Alveolar macrophages in pulmonary host defence the unrecognized role of apoptosis as a mechanism of intracellular bacterial killing.

Alveolar macrophages play an essential role in clearing bacteria from the lower airway, as the resident phagocyte alveolar macrophages must both phagocytose and kill bacteria, and if unable to do this completely must co-ordinate an inflammatory response. The decision to escalate the inflammatory response represents the transition between subclinical infection and the development of pneumonia. Alveolar macrophages are well equipped to phagocytose bacteria and have a large phagolysosomal capacity in which ingested bacteria are killed. The rate-limiting step in control of extracellular bacteria, such as Streptococcus pneumoniae, is the capacity of alveolar macrophages to kill ingested bacteria. Therefore, alveolar macrophages complement canonical microbicidal strategies with an additional level of apoptosis-associated killing to help kill ingested bacteria.

CD4 regulates susceptibility to Fas ligand- and tumor necrosis factor-mediated apoptosis.

The current knowledge of CD4 function is limited to its role as a necessary coreceptor in TCR-initiated signaling. We have investigated whether CD4 regulates additional T cell functions. Using human primary resting CD4+ T cells, we demonstrate that CD4 activation is sufficient to induce lymphocyte death. Immediately after CD4 cross-linking, CD4+ T cells are rendered susceptible to apoptosis mediated by TNF or FasL. This, together with the concomitant induction of FasL within the same population, results in significant CD4+ T cell death in vitro. The CD4-dependent induction of susceptibility to apoptosis that is mediated by TNF or FasL is protein synthesis independent but phosphorylation dependent. After CD4 activation, PKC regulates susceptibility to apoptosis mediated by FasL but not the induction of susceptibility to TNF-dependent apoptosis. Moreover, significant differences between CD3 and CD4 activation were observed with regards to the kinetics of induction of CD4+ T cell susceptibility to FasL- and TNF-mediated apoptosis. Altogether, these results provide a model with which to study the molecular mechanisms regulating lymphocyte survival after CD4 activation, and highlight the potential role of CD4 in controlling lymphocyte apoptosis under physiological conditions or in disease states such as HIV infection.

Rate of replenishment and microenvironment contribute to the sexually dimorphic phenotype and function of peritoneal macrophages.

Macrophages reside in the body cavities where they maintain serosal homeostasis and provide immune surveillance. Peritoneal macrophages are implicated in the etiology of pathologies including peritonitis, endometriosis, and metastatic cancer; thus, understanding the factors that govern their behavior is vital. Using a combination of fate mapping techniques, we have investigated the impact of sex and age on murine peritoneal macrophage differentiation, turnover, and function. We demonstrate that the sexually dimorphic replenishment of peritoneal macrophages from the bone marrow, which is high in males and very low in females, is driven by changes in the local microenvironment that arise upon sexual maturation. Population and single-cell RNA sequencing revealed marked dimorphisms in gene expression between male and female peritoneal macrophages that was, in part, explained by differences in composition of these populations. By estimating the time of residency of different subsets within the cavity and assessing development of dimorphisms with age and in monocytopenic Ccr2 -/- mice, we demonstrate that key sex-dependent features of peritoneal macrophages are a function of the differential rate of replenishment from the bone marrow, whereas others are reliant on local microenvironment signals. We demonstrate that the dimorphic turnover of peritoneal macrophages contributes to differences in the ability to protect against pneumococcal peritonitis between the sexes. These data highlight the importance of considering both sex and age in susceptibility to inflammatory and infectious diseases.

Salvage therapy for invasive aspergillosis.

Invasive aspergillosis (IA) makes a marked contribution to the mortality of immunocompromised hosts, especially those who have received cytotoxic chemotherapy for haematological malignancy or allogeneic haemopoietic stem cell transplantation. Salvage therapy, in the case of invasive fungal infection, generally refers to the treatment of infected individuals who are refractory or intolerant to initial therapy administered for at least 7 days. Although clinical trials of salvage therapy of IA have been undertaken, most were non-comparator studies or contained a non-randomized control group, and criteria for patient enrollment and the methods used to assess response were variable. Salvage therapy has produced relatively disappointing results, emphasizing the importance of early diagnosis and effective primary therapy for IA. Despite this, a number of agents have been studied in the treatment of IA and have demonstrated efficacy in a salvage setting. These include lipid-based formulations of amphotericin B, caspofungin, itraconazole, voriconazole, posaconazole and micafungin. Combinations of echinocandins with either azoles or amphotericin B have also been studied in small series. Further studies are required, ideally comparing newer agents and treatment strategies in randomized clinical trials, to clarify the optimal approach to salvage treatment of IA in this challenging group of patients.

In vivo analysis of Fas/FasL interactions in HIV-infected patients.

Recent insights into the pharmacological control of HIV replication and the molecular mechanisms of peripheral T cells homeostasis allowed us to investigate in vivo the mechanisms mediating T cell depletion in HIV-infected patients. Before the initiation of highly active antiretroviral therapy (HAART), a high degree of lymphoid tissue apoptosis is present, which is reduced upon HAART initiation (P < 0.001) and directly correlates with reduction of viral load and increases of peripheral T lymphocytes (P < 0.01). Because Fas/FasL interactions play a key role in peripheral T lymphocyte homeostasis, we investigated the susceptibility to Fas-mediated apoptosis in peripheral T lymphocytes and of FasL expression in lymphoid tissue before and during HAART. High levels of Fas-susceptibility found in peripheral CD4 T lymphocytes before HAART were significantly reduced after HAART, coinciding with decreases in viral load (P = 0.018) and increases in peripheral CD4 T lymphocyte counts (P < 0.01). However, the increased FasL expression in the lymphoid tissue of HIV-infected individuals was not reduced after HAART. These results demonstrate that lymphoid tissue apoptosis directly correlates with viral load and peripheral T lymphocyte numbers, and suggest that HIV-induced susceptibility to Fas-dependent apoptosis may play a key role in the regulation of T cell homeostasis in HIV-infected individuals.

The expression of Fas Ligand by macrophages and its upregulation by human immunodeficiency virus infection.

Fas/Fas Ligand (FasL) interactions play a significant role in peripheral T lymphocyte homeostasis and in certain pathological states characterized by T cell depletion. In this study, we demonstrate that antigen-presenting cells such as monocyte-derived human macrophages (MDM) but not monocyte-derived dendritic cells express basal levels of FasL. HIV infection of MDM increases FasL protein expression independent of posttranslational mechanisms, thus highlighting the virus-induced transcriptional upregulation of FasL. The in vitro relevance of these observations is confirmed in human lymphoid tissue. FasL protein expression is constitutive and restricted to tissue macrophages and not dendritic cells. Moreover, a significant increase in macrophage-associated FasL is observed in lymphoid tissue from HIV (+) individuals (P < 0.001), which is further supported by increased levels of FasL mRNA using in situ hybridization. The degree of FasL protein expression in vivo correlates with the degree of tissue apoptosis (r = 0.761, P < 0. 001), which is significantly increased in tissue from HIV-infected patients (P < 0.001). These results identify human tissue macrophages as a relevant source for FasL expression in vitro and in vivo and highlight the potential role of FasL expression in the immunopathogenesis of HIV infection.

Hypoxia determines survival outcomes of bacterial infection through HIF-1alpha dependent re-programming of leukocyte metabolism.

Hypoxia and bacterial infection frequently co-exist, in both acute and chronic clinical settings, and typically result in adverse clinical outcomes. To ameliorate this morbidity, we investigated the interaction between hypoxia and the host response. In the context of acute hypoxia, both S. aureus and S. pneumoniae infections rapidly induced progressive neutrophil mediated morbidity and mortality, with associated hypothermia and cardiovascular compromise. Preconditioning animals through longer exposures to hypoxia, prior to infection, prevented these pathophysiological responses and profoundly dampened the transcriptome of circulating leukocytes. Specifically, perturbation of HIF pathway and glycolysis genes by hypoxic preconditioning was associated with reduced leukocyte glucose utilisation, resulting in systemic rescue from a global negative energy state and myocardial protection. Thus we demonstrate that hypoxia preconditions the innate immune response and determines survival outcomes following bacterial infection through suppression of HIF-1α and neutrophil metabolism. The therapeutic implications of this work are that in the context of systemic or tissue hypoxia therapies that target the host response could improve infection associated morbidity and mortality.

Organizing pneumonia. Features and prognosis of cryptogenic, secondary, and focal variants.

BACKGROUND: Organizing pneumonia (OP) is a non-specific response to many types of lung injury. Clinicians frequently encounter pathology reports of OP in patients with no underlying condition (cryptogenic OP, also known as BOOP or bronchiolitis obliterans OP) or in association with drugs or nonpulmonary disease. The goals of this study are to describe the clinical course and outcomes in patients with 3 clinical variants of OP. METHODS: A retrospective study of patients with OP seen at the Mayo Clinic, Rochester, Minn, from January 1, 1984, through June 30, 1994, was conducted. Initial features were obtained from medical records. Chest radiographs and pathology specimens were reviewed for this study. Resolution, relapse, and survival were obtained from medical records and a follow-up patient questionnaire. RESULTS: Seventy-four patients had pathologically confirmed OP. Organizing pneumonia was classified into 3 clinical groups: symptomatic cryptogenic OP; symptomatic OP related to underlying hematologic malignant neoplasm, collagen vascular disease, or drugs (secondary OP); and asymptomatic OP presenting as a focal nodule (focal OP). Thirty-seven patients (50%) had cryptogenic OP and 27 patients (36%) had secondary OP. No difference was found between cryptogenic and secondary OP in type or severity of symptoms, signs, laboratory and pulmonary function tests, or radiologic or pathologic findings. Corticosteroids were given at a similar initial dose (prednisone, about 50 mg/d). Resolution of symptoms was more frequent in patients with cryptogenic OP than those with secondary OP. Relapse was infrequent in both of these groups. Five-year survival was higher in patients with cryptogenic OP (73%) than in secondary OP (44%), and respiratory-related deaths were more frequent in patients with secondary OP. Organizing pneumonia was an asymptomatic focal rounded opacity in 10 patients (14%), most often detected on chest radiograph and diagnosed on lung biopsy done for suspicion of lung cancer. Patients with focal OP required no treatment and had no relapse or respiratory-related deaths. CONCLUSIONS: Clinical classification of OP is useful to predict clinical course and outcome. Cryptogenic OP most often was a symptomatic bilateral lung process that had an overall favorable prognosis with prolonged corticosteroid therapy. Patients with secondary OP had a high mortality rate when the disease was associated with predisposing conditions or drugs. Patients with asymptomatic focal OP had an excellent prognosis.

Human herpesvirus 6 seronegativity before transplantation predicts the occurrence of fungal infection in liver transplant recipients.

BACKGROUND: Invasive fungal infection has a major impact on the morbidity and mortality of liver transplant recipients. Human herpesvirus (HHV)-6 infection after transplantation is associated with an immunosuppressive state and the development of cytomegalovirus disease. Because cytomegalovirus infection is a risk factor for invasive fungal infection after transplantation, we have examined whether HHV-6 and fungal infection are associated after transplantation. METHODS: Pretransplantation sera from 247 consecutive liver transplant recipients were analyzed for IgG to HHV-6. Thirty-three (13%) HHV-6-seronegative recipients were identified. Six of 33 (18%) seronegative recipients experienced fungal infection as compared with 15 of 214 (7%) seropositive recipients (P=0.034). RESULTS: In a univariate analysis of risk factors for fungal infection, pretransplantation seronegativity to HHV-6 (P=0.034), intraoperative cryoprecipitate requirements greater than the 75th percentile (P=0.035), reoperation (P=0.005), biliary stricturing postoperatively (P=0.046), and gastrointestinal or vascular complications postoperatively (P=0.030) were identified as significant risk factors. Moreover, in pairwise multivariate analysis, pretransplantation HHV-6 seronegativity remained a significant variable even in the presence of each of the other variables. CONCLUSIONS: These results suggest that HHV-6 seronegativity before transplantation is a valuable clinical marker that identifies patients at risk for developing fungal infection after transplantation.

Activation-induced CD4+ T cell death in HIV-positive individuals correlates with Fas susceptibility, CD4+ T cell count, and HIV plasma viral copy number.

The relevance of activation-induced cell death (AICD) of CD4+ T cells to AIDS pathogenesis is unknown. The present study investigates the relationship of AICD to a defined molecular mechanism regulating peripheral T cell homeostasis, Fas-mediated apoptosis, and clinical correlates of the pathogenesis of AIDS. Increased pokeweed mitogen (PWM)-induced AICD (22.8 versus 4.4%, p = 0.006) and Fas-mediated apoptosis (27.7 versus 12.0%, p = 0.002) of CD4+ T cells were observed in HIV+ versus HIV- individuals. Similarly, increased PWM-mediated AICD (16.2 versus 2.2%, p < 0.001) and Fas-mediated apoptosis (25.8 versus 7.6%, p = 0.005) were noted in CD8+ T cells from HIV+ versus HIV- individuals. PWM-induced AICD of CD4+ T cells was blocked (83% median specific inhibition) by Fas-blocking antibodies, whereas PWM-induced AICD of CD8+ T cells was Fas independent. Comparison between PWM- and anti-CD3-mediated AICD of CD4+ T cells indicated that PWM- and not CD3-induced AICD is Fas dependent. HIV+ individuals with an HIV RNA copy number of <30,000 copies/ml had lower PWM-induced AICD of CD4+ T cells than did those with an HIV RNA copy number of >30,000 copies/ml (5.7 versus 22.1%, p = 0.034), and PWM-induced AICD inversely correlated with CD4+ T cell count (R = -0.567, p = 0.043). Initiation of HAART decreased PWM-induced CD4+ T cell AICD from 24.4 to 9.4% posttreatment (p = 0.035). These results demonstrate that PWM-induced AICD of CD4+ T cells from HIV+ individuals is mediated by Fas/FasL, parallels the in vivo susceptibility of the CD4+ T cell to Fas-mediated apoptosis, and correlates with clinical markers of AIDS pathogenesis and response to HAART.

Hypercalcemia in a patient with hypoparathyroidism and Nocardia asteroides infection: a novel observation.

Hypercalcemia is associated with numerous chronic granulomatous processes and chronic infections. Increased production of 1,25-dihydroxyvitamin D by activated macrophages has been shown to be the cause in most cases. In this article, we describe a case of hypercalcemia related to infection with Nocardia asteroides. In a 34-year-old woman who previously had hypocalcemia, acute hypercalcemia developed coincident with Nocardia pericarditis. The hypercalcemia resolved after treatment of N. asteroides with sulfisoxazole. Parathyroid hormone and phosphorus levels were within normal limits, and total 25-hydroxyvitamin D levels were only mildly increased. After successful treatment of the Nocardia infection, the patient required supplemental calcium and vitamin D. Her hypercalcemia was temporally related to the duration of the N. asteroides infection. We believe this is the first reported case of hypercalcemia associated with N. asteroides infection.

Human herpesvirus 6.

Human herpesvirus (HHV) 6 is a beta-herpes, DNA virus. This virus shows closest homology with cytomegalovirus and HHV-7. Infection usually occurs in infants 6 to 24 months of age, and primary infection may result in roseola. HHV-6 infection in infants is the commonest cause of fever-induced seizures. Infection in adults is seen primarily in immunocompromised hosts with solid organ transplants or in those with human immunodeficiency virus infection. The virus is capable of pronounced interaction in vitro with cytomegalovirus and human immunodeficiency virus and induces immunosuppression and apoptosis. The importance of these interactions in vivo necessitates further investigation. HHV-6 infection may contribute to the pathogenesis of multiple sclerosis. HHV-6 may be diagnosed by viral culture, serology, or polymerase chain reaction.

Human herpesvirus 6: molecular biology and clinical features.

Human herpesvirus 6 (HHV-6) exists as distinct variants HHV-6A and HHV-6B. The complete genomes of HHV-6A and HHV-6B have been sequenced. HHV-6B contains 97 unique genes. CD46 is the cell receptor for HHV-6, explaining its broad tissue tropism but its restricted host-species range. HHV-6 utilizes a number of strategies to down-regulate the host immune response, including molecular mimicry by production of a functional chemokine and chemokine receptors. Immunosuppression is enhanced by depletion of CD4 T lymphocytes via direct infection of intra-thymic progenitors and by apoptosis induction. Infection is widespread in infants between 6 months and 2 years of age. A minority of infants develop roseola infantum, but undifferentiated febrile illness is more common. Reactivation from latency occurs in immunocompromised hosts. Organ-specific clinical syndromes occasionally result, but indirect effects including interactions with other viruses such as human immunodeficiency virus type 1 and human cytomegalovirus or graft dysfunction in transplant recipients may be more significant complications in this population. Recent advances in quantitative PCR are providing additional insights into the natural history of infection in paediatric populations and immunocompromised hosts.

The multiple roles of Fas ligand in the pathogenesis of infectious diseases.

Fas ligand (FasL) is a type II transmembrane protein that plays a critical role in immune homeostasis by binding to its receptor Fas (CD95) and inducing apoptosis. Fas/FasL dysregulation contributes to infectious disease pathogenesis. Microorganisms may inhibit Fas signal transduction to prolong intracellular survival and prevent killing by immune effector cells. FasL may be upregulated in directly infected cells to enhance killing of responding immune cells and facilitate immune evasion. The host response to infection may aim to induce apoptosis in directly infected cells, but immune cells that target directly infected cells can induce Fas-mediated apoptosis of uninfected bystander cells. FasL also contributes to the generation and regulation of the inflammatory response in infection. The multiple roles of FasL in infectious disease pathogenesis are discussed in the context of viral, bacterial and parasitic infections.

Apoptotic cell death in the pathogenesis of infectious diseases.

Apoptosis is a physiological process critical for tissue homeostasis. It is essential for the regulation of immune responses. A series of molecules transduce apoptoic signals and induce the characteristic morphological appearances of apoptotic cells. Infectious diseases modulate apoptosis and this contributes to disease pathogenesis. Infection with HIV results in enhanced levels of CD4 T-lymphocyte apoptosis in both directly infected cells and in uninfected bystander cells. A variety of HIV proteins including gp120 contribute to this process. A number of different pathways induce HIV-associated CD4 T-lymphocyte apoptosis and apoptosis of uninfected bystander cells is particularly associated with increased susceptibility to Fas. Other viruses including hepatitis viruses and the human herpesviruses also modulate apoptosis. Bacterial infection induces apoptosis which is frequently mediated by the direct activation of caspases in the absence of death receptor ligation. Bacterial induction of apoptosis may either be due to bacterial factors such as the invasin IpaB of Shigella flexneri or be the result of host immune responses which control infection as demonstrated in infections due to Mycobacterium spp. Apoptosis may be modulated by therapeutic strategies, such as antiretroviral therapy, and an improved understanding of infection-associated apoptosis modulation will aid the design of novel therapeutic approaches to control infectious diseases.

Monocyte and macrophage dysfunction as a cause of HIV-1 induced dysfunction of innate immunity.

HIV-1 can establish both long lived and productive infection of macrophages (Mϕ) but circulating monocytes are less permissive to infection. Multiple studies have identified extensive changes to monocyte and Mϕ phenotype, differentiation or function. These include alterations in Toll-like receptor signaling and resultant changes to cytokine responses, specific defects in phagocytosis and microbial killing and modulation of apoptotic responses, all of which may perturb the important role of these cells in innate immunity. Interpretation of contradictory data however, is complicated by the use of different experimental models and many of the reported effects may be an indirect consequence of HIV 1 infection that result from exposure to viral products or from disruption of cellular and cytokine networks in the immune system, rather than the direct consequence of productive HIV 1 infection. Future research should focus on refining experimental models and on elucidating the physiological mechanisms of monocyte/ Mϕ dysfunction during HIV 1 infection.