Neutrophil response to prolonged exercise in immune-competent and RAG2/gamma c null mice.

The two aims of this study were (i) to compare the effects of prolonged exercise on circulating neutrophil number and muscle myeloperoxidase (MPO) activity between RAG2/gamma c null and immune-competent mice, and (ii) to evaluate the general suitability of the lymphocyte-deficient RAG2/gamma c null strain for use in exercise models of immune regulation. RAG2/gamma c null (male and female) and C57BL/6 (congenic immune-competent, male) mice were assigned to either control (C) or treadmill exercise (EX, 22 m/min, 90 min, 6% grade) groups. EX mice were killed immediately (EX0) or 24 h (EX24) after exercise. RAG2/gamma c null males had significantly (P < 0.05) fewer circulating CD45+ cells and higher %CD45+ neutrophils than did C57BL/6 males, independent of exercise. A significant interaction was observed for the effects of exercise and gender on %CD45+ neutrophils in the blood. At EX24, gastrocnemius (Gastroc) MPO significantly increased in EX mice. Gastroc MPO activity was 44% and 35% higher in RAG2/gamma c null vs. C57BL/6 males, and in female vs. male RAG2/gamma c null mice, respectively. Heart MPO activity did not differ between strains or among treatments. We concluded that the Rag2/gamma c null strain is a suitable model for future investigations on immune regulation following acute exercise stress.

The modulation of B16BL6 melanoma metastasis is not directly mediated by cytolytic activity of natural killer cells in alcohol-consuming mice.

BACKGROUND: Previous studies in our laboratory indicate that alcohol consumption suppresses the metastasis of B16BL6 melanoma, whereas the cytolytic activity of natural killer (NK) cells is decreased in female C57BL/6 mice given 20% w/v alcohol in their drinking water. In the present study, we further evaluated the involvement of NK cells and alcohol consumption in the cytolytic activity of NK cells, the surface expression of NK phenotypic markers, and metastasis of B16BL6 melanoma in C57BL/6 beige (bgJ/bgJ) mutant mice, which possess inherently low NK-cell cytolytic activity. METHODS: Beige and control (bgJ/+) mice were given either water or 20% w/v of alcohol in drinking water for 6 1/2 to 7 weeks before assay for cytolytic activity, surface marker expression, and inoculation with B16BL6 melanoma intravenously or into the pinna of the ear. RESULTS: NK cytolytic activity was suppressed in beige mice, and alcohol consumption did not modulate further the cytolytic activity. Beige mice had a lower percentage of NK cells in the peripheral blood and spleen than control mice. Peripheral blood lymphocytes from beige mice also exhibited a reduced percentage of CD4+ T lymphocytes. Alcohol consumption similarly reduced the percentages of NK1.1- and LGL-1-expressing lymphocytes in the peripheral blood and spleen and reduced the percentage of CD8+ T lymphocytes in the peripheral blood in both control and beige mice. Tumor lung colonization was increased in beige mice relative to control mice after intravenous inoculation of B16BL6 melanoma. The increase was more pronounced in water-drinking beige mice than in control mice irrespective of alcohol consumption. Tumor lung colonization was significantly decreased (p < 0.05) by alcohol consumption in one experiment and partially decreased (p = 0.07) in the other. Mice that were inoculated into the pinna of the ear also exhibited a blunted antimetastatic response to alcohol consumption. CONCLUSIONS: These data suggest that the presence of the beige mutation diminishes the antimetastatic effect of alcohol consumption and that there is no interaction between alcohol consumption and NK-cell activity in the modulation of lung metastasis of B16BL6 melanoma cells.

Relationship between moderate intensity endurance training volume and natural killer cell cytolytic activity.

BACKGROUND: This study was designed to examine the relationship between endurance training volume and natural killer cell (NK) cytolytic activity. We hypothesized that a dose dependent relationship exists between forced treadmill training volume and training induced increases in NK cell cytolytic activity. EXPERIMENTAL DESIGN: female, Swiss Webster mice were assigned to treadmill control (TC) or treadmill trained groups (n = 10 per group). Trained mice ran at 12 m per min. (8 degrees grade) for: 15 (EX15), 30 (EX30), or 60 minutes (EX60) per day, five days per week for 11 weeks. Splenic NK cell activity was expressed as median lytic unit (LU), median LU per asialo GM1 (AsGM1+) cell, and median LU per spleen. RESULTS: Median NK activity was not significantly increased by training volume. A trend toward greater median LU per AsGM1+ cell was observed in EX30 group versus TC (p = 0.1). Training volumes less than or greater than this level produced smaller increases in NK cytolytic activity. CONCLUSIONS: These data provide preliminary evidence indicating that training induced increases in splenic NK cell cytolytic activity do not exhibit a dose dependent relationship with treadmill training volume.

Modulation of NK cell cytolytic activity by macrophages in chronically exercise-stressed mice.

This study was designed to investigate the effects of moderate-intensity endurance training on basal natural killer (NK) cell cytolytic activity in murine splenocytes that were enriched for 1) NK1.1+ cells or 2) macrophages and NK1.1+ cells. Mice were assigned to sedentary (Sed), treadmill control (TM), or treadmill-trained (Trn) groups. Splenocyte number, the percentages of NK1.1+, large granular lymphocytes (NK1.1+, LGL-1+), and other subpopulations did not change in Trn mice. Approximately 70% of cells enriched for NK1.1+ expressed this surface antigen. Lytic units (LU) expressed per LGL-1+ cell were significantly lower in Trn [83.9 +/- 3.2 (SE)] compared with Sed (109.5 +/- 7.5) and TM (101.3 +/- 6.4) groups. When macrophages remained in the in vitro assay, LU per LGL-1(+) cell did not differ across groups. The results indicate that highly enriched NK1.1+ cells from Trn mice had lower NK cell activity compared with Sed mice. No differences in NK cell activity were observed when cells were enriched for NK1.1+ cells and macrophages. These findings support the hypothesis that macrophage modulation of NK cells may be one mechanism contributing to augmented basal NK cell activity in endurance-trained individuals.

Ethanol modulates metastatic potential of B16BL6 melanoma and host responses.

The experimental metastatic potential (lung-colonizing ability) of B16BL6 melanoma cells was examined in C57BL/6 mice after exposure to ethanol in vitro and in vivo. In vitro, tumor cells were cultured with ethanol (0.3% v/v), or medium alone, for three passages at 5-day intervals. In vivo, B16BL6 melanoma was exposed to ethanol by administering ethanol (10% or 20% w/v) to mice following subcutaneous inoculation of tumor cells into the dorsal hip. All tumor cells were subsequently inoculated intravenously into the lateral tail vein of water-drinking mice to assess changes in metastatic phenotype. Tumor cells cocultured in vivo with ethanol produced significantly higher numbers of superficial lung colonies, compared with tumor cells cultured in control medium. Experimental metastasis of tumor cells obtained from 20% w/v ethanol-consuming mice was also significantly increased, compared with cells obtained from water-drinking mice. Metastasis of B16BL6 melanoma cells previously obtained from mice consuming 10% w/v ethanol did not differ from controls. In other experiments, water-drinking and ethanol-consuming (2.5%, 10%, and 20% w/v) mice were inoculated subcutaneously into the dorsal hip with B16BL6 melanoma cells, and monitored for tumor growth rate and survival time. In these experiments, survival times were significantly shorter in mice consuming 20% ethanol, compared with all other groups. Subcutaneous tumor growth rate was unaffected by ethanol consumption. Lung metastasis resulting from subcutaneous tumor implantation of B16BL6 melanoma was respectively inhibited, or absent, in 10% and 20% ethanol-consuming groups. Thus, tumor growth rate and incidence of lung metastases were not apparent determinants of decreased survival in 20% ethanol-consuming mice. The results of this study indicate that the experimental metastatic potential of B16BL6 melanoma is increased during exposure to ethanol; however, metastasis from subcutaneous tumor-bearing mice is suppressed. This latter finding is consistent with previous results in which spontaneous metastasis was also suppressed after inoculation of the tumor into the pinna of the ear. Although ethanol increases the ability of B16BL6 melanoma to colonize the lung after intravenous inoculation, this effect is abated in the presence of host factors in ethanol-consuming mice.

VersaClimbing elicits higher VO2max than does treadmill running or rowing ergometry.

Collegiate varsity oarswomen and coxswain (N = 11) completed maximal aerobic exercise tests on a treadmill, a rowing ergometer, and a simulated climbing machine. Successful completion of each test was evidenced by a plateau in oxygen consumption in response to increasing work rates. VO2max (l.min-1), and minute ventilation (VE, l.min-1) at VO2max were significantly greater (P < 0.05) during simulated climbing compared to treadmill running and rowing ergometry. Maximal heart rate (beats.min-1) was significantly greater (P < 0.05) during climbing and running than during rowing. Findings indicate that progressive, incremental, whole-body climbing exercise elicits significantly greater VO2max values for collegiate oarswomen and coxswain than does graded treadmill running or progressive rowing ergometry.

Mechanistic differences in NK cell cytolytic activity in treadmill-trained and chronic ethanol-consuming mice.

The present study was undertaken to investigate mechanisms contributing to differences in natural killer (NK) cell activity in moderately endurance-trained and ethanol-consuming mice. Independent of ethanol exposure, NK cell activity in nylon wool-nonadherent (NWNA) splenocytes is lower in trained than in sedentary control mice (Blank et al. J. Appl. Physiol. 72: 8-14, 1992). Reduced activity may result from a generalized loss of cytolytically active cells, redistribution of NK cells from the spleen to other body compartments, or disruption of paracrine regulation of NK cells after removal of nylon wool-adherent cells. To examine these possibilities, NK cell cytolytic activity was determined in nonenriched splenocytes from treadmill-trained and ethanol-consuming mice. Lymphocyte subpopulations in nonenriched splenocytes and NWNA splenocytes were also compared. Peripheral blood lymphocyte subpopulations were determined to examine combined effects of training and ethanol intake on regional distribution of lymphocytes in blood and spleen. NK cell activity in nonenriched splenocytes from trained water-drinking mice was not reduced compared with that in sedentary mice; rather, cytolytic activity was moderately enhanced (17% increase in lytic units, P < 0.05). Training did not change percentages of T-cells, B-cells, and NK [NK1.1+ and large granular lymphocytes (LGL-1+)] cells or the LGL/NK ratio in the spleen and blood. NK cell cytolytic activity was significantly reduced in nonenriched splenocytes from ethanol-consuming mice, independent of training. These findings support the hypothesis that moderate-intensity endurance training influences splenic NK cell function by modulating paracrine regulation of NK cells.(ABSTRACT TRUNCATED AT 250 WORDS)

Factors involved in modulation of NK cell activity by ethanol consumption.

Natural killer (NK) cells are large granular lymphocytes (LGL) whose functional characteristics play an important immunological role in tumor surveillance, bacterial and viral infections, baematopoiesis, and tissue transplantation. NK cell cytotoxicity is mediated through non-MHC-restricted lysis and by antibody-dependent cellular lysis against NK-sensitive and insensitive targets. Non-MHC restricted lysis of targets can occur when NK cells are in a non-activated state (inherent NK cell cytolytic activity) and following activation by interleukin 2 (IL-2).

Plasma corticosterone response to chronic ethanol consumption and exercise stress.

Acute exposure to ethanol induces a stress response in mice that is manifested by increased plasma corticosterone (PC) concentration. However, during chronic intake of 7.5% w/v ethanol, diurnal fluctuation of PC is dampened. Whether chronic consumption of 20% w/v ethanol alters normal diurnal fluctuation of plasma glucocorticoids is not known. Investigating the PC response in 20% w/v ethanol-consuming mice is of interest because glucocorticoids are known suppressants of natural killer (NK) cell activity and increased concentration or altered diurnal fluctuation of PC may have a modulatory role on NK cells in these mice. Mice given 20% w/v ethanol for at least 7 days and for as long as 10 weeks have suppressed splenic NK cell cytolytic activity. Thus, the purpose of this study was to examine whether mice given 20% w/v ethanol exhibited normal concentrations and diurnal variation of PC. To further define the glucocorticoid response in chronic ethanol-consuming mice, PC concentration was evaluated in response to a secondary stress of physical exercise. After 1 week, ethanol-consuming mice exhibited abnormal diurnal PC periodicity that was progressively dampened during the remaining 9 weeks. Acute physical exercise during Week 1 induced a 2-fold increase in PC concentration compared with pre-exercise values, a response that was independent of ethanol intake. After 6 and 10 weeks, the postexercise PC concentration was attenuated in ethanol-consuming compared with water-drinking mice. It was concluded that suppressed NK cell activity typically observed with this model of chronic ethanol intake is not directly associated with dampened diurnal fluctuation in PC.

Ethanol-induced changes in peripheral blood and splenic natural killer cells.

Previous work demonstrated that splenic natural killer (NK) cell cytolytic activity was suppressed in ethanol-consuming mice within 1 week and for as long as 10 weeks concurrent with ethanol intake. However, it is unknown if suppression of NK cell activity with ethanol consumption results from regional losses in NK cells. The present experiments were designed to examine the effects of ethanol on the percentage and total number of NK cells in the spleen and blood. Data indicate that, after 4 weeks of ethanol intake, NK cell activity of peripheral blood lymphocytes was also suppressed (50% of controls). With short-term (2-week) exposure, the percentage of NK cells in the blood and the spleen remained relatively constant. In the spleen, changes in the number of NK cells reflected generalized fluctuations in lymphocyte numbers rather than variation in the percentage of NK cells. NK cells were most sensitive to the effects of ethanol during long-term (8-week) ethanol intake, as demonstrated by significant reductions in the total percentage of NK1.1+ cells in the blood, and the LGL-1+ subset in the blood and spleen. These results provide insight into the mechanism of NK modulation by ethanol. Because NK cells were not selectively depleted after 2-4 weeks of ethanol intake, loss of cytolytically active NK cells cannot explain the suppression of in vitro cytolytic activity of cells from ethanol-consuming mice. However, with prolonged ethanol intake (8 weeks), the depletion of NK cells, specifically the LGL-1+ subset, may contribute to the overall suppression of NK cell cytolytic function.

Exercise ventilatory response to upright and aero-posture cycling.

The aero-cycling posture has become increasingly popular among cyclists. Because of the potential for altered lung mechanics in the aero-cycling posture, the ventilatory and gas exchange profiles of 10 moderately trained males (31.1 +/- 6.3 yr, mean +/- SD) exercising on a cycle ergometer in the upright posture (UC) and aero-cycling (AC) posture were studied. Exercise consisted of 3-min work stages with 50-W incremental changes until volitional exhaustion. Ventilatory and gas exchange responses to exercise were averaged each minute. Maximal oxygen consumption (54.3 +/- 6.3 vs 53.4 +/- 6.9 ml.kg-1.min-1) in the UC and AC were not significantly different. Maximal values for ventilation (132.5 +/- 32.3 vs 128.0 +/- 28.7 l.min-1), tidal volume (2.7 +/- 0.5 vs 2.6 +/- 0.3 1.br-1), breathing frequency (50.0 +/- 8.0 vs 47.0 +/- 5.0 br.min-1), and submaximal ventilatory and heart rate responses in the UC were not significantly different compared with the AC. No significant difference existed between postures for mean inspiratory flow and inspiratory duty cycle at comparable submaximal power outputs. These results indicate that at absolute power outputs, ventilatory responses, as determined by respiratory pattern and timing, and metabolic cost, were similar in the UC and AC. Thus, the AC does not impair physiologic responses to high intensity exercise.

Alcohol consumption suppresses metastasis of B16-BL6 melanoma in mice.

Female C57BL/6 mice were fed a defined, pelleted diet and given 10% w/v or 20% w/v ethanol in their drinking water. Natural killer (NK) cell cytolytic activity was compared between water-drinking and ethanol-consuming mice and in mice that were also treated with polyinosinic-polycytidylic acid (poly I:C) to augment NK cell activity or with anti-NK1.1 antibody to decrease activity. NK cell cytolytic activity was not altered in mice given 10% ethanol, but was decreased in mice given 20% ethanol compared to water-drinking mice. Poly I:C treatment increased and anti-NK1.1 antibody treatment decreased NK cell activity in both water-drinking and 20% ethanol-consuming mice. Experimental and spontaneous metastases of B16-BL6 melanoma were evaluated as a function of the duration of ethanol consumption before tumor inoculation and as a function of altered NK cell activity. Experimental metastasis was inhibited after 4 and also after 6.5 weeks of ethanol exposure. Poly I:C treatment inhibited tumor lung colonization irrespective of ethanol consumption. Anti-NK1.1 antibody treatment increased metastasis, although to a lesser degree in mice consuming 10% ethanol. Spontaneous metastasis was inhibited in mice consuming 10% ethanol for 4 weeks, and in mice consuming 20% ethanol for 1 and 4 weeks before melanoma inoculation.

Modulation of NK cell activity by moderate intensity endurance training and chronic ethanol consumption.

Chronic ethanol consumption can suppress natural killer (NK) cell activity. Exercise after ethanol administration may enhance blood ethanol clearance, which may benefit the immune response. This study examined the effects of moderate intensity endurance training and chronic ethanol consumption (20% wt/vol) on splenic NK cell activity. Mice were assigned to one of four groups: sedentary, water drinking (SED-H2O); sedentary, ethanol consuming (SED-EtOH); trained, water drinking (TR-H2O), and trained, ethanol consuming (TR-EtOH). TR groups ran 60 min/day, 5 days/wk, at 12 m/min for 10 wk. Mice were killed 48 h after exercise. Baseline NK cell activity was suppressed 30% in TR and EtOH groups compared with SED-H2O controls. Activation with recombinant human interleukin-2 increased cytolytic activity in all groups four- to fivefold. These results indicate that training did not abrogate the effects of chronic ethanol consumption on NK cell activity. Furthermore, moderate endurance training may contribute to suppressed nylon wool-enriched NK cell activity in murine splenocytes for as long as 48 h after exercise.

Suppression of natural killer cell activity by ethanol consumption and food restriction.

Effects of 4-week food restriction and ethanol consumption on natural killer (NK) cell activity and carcass composition were evaluated. Female, C57BL/6 mice given water (H2O) or ethanol (20% w/v, ETOH) ad libitum were placed in one of three dietary groups: unrestricted (UNR), moderately restricted (MR, 2.2 g/day), or severely restricted (SR, 1.8 g/day). Food restriction alone (MR, SR) significantly reduced body, spleen, and thymus weights; carcass lipid content (SR only); spleen cell number; and baseline and interleukin-2 (rIL-2) stimulated NK cell activities. Ethanol consumption was unaffected by food restriction and in restricted mice it did not suppress food intake. Thus, average calories derived from ethanol increased from 30% (UNR) to 40% (SR) with the degree of food restriction in these groups. Mice given ethanol and restricted food intake had at least as heavy or heavier body, spleen, and thymus weights than water-drinking (H2O) counterparts. Spleen cell number was reduced in ethanol-consuming (ETOH), food restricted groups compared with UNR H2O control. Baseline NK cell activity was suppressed 50% to 90% in all ETOH and food-restricted groups. rIL-2 stimulated NK cell activity was suppressed 18% to 76% in food restricted mice independent of ethanol intake. These results indicate that supplementary ethanol calories did not enhance NK cell activity in UNR ETOH mice, nor did they protect splenic NK cell activity from the suppressant effects of food restriction. Ethanol consumption significantly increased carcass lipid content in all groups compared with their H2O counterparts. This increase was largely responsible for the preservation of body weight in ETOH mice especially during food restriction.

Influence of ethanol consumption on natural killer cell activity in mice.

Alcohol is a known suppressant to the immune system, and alcoholics frequently have impaired humoral and cell-mediated immunity. Several studies indicate that alcohol modulates natural killer (NK) cell activity. NK cells provide important defense against certain infectious diseases, spontaneously arising tumors and, in particular, to blood-borne metastasizing tumor cells. Evaluation of the effects of alcohol on NK cells is complicated by many factors including: the level and duration of alcohol abuse, polydrug use, the subject's age, and nutritional and health status. This study examined the effects of 1 and 2 weeks of alcohol consumption on baseline and interleukin 2 (IL-2) stimulated murine NK cell activity. Well nourished female C57BL/6 mice were given continuous access to 20% w/v ethanol as the sole fluid source and consumed about 40% of their total caloric intake as ethanol. Splenic baseline and IL-2 stimulated NK cell activity were significantly lower in ethanol-consuming groups compared to control groups after the 1- and 2-week test periods. The average daily intake of ethanol, blood alcohol concentration, and the percentage of ethanol-derived calories were not associated with the decreased NK cell activity of the experimental animals; nor did any other measured parameter appear to serve as an indicator of ethanol modulation of splenic NK cell activity. Whether this immunosuppression results from the "direct" modulation of ethanol or from indirect factors is presently unknown.

Experimental allergic encephalomyelitis in the rat: response to encephalitogenic proteins and peptides.

Lewis rats were used to determine the encephalitogenic activity of myelin basic protein of different species and of 45-residue fragments of basic protein. Basic protein from guinea pigs was more active than that from rats, and the fragments from the two species showed the same order of activity. Bovine basic protein was the least active of the intact proteins, and the respective fragment was inactive. Studies of serum-binding capacity did not support the hypothesis that blocking antibody played a role in this biological variation, whereas consideration of the amino acid sequences of the three fragments suggested that differences in primary structure, operating either at the sensitization or the effector phase of the immune response, could account for the variation.