the anti-IL9 antibody group, the level of PD-1 and TIGIT on Tregs decreased, and the level of granzyme B in CD8⁺ T cells increased significantly. The level of IL-9 in the anti-IL9 antibody group decreased statistically. And MEC-1 cells showed increased early apoptotic rate in the anti-IL9 antibody group statistically. Conclusion In CLL, ILC2 affects CD8⁺ T cells and Tregs through IL-9, which weakens the anti-tumor effect of CD8⁺ T cells, enhances the immunosuppressive effect of Tregs, and plays a role in the occurrence and development of CLL disease.
Humans ; Leukemia, Lymphocytic, Chronic, B-Cell/immunology* ; CD8-Positive T-Lymphocytes/immunology* ; T-Lymphocytes, Regulatory/immunology* ; Middle Aged ; Male ; Female ; Interleukin-9/blood* ; Aged ; Granzymes/metabolism* ; Perforin/metabolism* ; Immunity, Innate ; Adult ; Lymphocytes/immunology*

No abstract available.
Asian Continental Ancestry Group/*genetics ; Base Sequence ; Bone Marrow Cells/cytology/pathology ; Cytomegalovirus Infections/diagnosis ; Epstein-Barr Virus Infections/diagnosis ; Female ; Flow Cytometry ; Heterozygote ; Humans ; Infant ; Killer Cells, Natural/cytology/immunology ; Lymphohistiocytosis, Hemophagocytic/*diagnosis/genetics ; Perforin/*genetics ; Phagocytosis ; Polymorphism, Single Nucleotide ; Republic of Korea ; Sequence Analysis, DNA

PURPOSE: Cutaneous lymphocyte-associated antigen (CLA)-expressing CD8+T cells have been known to play an important role in the pathogenesis of atopic dermatitis (AD). However, the mechanisms underlying the loss of self-tolerance remain unclear. Regulatory T cells (Tregs) play a key role in the development of homeostasis in the immune system. We, therefore, hypothesized that a reduced ability of Tregs to inhibit autologous CD8+CLA+T cells might be underlying mechanism in AD. MATERIALS AND METHODS: CD8+CLA+T cells and Tregs were obtained from the peripheral blood of AD patients and control volunteers. The frequencies of CD8+CLA+T cells were evaluated. The proliferative responses of CD8+CLA+T cells were assessed by flow cytometry, and the levels of transforming growth factor-beta1 (TGF-beta1) and interleukin-10 (IL-10) in culture supernatants were detected by enzyme-linked immunosorbent assay. RESULTS: Our results revealed higher frequency and increased expression of perforin and granzyme-B in peripheral CD8+CLA+T cells in AD, and lower inhibitory ability of Tregs on proliferation of CD8+CLA+T cells in AD. Meanwhile, the levels of TGF-beta1 produced by Tregs were significantly lower in AD, and anti-TGF-beta1 abolished such suppression. CONCLUSION: The attenuated inhibitory ability of Tregs on hyper-activated autologous CD8+CLA+T cells, mediated by TGF-beta1, plays an important role in the pathogenesis of AD.
Adult ; Aged ; CD8-Positive T-Lymphocytes/drug effects/*immunology ; Case-Control Studies ; Cell Proliferation ; Cell Separation ; Dermatitis, Atopic/*immunology/pathology ; Female ; Granzymes/metabolism ; Humans ; Interleukin-10/metabolism ; Lymphocyte Count ; Male ; Perforin/metabolism ; Skin/*immunology/pathology ; T-Lymphocytes, Cytotoxic/drug effects/immunology ; T-Lymphocytes, Regulatory/drug effects/*immunology ; Transforming Growth Factor beta1/pharmacology

OBJECTIVE: To explore the expression of perforin and granzyme-B in peripheral blood lymphocyte (PBL) in patients with prostate cancer (PCa) and the clinical significance. METHODS: The expressions of perforin and granzyme-B in PBL were detected by fluorescence quantitative reverse transcription polymerase chain reaction. The results of perforin and granzyme-B expression were compared among patients with PCa (n=60), patients with BPH (benign prostatic hyperplasia, n=40) and healthy controls (n=20). RESULTS: Th e expressions of perforin and granzyme-B in patients with PCa were significantly lower than that in patients with BPH or that in the healthy controls (P<0.05), respectively. Furthermore, in PCa patients with low pathological grade, the expressions of perforin and granzyme-B in PBL was statistically higher than that in patients with high pathological grade (P<0.05). The expressions of perforin and granzyme-B in PCa patients at high clinical stage was statistically lower than that in PCa patients at low clinical stage (P<0.05). CONCLUSION The results of this study suggest that development and progression of PCa might be associated with poor immune status of patients.
Case-Control Studies ; Granzymes ; metabolism ; Humans ; Lymphocytes ; enzymology ; Male ; Perforin ; metabolism ; Prostatic Hyperplasia ; Prostatic Neoplasms ; immunology

This study was aimed to investigate the expression level of perforin in cord blood NK cells and the relation of perforin expression after IL-2, IL-15 stimulation to cytotoxicity of NK cells. NK cells were isolated from cord blood MNC by depleting CD3(+) cells and then enriching CD56(+) cells using immunomagnetic separation (CD3 and CD56 cell isolation kit, autoMACS, miltenyi). The purity was analysed by flow cytometry. According to the different combination of cytokines, there were two groups: group A (IL-2) and group B (IL-2 + IL-15). The cytotoxicity and perforin expression rate of fresh and different cultured CB-NK cells against K562/Jurkat cell lines were estimated by LDH release test (cytotoxic 96 non-radioactive cytotoxicity assay). The results showed that the purity of NK cells after separation was more than 90%. The cytotoxicity towards both tumor lines in group B was higher than that in group A (p < 0.05), and cytotoxicity in group A was higher than that of fresh NK cells (p < 0.05). Perforin expression rate of group A (84.55%) was higher than that of fresh NK cells (67.21%) (p < 0.05), and there was no significant difference between group A and B (84.55% versus 87.22%) Cytotoxic activity of CB-NK cells was positively correlated with perforin expression rate (r = 0.886, p < 0.05). It is concluded that IL-2 can enhance cytotoxicity of CB/BM-NK cells by increasing the perforin expression.
CD56 Antigen ; metabolism ; Cells, Cultured ; Cytotoxicity, Immunologic ; Fetal Blood ; cytology ; Flow Cytometry ; Humans ; Interleukin-15 ; pharmacology ; Interleukin-2 ; pharmacology ; K562 Cells ; Killer Cells, Natural ; cytology ; immunology ; metabolism ; Perforin ; metabolism

BACKGROUND AND OBJECTIVEThe actual evaluation of immunological function is significant for studing the tumor development and devising a treatment in time. The aim of this study is to evaluate the immunological function of advanced lung cancer patients systematically, and to discuss the clinical significance.

METHODSThe nucleated cell amounts of advanced lung cancer patients and the healthy individuals were counted. The immune cell subsets and the levels of IL-4, INF-gamma, perforin and granzyme in CD8+T cells by the flow cytometry were measured. The proliferation activity and the inhibition ratio of immune cells to several tumor cell lines were evaluated by MTT assay.

RESULTSThe absolute amounts and subsets of T, B, NK cells of advanced lung cancer patients were lower than the healthy individuals (P < 0.05); However, the proportion of regulatory T cells of advanced lung cancer patients (4.00 +/- 1.84)% was lower than the healthy individuals (1.27 +/- 0.78)% (P < 0.05). The positive rates of IFN-gamma perforin, granzyme in CD8+T cells decreased while them in IL-4 did not in the advanced lung cancer patients compared to the healthy control group (P < 0.05). The proliferation activity of immune cells, the positive rate of PPD masculine and the inhibition ratio to tumor cells in the advanced lung cancer patients was lower than the healthy subsets obviously (P < 0.05).

CONCLUSIONThere was a significant immune depression in the advanced lung cancer patients compared to the healthy individuals.

Adult ; Aged ; CD8-Positive T-Lymphocytes ; immunology ; metabolism ; Cell Line, Tumor ; Female ; Flow Cytometry ; Granzymes ; metabolism ; Humans ; Interferon-gamma ; metabolism ; Interleukin-4 ; metabolism ; Lung Neoplasms ; immunology ; pathology ; Male ; Middle Aged ; Perforin ; metabolism

OBJECTIVETo investigate the expressions of perforin (PF), granzyme B (GrB), granulysin (GNLY), TNF-alpha and IFN-gamma in peripheral CD8+ T lymphocytes and their correlation to infection status in patients with chronic hepatitis B (CHB).

METHODSALT, AST, TB and HBV DNA copy were detected to evaluate the infection status in CHB patients, with healthy volunteers serving as the control group. According to the infection status, the CHB patients were divided into 4 groups, namely normal hepatic function and high HBV DNA level group, normal hepatic function and low HBV DNA level group, abnormal hepatic function and high HBV DNA level group and abnormal hepatic function and low HBV DNA level group. The expressions of some immune effector molecules in CD8+T cells were detected by flow cytometry, and the correlations between these immune effector molecules and the infection status were analyzed.

RESULTSThe expressions of GrB, TNF-alpha and IFN-gamma in normal hepatic function and low HBV DNA level group were significantly higher than those in abnormal hepatic function and high HBV DNA level group (P<0.05). The expression of IFN-gamma in normal hepatic function and high HBV DNA level group was significantly higher than that in abnormal hepatic function and high HBV DNA level group (P<0.05). The expressions of PF and GNLY were similar among all the 4 groups. Positive correlations were noted between GrB, PF, GNLY, TNF-alpha and IFN-gamma.

CONCLUSIONGrB, TNF-alpha and IFN-gamma in peripheral CD8+ T cells are inversely correlated to hepatic dysfunction and HBV DNA level in CHB patients.

Adult ; CD8-Positive T-Lymphocytes ; immunology ; metabolism ; Case-Control Studies ; DNA, Viral ; blood ; Female ; Granzymes ; blood ; Hepatitis B, Chronic ; blood ; immunology ; Humans ; Interferon-gamma ; blood ; Liver ; physiopathology ; virology ; Male ; Middle Aged ; Perforin ; blood ; Tumor Necrosis Factor-alpha ; blood ; Young Adult

OBJECTIVETo investigate the expression profile of immune effector molecules in peripheral natural killer cells (NK) in patients with chronic hepatitis virus B.

METHODSAccording to the infection status, patients were divided into four experiment groups: normal hepatic function and high HBV DNA level group, normal hepatic function and low HBV DNA level group, abnormal hepatic function and high HBV DNA level group and abnormal hepatic function and low HBV DNA level group. The expression of perforin (PF), granzyme B (Gr B), granulysin (GNLY), tumor necrosis factor alpha (TNFa) and interferon gamma (IFNr) in NK cells were detected by flow cytometer.

RESULTSCompared with control group (31.50%+/-27.64%), the expression of GNLY was significantly increased in normal hepatic function and high HBV DNA level group (59.74%+/-30.82%) and normal hepatic function and low HBV DNA level group (61.89%+/-33.30%); the expression of IFNr in normal hepatic function and high HBV DNA level group (39.89%+/-21.30%) and abnormal hepatic function and high HBV DNA level group (37.54%+/-18.79%) was lower than that in normal control group (57.38%+/-23.69%); the expression of PF, GrB, GNLY in abnormal hepatic function and high HBV DNA level group (35.47%+/-29.64%, 66.55%+/-22.92%, 42.03%+/-33.17%) was lower than that in normal hepatic function and high HBV DNA level group (56.98%+/-38.34%, 81.53%+/-19.58%, 59.74%+/-30.82%) and normal hepatic function and low HBV DNA level groups (62.95%+/-31.98%, 84.51%+/-14.57%, 61.89%+/-33.3%); there were positive correlations between ef PF, Gr B, GNLY, TNFa, and IFNr.

CONCLUSIONThe expression of IFNr in NK cells from patients with high HBV DNA replication level is lower than that in normal control group; the expression of PF, Gr B and GNLY in NK cells from patients with normal hepatic function is higher than that in NK cells from patients with abnormal hepatic function.

Adolescent ; Adult ; Antigens, Differentiation, T-Lymphocyte ; metabolism ; Case-Control Studies ; Cytokines ; metabolism ; DNA, Viral ; blood ; Female ; Flow Cytometry ; Gene Expression Profiling ; Granzymes ; metabolism ; Hepatitis B virus ; genetics ; immunology ; Hepatitis B, Chronic ; immunology ; pathology ; Humans ; Killer Cells, Natural ; immunology ; metabolism ; Liver Function Tests ; Male ; Middle Aged ; Perforin ; metabolism ; Virus Replication ; Young Adult