Regulation of intestinal immunity and tissue repair by enteric glia.

Tissue maintenance and repair depend on the integrated activity of multiple cell types1. Whereas the contributions of epithelial2,3, immune4,5 and stromal cells6,7 in intestinal tissue integrity are well understood, the role of intrinsic neuroglia networks remains largely unknown. Here we uncover important roles of enteric glial cells (EGCs) in intestinal homeostasis, immunity and tissue repair. We demonstrate that infection of mice with Heligmosomoides polygyrus leads to enteric gliosis and the upregulation of an interferon gamma (IFNγ) gene signature. IFNγ-dependent gene modules were also induced in EGCs from patients with inflammatory bowel disease8. Single-cell transcriptomics analysis of the tunica muscularis showed that glia-specific abrogation of IFNγ signalling leads to tissue-wide activation of pro-inflammatory transcriptional programs. Furthermore, disruption of the IFNγ-EGC signalling axis enhanced the inflammatory and granulomatous response of the tunica muscularis to helminths. Mechanistically, we show that the upregulation of Cxcl10 is an early immediate response of EGCs to IFNγ signalling and provide evidence that this chemokine and the downstream amplification of IFNγ signalling in the tunica muscularis are required for a measured inflammatory response to helminths and resolution of the granulomatous pathology. Our study demonstrates that IFNγ signalling in enteric glia is central to intestinal homeostasis and reveals critical roles of the IFNγ-EGC-CXCL10 axis in immune response and tissue repair after infectious challenge.

Impact of diabetes on mechanisms of immunity against Mycobacterium tuberculosis.

Tuberculosis (TB), caused by infection with mycobacterium tuberculosis, is a major source of morbidity and mortality in Pakistan. Diabetes caused by imbalance in glycaemic control is also highly prevalent in the country. The coincidence of both diseases results in worsening outcomes of TB, making treatment and management more difficult. Both innate and adaptive arms of the host immune response are required for protection against M. tuberculosis infection. Host immunity is modified in diabetes mellitus type 2 where key pathways such as, the T cell driven interferon-gamma responses to M. tuberculosis antigens and other T cell and macrophage activating cytokines are suppressed. This makes diabetes with TB a more severe disease and results in worse treatment outcomes. Effective coordination between T cells and host macrophages is required for control of TB infection. Therefore, early identification of diabetes and management of hyperglycaemia during TB treatment is essential for favourable outcomes.

Role of TNF α, IL-6 and CXCL10 in Dengue disease severity.

BACKGROUND AND OBJECTIVES: Dengue virus infections (Dengue) have become increasingly common in Pakistan and can result in case fatalities if not managed appropriately. Patients with Dengue virus infection may be asymptomatic or present with Dengue fever (DF), Dengue with warning signs (DWS) or severe Dengue (SD). Severity in Dengue is coincident with an exacerbated production of lymphocyte-induced cytokines and chemokines which are associated with plasma leakage. We investigated the association of circulating levels of cytokines such as Interleukin (IL)-6, tumor necrosis factor (TNF)-alpha and CXCL-10 in Dengue patients with differing severity of disease. MATERIALS AND METHODS: Dengue infection was confirmed by testing for human IgM to the Dengue virus. Dengue patients (n=58) and healthy controls (n=33) were recruited. Dengue patients were grouped into those with DF (n=39), DWS (n=15) and SD (n=4). Serum IL-6, TNFα and CXCL10 levels were tested by ELISA. The Mann Whitney U test was used for statistical analysis. RESULTS: Circulating levels of TNFα (p≤0.001) and CXCL10 (p≤0.001) levels were increased in Dengue patients as compared with controls. When patients were stratified for disease severity, it was observed that CXCL10 was increased in DWS as compared to DF (p=0.046). IL-6 levels were increased in patients with SD as compared to those with DWS (p=0.044). TNFα levels were not found to differ between different groups of Dengue patients. CONCLUSION: Raised CXCL10 and TNFα levels were associated with increased clinical severity of Dengue infection and probably increased disease progression due to excessive inflammation and increased vascular changes in the patients.

Increasing usage of rapid diagnostics for Dengue virus detection in Pakistan.

OBJECTIVE: To evaluate the trends in usage of dengue virus diagnostics in Pakistan. METHODS: This retrospective study was conducted at the Aga Khan University Hospital, Karachi, and comprised data for specimens tested for dengue virus from January 2012 to December 2015. Test for dengue virus ribonucleic acid by reverse transcription polymerase chain reaction, dengue virus antigen by immunochromatic assay and for human immunoglobulin M against dengue virus by enzyme-linked immunosorbent assay were reviewed. SPSS 17 was used for data analysis. RESULTS: Overall, 33,577 specimens tested for dengue virus. Of them, 11,995 (35.7%) were positive. among them, 1,039(8.66%) were reported in 2012; 5,791(48.28%) in 2013; 1,027(8.56%) in 2014; and 4,138(34.49%) in 2015. In 2012, 966(93%) of the positive samples were diagnosed by immunoglobulin M-based method and 73(7%) by non-structural protein-1 antigen. In 2013, 4,401(76%) samples were tested positive by immunoglobulin M, 1,332(23%) by antigen and 58(1%) by polymerase chain reaction. The trend continued in 2014, but in 2015, 2,111(51%) of all dengue positive tests were determined by antigen testing, 1,969(47.6%) by immunoglobulin M and 58(1.4%) by polymerase chain reaction. CONCLUSIONS: There was a shift in usage of direct virus identification for rapid diagnosis of dengue virus compared with host immunoglobulin M testing.

Increased Mycobacterium tuberculosis antigen-induced gene expression of interferon-gamma, tumor necrosis factor alpha and interleukin-6 in patients with diabetes.

INTRODUCTION: Pakistan ranks fifth in high tuberculosis (TB)-burden countries and seventh among countries with high prevalence rates of diabetes mellitus (DM). DM is a risk factor for TB and worsens disease outcomes. Furthermore, Mycobacterium tuberculosis (MTB) infection can induce glucose intolerance and worsen glycemic control in diabetes. Suppressor of cytokine signaling (SOCS)-1 and -3 molecules regulate cytokine signaling and are important in maintaining an immune balance. In TB, interleukin (IL)-6 upregulation induces SOCS3, which is also a negative regulator of insulin signaling. This research focuses on the mechanism by which SOCS1 and SOCS3 affect insulin resistance and increased susceptibility to TB. METHODS: We studied gene expression in peripheral blood cells of patients with diabetes (n=10) and healthy endemic controls (EC, n=11) both with and without MTB infection. Mycobacterial antigen (PPD) and mitogen-stimulated SOCS1, SOCS3, interferon-gamma (IFN-γ), IL-6, and tumor necrosis factor alpha (TNFα) mRNA expression levels were determined using real-time polymerase chain reaction. RESULTS: MTB antigen-stimulated mRNA levels of IFN-γ was 10-fold higher, SOCS1 was 4 times greater, TNFα was 10-fold higher, and IL-6 was 2-fold greater in patients with DM than in ECs. Overall levels of PPD-stimulated IL-6 was higher in patients with DM than in ECs (p=.036). Mitogen-induced mRNA levels of IFN-γ were 30-fold higher, SOCS3 was 20 fold higher, and SOCS1 was 4-fold higher in patients with DM than in ECs. CONCLUSION: Increased proinflammatory cytokine production in response to MTB antigens in diabetes would lead to exacerbated pathology and reduced inflammatory control at the site of MTB infection. This would in turn hamper the resolution of inflammation, resulting in unfavorable disease outcomes.