Apoptosis Rate Of Lymphocytes Research Articles

Experimental study of inhibition of ZGDHu-1 on T lymphocytes activation

Objective To investigate the effects of ZGDHu-1 on T lymphocytes activation in vitro to elucidate its immunosuppressive effects. Methods Lymphocytes isolated from healthy persons were stim-ulated with phytohanmagglutinin(PHA) and different experimental groups were set by cocultured for 24 h, 48 h with ZGDHu-1 or with ZGDHu-1 and Cyclosporin A(CSA). To assess the proliferation and apoptosis of T lymphocytes, we detected CD3~+ CD69~+ , CD3~+ CD25~+ , CD4~+ CD25~+ , CD8~+ CD25~+ and CD3~+ Fas~+, CD4~+ Fas~+ , CD8~+ Fas~+ with flow cytometry. The early apoptosis rate of lymphocytes was analyzed with flow cytometry. Culture supernatant IL-2 and TGF-β1 were detected with ELISA. Results ZGDHu-1 decreased PHA activative CD3~+ CD69~+, CD3~+ CD25~+, CD4~+ CD25~+ and CD3~+ Fas~+, CD4~+ Fas~+, Annexin V~+/ PI~- and inhibited IL-2 secretion and promoted TGF-β1 secretion respectively. ZGDHu-1 has synergistic effect with CSA to be more obvious. Conclusion ZGDHu-1 can inhibit T lymphocytes activation and de-creased apeptosis of T lymphocytes. ZGDHu-1 has synergistic effect with CSA to be obvious. Key words: ZGDHu-1; Cyclosporin A; T cells activation; Apoptosis; Flow cytometry

Lymphocyte apoptosis after major abdominal surgery is not influenced by anesthetic technique: a comparative study of general anesthesia versus combined general and epidural analgesia

Study Objective To examine the influence of abdominal colectomy with combined general anesthesia and epidural analgesia versus general anesthesia on apoptosis of circulating lymphocytes. Design Prospective, randomized, clinical comparison study. Setting Tertiary-care general hospital. Patients 40 ASA physical status I and II patients undergoing elective open colectomy for nonmetastatic colon carcinoma. Interventions Patients were randomly allocated to two groups to receiver either general anesthesia alone (Group G) or general anesthesia combined with epidural analgesia (Group C). Group C comprised 21 patients while 19 patients constituted Group G. All patients underwent median longitudinal laparotomy. Measurements Blood samples were collected preoperatively and 24 hours postoperatively for measurement of lymphocyte apoptosis, serum cortisol, C-reactive protein (CRP), and erythrocyte sedimentation rate (ESR). Main Results There were no differences between the two groups in age, weight, or duration of surgery. No significant alterations in total lymphocyte counts, as well as in lymphocyte subpopulations (early apoptotic, late apoptotic, viable, and necrotic), were observed between the general and combined anesthesia groups. Cortisol, ESR, and CRP were significantly increased postoperatively in both groups. Group C presented with lower serum cortisol levels postoperatively than Group G (b = −5.38, CI95%: −8.72 to −2.05, P = 0.002). Conclusions Epidural block could not suppress postoperative lymphocyte apoptosis, increases in cortisol, CRP, or ESR compared with general anesthesia.

Effect of carbachol on the expression of la on monocyte and lymphocyte apoptosis rate of the burned rats with delayed resuscitation

Objective To explore the changing trend of Ia on monocyte, lymphocyte apoptosis rate, TNF-α and IL-6 in abdominal aorta of burned rats with delayed resuscitation and the influence of application of carbachol on them. Methods Adult male Wistar rats were randomly divided into normal control group(n =8), scald group(n =48) and scald with carbachol treatment group(n =48). In latter two groups, rats were inflicted with 30% total body surface area (TBSA) full-thickness scald and delayed fluid resuscitation. All scald rats were sacrificed at the 6th hours or 1st, 2nd, 3rd, 7th, 14th day after scald, with 8 rats at each time point. Expression of Ia antigen on monocyte and lymphocyte apoptosis rate were determined by direct immunofluorescence on a flow cytometer, and TNF-α and IL-6 was measured by ELISA. Results Expression of la on monocyte was obviously lower than that of controls. The lowest levels were recorded on the 6th hours and 1st day after scald. Subsequently, Ia was elevated gradually, but still lower than that of normal rats(P <0. 01). After administration of carbachol, Ia expression was obviously promoted, compared with the simple scald group (P <0. 01). Lymphocyte apoptosis rate, TNF-α and IL-6 was higher than that of controls(P <0. 01). After administration of cavachol, , lymphocyte apoptosis rate and TNF-α and IL-6 was obviously down-regulated on the 6th hours, 1st day, 2nd day and 3rd day after scald injury, compared with the simple scald group (P < 0. 01 or 0. 05). Conclusion After severe burn with delayed fluid resuscitation, there is a low la expression, high lymphocyte apoptosis rate and increased releasing of proinflammatory cytokine. Immune function was suppressed. Carbacho] could improve the immune function of scald rats with delayed fluid resuscitation. Key words: CarbachoL/PD ; Bums/DT/IM

Lack of a time‐dependent effect of melatonin on radiation‐induced apoptosis in cultured rat lymphocytes

Ionizing radiation is widely used for the treatment of solid tumors and it is thought to act by directly targeting tumor clonogens, also known as stem cells. Apoptosis is a genetically programmed mechanism of cell death often characterized by internucleosomal DNA cleavage. Although it has been previously shown that lymphocytes readily undergo apoptosis in patients receiving anticancer drugs or treatment with ionizing radiation, this is the first study to investigate the influence of radiotherapy and melatonin on apoptosis in rat lymphocytes at two different times of the day. Melatonin, a free radical scavenger, is an endogenous neurohormone predominantly synthesized in and secreted by the pineal gland. It has been shown that melatonin inhibits apoptosis in normal cells but it increases the rate of apoptosis in various cancer cells. Therefore, in the present study, the effect of melatonin on apoptosis in cultured lymphocytes was studied after total body irradiation (TBI) was given to rats in the morning (1 HALO) or evening (13 HALO) with morphological and DNA fragmentation analysis. Two-way analysis of variance (ANOVA) revealed that radiation increased the rate of apoptosis in rat lymphocytes after TBI, and melatonin treatment did not reduce the rate of apoptosis after TBI at either time point. We conclude that the lack of an effect of melatonin on the apoptosis rate in rat lymphocytes might be due to the dose-dependent effect of melatonin, the time course of apoptosis investigated, or the cell type in which apoptosis was examined.

Evidence for Increased Apoptosis of Duodenal Intraepithelial Lymphocytes in Cow's Milk Sensitive Enteropathy

Enterocyte apoptosis induced by activated intraepithelial lymphocytes (IELs) is increased in celiac disease (CD). A lesser increment in intraepithelial lymphocytes associated with normal villous structure is also characteristic of cow's milk sensitive enteropathy (CMSE), but no information is available about the apoptosis rate of this condition. Endoscopic biopsy samples of 21 children with untreated CMSE were compared with samples from 15 children with CD and 18 controls. Apoptosis was analyzed using the TUNEL technique and the M30 antibody from duodenal bulb and mid-duodenum samples. Ki-67 was used to detect the proliferation rate and CD3(+) to count the overall number of T lymphocytes. In the mid-duodenum, CMSE patients showed a significantly increased intraepithelial and lamina propria density of TUNEL(+) cells, but no difference in the density of M30(+) epithelial cells was seen compared with controls. In CD subjects, TUNEL(+) counts in mid-duodenal villous epithelium were increased. CD3(+) intraepithelial lymphocytes were increased in both CMSE and CD and correlated with TUNEL(+) and M30(+) counts among all patients. No increase in proliferation rate was seen in CMSE, and no significant correlations between apoptosis and proliferation rates were detected. The observed increase in TUNEL(+) counts and the absence of any increase in M30(+) cells suggest that the apoptosis rate of intraepithelial lymphocytes, but not of enterocytes, is increased in the small IELs in CMSE. However, the number of intraepithelial lymphocytes is still elevated in CMSE, indicating that a disturbance of homeostasis of intraepithelial lymphocytes is important in its pathogenesis.

Increased CD8+ T cell apoptosis in scleroderma is associated with low levels of NF-kappa B.

Our objectives were (1) to compare lymphocyte subpopulation apoptosis rates in SSc patients versus healthy controls and (2) to compare Bcl-2 and NF-kappa B expression in cultured CD8 lymphocytes of SSc patients versus controls. Peripheral blood samples were obtained from 27 SSc patients meeting the American College of Rheumatology criteria for SSc and 28 healthy individuals. Mononuclear cells were isolated by Ficoll-Hypaque density gradient separation and cultured for 48 hr. For determination of apoptosis within specific cell populations, samples were labeled with PE-conjugated monoclonal antibody to CD8, CD4, and a FITC-conjugated monoclonal antibody to Annexin V. Flow cytometry was carried out with a FACS operating with Cellquest software. CD8+ lymphocytes were positively selected with magnetic microbeads conjugated to antihuman CD8. CD8 T cells were separated, then incubated with activation for 48 hr, and NF-kappa B and Bcl-2 analysis was carried out using Western immunoblotting. The CD4:CD8 ratio was increased in SSc compared to controls (2.6 +/- 1.13 vs.1.87 +/- 0.76; P = 0.018). The spontaneous apoptosis rate of SSc CD8 lymphocytes was increased compared to that of controls of (21.9 +/- 13.7 vs. 13.3 +/- 9.9; P = 0.019). No difference was found in the rate of CD4 apoptosis of SSc patients versus controls (9.8 +/- 5.2 vs. 7.18 +/- 4.89%; P = ns). The expression of NF-kappa B in SSc CD8 lymphocytes was decreased compared with that of CD8 lymphocytes from healthy controls (144 +/- 13 vs. 188 +/- 11; P = 0.018). Whereas expression of Bcl-2 was similar in activated CD8+ T cells of SSc patients and healthy controls, CD8+ T cell apoptosis rate was found to be in reverse correlation with expression of NF-kappa B in these cells ( r = - 0.53, P = 0.029). The increased CD4:CD8 ratio in SSC may result from increased CD8+ T cell apoptosis. Increased SSc CD8 T cell apoptosis is associated with low levels of NF-kappa B.

B cell apoptosis accelerates the onset of murine lupus.

To investigate whether the increased rate of lymphocyte apoptosis in systemic lupus erythematosus is involved in the onset of the disease, apoptotic or necrotic T or B lymphocytes from various cell lines were injected intraperitoneally into pre-autoimmune (NZBxNZW)F1 mice (BW) and non-autoimmune BALB/c mice. The intraperitoneal production of cytokines and chemokines, the specific T cell response in the spleen, and the production of anti-histone and anti-dsDNA Ab were investigated. The onset of the disease was characterized by creatinine levels and evaluation of glomerular IgG deposits. In BW, but not in BALB/c mice, injection of apoptotic and not necrotic cells up-regulated IL-6 and IL-10 in resident macrophages. Administration of apoptotic cells augmented the number of Th2 and B lymphocytes recruited in the peritoneal cavity. Only the treatment with apoptotic B cells promoted a systemic Th2 autoimmune response to H2 histones, associated with earlier occurrence of high levels of anti-dsDNA autoantibodies, higher creatinine levels and more numerous glomerular IgG deposits than in BW controls not injected with apoptotic B cells. In genetically susceptible mice exposure to apoptotic of B, but not T, lymphocytes can elicit a Th2 response to H2 histones that helps B cell production of anti-dsDNA Ab and finally triggers the onset of lupus.

Whole body hyperthermia induces apoptosis in subpopulations of blood lymphocytes

Whole Body Hyperthermia (WBH) has been shown to induce alterations of lymphocyte subpopulations in peripheral blood: T-cells decrease and NK-cells increase in number in the course of this therapy. As elevated temperature induces programmed cell death in healthy lymphocytes in vitro, we intended to determine the role of lymphocyte apoptosis in WBH by measuring the rate of apoptosis in blood lymphocytes in the course of this treatment. Blood was taken from cancer patients, treated with whole body hyperthermia and chemotherapy, before, during and the day after treatment. Apoptosis rates of the whole lymphocyte population, as well as, of B-, T-, CD4 + -T-, CD8 + -T-, and Natural-Killer (NK)-cell-subpopulations were determined by staining with AnnexinV-FITC and FACS flow analysis. A significant rise of apoptosis in the whole lymphocyte population, in CD4 + -T- and in CD8 + -T-cells occurred during treatment. In contrast, an elevated rate of apoptosis in NK-cells was observed 20 hours after termination of WBH. These differences were similar when the cells were incubated at 37 degrees C for 24 hours. Our results suggest, that apoptosis is one reason for the previously described decrease of T-cells during WBH and of NK-cells after WBH, and that the hyperthermia-related apoptosis-inducing mechanism is different in T-cells and NK-cells.

Anti-phospholipid antibodies and CD5+ B cells in HIV infection.

This cross-sectional study evaluates the correlation between anti-phospholipid antibodies and CD5+ B cells in 110 patients infected with HIV-1. There were 89.1% of the patients who had IgG antibodies against cardiolipin and phosphatidylserine. The prevalence of IgM and IgA antibodies was < 22%. AIDS was associated with lower frequencies of IgM antibodies against cardiolipin (P = 0.05) and IgG-antibodies against cardiolipin and phosphatidylserine (P = 0.011). Drug users had higher IgM antibodies against phospholipids than patients from other risk groups (P = 0.02). A history of thromboembolic events was not accompanied by higher levels of anti-phospholipid antibodies (P > 0.2). No correlation between anti-phospholipid antibodies and CD5+ B cells was detected. Percentage part of CD5+ B lymphocytes was elevated in all patients and absolute CD4+ T lymphocyte counts and HIV p24 antigen were inversely correlated. In advanced disease a significant reduction of anti-phospholipid antibodies was contrasted with persistent elevation of CD5+ B lymphocytes. These observations may reflect immunological dysfunction involving apoptosis and endothelial damage rather than polyclonal B cell hyperstimulation. A possible explanation would be that in HIV infection an increased rate of spontaneous apoptosis in peripheral blood lymphocytes is accompanied by functional and structural changes of mitochondria. Therefore, structurally altered mitochondrial phospholipids could serve as antigen to induce specific humoral immune responses.

Cell death in HIV pathogenesis and its modulation by retinoids.

Patients infected with the human immunodeficiency virus exhibit a progressive decline in the CD4 T-cell number, resulting in immunodeficiency and increased susceptibility to opportunistic infections and malignancies. Although CD4 T cell production is impaired in patients infected with HIV, there is now increasing evidence that the primary basis of T cell depletion is accelerated apoptosis of CD4 and CD8 T cells. The rate of lymphocyte apoptosis in HIV infection correlates inversely with the progression of the disease: it is low in long-term progressors and in patients undergoing highly active antiretroviral therapy. Interestingly, only a minor fraction of apoptotic lymphocytes are infected by HIV, indicating that the enhanced apoptosis does not necessarily always serve to remove the HIV+ cells and results from mechanisms other than direct infection. Thus, understanding and influencing the mechanisms of HIV-associated lymphocyte apoptosis may lead to new therapies for HIV disease. In this paper the potential effects of retinoids on CD4 T cell apoptosis is discussed.

Apoptosis and the CD95 system in HIV disease: impact of highly active anti-retroviral therapy (HAART)

Apoptosis markers and the rate of lymphocyte apoptosis were followed in the peripheral blood of HIV infected persons at various stages of disease. Our study suggests that the early increase in memory cells following therapy may also be due to a significant decrease in apoptosis in this subset. The intrinsic resistance to apoptosis in the naive subset appears to be maintained following HIV infection and is not modified following highly active anti-retroviral treatment (HAART).

Female cats have lower rates of apoptosis in peripheral blood lymphocytes than male cats: Correlation with estradiol-17β, but not with progesterone blood levels

During earlier study, we quantified by flow cytometry the rate of apoptotic feline lymphocytes after overnight culture. We found evidence that the sex of the animals influences the rate of apoptosis, intact females showed lower rates of apoptosis in lymphocytes cultured overnight than castrated male cats. This observation was also confirmed for cats that were previously experimentally infected with the feline immunodeficiency virus (FIV). In an attempt to find an explanation for these sexually determined differences, plasma estradiol-17β and progesterone levels were measured by radio-immuno assay in the blood of these cats. The hormone levels were analyzed with respect to the rate of lymphocyte apoptosis. As expected, castrated males had lower blood levels of estradiol and progesterone than females. However, no overall correlation was found between hormone blood levels and rate of apoptosis under non-stimulating conditions. Interestingly, the rate of apoptosis found in lymphocytes collected from females and stimulated overnight in phytohaemaglutinin-containing medium, showed a strong negative correlation with the estradiol levels in the blood of these cats. To our knowledge, this is the first confirmation that estradiol in physiological concentrations may protect peripheral lymphocytes from apoptosis after stimulation. No correlation was found in male cats. In conclusion, these observations broaden the list of sexually determined differences of the immune system, sex and sex hormones predispose males and females for certain immune responses and dysfunctions. The present observations have to be taken into account when designing or interpreting experiments on apoptosis and, for example, evaluating the influence of a preexisting FIV infection on the rate of apoptosis. It would be highly advisable to include only spayed cats in studies on the immune system so as to minimize the influence of sex hormones.