Induction of GITRL expression in human keratinocytes by Th2 cytokines and TNF-α: implications for atopic dermatitis.

BACKGROUND: Glucocorticoid-induced TNF receptor-related protein ligand (GITRL), a ligand for the T cell co-stimulatory molecule GITR, is expressed by keratinocytes and involved in chemokine production. The expression of GITRL in skin inflammation remains unknown. OBJECTIVES: This study investigated cytokine regulation of keratinocyte GITRL expression. METHODS: Glucocorticoid-induced TNF receptor expression was evaluated in cytokine-treated human epidermal keratinocytes (HEK)s, murine PAM 212 cell line, murine and human skin explants by real time PCR, flow cytometry and immunostaining. Functional responses to GITR fusion protein were examined by real time PCR and ELISA. GITRL expression in AD and psoriasis was studied by immunohistochemistry. RESULTS: Skin biopsies from STAT6VT transgenic mice, which develop spontaneous atopic skin inflammation, were found by immunofluoresence, to have increased keratinocyte GITRL expression. Exposure to Th2 cytokines augmented GITRL mRNA expression in the murine PAM 212 keratinocytic cell line and murine skin explants. In contrast, GITRL mRNA and protein expression was only increased in HEKs and human skin explants in the presence of the combination of TNF-α and Th2 cytokines. A synergistic effect of Th2 cytokines and GITR fusion protein on production of CCL17, the Th2 chemokine, by murine keratinocytes was demonstrated. Immunohistochemical staining showed that acute AD lesions have increased expression of GITRL compared with normal skin, chronic AD lesions and psoriatic plaques. CONCLUSIONS AND CLINICAL RELEVANCE: Our studies demonstrate that GITRL expression is augmented by Th2 cytokines and TNF-α in keratinocytes. Increased GITRL expression in acute AD skin lesions is shown. This observation suggests a link between cytokine-regulated keratinocyte GITRL expression and its role in inflammatory responses in AD.

Immune signatures underlying post-acute COVID-19 lung sequelae.

Severe coronavirus disease 2019 (COVID-19) pneumonia survivors often exhibit long-term pulmonary sequelae, but the underlying mechanisms or associated local and systemic immune correlates are not known. Here, we have performed high-dimensional characterization of the pathophysiological and immune traits of aged COVID-19 convalescents, and correlated the local and systemic immune profiles with pulmonary function and lung imaging. We found that chronic lung impairment was accompanied by persistent respiratory immune alterations. We showed that functional severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)­specific memory T and B cells were enriched at the site of infection compared with those of blood. Detailed evaluation of the lung immune compartment revealed that dysregulated respiratory CD8+ T cell responses were associated with the impaired lung function after acute COVID-19. Single-cell transcriptomic analysis identified the potential pathogenic subsets of respiratory CD8+ T cells contributing to persistent tissue conditions after COVID-19. Our results have revealed pathophysiological and immune traits that may support the development of lung sequelae after SARS-CoV-2 pneumonia in older individuals, with implications for the treatment of chronic COVID-19 symptoms.

A signal transducer and activator of transcription (Stat)4-independent pathway for the development of T helper type 1 cells.

The differentiation of T helper (Th) cells is regulated by members of the signal transducer and activator of transcription (STAT) family of signaling molecules. We have generated mice lacking both Stat4 and Stat6 to examine the ability of Th cells to develop in the absence of these two transcription factors. Stat4, Stat6(-/-) lymphocytes fail to differentiate into interleukin (IL)-4-secreting Th2 cells. However, in contrast to Stat4(-/-) lymphocytes, T cells from Stat4, Stat6(-/-) mice produce significant amounts of interferon (IFN)-gamma when activated in vitro. Although Stat4, Stat6(-/-) lymphocytes produce less IFN-gamma than IL-12-stimulated control lymphocytes, equivalent numbers of IFN-gamma-secreting cells can be generated from cultures of Stat4, Stat6(-/-) lymphocytes activated under neutral conditions and control lymphocytes activated under Th1 cell-promoting conditions. Moreover, Stat4, Stat6(-/-) mice are able to mount an in vivo Th1 cell-mediated delayed-type hypersensitivity response. These results support a model of Th cell differentiation in which the generation of Th2 cells requires Stat6, whereas a Stat4-independent pathway exists for the development of Th1 cells.

Pivotal role of signal transducer and activator of transcription (Stat)4 and Stat6 in the innate immune response during sepsis.

Signal transducer and activator of transcription (Stat)4 and Stat6 are transcription factors that provide type 1 and type 2 response, respectively. Here, we explored the role of Stat4 and Stat6 in innate immunity during septic peritonitis. Stat4-/- and Stat6-/- mice were resistant to the lethality compared with wild-type (WT) mice. At the mechanistic level, bacterial levels in Stat6-/- mice were much lower than in WT mice, which was associated with increased peritoneal levels of interleukin (IL)-12, tumor necrosis factor (TNF)-alpha, macrophage-derived chemokine (MDC), and C10, known to enhance bacterial clearance. In Stat4-/- mice, hepatic inflammation and injury during sepsis were significantly ameliorated without affecting local responses. This event was associated with increased hepatic levels of IL-10 and IL-13, while decreasing those of macrophage inflammatory protein (MIP)-2 and KC. Sepsis-induced renal injury was also abrogated in Stat4-/- mice, which was accompanied by decreased renal levels of MIP-2 and KC without altering IL-10 and IL-13 levels. Thus, Stat6-/- and Stat4-/- mice appeared to be resistant to septic peritonitis by enhancing local bacterial clearance and modulating systemic organ damage, respectively, via balancing cytokine responses. These results clearly highlight an important role of local type 1 and systemic type 2 cytokine response in protective immunity during sepsis, which can be regulated by Stat proteins.

Encoding a superantigen by Staphylococcus aureus does not affect clinical characteristics of infected atopic dermatitis lesions.

BACKGROUND: Bacterial infection with Staphylococcus aureus is a known trigger for the worsening of atopic dermatitis (AD). Staphylococcal superantigens have been theorized to make a potential contribution to this worsening of AD seen with infection. OBJECTIVES: We sought to assess whether encoding a superantigen by S. aureus affects the inflammatory characteristics of impetiginized AD skin lesions. METHODS: Fifty-two children with clinically impetiginized lesions of AD which were positive for S. aureus were enrolled in this study. A lesion was graded clinically using the Eczema Area and Severity Index (EASI), and then wash fluid was obtained from the lesion for quantitative bacterial culture, and measurement of bacterial products lipoteichoic acid and staphylococcal protein A and cytokines. The staphylococcal isolate was tested for antibiotic susceptibilities and the presence of a superantigen. RESULTS: Fifty-four per cent (28 of 52) of the staphylococcal isolates encoded a superantigen. The presence of a superantigen had no significant effect on EASI score, amounts of bacterial products or inflammatory cytokines in the AD lesion. CONCLUSIONS: These studies suggest that the expression of a superantigen by S. aureus alone does not play an important role in the increased skin inflammation associated with staphylococcal infection in childhood AD.

Immunoglobulin E production in the absence of interleukin-4-secreting CD1-dependent cells.

A lymphocyte population that expresses surface markers found on T cells and natural killer (NK) cells secretes large amounts of interleukin-4 (IL-4) immediately after T cell receptor ligation. These NK-like T cells are thus thought to be important for the initiation of type 2 T helper cell (TH2) responses. CD1-deficient mice were found to lack this lymphocyte subset, but they could nevertheless mount a protypical TH2 response; after immunization with antibody to immunoglobulin D (IgD), CD1-deficient mice produced IgE. Thus, although dependent on CD1 for their development, IL-4-secreting NK-like T cells are not required for TH2 responses.

Deleted HTLV-I provirus in blood and cutaneous lesions of patients with mycosis fungoides.

Mycosis fungoides, a rare form of cutaneous T cell leukemia/lymphoma, is suspected of having a viral etiology on the basis of certain similarities to adult T cell leukemia, which is associated with human T cell leukemia/lymphoma virus type I (HTLV-I) infection. Cell lines were established from peripheral blood mononuclear cells (PBMC) of an HTLV-I-seronegative patient with mycosis fungoides. DNA hybridization analysis revealed the presence of HTLV-I-related sequences with unusual restriction endonuclease sites. Sequence analysis of subcloned fragments demonstrated the presence of a monoclonally integrated provirus with a 5.5-kilobase deletion involving large regions of gag and env and all of pol. Additional evidence for the presence of deleted proviruses was found by polymerase chain reaction (PCR) amplification of DNA from cutaneous lesions of five other HTLV-I-seronegative patients. The findings suggest that HTLV-I infection may be involved in the etiology of at least certain cases of mycosis fungoides.

The role of mononuclear phagocytes in HTLV-III/LAV infection.

Cells with properties characteristic of mononuclear phagocytes were evaluated for infectivity with five different isolates of the AIDS virus, HTLV-III/LAV. Mononuclear phagocytes cultured from brain and lung tissues of AIDS patients harbored the virus. In vitro-infected macrophages from the peripheral blood, bone marrow, or cord blood of healthy donors produced large quantities of virus. Virus production persisted for at least 40 days and was not dependent on host cell proliferation. Giant multinucleated cells were frequently observed in the macrophage cultures and numerous virus particles, often located within vacuole-like structures, were present in infected cells. The different virus isolates were compared for their ability to infect macrophages and T cells. Isolates from lung- and brain-derived macrophages had a significantly higher ability to infect macrophages than T cells. In contrast, the prototype HTLV-III beta showed a 10,000-fold lower ability to infect macrophages than T cells and virus production was one-tenth that in macrophage cultures infected with other isolates, indicating that a particular variant of HTLV-III/LAV may have a preferential tropism for macrophages or T cells. These results suggest that mononuclear phagocytes may serve as primary targets for infection and agents for virus dissemination and that these virus-infected cells may play a role in the pathogenesis of the disease.

Angiogenic properties of Kaposi's sarcoma-derived cells after long-term culture in vitro.

Cells derived from lung biopsies and pleural effusions from AIDS patients with Kaposi's sarcoma (KS) of the lungs were established in long-term culture with the aid of conditioned medium from HTLV-II-transformed T cells (HTLV-II CM). These AIDS-KS cells were similar to the so-called spindle cells in KS lesions and had some of their features. They produced factors that supported their own growth (autocrine) and the growth of other cells (paracrine), including umbilical vein endothelium and fibroblasts. That the AIDS-KS cells also expressed potent angiogenic activity was demonstrated by the chorioallantoic membrane assay and by subcutaneous inoculation of AIDS-KS cells into nude mice, which resulted in the development of angiogenic lesions composed of mouse cells and showing histological features similar to those of human KS lesions. These data suggest that AIDS-associated KS and possibly other types of KS may be initiated by signals that induce the growth of particular cells (spindle cells of lymphatic or vascular origin) and the expression of autocrine and paracrine activities.

Immunopathologic studies of systemic lupus erythematosus. II. Antinuclear reaction of gamma-globulin eluted from homogenates and isolated glomeruli of kidneys from patients with lupus nephritis.

The gammaG-globulin eluted at acid pH from kidney cortex homogenates and isolated glomeruli of five of six patients with lupus nephritis was found to exhibit antinuclear activity, which was not dependent on presence of fresh human serum. Specificity, as demonstrated by absorption of antinuclear activity, was related to nucleoprotein in three glomerular acid eluates and to DNA in two acid eluates as well as in a deoxyribonuclease digest of disrupted glomeruli in one patient. Antinuclear activity was not found in acid eluates of kidneys from two patients with chronic liver disease and chronic discoid lupus, respectively, and one with lupus nephritis. These patients had a low titer of serum antinuclear factor and lesser amounts of kidney bound immunoglobulins. The presence of antinuclear activity in eluates of kidneys appeared to correlate with the amount of glomerular bound immunoglobulin and the level of antinuclear antibodies in serum. These findings suggest that in lupus nephritis, part of the glomerular bound immunoglobulin is derived from serum antinuclear factors possibly deposited as immune complexes.

Immunopathologic studies of systemic lupus erythematosus (SLE). I. Tissue-bound immunoglobulins in relation to serum antinuclear immunoglobulins in systemic lupus and in chronic liver disease with LE cell factor.

We studied the composition of tissue-bound immunoglobulins and of antinuclear factors by immunofluorescent techniques in five patients with systemic lupus and two with chronic liver disease associated with positive LE cell tests. Renal glomeruli in all seven demonstrated deposits of bound gammaG-globulin and complement, although the presence of gammaA- and gammaM-immunoglobulins was variable. Blood vessel walls contained primarily gammaG-globulin and complement in the systemic lupus patients, but such deposits were absent from vessels in the two with chronic liver disease. We observed antinuclear factors, demonstrated by immunofluorescence, in all three immunoglobulin classes. In six of the seven patients, evidence was obtained of a correspondence between the classes of bound immunoglobulins in glomeruli and vessels and the serum titers of antinuclear immunoglobulins. These observations are consistent with the concept that immunoglobulin deposits in tissues may be derived at least in part from antinuclear factors. Neither bound immunoglobulins nor complement was observed in liver parenchyma of the two patients with chronic liver disease or in two patients with systemic lupus and liver pathology. It thus seems doubtful that serum antibodies play a primary role in the pathogenesis of forms of chronic liver disease associated with positive LE cell tests.

Stat proteins control lymphocyte proliferation by regulating p27Kip1 expression.

The proliferation of lymphocytes in response to cytokine stimulation is essential for a variety of immune responses. Recent studies with signal transducer and activator of transcription 6 (Stat6)-deficient mice have demonstrated that this protein is required for the normal proliferation of lymphocytes in response to interleukin-4 (IL-4). In this report, we show that the impaired IL-4-induced proliferative response of Stat6-deficient lymphocytes is not due to an inability to activate alternate signaling pathways, such as those involving insulin receptor substrates, or to a failure to upregulate IL-4 receptor levels. Cell cycle analysis showed that the percentage of Stat6-deficient lymphocytes that transit from the G1 to the S phase of the cell cycle following IL-4 stimulation is lower than that of control lymphocytes. Although the regulation of many genes involved in the control of cytokine-induced proliferation is normal in Stat6-deficient lymphocytes, protein levels of the cdk inhibitor p27Kip1 were found to be markedly dysregulated. p27Kip1 is expressed at significantly higher levels in Stat6-deficient lymphocytes than in control cells following IL-4 stimulation. The higher level of p27Kip1 expression seen in IL-4-stimulated Stat6-deficient lymphocytes correlates with decreased cdk2-associated kinase activity and is the result of the increased accumulation of protein rather than altered mRNA expression. Similarly, higher levels of p27Kip1 protein expression are also seen following IL-12 stimulation of Stat4-deficient lymphocytes than are seen following stimulation of control cells. These data suggest that Stat proteins may control the cytokine-induced proliferative response of activated T cells by regulating the expression of cell cycle inhibitors so that cyclin-cdk complexes may function to promote transition from the G1 to the S phase of the cell cycle.

STAT Transcription Factors in T Cell Control of Health and Disease.

The Jak-STAT pathway is one of many pleiotropic signaling pathways that plays an important role in organismal development and in response to changing environmental cues. As a key signaling cascade for cytokines and growth factors, Jak-STAT plays central role in the innate and adaptive immune system. Cytokines control the stability, commitment, and maturation of cytotoxic and helper T cells, parts of the adaptive immune system that mediate immunity to pathogens and are linked to inflammatory diseases. Dysregulation of Jak-STAT protein expression or function leads to autoimmunity, allergic diseases, and cancer. Because of their central role in these responses, Jak and STAT molecules have been targeted to develop therapeutics. This review extensively discusses the mechanism of how Jak-STAT signaling in T cells defines our immune responses in the battle against foreign pathogens.

A classification of HTLV-III infection based on 75 cases seen in a suburban community.

Since 1981, 75 patients have been seen at our hospital with human T-cell lymphotropic virus type III (HTLV-III) infection. We have classified their clinical presentation into Groups 0 to 6. Groups 0 to 3 all have antibody to the Mr 41,000 protein of HTLV-III. Group 0 has no evident disease (9 patients), Group 1 has lymphadenopathy with or without exaggerated infection (16 patients), Group 2 has persistent lymphadenopathy with chronic hepatitis B surface antigenemia or profound hypergammaglobulinemia (7 patients), Group 3 has oral candidiasis with or without lymphadenopathy (7 patients). In Group 4 are acquired immunodeficiency syndrome (AIDS) adults or children (32 patients). Group 5 is a special classification for immunocompromised patients. Group 6 patients have lymphomas and Mr 41,000 protein antibody. Four children were classified separately. Three patients in Group 3 developed Group 4 disorders (AIDS). Four patients in Group 4 developed Group 6 disorders. HTLV-III infection spread in families (8 of 36), all from infected mothers to children. In 17 sexual partners, 6 were found to be infected. Five of 6 infected partners were homosexuals. We saw an inordinate number of transfusional AIDS (4 of 29) and 1 of 46 other disorders. Two infants also presented with severe intracranial defects, one with microcephaly and one with cranial calcifications and lucency. HTLV-III is spreading with alarming speed.

Regulation of T helper cell differentiation by STAT molecules.

It is now well appreciated that the cytokines interleukin (IL)-12 and IL-4 are important for the generation of Th1 and Th2 cells, respectively. Only recently, however, have the molecular mechanisms by which these cytokines affect Th cell differentiation begun to be defined. Previous work from our laboratory has demonstrated that members of the signal transducer and activator of transcription (STAT) gene family are critical for the differentiation of Th cell subsets. In particular, Stat4-deficient mice show an impairment in the generation of Th1 cells, whereas Stat6-deficient animals do not generate Th2 cells. We have now generated Stat4-Stat6 double-deficient mice to determine whether STAT-independent pathways exist for the development of Th cell subsets. It is surprising that Th1 but not Th2 cells can be generated from double-deficient mice in vitro and these animals are able to mount an in vivo Th1 cell-mediated immune response. These results suggest a model of Th cell differentiation in which Stat4 and Stat6 have different roles in the development of Th cell subsets.

Steroids for Pneumocystis carinii pneumonia and respiratory failure in the acquired immunodeficiency syndrome. A reassessment.

Patients with acquired immunodeficiency syndrome who have Pneumocystis carinii pneumonia (PCP) and respiratory failure have a high mortality. Previous reports have suggested that corticosteroids administered in conjunction with antibiotics improve the outcome in these patients. We reviewed our experience with adjunctive corticosteroids in 20 patients with acquired immunodeficiency syndrome and respiratory failure due to PCP to determine if this was the case. Fourteen patients responded to therapy with initial reversal of their respiratory failure. However, nine of these relapsed with recurrence of respiratory failure after steroid therapy was withdrawn. Eight (40%) of the patients remained alive and well 3 months or more following treatment. When the analysis was restricted to patients requiring intubation, only 25% were alive 3 months later. Despite good initial response to steroids in PCP and respiratory failure, survival remains limited. Controlled trials are needed to define better the role of steroid treatment in these patients.

Cell-associated HIV-1 messenger RNA and DNA in T-helper cell and monocytes in asymptomatic HIV-1-infected subjects on HAART plus an inactivated HIV-1 immunogen.

OBJECTIVE: We examined the effect of an HIV-1-specific immune-based therapy on cell-associated HIV-1 DNA and RNA. DESIGN: Five HIV-1-infected subjects receiving HIV-1 immunogen plus HAART were compared with three HIV-1-infected subjects who received incomplete Freund's adjuvant (IFA) plus HAART. METHODS: Cell-associated HIV-1 RNA or DNA in lymphocytes and monocytes was determined using a dual immunophenotyping/in situ hybridization assay with or without in situ PCR amplification. RESULTS: Cell-associated HIV-1 RNA in CD4 cells correlated with plasma RNA overall. CD4, HIV-1 gag-pol messenger (m)RNA+ cells decreased in the immunogen plus HAART group compared with the IFA plus HAART group. Decreases in HIV-1 DNA+ CD4 cells were observed in the immunogen plus HAART compared with the IFA plus HAART group. Decreases in HIV-1 gag-pol mRNA+ monocytes were observed in the immunogen plus HAART group compared with the IFA plus HAART group. Consistent with the findings in CD4 cells, decreases in HIV-1 DNA+ monocytes were observed in the immunogen plus HAART group compared with the IFA plus HAART group. CONCLUSIONS: These preliminary observations support the rationale for examining the combination of immune-based therapies and antiretroviral drugs for effective HIV-1 control.

Relation of personality and attentional factors to cognitive deficits in human immunodeficiency virus-infected subjects.

In view of the evidence that patients with human immunodeficiency virus infection experience reactive depression and anxiety, it is important to determine whether these factors might account for some of the cognitive deficiencies observed in this group, as is often the case in psychiatric populations. An extensive battery of cognitive, personality, and attention tests was administered to 26 patients who tested positive for the human immunodeficiency virus. In this group were patients who demonstrated no symptoms, patients who had acquired immunodeficiency syndrome-related complex, and patients who had acquired immunodeficiency syndrome. Pearson Product Moment correlations were computed between scores on the three types of measures. The results of this correlational study suggest that cognitive decline in patients infected with human immunodeficiency virus is independent of mood and attentional changes.

RANTES and MIP-1beta mRNA expression in human peripheral blood mononuclear cells: transcript quantification using NASBA technology.

The importance of chemokines in the immune response, as well as in a range of specific disease states, is becoming increasingly apparent. The role of CC- (or beta-) chemokines and their receptors in the pathology and mechanisms of HIV-1 infection has served to intensify interest in these factors. Although the functionality of these factors resides in their protein forms, assays for the detection and quantification of these protein factors in clinical samples are not readily available. Consequently, we designed NASBA-based assays for the quantification of the mRNA encoding two members of the CC-chemokine family: RANTES and MIP-1beta. The NASBA-based assays are extremely sensitive, accurate, and reproducible across a dynamic range of at least four orders of magnitude. Inter-assay performance is comparable to intra-assay performance. We applied these methods to the analysis of normal human PBMC and PBMC from HIV-1 infected individuals. Although MIP-1beta mRNA levels are higher than RANTES levels in both populations, RANTES levels in HIV-1+ patients are higher than in normal individuals. The utility of these assays in longitudinal studies of specific subpopulations of cells, as well as their potential use in clinical diagnostics, is discussed.

Bacteremia and fungemia complicating neoplastic disease. A study of 364 cases.

During a 14 month period there were 364 episodes of bacteremia and fungemia at Memorial Sloan-Kettering Cancer Center. The first nine months of the study were retrospective, and the next five prospective. In patients with leukemia or lymphoma (group 1), Escherichia coli, Pseudomonas aeruginosa, Klebsiella pneumoniae and Staphylococcus aureus were the most frequently isolated organisms. The mortality in this group was 40.5 per cent. In the patients with solid tumor (group 2), Esch. coli, Staph. aureus, Bacteroides sp. and Candida sp. were most frequent. Mortality was 27.8 per cent. The source of infection in both groups was often indeterminate. High mortality was associated with pulmonary and intraabdominal infection and with Ps. aeruginosa, K. pneumoniae or polymicrobic sepsis. Factors of prognostic significance were the causative microorganism, source of infection and shock. Although mortality was higher in patients with leukopenia than in those with normal leukocyte counts, the differences were not significant. The mortality in this series was low considering the severity of the underlying diseases and the immunosuppressed state of many of the patients. In a prospective, randomly controlled study, mortality was further diminished by infectious disease consultation at the time the positive blood culture was reported. Severe fungal superinfection, predominantly aspergillosis and candidiasis, was found in 52 per cent of the autopsy patients with leukemia or lymphoma (group 1), but in only 8 per cent of those with solid tumors (group 2).