Treatment with Interleukin-7 Restores Host Defense against Pneumocystis in CD4+ T-Lymphocyte-Depleted Mice.

Pneumocystis pneumonia (PCP) is a major cause of morbidity and mortality in patients with HIV infection. CD4(+) T lymphocytes are critical for host defense against this infection, but in the absence of CD4(+) T lymphocytes, CD8(+) T lymphocytes may provide limited host defense. The cytokine interleukin-7 (IL-7) functions to enhance lymphocyte proliferation, survival, and recruitment of immune cells to sites of infection. However, there is little known about the role of IL-7 in PCP or its potential use as an immunotherapeutic agent. We hypothesized that treatment with recombinant human IL-7 (rhIL-7) would augment host defense against Pneumocystis and accelerate pathogen clearance in CD4-depleted mice. Control and CD4-depleted mice were infected with Pneumocystis, and rhIL-7 was administered via intraperitoneal injection. Our studies indicate that endogenous murine IL-7 is part of the normal host response to Pneumocystis murina and that administration of rhIL-7 markedly enhanced clearance of Pneumocystis in CD4-depleted mice. Additionally, we observed increased recruitment of CD8(+) T lymphocytes to the lungs and decreased apoptosis of pulmonary CD8(+) T lymphocytes in rhIL-7-treated animals compared to those in untreated mice. The antiapoptotic effect of rhIL-7 was associated with increased levels of Bcl-2 protein in T lymphocytes. rhIL-7 immunotherapy in CD4-depleted mice also increased the number of gamma interferon (IFN-γ)-positive CD8(+) central memory T lymphocytes in the lungs. We conclude that rhIL-7 has a potent therapeutic effect in the treatment of murine Pneumocystis pneumonia in CD4-depleted mice. This therapeutic effect is mediated through enhanced recruitment of CD8(+) T cells and decreased apoptosis of lung T lymphocytes, with a preferential action on central memory CD8(+) T lymphocytes.

Chemokine receptor 5 and its ligands in the immune response to murine tuberculosis.

SETTING: The ability of chemokines such as macrophage inflammatory protein (MIP)-1alpha, MIP-1beta, and regulated-upon-activation, normal T cell expressed and secreted (RANTES), to attract and activate T cells and monocytes, the building blocks of the granuloma, suggests that these chemokines may have a role in modulating immune responses to Mycobacterium tuberculosis infection. OBJECTIVE: We hypothesized that the chemokine receptor 5 (CCR5) ligands, MIP-1alpha, MIP-1beta and RANTES, are virulence correlates in M. tuberculosis infection and are indispensable to granuloma formation. DESIGN: The ability of virulent (H37Rv) and avirulent (H37Ra) strains of M. tuberculosis to induce chemokine production in vivo and in vitro was determined at protein and mRNA levels. We also compared bacterial burden, and granuloma numbers and size in H37Rv-infected CCR5-/- or wild-type C57BL/6 mice. RESULTS: In vivo, lung mRNA and protein measurements of MIP-1alpha, MIP-1beta and RANTES indicate significantly higher (p<0.05) values (days 14-28) in the H37Rv-infected than the H37Ra-infected mice. This is consistent with a higher infection burden of the virulent strain. However, in vitro alveolar macrophage stimulation by H37Rv or H37Ra yielded no significant differences in production of the three chemokines at all time points. Histological analysis of granulomas did not show any significant differences in granuloma numbers, size and M. tuberculosis growth in CCR5-/- compared to wild-type mice. CONCLUSIONS: The production of the CCR5 ligands, MIP-1alpha, MIP-1beta, and RANTES, does not clearly correlate with virulence of M. tuberculosis. These ligands and their receptors may not be indispensable to the development of granulomas in murine tuberculosis.

Acute alcohol intoxication during hemorrhagic shock: impact on host defense from infection.

BACKGROUND: Acute alcohol intoxication is a frequent underlying condition associated with traumatic injury. Our studies have demonstrated that acute alcohol intoxication significantly impairs the immediate hemodynamic, metabolic, and inflammatory responses to hemorrhagic shock. This study investigated whether acute alcohol intoxication during hemorrhagic shock would alter the outcome from an infectious challenge during the initial 24 hr recovery period. METHODS: Chronically catheterized male Sprague Dawley rats were randomized to acute alcohol intoxication (EtOH; 1.75 g/kg bolus followed by a constant 15 hr infusion at 250-300 mg/kg/hr) or isocaloric isovolemic dextrose infusion (dex; 3 ml + 0.375 ml/hr). EtOH and dex were assigned to either fixed-volume (50%) hemorrhagic shock followed by fluid resuscitation with Ringer's lactate (EtOH/hem, dex/hem) or sham hemorrhagic shock (EtOH/sham, dex/sham). Indexes of circulating neutrophil function (apoptosis, phagocytosis, oxidative burst) were obtained at baseline, at completion of hemorrhagic shock, and at the end of fluid resuscitation. Bacterial clearance, lung cytokine expression, and myeloperoxidase activity were determined at 6 and 18 hr after an intratracheal challenge with Klebsiella pneumoniae (10 colony-forming units). RESULTS: Mean arterial blood pressure was significantly lower in acute alcohol intoxication-hemorrhagic shock animals throughout the hemorrhagic shock. In sham animals, acute alcohol intoxication alone did not produce significant changes in neutrophil apoptosis or phagocytic activity but significantly suppressed phorbol myristic acid (PMA)-stimulated oxidative burst. Hemorrhagic shock produced a modest increase in neutrophil apoptosis and suppression of neutrophil phagocytic capacity but significantly suppressed PMA-stimulated oxidative burst. Acute alcohol intoxication exacerbated the hemorrhagic shock-induced neutrophil apoptosis and the hemorrhagic shock-induced suppression of phagocytosis without further affecting PMA-stimulated oxidative burst. Fluid resuscitation did not restore neutrophil phagocytosis or oxidative burst. Acute alcohol intoxication decreased (-40%) 3-day survival from K. pneumoniae in hemorrhagic shock animals, impaired bacterial clearance during the first 18 hr postinfection, and prolonged lung proinflammatory cytokine expression. CONCLUSIONS: These results demonstrate that the early alterations in metabolic and inflammatory responses to hemorrhagic shock produced by acute alcohol intoxication are associated with neutrophil dysfunction and impaired host response to a secondary infectious challenge leading to increased morbidity and mortality.

CD4+ T cell-independent vaccination against Pneumocystis carinii in mice.

Host defenses are profoundly compromised in HIV-infected hosts due to progressive depletion of CD4+ T lymphocytes. Moreover, deficient CD4+ T lymphocytes impair vaccination approaches to prevent opportunistic infection. Therefore, we investigated a CD4+ T cell-independent vaccine approach to a prototypic AIDS-defining infection, Pneumocystis carinii (PC) pneumonia. Here, we demonstrate that bone marrow-derived dendritic cells (DCs) expressing the murine CD40 ligand, when pulsed ex vivo by PC antigen, elicited significant titers of anti-PC IgG in CD4-deficient mice. Vaccinated animals demonstrated significant protection from PC infection, and this protection was the result of an effective humoral response, since adoptive transfer of CD4-depleted splenocytes or serum conferred this protection to CD4-deficient mice. Western blot analysis of PC antigen revealed that DC-vaccinated, CD4-deficient mice predominantly reacted to a 55-kDa PC antigen. These studies show promise for advances in CD4-independent vaccination against HIV-related pathogens.

Interleukin-17 and lung host defense against Klebsiella pneumoniae infection.

Bacterial pneumonia remains an important cause of morbidity and mortality worldwide, especially in immune-compromised patients. Cytokines and chemokines are critical molecules expressed in response to invading pathogens and are necessary for normal lung bacterial host defenses. Here we show that interleukin (IL)-17, a novel cytokine produced largely by CD4+ T cells, is produced in a compartmentalized fashion in the lung after challenge with Klebsiella pneumoniae. Moreover, overexpression of IL-17 in the pulmonary compartment using a recombinant adenovirus encoding murine IL-17 (AdIL-17) resulted in the local induction of tumor necrosis factor-alpha, IL-1beta, macrophage inflammatory protein-2, and granulocyte colony-stimulating factor (G-CSF); augmented polymorphonuclear leukocyte recruitment; and enhanced bacterial clearance and survival after challenge with K. pneumoniae. However, simultaneous treatment with AdIL-17 provided no survival benefit after intranasal K. pneumoniae challenge. These data show that IL-17 may have a role in priming for enhanced chemokine and G-CSF production in the context of lung infection and that optimally timed gene therapy with IL-17 may augment host defense against bacterial pneumonia.

Requirement of interleukin 17 receptor signaling for lung CXC chemokine and granulocyte colony-stimulating factor expression, neutrophil recruitment, and host defense.

Bacterial pneumonia is an increasing complication of HIV infection and inversely correlates with the CD4(+) lymphocyte count. Interleukin (IL)-17 is a cytokine produced principally by CD4(+) T cells, which induces granulopoiesis via granulocyte colony-stimulating factor (G-CSF) production and induces CXC chemokines. We hypothesized that IL-17 receptor (IL-17R) signaling is critical for G-CSF and CXC chemokine production and lung host defenses. To test this, we used a model of Klebsiella pneumoniae lung infection in mice genetically deficient in IL-17R or in mice overexpressing a soluble IL-17R. IL-17R-deficient mice were exquisitely sensitive to intranasal K. pneumoniae with 100% mortality after 48 h compared with only 40% mortality in controls. IL-17R knockout (KO) mice displayed a significant delay in neutrophil recruitment into the alveolar space, and had greater dissemination of K. pneumoniae compared with control mice. This defect was associated with a significant reduction in steady-state levels of G-CSF and macrophage inflammatory protein (MIP)-2 mRNA and protein in the lung in response to the K. pneumoniae challenge in IL-17R KO mice. Thus, IL-17R signaling is critical for optimal production of G-CSF and MIP-2 and local control of pulmonary K. pneumoniae infection. These data support impaired IL-17R signaling as a potential mechanism by which deficiency of CD4 lymphocytes predisposes to bacterial pneumonia.

Pulmonary hypertension associated with HIV infection.

Pulmonary hypertension occurs with increased frequency among patients with human immunodeficiency virus (HIV) infection. Although the pathogenesis of HIV-associated pulmonary hypertension remains unknown, it appears to occur independently of other risk factors associated with pulmonary vasculopathy, such as chronic hepatitis C infection and intravenous drug use. Signs and symptoms are typical of those immunocompetent patients with primary pulmonary hypertension, but because many HIV-infected patients are receiving intensive medical supervision, the diagnosis of pulmonary hypertension is often made at an earlier stage. Acute responses to epoprostenol are similar to those among non-HIV-infected individuals, but the benefits of long-term, intravenous treatment with epoprostenol in HIV-infected patients is unknown. Future investigations should define the true incidence of pulmonary hypertension and the long-term effects of epoprostenol on survival among HIV-infected individuals. Physicians should be alert to possible pulmonary hypertension in persons infected with HIV.

Effect of alcohol consumption on host release of interleukin-17 during pulmonary infection with Klebsiella pneumoniae.

BACKGROUND: A link between alcohol abuse and bacterial pneumonia has been recognized for centuries, but mechanisms to explain this relationship are unclarified. Interleukin-17 (IL-17) is a lymphocyte-derived cytokine that is part of the inflammatory cytokine cascade. Previous studies from our laboratory indicated that IL-17 is released in lung tissue in a murine model of bacterial pneumonia caused by Klebsiella pneumoniae. The effects of alcohol consumption on pulmonary release of IL-17 are unknown. METHODS: Mice were maintained on 20% ethanol in drinking water or on a control diet without alcohol. After 2 weeks, alcohol and control mice were challenged with intratracheal K. pneumoniae. Mice were followed for survival after bacterial challenge, neutrophil recruitment was assayed as myeloperoxidase, and IL-17 was measured in lung lavage fluid by enzyme-linked immunosorbent assay. In additional experiments, splenocytes from control mice were incubated with ethanol in vitro, and release of IL-17 was measured in culture supernatants. Finally, control and alcohol mice received intrapulmonary gene transfer of E-1-deleted adenovirus containing the murine IL-17 gene. These mice were then challenged with K. pneumoniae and followed for survival and neutrophil recruitment. RESULTS: In these studies, we demonstrate that a 2-week history of ethanol consumption in mice suppresses release of IL-17 into lung tissue, decreases neutrophil recruitment, and increases mortality from experimental K. pneumonia. In vitro experiments confirm a direct suppressive effect of ethanol on the release of IL-17 from splenocytes. In vivo administration of the IL-17 gene in an adenoviral vector to alcohol-consuming mice results in release of IL-17 into lavage fluid and normalizes neutrophil recruitment and mortality after bacterial challenge. CONCLUSIONS: The results of these experiments strongly implicate IL-17 as an important pathway for the immunosuppression associated with alcohol abuse and support gene therapeutic approaches to augment immune function in the alcoholic host or to treat infections associated with alcoholism.

Inhibition of hematopoietic progenitor cell proliferation by ethanol in human immunodeficiency virus type 1 tat-expressing transgenic mice.

BACKGROUND: A number of hematological abnormalities are associated with both human immunodeficiency virus type 1 (HIV-1) infection and alcohol abuse. There is little information on how alcohol abuse might further influence the survival and growth of hematopoietic progenitors in HIV-infected individuals in the presence of immune system abnormalities and anti-HIV drugs. Because there is evidence that viral transactivator Tat itself can induce hematopoietic suppression, in this study we examined the role of ethanol as a cofactor in transgenic mice that expressed HIV-1 Tat protein. METHODS: Tat transgenic mice and nontransgenic littermates were given ethanol (20% v/v) and the anti-HIV drug 3'-azido-3'-deoxythymidine (AZT; 1 mg/ml) in drinking water. Immunosuppression in mice was induced by weekly intraperitoneal injections of anti-CD4 antibody. Hematopoiesis was examined by erythroid colony forming unit (CFU-E) and granulocyte/macrophage colony-forming unit (CFU-GM) assays of the bone marrow progenitor cells. RESULTS: Administration of ethanol for 7 weeks resulted in a 50% decrease in the proliferative capacity of CFU-E- and CFU-GM-derived progenitors from transgenic mice compared with that of ethanol-treated nontransgenic controls. Similar decreases also were observed in transgenic mice treated with AZT or a combination of AZT and ethanol. Furthermore, ethanol and AZT were significantly more toxic to the granulopoietic progenitors (40-50% inhibition) than to the erythropoietic progenitors (10-20% inhibition) in Tat transgenic mice. Although a 10 day exposure of Tat transgenic and nontransgenic mice to a combination of ethanol and AZT had no suppressive effect on the erythropoietic and granulopoietic progenitor cells, there was a marked decrease (40-60%) in CFU-GM in mice made immunodeficient by CD4+ T-lymphocyte depletion. The ethanol-treated Tat transgenic mice but not the nontransgenic litter-mates also showed a significant decrease (25%) in CFU-GM. CONCLUSION: Our in vivo study strongly suggests that ethanol ingestion in HIV-1-infected individuals, particularly those on antiretroviral drugs, might increase bone marrow toxicity and contribute to HIV-1-associated hematopoietic impairment.

Gene therapy to modify pulmonary host defenses.

Respiratory infections remain a significant public health problem and are presently the 6th leading cause of death in the United States. Antibiotic-resistant organisms are encountered increasingly in both community-acquired and nosocomial infections. Despite progress in antibiotic development, biological-response modifiers may have increasing application to augment pulmonary host defenses against either drug-resistant infections or in high-risk hosts. Toward this end, gene therapy proposes to deliver biologicals as nucleic acids rather than protein. Gene therapy has the potential advantage of targeting the biological to specific cells or tissue compartments, and a more favorable pharmacokinetic profile. Data on gene delivery and efficacy in preclinical models of pulmonary infection are presented and discussed.

CD4+ lymphocyte responses to pulmonary infection with Mycobacterium tuberculosis in naïve and vaccinated BALB/c mice.

The Biostack experiments I and II were flown on board the Apollo 16 and 17 command modules in order to obtain information on the biological damage produced by the bombardment of heavy high-energy (HZE) particles of cosmic radiation during spaceflight. Such data are required for estimating radiation hazards in manned spaceflight. Seven biological systems in resting state (Bacillus subtilis spores, Colpoda cucullus cysts, Arabidopsis thaliana seeds, and eggs of Artemia salina, Tribolium castaneum and of Carausius morosus) were accommodated in the two Biostacks. By using a special sandwich construction of visual track detectors and layers of biological objects, identification of each hit biological object was achieved and the possible biological damage correlated with the physical features of the responsible HZE-particle. In the different systems the degree of damage depended on whether the hit cell was replaceable or not. A high sensitivity to HZE-particle bombardment was observed on Artemia salina eggs; 90% of the embryos, which were induced to develop from hit eggs, died at different developmental stages. Malformations of the abdomen or the extremities of the nauplius were frequently induced. In contrast, the growth of hit Vicia faba radiculae and the germination of hit Arabidopsis thaliana seeds and hit Bacillus subtilis spores were not influenced remarkably. But there was an increase in multicaulous plants and a reduction in the outgrowth of the bacteria] spores. In addition, information was obtained on the fluence of the HZE-particles, on their spectrum of charge and energy loss, and on the absorption by the Apollo spacecraft and the Biostack material itself. This will help to improve knowledge concerning radiation conditions inside of spacecrafts, necessary to secure a The Biostack experiments I and II were flown on board the Apollo 16 and 17 command modules in order to obtain information on the biological damage produced by the bombardment of heavy high-energy (HZE) particles of cosmic radiation during spaceflight. Such data are required for estimating radiation hazards in manned spaceflight. Seven biological systems in resting state (Bacillus subtilis spores, Colpoda cucullus cysts, Arabidopsis thaliana seeds, and eggs of Artemia salina, Tribolium castaneum and of Carausius morosus) were accommodated in the two Biostacks. By using a special sandwich construction of visual track detectors and layers of biological objects, identification of each hit biological object was achieved and the possible biological damage correlated with the physical features of the responsible HZE-particle. In the different systems the degree of damage depended on whether the hit cell was replaceable or not. A high sensitivity to HZE-particle bombardment was observed on Artemia salina eggs; 90% of the embryos, which were induced to develop from hit eggs, died at different developmental stages. Malformations of the abdomen or the extremities of the nauplius were frequently induced. In contrast, the growth of hit Vicia faba radiculae and the germination of hit Arabidopsis thaliana seeds and hit Bacillus subtilis spores were not influenced remarkably. But there was an increase in multicaulous plants and a reduction in the outgrowth of the bacteria] spores. In addition, information was obtained on the fluence of the HZE-particles, on their spectrum of charge and energy loss, and on the absorption by the Apollo spacecraft and the Biostack material itself. This will help to improve knowledge concerning radiation conditions inside of spacecrafts, necessary to secure a The Biostack experiments I and II were flown on board the Apollo 16 and 17 command modules in order to obtain information on the biological damage produced by the bombardment of heavy high-energy (HZE) particles of cosmic radiation during spaceflight. Such data are required for estimating radiation hazards in manned spaceflight. Seven biological systems in resting state (Bacillus subtilis spores, Colpoda cucullus cysts, Arabidopsis thaliana seeds, and eggs of Artemia salina, Tribolium castaneum and of Carausius morosus) were accommodated in the two Biostacks. By using a special sandwich construction of visual track detectors and layers of biological objects, identification of each hit biological object was achieved and the possible biological damage correlated with the physical features of the responsible HZE-particle. In the different systems the degree of damage depended on whether the hit cell was replaceable or not. A high sensitivity to HZE-particle bombardment was observed on Artemia salina eggs; 90% of the embryos, which were induced to develop from hit eggs, died at different developmental stages. Malformations of the abdomen or the extremities of the nauplius were frequently induced. In contrast, the growth of hit Vicia faba radiculae and the germination of hit Arabidopsis thaliana seeds and hit Bacillus subtilis spores were not influenced remarkably. But there was an increase in multicaulous plants and a reduction in the outgrowth of the bacteria] spores. In addition, information was obtained on the fluence of the HZE-particles, on their spectrum of charge and energy loss, and on the absorption by the Apollo spacecraft and the Biostack material itself. This will help to improve knowledge concerning radiation conditions inside of spacecrafts, necessary to secure a maximum possible protection to the astronauts.

Murine CD4+ T lymphocyte subsets and host defense against Pneumocystis carinii.

The recruitment of specific subsets of CD4(+) T lymphocytes to the lungs in response to Pneumocystis carinii was investigated. For mice inoculated with P. carinii, an ELISPOT assay was used to calculate the numbers of lymph node and lung tissue CD4(+) cells that secreted interferon (IFN)-gamma (Th1 cytokine) and interleukin (IL)-4 (Th2 cytokine) after concanavalin A stimulation. An ELISA was used to assay culture supernatants for cytokine concentrations. Precursor frequency of both IFN-gamma- and IL-4-secreting cells was increased in lymph nodes at 1 week, whereas increases in Th1 and Th2 cells in lung tissue were delayed 3 weeks before declining. The frequency of IL-4-secreting cells always was greater than the frequency of IFN-gamma secreting cells. These results demonstrate an early T lymphocyte response in draining lymph nodes, followed by later recruitment of Th1 and Th2 lymphocytes into lung tissue. The overall CD4(+) T cell response to P. carinii involves both Th1 and Th2 subsets, but the response is Th2 dominant in both lymph node and lung tissue.

Inhibition of TNF-alpha processing and TACE-mediated ectodomain shedding by ethanol.

Alcohol (EtOH) is a well-documented immunosuppressant. Acute EtOH-induced immunosuppression is partially due to suppression of tumor necrosis factor alpha (TNF-alpha) secretion. We investigated the mechanism of acute EtOH-induced TNF-alpha suppression in two monocytic cell lines, Mono Mac 6 and DRM. EtOH inhibited TNF-alpha secretion in a dose-dependent manner. However, TNF-alpha transcription was not affected by EtOH. Enzyme-linked immunosorbent assay and confocal microscopy showed that EtOH treatment increased cell-associated TNF-alpha. Ectodomain shedding of TNF-alpha from the cell surface is mediated by TNF-alpha converting enzyme (TACE). In contrast with TNF-alpha, EtOH did not inhibit interleukin-8 (IL-8) secretion, which does not require shedding. Furthermore, TNF p75 receptor shedding, a biomarker for TACE activity, was inhibited by EtOH in both cell lines. EtOH also inhibited TNF p75 receptor shedding in TACE-reconstituted fibroblasts, suggesting that EtOH inhibits the shedding process. These data show that acute EtOH exposure can posttranscriptionally suppress TNF-alpha production, resulting in specific defects in immune defense.

Immunomodulation and adenoviral gene transfer to the lungs of nonhuman primates.

Previous data from our laboratory and others have demonstrated a critical role for the CD4+ T lymphocyte in in vivo immune responses to recombinant adenoviral vectors. In rodent models, this subset of T cells is required for T cell proliferation, subsequent cytotoxic T cell generation, and production of anti-adenoviral antibodies by B cells. Both depleting and nondepleting anti-CD4 antibodies can attenuate these immune responses to recombinant adenovirus. On the basis of these data, we hypothesized that a nondepleting CDR-engrafted anti-human CD4 antibody (OKT4A) with cross-reactivity to rhesus macaques would attenuate both T and B cell responses to intrapulmonary administration of recombinant adenovirus and permit prolonged reporter gene expression and permit secondary gene transfer. Juvenile rhesus macaques were treated with PBS or OKT4A antibody (10 mg/kg) daily beginning 1 day prior to and up to 11 days after gene transfer. OKT4A resulted in significant attenuation of lymphocyte recruitment into the lung, lymphocyte-proliferative responses to both adenovirus capsid proteins and transgene protein, and adenovirus-induced interferon-gamma elaboration in whole blood and hilar lymph nodes. However, OKT4A was ineffective in attenuating adenovirus-induced IL-4 production in whole blood or hilar lymph nodes, generating neutralizing anti-adenoviral antibodies, or permitting secondary gene transfer. As all the monkeys in this protocol had baseline-detectable anti-adenoviral antibodies by ELISA that were nonneutralizing, analogous to most patients with cystic fibrosis, we postulate that anti-CD4 did not block the proliferation of memory B cells. Moreover, these data suggest that for transient immunomodulation to be successful, strategies need to focus specifically on B cell activation independent of CD4+ T cell help.

Orthotopic human lung carcinoma xenografts in BALB/c mice immunosuppressed with anti-CD4 monoclonal antibodies and chronic alcohol consumption.

BACKGROUND: The role of the immune system in the surveillance of the body for cancer cells is well established. Human tumor cells do not survive in mice with intact immune systems, but they propagate in athymic nude mice. Presumably, the lack of a thymus gland and consequent loss of T lymphocytes results in a seriously compromised immune system without adequate cell-mediated immunity and tumor surveillance. In patients infected with the human immunodeficiency virus (HIV), a progressive loss of cell-mediated immunity is associated with the development of malignancies and opportunistic infections. This effect may be exacerbated in patients who chronically consume alcohol. METHODS: Normal and alcoholic BALB/c mice were treated with a monoclonal antibody to deplete CD4(+) lymphocytes before orthotopic implantation of human lung adenocarcinoma xenografts. Tumor volume and weight were measured and compared between groups. RESULTS: The authors' data show that a single treatment of anti-CD4 antibody causes almost complete depletion of CD4(+) lymphocytes and permits the formation of large intrapulmonary human nonsmall lung carcinoma xenografts in 100% of treated mice. All control animals injected with heat-denatured antibody failed to produce tumors. Chronic alcohol consumption by CD4-depleted mice resulted in larger tumors, compared with mice that did not receive ethanol in their diet (P = 0.05). CONCLUSIONS: Depletion of CD4(+) lymphocytes allows for the orthotopic growth of human lung adenocarcinoma xenografts in BALB/c mice. Furthermore, the consumption of alcohol reduces the ability of the impaired immune system to reject tumors.

IFN-gamma and CD8+ T cells restore host defenses against Pneumocystis carinii in mice depleted of CD4+ T cells.

Host defenses against infection are profoundly compromised in HIV-infected hosts due to progressive depletion of CD4+ T lymphocytes and defective cell-mediated immunity. Although recent advances in antiretroviral therapy can dramatically lower HIV viral load, blood CD4+ T lymphocytes are not restored to normal levels. Therefore, we investigated mechanisms of host defense other than those involving CD4+ T lymphocytes against a common HIV-related opportunistic infection, Pneumocystis carinii (PC) pneumonia. Using CD4-depleted mice, which are permissive for chronic PC infection, we show that up-regulation of murine IFN-gamma by gene transfer into the lung tissue results in clearance of PC from the lungs in the absence of CD4+ lymphocytes. This resolution of infection was associated with a >4-fold increase in recruited CD8+ T lymphocytes and NK cells into the lungs. The role of CD8+ T cells as effector cells in this model was further confirmed by a lack of an effect of IFN-gamma gene transfer in scid mice or mice depleted of both CD4+ and CD8+ T cells. Cytokine mRNA analysis revealed that recruited, lung-derived CD8+ T cells had greater expression of IFN-gamma message in animals treated with the IFN-gamma gene. These results indicate that CD8+ T cells are capable of clearing PC pneumonia in the absence of CD4+ T cells and that this host defense function of CD8+ T cells, as well as their cytokine repertoire, can be up-regulated through cytokine gene transfer.

Alveolar macrophage release of tumor necrosis factor-alpha in chronic alcoholics without liver disease.

Alcohol is an immunosuppressive drug, and chronic abuse has been associated with increased susceptibility to a variety of infections, including bacterial pneumonia and tuberculosis. Alveolar macrophages are the resident phagocytes of the lung and play a central role in lung host defenses against infection ranging from direct antibacterial activity to the release of proinflammatory cytokines such as tumor necrosis factor-alpha (TNFalpha). TNFalpha, in particular, plays a key role in the development of the early inflammatory response. In this study, we investigated the effects of chronic alcohol consumption on alveolar macrophage release of TNFalpha in vitro. We prospectively studied lipopolysaccharide (LPS)-stimulated release of TNFalpha from alveolar macrophages obtained from bronchoalveolar lavage fluid (BALF) in 22 alcoholic (18 smokers, 4 nonsmokers) and 7 nondrinking healthy volunteers (3 smokers, 4 nonsmokers). The total number of cells recovered by bronchoalveolar lavage (BAL) and their differential distribution were not significantly different in alcoholics versus controls (43 +/- 8 x 10(6) and 39 +/- 13 x 10(6), respectively). However, the total number of cells recovered from BALF was significantly higher in smokers (51 +/- 8 x 10(6)) than in nonsmokers (19 +/- 5 x 10(6)). Spontaneous (basal) release of TNFalpha by alveolar macrophages was the same in alcoholics and controls. In contrast, LPS-stimulated release of TNFalpha was significantly suppressed in alcoholics compared with that of controls (1343 +/- 271 vs. 3806 +/- 926 U TNF/ml/10(6) cells, respectively, p < 0.015). When controlled for smoking, LPS-stimulated TNFalpha production was suppressed in alcoholic nonsmokers (563 +/- 413 U TNF/ml/10(6)) compared with control nonsmokers (5113 +/- 1264 U TNF/ml/10(6)). LPS-stimulated TNFalpha production was also less in control smokers (2063 +/- 386 U TNF/ml/10(6) cells) than in control nonsmokers (5113 +/- 1264 U TNF/ml/10(6) cells). There was no difference in TNFalpha production between smoking alcoholics and smoking control subjects. We conclude that chronic alcohol consumption significantly suppresses LPS-stimulated alveolar macrophage production of TNFalpha. This effect is obscured if the subject also smokes. Because TNFalpha production is an important element in host defense, this may explain, in part, the susceptibility of chronic alcohol abusers to a variety of infections.

Alcohol decreases T-lymphocyte migration into lung tissue in response to Pneumocystis carinii and depletes T-lymphocyte numbers in the spleens of mice.

Previous work from our laboratory has shown that chronic alcohol consumption in mice creates immunosuppression sufficient to permit infection with the opportunistic pathogen Pneumocystis carinii. Host defense against P. carinii is critically dependent upon host T lymphocytes. In these experiments, we address the effect of chronic alcohol consumption on recruitment of T lymphocytes into infected lung tissue and on lymphocytes in host lymphoid tissue. We find that mice administered alcohol in drinking water and then inoculated with P. carinii show significantly decreased recruitment of CD4+ and CD8+ T lymphocytes into lung tissue in comparison with control mice. Additional experiments show significant depletion of CD4+ lymphocytes in spleens from alcohol mice and decreased numbers of activated T lymphocytes. Analysis of surface expression of the adhesion molecules LFA-1, VLA-4, and ICAM-1 show no significant differences in lymphocytes from alcohol-consuming mice, and lymphocyte chemotaxis in vitro is also unaltered. We conclude that chronic consumption of alcohol impairs lung recruitment of lymphocytes in response to an infectious challenge. This impaired lymphocyte recruitment may be a consequence of depletion of T lymphocytes in host lymphoid tissue. Impaired recruitment of lymphocytes may explain the increased morbidity and mortality of pulmonary infections in alcoholic subjects.