Intestinal Lymphocyte Apoptosis Research Articles

Electroacupuncture at "Zusanli"(ST36) protects intestinal mucosal immune barrier by suppre-ssing apoptosis of intestinal lymphocytes and regulating expression of Bcl-2 and Bax in sepsis rats

To observe the effect of electroacupuncture (EA) at "Zusanli" (ST36) on apoptosis of intestinal T lymphocytes, translocation of intestinal bacteria and expression of intestinal Bcl-2 and Bax proteins and intestinal mucosal immune barrier in sepsis rats, so as to explore its underlying mechanism in relieving sepsis. SD rats were randomly divided into sham operation (n=6), model (n=15), non-meridian and non-acupoint (non-acupoint, n=15) and acupoint EA(n=15) groups by using random number table method. The sepsis model was established by using cecal ligation and perforation(CLP) method. EA (2 Hz, 2 mA) was applied to bilateral ST36 or non-acupoint for 30 min one hour after modeling, once every day for 3 days. The rats' general conditions and fatality rate in 3 days after modeling were recorded. The liver, spleen and mesenteric lymph nodes were taken for bacterial culture to detect the translocation rate of intestinal bacteria. The small intestinal tissue was taken for observing histopathological changes (Chiu's score: 0-5 points) after HE staining, and for determining the expression levels of Bcl-2 and Bax proteins using Western blot. The intestinal mucosa was sampled for detecting the apop-tosis (apoptotic index) of lymphocytes by using terminal deoxynucleoitidyl transferase-mediated deoxyuridine triphosphate biotin nick end labeling (TUNEL) assay, and the counts of CD4+ and CD8+T cells using flow cytometry. The contents of IL-4 in the small intestine and that of secretory IgA (sIgA) in the small intestinal mucus were determined by using ELISA. After modeling, of the 15 rats in each of the 3 groups, 7, 7 and 2 in the model, non-acupoint and EA groups were dead in the first 3 days, with the fatality rate being 46.67% (7/15), 46.67% (7/15) and 13.33% (2/15), respectively (being obviously lower in the EA group than in the former two groups, P<0.05). Compared with the sham operation group, the incidence of intestinal bacterial translocation, apoptotic index, Chiu's score, and Bax expression were significantly increased (P<0.05), and the percentages of CD4+ and CD8+T cells, IL-4 and sIgA contents and Bcl-2 expression considerably decreased (P<0.05) in the model group. In comparison with the model group, modeling-induced increase of incidence of bacterial translocation, apoptotic index and Bax expression, and decrease of percentages of CD4+ and CD8+T cells, IL-4 and sIgA contents and Bcl-2 expression were reversed (P<0.05) in the EA group. EA at ST36 can reduce death rate and intestinal bacteria translocation incidence in sepsis rats, which may be related to its functions in regulating the expression of intestinal Bcl-2 and Bax proteins and inhibiting the apoptosis of intestinal mucosal T lymphocytes, thereby protecting the immune barrier function of intestinal mucosa to reduce the intestinal permeability.

Lactoferrin Effect on Lymphocyte Cytokines and Apoptosis Is Independent of Exercise

Strenuous exercise increases apoptosis of intestinal lymphocytes (IL). Bovine lactoferrin (bLf), a protein found in milk products, affects lymphocyte apoptosis and the expression of TH1 and TH2 cytokines. The purpose was to determine whether bLf affects apoptosis and TH1 (tumor necrosis factor alpha) and TH2 (interleukin-10) cytokine expression in IL of mice given strenuous exercise. : Female C57BL/6 mice (n = 89), given three bouts of treadmill exercise, were killed either immediately, 24 h after the last bout, or before initiation of exercise; within exercise conditions, mice were fed control (0% bLf) or bLf-supplemented (2% bLf) diet for 12 d until sacrifice. IL were enumerated, and apoptosis and cytokine expression were determined by Western blot analysis; markers of stress (corticosterone and iso-prostanes) were measured in the plasma by radioimmunoassay and direct immunoassay. Exercise increased IL loss (P < 0.05) and the expression of caspase 3 (P < 0.001), heat shock protein 70 (P < 0.01), and interleukin-10 (P < 0.05) in mouse IL; bLf did not alter these responses. However, bLf reduced tumor necrosis factor alpha expression in mouse IL (P < 0.05), possibly through decreased nuclear factor kappaB expression (P < 0.05) in the supplemented group. Dietary bLf does not affect IL apoptosis after exercise but may confer intestinal protection through changes in cytokine expression, independent of exercise.

Dietary bovine lactoferrin does not alter intestinal lymphocyte apoptosis but decreases TNF‐α production during exercise induced intestinal inflammation

Strenuous exercise can induce physiological alterations (e.g., ischemia‐reperfusion injury, generation of free radicals, increased stress hormones, apoptosis) which increase the likelihood of intestinal inflammation. Bovine lactoferrin (bLf), a common dietary protein which affects cytokine expression and lymphocyte apoptosis, is a potential therapeutic tool against the development of intestinal inflammation. The purpose of this study was to determine the effects of bLf on intestinal lymphocyte (IL) apoptosis and cytokine production following strenuous exercise. C57BL/6 female mice (n=89) were given three repeated bouts of exhaustive exercise, plasma collected and IL isolated. Exercise resulted in IL loss (p&lt;0.05) attributed to increased expression of pro‐apoptotic caspase‐3 (p&lt;0.001) and decreased expression of antiapoptotic Bcl‐2 (p&lt;0.05) in IL. Elevations in corticosterone (p&lt;0.05) and 8‐isoprostane F2α (p&lt;0.05) in plasma and HSP 70 (p&lt;0.01) in IL were also observed following exercise. Dietary bLf did not alter these exercise induced effects but did decrease TNF‐α expression (p&lt;0.05), possibly through reduced levels of NFκB (p&lt;0.05) in IL; there was no change in IL‐10. Thus, dietary bLf does not appear to alter exercise induced IL cell loss and apoptosis but may confer some intestinal protection by decreasing the TNF‐α (inflammatory) cytokine response. Research supported by NSERC of Canada.

Effect of short-term dietary intake of bovine lactoferrin on intestinal lymphocyte apoptosis in healthy mice

Objective The objective of this study was to describe the effects of short-term dietary exposure of bovine lactoferrin (Lf) on intestinal lymphocyte apoptosis and expression of tumor necrosis factor-α (TNF-α) in healthy mice. Methods Female C57BL/6 mice were randomly assigned to three treatment groups: 0% Lf ( n = 16), 0.2% Lf ( n = 16), and 2.0% Lf ( n = 15). Bovine Lf was administered orally, as part of the diet, for 4 consecutive days. Intestinal lymphocytes (ILs) were isolated and analyzed for percentages of CD4, CD8, apoptotic CD4, and apoptotic CD8 cells using flow cytometry. Pro- (caspase-3) and anti- (Bcl-2) apoptotic protein expressions and TNF-α expression in ILs were determined by Western blotting. Results There were significant increases in the percentages of CD4 ( P = 0.02) and apoptotic CD4 ( P = 0.02) ILs in bovine Lf–fed compared with control mice. Percentages of CD8 and apoptotic CD8 cells and expression of caspase-3 and Bcl-2 in ILs did not differ significantly by diet group. In contrast, the expression of TNF-α was significantly lower in Lf-fed versus control mice ( P = 0.01). Conclusion Short-term dietary Lf decreased TNF-α expression in ILs and increased apoptosis of CD4 ILs in healthy mice.

In vivo corticosterone administration at levels occurring with intense exercise does not induce intestinal lymphocyte apoptosis in mice

Intestinal lymphocyte apoptosis can occur following physiological and pathophysiological stress as well as exhaustive exercise. In this study we investigated whether corticosterone (CORT) administration at physiological concentrations observed following strenuous exercise induces intestinal lymphocyte apoptosis and cell loss in mice. CORT injection (14 mg/kg; i.p.) caused a four-fold increase in plasma CORT concentrations, but did not affect intestinal lymphocyte cell loss or alter baseline intestinal lymphocyte apoptosis, as measured by phosphatidylserine externalization, cell viability, mitochondrial membrane depolarization, caspase 3, Bcl-2 and cytosolic cytochrome c protein levels. These findings indicate that CORT at levels observed following strenuous exercise is not involved in intestinal lymphocyte apoptosis and cell loss.

N-Acetyl-L-Cysteine Inhibits Exercise-Induced Lymphocyte Apoptotic Protein Alterations

To investigate the effect of strenuous exercise and antioxidant administration on pro- and antiapoptotic protein expression in intestinal lymphocytes. Female C57BL/6 mice (N = 52) were randomly assigned to receive N-acetyl-L-cysteine (NAC; 1 g.kg(-1)) or saline (SAL) 30 min before treadmill exercise (EX) for 90 min and 2 degrees slope (30 min at 22 m.min(-1), 30 min at 25 m.min(-1), and 30 min at 28 m.min(-1)) and sacrificed immediately (Imm) or 24 h (24 h) after exercise. Control mice were exposed to treadmill noise and vibration without running (nonexercised). Intestinal lymphocytes (IL) were isolated and pro- and antiapoptotic protein expression was evaluated by Western blot analysis. IL protein levels of proapoptotic (caspase 3 and cytosolic cytochrome c) and antiapoptotic (Bcl-2) were significantly different among groups. Relative to nonexercised mice, protein levels of caspase 3 (P < 0.001) and cytosolic cytochrome c (P < 0.005) were significantly elevated, whereas Bcl-2 (P < 0.05) was significantly lower immediately after exercise in mice receiving saline (EX + SAL + Imm) but not in animals receiving NAC (EX + NAC + Imm) or both 24 h postgroups (EX + SAL + 24 h and EX + NAC + 24 h). These results suggest that oxidative stress acting through a mitochondrial pathway may play a role in intestinal lymphocyte apoptosis after strenuous exercise.

N-Acetyl-l-cysteine prevents exercise-induced intestinal lymphocyte apoptosis by maintaining intracellular glutathione levels and reducing mitochondrial membrane depolarization

Intense exercise leads to post-exercise lymphocytopenia and immunosuppression, possibly by triggering lymphocyte apoptosis. To test the role of oxidative stress on exercise-induced lymphocyte apoptosis, we administered the antioxidant N-acetyl-l-cysteine (NAC) and measured apoptosis in intestinal lymphocytes (IL) from exhaustively exercised animals. Eighty-seven female C57BL/6 mice were randomly assigned to receive NAC (1g/kg) or saline 30min prior to treadmill exercise for 90min at 2° slope (30min at 22mmin−1, 30min at 25mmin−1, and 30min at 28mmin−1) and sacrificed immediately (Imm) or 24hours (24h) after cessation of exercise. Control mice (nonexercised) were exposed to treadmill noise and vibration without running. Exercise increased IL phosphatidylserine externalization (p<0.001), mitochondrial membrane depolarization (p<0.05), and decreased intracellular glutathione concentrations (p<0.05) immediately following exercise in saline relative to nonexercised mice. At 24h post-exercise, saline injected mice had fewer total (p<0.001) and CD3+ (p<0.005) IL compared to nonexercised animals. NAC injection in mice maintained intracellular glutathione levels, prevented phosphatidylserine externalization, mitochondrial membrane depolarization, and loss of IL immediately and 24h after exercise. These data suggest that lymphocyte apoptosis precedes post-exercise lymphocytopenia and may be due to oxidative stress.