Immune responses in adult female volunteers during the bed-rest model of spaceflight: antibodies and cytokines.

BACKGROUND: It is unknown whether a prolonged period of bed rest will affect human immune responses, particularly in female subjects. OBJECTIVE: We sought to measure immune responses in adult female subjects exposed to prolonged bed rest. METHODS: Adult (25-40 years) female volunteers (n = 24) were maintained in a supine (6 degrees tilt) head-down bed-rest (HDBR) position for 60 days: 8 with HDBR only, 8 with HDBR and regular muscular exercise, and 8 with HDBR and dietary protein supplementation. Subjects were immunized with bacteriophage phiX-174. Antibody production and plasma cytokine levels were determined. RESULTS: The rate of primary antibody production of the HDBR plus exercise group increased faster (P = .01) and to a higher level versus that of the HDBR-only group (P = .03) and that of the HDBR plus diet group (trend P = .08). The rates of secondary antibody production between the 3 groups were similar, but the level of antibody in the HDBR plus exercise group remained higher versus that in the HDBR-only group (P = .03). Both the HDBR (P = .001) and HDBR plus diet (P = .02) groups had time-related progressive increases in TNF-alpha receptor levels, but the HDBR plus exercise group remained at baseline. The HDBR plus exercise group experienced an acute increase in IL-1 receptor antagonist levels versus the HDBR (P = .02) and the HDBR plus diet (P = .02) groups, with similar increases in RANTES levels. CONCLUSIONS: The exercise countermeasure accelerated primary antibody production and increased antibody levels to bacteriophage phiX-174 and also opposed the potentially harmful effects of increased TNF-alpha levels caused by prolonged bed rest, possibly by activating the anti-inflammatory cytokine IL-1 receptor antagonist and the chemotactic factor RANTES.

Effects of exposure of mice to hindlimb unloading on leukocyte subsets and sympathetic nervous system activity.

The hindlimb unloading (HU) rodent model was developed to simulate some of the aspects of spaceflight conditions. Our previous studies showed that exposure to HU for 48 h (h) followed by bacterial challenge, reduces the ability of mice to resist infection. The purpose of this study was to investigate the physiological changes in mice during the 48 h of exposure to HU to understand the mechanisms involved in the increased susceptibility to infection observed in mice subjected to these conditions. Female Swiss Webster mice were hindlimb-unloaded during 48 h. Blood samples, spleen and peritoneal cells were removed before and after 18 or 48 h of HU-exposure. Leukocyte subset analysis was performed in spleen and peritoneal cells by flow cytometry, and catecholamine levels were measured in plasma and whole spleen by a catecholamine enzyme immunoassay. Catecholamine levels measured in plasma and spleen were significantly greater in mice exposed to HU compared to control. This increase coincided with significant reductions in spleen size in the HU group. Flow cytometric analyses showed a significant reduction of splenic CD19 + B-cells and NK1.1+ cells in mice exposed to HU with a concomitant increase in T-cells. These results suggest that exposure to HU increases the activity of the sympathetic nervous system (SNS) and induces lymphocyte sub-population changes that may contribute to the deregulation of immunity seen in mice exposed to HU and, more importantly may predispose the otherwise healthy host to the subsequent reduced ability to resist infections.

Antibodies in cold stressed mice recognize a surface protein in Toxoplasma gondii tachyzoites.

Physical or psychological stressors are known to have significant consequences for immune function and the outcome of disease in human and animal models. In mice, cold water stress (CWS) has been shown to delay control of acute infection and reactivation of latent infections. Increased levels of parasite-specific IgG and IgM antibodies are observed when CWS is applied in the chronic phase. The present study examined the effects of a physical stressor, CWS, on tachyzoites antigens of Toxoplasma gondii, with particular emphasis on a low molecular weight antigen, 5 kDa, which seems to be recognized by antibodies from mice subjected to CWS in the chronic phase. This antigen is not recognized by antibodies from infected mice not subjected to CWS. Sera obtained from stressed and infected (CWS + INF) mice subjected to CWS during the chronic phase (CWS + INF + CWS) were used to harvest anti-5-kDa antibodies for immunolocalization studies. Tachyzoite lysate preparations were electrophoretically separated and transferred to nitrocellulose membranes. Strips of nitrocellulose containing tachyzoite antigens in the 4-10-kDa range were used to select for anti-5-kDa antibodies. Harvested anti-5-kDa localized this antigen on the surface of tachyzoites. This antigen was not present in bradyzoite preparations. Treatment with phosphatidylinositol-specific phospholipase C showed this antigen was not anchored to the cell membrane through glycosyl-phosphatidylinositol. Strong antibody responses in stressed animals during the chronic phase are associated with parasite reactivation. The 5-kDa antigen constitutes a unique immunogenic component of T. gondii, with significant diagnostic potential for identifying reactivation of latent infections.

Active hexose correlated compound activates immune function to decrease bacterial load in a murine model of intramuscular infection.

BACKGROUND: Infection is a serious, costly, and common complication of surgery and constitutes the principal cause of late death in patients undergoing surgery. The objective of this study was to clarify the mechanisms by which active hexose correlated compound (AHCC) increases survival in a murine model of intramuscular infection. METHODS: Food-deprived mice receiving either AHCC or excipient were infected with bacteria. Kinetics of bacterial load, white blood cell counts, cytokine levels, and antibody levels were compared between groups. RESULTS: AHCC-treated mice had reduced bacterial load at day 5 and cleared bacteria entirely at day 6. Levels of interleukin-12, tumor necrosis factor-alpha, and interleukin-6 peaked earlier in this group (day 3) compared with controls (day 5). Increased percentages of peripheral lymphocytes and monocytes and decreased numbers of polymorphonuclear cells were detected in the AHCC group. CONCLUSIONS: AHCC appears to induce an early activation of the immune response, leading to an effective clearance of bacteria and rapid recovery.

Kinetics of systemic cytokine and brain chemokine gene expression in murine toxoplasma infection.

Toxoplasma gondii often migrates to the central nervous system in immunocompromised patients, where it induces a severe inflammation referred to as Toxoplasma encephalitis. The mechanisms involved in control of parasite multiplication and prevention of Toxoplasma encephalitis remain unclear. The objective of the present study was to characterize the inflammatory response in the brains of mice during acute T. gondii infection, with emphasis on the expression of chemokine receptors. Susceptible C57BL/6 mice were orally infected with 10 cysts of the low-virulent ME49 strain of T. gondii. Levels of cytokines (TNF-alpha, IFN-gamma, IL-10, IL-6, and IL-12p70) and chemokines (CCL/2MCP-1) were measured in plasma at 5, 10, 15, 20, and 30 days after infection. In addition, the mRNA expression of chemokines (CCL5/RANTES, CCL2/MCP-1, CCL4/MIP-1beta) and chemokine receptors (CCR1, CCR2, CCR5, CCR7, CCR8, CXCR4, and CXR5) were measured in brain tissues at the same time points. Plasma levels of IFN-gamma and CCL2/MCP-1 were highly expressed at day 5, whereas TNF-alpha had a moderate increase at day 5, peaked at day 10, and returned to normal levels by day 30. Plasma levels of IL-10, IL-6, and IL-12p70 were not detected throughout the study. Analyses of mRNA expression of chemokines and chemokine receptors in the brain showed that CCL5/ RANTES, CCR7, CXCR4, and CXCR5 were upregulated, peaking after 10 days of T. gondii infection. IgM-specific antibody levels increased at day 5 and peaked at days 10 and 30, whereas IgG levels increased at day 10 and continued to increase thereafter, reaching maximum levels at day 30 postinfection (PI). Our results suggest that T. gondii infection is controlled at local and systemic levels, and that proinflammatory proteins and their receptors may be acting coordinately to induce stage conversion and prevent parasite multiplication and development of Toxoplasma encephalitis. The early production of IFN-gamma and the delayed expression of CXCR4 and CXCR5 indicate that T. gondii induces an early robust cellular immune response, followed by a strong and sustained antibody-mediated immunity.

Enhancement of in vitro growth of pathogenic bacteria by norepinephrine: importance of inoculum density and role of transferrin.

Norepinephrine is a stress hormone that enhances bacterial growth. We examined the effects of a small inoculum on the norepinephrine-induced growth of species previously reported to be unaffected by norepinephrine. The results indicated that a reduced inoculum density is essential for observing norepinephrine-induced effects. Additional studies using serum-free media suggested that transferrin plays a role in norepinephrine-induced growth.

Active hexose correlated compound (AHCC) enhances resistance to infection in a mouse model of surgical wound infection.

BACKGROUND: Infection is the most common postoperative complication within the surgical wound and during severe trauma. In spite of the use of modern sterile techniques and prophylaxis, infection continues to be a leading cause of death in these patients. Therefore, it has become crucial to develop new alternatives to prevent the effects of trauma and other complications on the immune system and improve resistance to infection. The objective of this study was to test the prophylactic effects of oral administration of active hexose correlated compound (AHCC), a natural immunoenhancer, on survival in a mouse model of surgical soft tissue infection. METHODS: The model involves the intramuscular administration of a 50% lethal dose (LD50) of K. pneumoniae to mice that have restricted food intake for 24 hours prior to and six hours after infection and simulates local infection and food deprivation that often occur during trauma or surgical procedures. In the present study, AHCC was administrated orally to Swiss Webster mice for eight days prior to and during the infection period. Survival, time of death, LD50, and clearance of bacteria of this group were compared with those control mice receiving the excipient alone. RESULTS: Survival and mean time to death were increased significantly in the AHCC-treated group; the LD50 was greater in mice receiving AHCC than in mice receiving the excipient. Mice receiving AHCC were better able to clear bacteria from their systems than were control animals. CONCLUSIONS: The results suggest that AHCC protects mice in this model by restoring the immune and other systems negatively affected by trauma, infection, and food deprivation. More studies are necessary to determine the intrinsic mechanisms involved in this model and whether AHCC can prevent infection or improve survival in human beings with severe trauma or undergoing surgical procedures.

Effects of space flight conditions on the function of the immune system and catecholamine production simulated in a rodent model of hindlimb unloading.

UNLABELLED: The rodent model of hindlimb unloading has been successfully used to simulate some of the effects of space flight conditions. Previous studies have indicated that mice exposed to hindlimb-unloading conditions have decreased resistance to infections compared to restrained and normally housed control mice. OBJECTIVE: The purpose of this study was to clarify the mechanisms involved in resistance to infection in this model by examining the effects of hindlimb unloading on the function of the immune system and its impact on the production of catecholamines. METHODS: Female Swiss Webster mice were hindlimb-unloaded during 48 h and the function of the immune system was assessed in spleen and peritoneal cells immediately after this period. In addition, the kinetics of catecholamine production was measured throughout the hindlimb-unloading period. RESULTS: The function of the immune system was significantly suppressed in the hindlimb-unloaded group compared to restrained and normally housed control mice. Levels of catecholamines were increased in the hindlimb-unloaded group and peaked at 12 h following the commencement of unloading. CONCLUSION: These results suggest that physiological responses of mice are altered early after hindlimb unloading and that catecholamines may play a critical role in the modulation of the immune system. These changes may affect the ability of mice to resist infections.

Active hexose correlated compound enhances the immune function of mice in the hindlimb-unloading model of spaceflight conditions.

Hindlimb unloading is a ground-based model that simulates some of the aspects of spaceflight conditions, including lack of load bearing on hindlimbs and a fluid shift to the head. It has been shown that treatment with active hexose correlated compound (AHCC) restores resistance to infection in mice maintained under hindlimb-unloading conditions. The present study was designed to clarify the mechanisms by which AHCC enhances resistance to infection in this model. We hypothesized that oral administration of AHCC will enhance the function of the immune system, which could lead to the increased resistance to infection observed in this model. AHCC or the excipient was orally administered to mice, and the function of the immune system was assessed in spleen and peritoneal cells isolated from those groups. The results of the present study showed that administration of AHCC for 1 wk before and throughout the second day of the hindlimb-unloading period enhanced the function of the immune system assessed by spleen cell proliferation and cytokine production in spleens and nitric oxide and cytokine production in peritoneal cells. These findings suggest that AHCC can be used as a potent immunoenhancer, especially in cases in which the immune system is suppressed by any condition, including diseases such as human immunodeficiency virus infection and cancer.

Effects of cold stress on spleen cell proliferation and cytokine production during chronic Toxoplasma gondii infection.

BACKGROUND: Cell-mediated immunity is critical for controlling infection and preventing reactivation during the chronic phase of Toxoplasma gondii infection. In people suffering from AIDS, T. gondii is one of the major opportunistic infectious agents. Mechanisms regulating rapid development of clinical signs in previously asymptomatic patients remain unclear; however, cofactors such as stress are suspected to play a role in the susceptibility to opportunistic infections. OBJECTIVE: This study examined the role of cold stress (CS) in splenocyte function during chronic T. gondii infection. METHODS: Control mice and mice previously infected orally with T. gondii were subjected to CS during the chronic phase (CSchr), i.e. 90 days after infection, and in vitro cell proliferation and cytokine production were measured before (day 0) and 1, 15 and 25 days after CSchr. Splenocyte proliferation and cytokine production were measured after in vitro stimulation with concanavalin A (Con-A), anti-CD3 antibody (A-CD3) and Toxoplasma lysate antigen. RESULTS: CSchr enhanced splenocyte proliferation in cells stimulated with Con-A and A-CD3, but it suppressed proliferation in cells stimulated with T. gondii antigens. Increased levels of interferon (IFN)-gamma were detected independent of the type of stimulation after CSchr and remained high throughout the experiment. CS had similar results in noninfected animals. CONCLUSION: Although an overall increase in splenocyte function occurred after nonspecific stimulation, CS suppressed primed spleen cells from responding to T. gondii antigens which could lead to reactivation of latent infection. The increase in IFN-gamma after CSchr could be a result of spleen cells being primed by released parasites by this stressor. IFN-gamma is critical in the control of parasite reactivation.

Catecholamines and in vitro growth of pathogenic bacteria: enhancement of growth varies greatly among bacterial species.

The purpose of this study was to examine the effects of catecholamines on in vitro growth of a range of bacterial species, including anaerobes. Bacteria tested included: Porphyromonas gingivalis, Bacteriodes fragilis, Shigella boydii, Shigella sonnie, Enterobacter Sp, and Salmonella choleraesuis. The results of the current study indicated that supplementation of bacterial cultures in minimal medium with norepinephrine or epinephrine did not result in increased growth of bacteria. Positive controls involving treatment of Escherichia coli with catecholamines did result in increased growth of that bacterial species. The results of the present study extend previous observations that showed differential capability of catecholamines to enhance bacterial growth in vitro.

Active hexose correlated compound enhances resistance to Klebsiella pneumoniae infection in mice in the hindlimb-unloading model of spaceflight conditions.

Previous studies have demonstrated that resistance to infection is decreased in Swiss Webster female mice maintained in the hindlimb-unloading model (Aviles H, Belay T, Fountain K, Vance M, and Sonnenfeld G. J Appl Physiol 95: 73-80, 2003; Belay T, Aviles H, Vance M, Fountain K, and Sonnenfeld G. J Allergy Clin Immunol 110: 262-268, 2002). This is a model of some of the aspects of spaceflight conditions, including lack of load bearing on hindlimbs and a fluid shift to the head. Active hexose correlated compound (AHCC), extracted from Basidiomycete mushrooms, has been shown to induce enhancement of immune responses, including enhanced natural killer activity. In the present study, AHCC was orally administered to mice to determine whether the treatment could decrease immunosuppression and mortality of mice maintained in the hindlimb-unloaded model and infected with Klebsiella pneumoniae. The results of the present study showed that administration of AHCC by gavage for 1 wk (1 g/kg body wt) before suspension and throughout the 10-day suspension period yielded significant beneficial effects for the hindlimb-unloaded group, including 1). decreased mortality, 2). increased time to death, and 3). increased ability to clear bacteria. The results suggest that AHCC can decrease the deleterious effects of the hindlimb-unloading model on immunity and resistance to infection.

Increased susceptibility to Pseudomonas aeruginosa infection under hindlimb-unloading conditions.

It has been reported that spaceflight conditions alter the immune system and resistance to infection [Belay T, Aviles H, Vance M, Fountain K, and Sonnenfeld G. J Allergy Clin Immunol 170: 262-268, 2002; Hankins WR and Ziegelschmid JF. In: Biomedical Results of Apollo. Washington, DC: NASA, 1975, p. 43-81. (NASA Spec. Rep. SP-368)]. Ground-based models, including the hindlimb-unloading model, have become important tools for increasing understanding of how spaceflight conditions can influence physiology. The objective of the present study was to determine the effect of hindlimb unloading on the susceptibility of mice to Pseudomonas aeruginosa infection. Hindlimb-unloaded and control mice were subcutaneously infected with 1 LD50 of P. aeruginosa. Survival, bacterial organ load, and antibody and corticosterone levels were compared among the groups. Hindlimb unloading had detrimental effects for infected mice. Animals in the hindlimb-unloaded group, compared with controls, 1). showed significantly increased mortality and reduced time to death, 2). had increased levels of corticosterone, and 3). were much less able to clear bacteria from the organs. These results suggest that hindlimb unloading may induce the production of corticosterone, which may play a critical role in the modulation of the immune system leading to increased susceptibility to P. aeruginosa infection.

Effects of the hindlimb-unloading model of spaceflight conditions on resistance of mice to infection with Klebsiella pneumoniae.

BACKGROUND: It has been well documented in several studies that many immunologic parameters are altered in experimental animals and human subjects who have flown in space. However, it is not fully known whether these immunologic changes could result in increased susceptibility to infection. Hindlimb (antiorthostatic) unloading of rodents has been used successfully to simulate some of the effects of spaceflight on physiologic systems. OBJECTIVE: The objective of this study was to determine the effect of hindlimb unloading on the outcome of Klebsiella pneumoniae infection in mice. METHODS: Hindlimb-unloaded, hindlimb-restrained, and control mice were intraperitoneally infected with one 50% lethal dose of K pneumoniae 2 days after suspension. Mortality and bacterial load in several organs were compared among the groups. RESULTS: Unloaded mice showed significantly increased mortality and reduced mean time to death compared with that seen in the control groups. Kinetics of bacterial growth with smaller infective doses revealed that control mice were able to clear bacteria from the organs after 30 hours. In contrast, unloaded mice had continued bacterial growth at the same time point. CONCLUSION: The results of this study suggest that hindlimb unloading might enhance the dissemination of K pneumoniae, leading to increased mortality. The complex physiologic changes observed during hindlimb unloading, including stress, have a key role in the pathophysiology of this infection.

Stress, suspension and resistance to infection.

Immune function is altered in stressful situations, including space flight. This may result in increased risk of infection. Antiorthostatic suspension has been used to study the effects of space flight-like conditions on immunity. The mechanisms of promoting infection in stressful situations have not been defined, but catecholamines could play a role. In the present study gram negative bacteria grown with catecholamines showed enhanced bacterial growth compared to controls. Additionally, antiorthostatically suspended mice infected with Klebsiella pneumoniae showed decreased survival compared to restrained or normally caged controls. Therefore, stress-induced enhanced bacterial growth and immunosuppression could play a role in suspension-induced enhanced mortality due to infection.

Identificación de kinetoplastideos aislados en el Ecuador mediante el uso de la técnica de Reacción en Cadena de Polimerasa (PCR)

La amplificación del DNA del kinetoplasto, cada vez tiene mayor aplicación en la detección Leishmania y diagnóstico de leishmaniasis. En el presente estudio se emplea un par de sondas: KIN 1 y KIN 2, que se amplificaron en todas las especies de Leishmania, tanto las cepas de referencia como las aisladas a partir de lesiones en humanos, en quienes se observó un segmento de 470 pares de bases (pb), mientras que en Trypanosoma cruzi, un segmento de 520 pb. Es importante recalcar la necesidad de validar la utilidad de esta prueba en el diagnóstico clínico, epidemiológico y entomológico de esta enfermedad.