Heat-killed Mycobacterium Tuberculosis Research Articles

Early Deaths During Tuberculosis Treatment Are Associated With Depressed Innate Responses, Bacterial Infection, and Tuberculosis Progression

Up to 14% of Malawian adults die during the intensive phase of tuberculosis treatment. In a prospective cohort of 199 Malawian adults with microbiologically confirmed pulmonary tuberculosis, clinical and laboratory parameters were compared between those who died or deteriorated with those who had an uneventful recovery. Baseline tumor necrosis factor alpha responses to stimulation with heat-killed Mycobacterium tuberculosis and lipopolysaccharide were reduced among the 22 patients with poor outcome (P = .017). Low body mass index (P = .002) and elevated respiratory rate (P = .01) at tuberculosis diagnosis independently predicted poor outcome. Validation of a clinical score identifying high-risk individuals is warranted, together with further investigation of immunological derangements.

The Role of TLR4 Activation in Photoreceptor Mitochondrial Oxidative Stress

Herein the authors investigated whether the activation of Toll-like receptors (TLRs) in the innate immune response causes retinal photoreceptor oxidative stress and mitochondrial DNA (mtDNA) damage. On day 5 after injection of complete Freund's adjuvant containing heat-killed Mycobacterium tuberculosis (CFA), retinas were submitted to polymerase chain reaction (PCR) array focused on the TLR signaling, or apoptosis, pathway. CFA-mediated TLR4 activation, oxidative stress, and mtDNA damage were determined in B10.RIII and knockout (KO) mice (recombination activation gene [Rag] 1(KO), TLR4(KO), myeloid differentiation primary response gene 88 [MyD88](KO), tumor necrosis factor [TNF]-α(KO), or caspase 7(KO) mice) using quantitative real-time PCR, enzyme-linked immunosorbent assay, Western blot analysis, and immunohistochemistry. The mycobacterial DNA load on the retina, brain, liver, and spleen was determined by real-time PCR after intracardiac perfusion. PCR array demonstrated the upregulation of TLRs and their signaling molecules in retinas of CFA-injected mice compared with those of control animals without inflammatory cell infiltration in the retina and uvea. Mycobacterial DNA was detected in the retinas of CFA-injected mice. Retinas of CFA-injected animals showed oxidative stress and mtDNA damage, primarily in the photoreceptor inner segments. Upregulated TLR4 was localized with CD11b(+)MHCII(+) cells but not with GFAP(+) astrocytes. This oxidative stress/damage was similar in CFA-injected Rag1(KO) mice compared with wild-type controls. Such damage was absent in the retinas of CFA-injected TLR4(KO), MyD88(KO), and TNF-α(KO) mice. CFA-mediated inducible nitric oxide synthase expression in the retina was significantly decreased in TNF-α(KO) mice. Retinal photoreceptors are susceptible to mitochondrial oxidative stress/mtDNA damage in robust TLR4-mediated innate immune response.

Rapid, Cost-Effective Application of Tibilia TB Rapid Test for Culture Confirmation of Live and Heat-Killed Mycobacterium tuberculosis

Tuberculosis (TB) diagnostic laboratories face many challenges in their obligation to provide rapid, accurate reports that can help clinicians to make therapeutic decisions. One important procedure in TB diagnosis is the confirmation of an acid-fast isolate as Mycobacterium tuberculosis. This is

Nicotine‐induced differential modulation of autoimmune arthritis in the Lewis rat involves changes in interleukin‐17 and anti–cyclic citrullinated peptide antibodies

Rheumatoid arthritis (RA) is a debilitating autoimmune disease, and smoking is an important environmental factor in a subset of RA patients. A role of the cholinergic antiinflammatory pathway in autoimmune inflammation is increasingly being realized. Nicotine is a major component of cigarette smoke, and it stimulates the α7 nicotinic acetylcholine receptors. Therefore, defining the mechanisms underlying the immunomodulatory effects of nicotine on arthritis is of high relevance. The purpose of this study was to address this issue using the rat adjuvant-induced arthritis (AIA) model of human RA. Lewis rats were immunized subcutaneously with heat-killed Mycobacterium tuberculosis H37Ra for disease induction. Rats were treated with nicotine intraperitoneally either before (pretreatment) or after (posttreatment) the onset of AIA. Control rats received the vehicle (buffer) in place of nicotine. The severity of arthritis was assessed and graded. The draining lymph node cells were tested for T cell proliferative and cytokine responses against the disease-related antigen mycobacterial heat-shock protein 65. The sera were tested for anti-cyclic citrullinated peptide (anti-CCP) antibodies and anti-mycobacterial Hsp65 antibodies. Nicotine pretreatment aggravated the arthritis, whereas nicotine posttreatment suppressed the disease. This altered severity of AIA directly correlated with the levels of the anti-CCP antibodies, of the Th1/Th17 cytokines, and of the corresponding dendritic cell-derived cytokines. The majority of these effects on cellular responses could be replicated in vitro. Nicotine-induced modulation of AIA involves specific alterations in the disease-related cellular and humoral immune responses in AIA. These results are of significance in advancing our understanding of the pathogenesis of RA.

A GROUP OF LOW MOLECULAR WEIGHT (

Heat-killed Mycobacterium tuberculosis (Mtb) H37Rv–derived antigens were purified by electroelution, and their efficacy was evaluated in conferring protection against Tb in guinea pigs in order to identify those protein antigens as a subunit vaccine. A total sonicate extract of Mtb H37Rv (MTSE) was prepared by growing bacteria on Lowenstein–Jensen (LJ) solid medium to its mid-log phase, harvested, and subjected to ultrasonication. Fractions of MTSE were resolved on 10% polyacrylamide gel electrophoresis into as high (HMW, >60 kDa), middle (MMW, 31-59 kDa), and low molecular weight (LMW, <30 kDa) antigens and were purified by electroelution. The protection efficacy of each purified fraction was assessed in a placebo-controlled study by injecting them separately into a group of two animals. After two booster doses, animals were challenged on day 84, and autopsy of all animals was performed on day 126. On autopsy, animals were scored for formation of tubercles in lungs and their dissemination to organs such as spleen, liver, kidney, and lymph nodes. Viable counts (determined as colony forming unit or CFU) of lung and spleen from each animal were performed on LJ solid medium. Results of this study show that CFU of animals that received LMW antigens were significantly low with respect to control animals when compared to MMW and HMW. In conclusion, LMW of Mtb has greater potential in prevention of tubercle formation and thus could be viewed as a possible vaccine candidate in designing alternative subunit vaccines.

Hypercytokinaemia accompanies HIV–tuberculosis immune reconstitution inflammatory syndrome

Increased access to combination antiretroviral therapy in areas co-endemic for tuberculosis (TB) and HIV-1 infection is associated with an increased incidence of immune reconstitution inflammatory syndrome (TB-IRIS) whose cause is poorly understood. A case-control analysis of pro- and anti-inflammatory cytokines in TB-IRIS patients sampled at clinical presentation, and similar control patients with HIV-TB prescribed combined antiretroviral therapy who did not develop TB-IRIS. Peripheral blood mononuclear cells were cultured in the presence or absence of heat-killed Mycobacterium tuberculosis for 6 and 24 h. Stimulation with M. tuberculosis increased the abundance of many cytokine transcripts with interleukin (IL)-1β, IL-5, IL-6, IL-10, IL-13, IL-17A, interferon (IFN)-γ, granulocyte-macrophage colony-stimulating factor (GM-CSF) and tumour necrosis factor (TNF) being greater in stimulated TB-IRIS cultures. Analysis of the corresponding proteins in culture supernatants, revealed increased IL-1β, IL-2, IL-6, IL-8, IL-10, IL-12p40, IFN-γ, GM-CSF and TNF in TB-IRIS cultures. In serum, higher concentrations of TNF, IL-6, and IFN-γ were observed in TB-IRIS patients. Serum IL-6 and TNF decreased during prednisone therapy in TB-IRIS patients. These data suggest that cytokine release contributes to pathology in TB-IRIS. IL-6 and TNF were consistently elevated and decreased in serum during corticosteroid therapy. Specific blockade of these cytokines may be rational approach to immunomodulation in TB-IRIS.

Anti-arthritic activities of Annona muricata L. leaves extract on complete Freund's adjuvant (CFA) – induced arthritis in rats

Annona muricata L. (Annonaceae) (AML) has been used as folkloric herbal medicine in many regions throughout the world in treating fever, cough, diarrhea, sedative, rash, ring worm and lactation for women afterbirth (1). The present study was carried out to investigate the anti-arthritic effects of AML ethanolic extract in complete Freund's adjuvant (CFA)-induced arthritis in rats. Edema in the left hind paw of rat was induced by the subcutaneous injection of 0.1ml of heat-killed Mycobacterium tuberculosis in mineral oil (5mg/ml). AML (3, 10, 30 and 100mg/kg) or indomethacin (10mg/kg) as positive control was administered orally once a day between days 1 and 14 post-CFA injection. Edema volume in each paw was measured by using a plethysmometer for 14 days. Levels of tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1β) in local tissue were measured using enzyme-linked immunosorbent assay (ELISA) on day 14 post-CFA injection. The result exhibited that AML at all doses (3, 10, 30 and 100mg/kg) were significantly (p&lt;0.05) reduced the edema by 48.39, 66.67, 79.57 and 72.04% respectively. AML at higher doses 30 and 100mg/kg significantly suppressed the local tissue TNF-α level by 42.81 and 51.82% and IL-1β level by 35.57 and 39.79% respectively on day 14 post-CFA injection. For TNF-α, the efficacies of the extract at doses 30 and 100mg/kg were higher compared to the effect of indomethacin (51.82%). In conclusion, the findings suggest that AML present notable anti-arthritic activities that may be mediated by suppressing pro-inflammatory cytokines.

Interleukin-25 Expressed by Brain Capillary Endothelial Cells Maintains Blood-Brain Barrier Function in a Protein Kinase Cϵ-dependent Manner

Interleukin (IL)-25, a member of the IL-17 family of cytokines, is expressed in the brains of normal mice. However, the cellular source of IL-25 and its function in the brain remain to be elucidated. Here, we show that IL-25 plays an important role in preventing infiltration of the inflammatory cells into the central nervous system. Brain capillary endothelial cells (BCECs) express IL-25. However, it is down-regulated by inflammatory cytokines, including tumor necrosis factor (TNF)-alpha, IL-17, interferon-gamma, IL-1beta, and IL-6 in vitro, and is also reduced in active multiple sclerosis (MS) lesions and in the inflamed spinal cord of experimental autoimmune encephalomyelitis, an animal model of MS. Furthermore, IL-25 restores the reduced expression of tight junction proteins, occludin, junction adhesion molecule, and claudin-5, induced by TNF-alpha in BCECs and consequently repairs TNF-alpha-induced blood-brain barrier (BBB) permeability. IL-25 induces protein kinase Cepsilon (PKCepsilon) phosphorylation, and up-regulation of claudin-5 is suppressed by PKCepsilon inhibitor peptide in the IL-25-stimulated BCECs. These results suggest that IL-25 is produced by BCECs and protects against inflammatory cytokine-induced excessive BBB collapse through a PKCepsilon-dependent pathway. These novel functions of IL-25 in maintaining BBB integrity may help us understand the pathophysiology of inflammatory brain diseases such as MS.

Studies on the activating capacity of heat killed M. tuberculosis and its culture filtrate proteins on polymorphonuclear neutrophils and peripheral mononuclear cells from tuberculosis patients

The efficacy of heat-killed Mycobacterium tuberculosis (HKMtb) and its culture filtrate proteins (CFP) to activate blood mononuclear cells (MC) and polymorphonuclear neutrophils (PMN) from tuberculosis patients was investigated. Respiratory burst, NO-derived metabolites, IL-2, IL-12 and TNF-α production of stimulated cells from 16 HIV- tuberculosis patients and 12 healthy controls were analyzed. Increased amounts of TNF-α in supernatants from baseline and stimulated polymorphonuclear and mononuclear cells of tuberculosis patients were detected when compared with controls, except for CFP stimulated neutrophils. Augmented IL-2 and IL-12 levels were observed in supernatants of both stimulated MC and PMN from TBP while no differences were found in control supernatants. The patients had a lower respiratory burst response than the controls, for both cell types, regardless of the stimulus employed. Higher nitrite concentrations were found in HKMtb- and CFP-stimulated mononuclear supernatants from patients, compared with controls.The obtained data of the stimulated cellular responses provides us information about the protective immunity against Mycobacterium tuberculosis and some resources to obtain a more efficient anti-tuberculous therapy.

Role of IL-23 in mobilization of immunoregulatory nitric oxide- or superoxide-producing Gr-1 + cells from bone marrow

Spleens of mice injected with heat-killed Mycobacterium tuberculosis increase their Gr-1 + cell content and develop a system of interactive Ly-6G + and Ly-6G −Gr-1 + populations or “G reg ” subsets, which, upon stimulation by activated T cells, produce immunoregulatory superoxide (O 2 −) and nitric oxide (NO), respectively. The balance between immunosuppressive NO and its antagonist O 2 − regulates T cell expansion, similar to regulation of vasodilation. Reduction of NO levels by O 2 − is required for efficient T cell expansion and development of autoimmunity. We studied the source of Gr-1 + cells in bone marrow (BM), where their levels were higher than in spleen, with both G reg subsets expressing strong activity. In the spleens of primed IL-23 −/− mice, Ly-6G + cells remained at naïve levels and produced no O 2 −. The complementary Ly-6G −Gr-1 + splenocytes and their suppressive activity were partially reduced. Surprisingly, Gr-1 + cell levels in BM of IL-23 −/− mice were increased, as were their O 2 − and NO production. Transfer of primed BM cells partially restored regulatory function in the spleen of IL-23 −/− recipients. The results suggest that IL-23 is involved in mobilization of O 2 −- and NO-producing Gr-1 + cells from BM, which may contribute to its widely studied role in (auto)immunity.

Huo-Luo-Xiao-Ling Dan modulates antigen-directed immune response in adjuvant-induced inflammation

Ethnopharmacological relevance HLXL is a traditional Chinese medicine that has long been used in folk medicine for the treatment of chronic inflammatory diseases. However, the precise immunological mechanisms by which HLXL mediates its anti-inflammatory activity are not fully defined. Aim of the study To determine the effects of HLXL on antigen-specific immune parameters in adjuvant-induced inflammation model in the Lewis rat. Materials and methods Rats were fed daily with either HLXL (2.3 g/kg) or vehicle (water) beginning 3 days before subcutaneous injection of heat-killed Mycobacterium tuberculosis H37Ra (Mtb), and then continued for another 6 days. After 9 days of Mtb injection, the draining lymph node cells were tested for T cell proliferative and cytokine responses against mycobacterial heat-shock protein 65 (Bhsp65). Moreover, sera were tested for anti-Bhsp65 antibodies and nitric oxide (NO). Results HLXL-treated rats showed reduced T cell proliferative response to Bhsp65 compared to control rats. Furthermore, HLXL suppressed IL-17 response but enhanced IL-10 response without much effect on IFN-γ. HLXL treatment also reduced the levels of anti-Bhsp65 antibodies but not that of NO. Conclusions HLXL feeding modulated both the cellular and the humoral immune response to Bhsp65 favoring an anti-inflammatory milieu for the suppression of adjuvant-induced inflammation.

Proinflammatory role of the Th17 cytokine interleukin‐22 in collagen‐induced arthritis in C57BL/6 mice

To investigate the role of interleukin-22 (IL-22) in collagen-induced arthritis (CIA), an animal model of rheumatoid arthritis. C57BL/6 mice were immunized with type II collagen (CII) in Freund's incomplete adjuvant with added Mycobacterium tuberculosis, and levels of IL-22 and its specific receptor, IL-22 receptor type I (IL-22RI), were measured in sera and tissue by enzyme-linked immunosorbent assay and real-time quantitative polymerase chain reaction analysis. Clinical and histologic signs of arthritis were recorded and compared with those in C57BL/6 mice deficient in the IL-22 gene (IL-22(-/-)). Humoral and cellular immune responses against CII were analyzed. In vitro osteoclastogenesis assays were performed on splenocytes. Upon immunization with CII in Freund's incomplete adjuvant plus heat-killed Mycobacterium tuberculosis, sera from C57BL/6 mice were found to contain high levels of IL-22, and the specific IL-22RI was expressed in lymphoid tissue, including splenocytes. IL-22(-/-) mice were less susceptible to CIA than were wild-type mice, as evidenced by their decreased incidence of arthritis and decreased pannus formation. Remarkably, the less severe form of arthritis in IL-22(-/-) mice was associated with increased production of CII-specific and total IgG antibodies, whereas cellular CII responses were unchanged. In vitro, IL-22 was found to promote osteoclastogenesis, a process that might contribute to its proinflammatory activity in CIA. Endogenous IL-22 plays a proinflammatory role in CIA in C57BL/6 mice. Our data also indicate that IL-22 promotes osteoclastogenesis and regulates antibody production.

Tolerization with Hsp65 induces protection against adjuvant‐induced arthritis by modulating the antigen‐directed interferon‐γ, interleukin‐17, and antibody responses

Pretreatment of Lewis rats with soluble mycobacterial Hsp65 affords protection against subsequent adjuvant-induced arthritis (AIA). This study was aimed at unraveling the mechanisms underlying mycobacterial Hsp65-induced protection against arthritis, using contemporary parameters of immunity. Lewis rats were given 3 intraperitoneal injections of mycobacterial Hsp65 in solution prior to the initiation of AIA with heat-killed Mycobacterium tuberculosis. Thereafter, mycobacterial Hsp65-specific T cell proliferative, cytokine, and antibody responses were tested in tolerized rats. The roles of anergy and the indoleamine 2,3 dioxygenase (IDO)-tryptophan pathway in tolerance induction were assessed, and the frequency and suppressive function of CD4+FoxP3+ Treg cells were monitored. Also tested was the effect of mycobacterial Hsp65 tolerization on T cell responses to AIA-related mycobacterial Hsp70, mycobacterial Hsp10, and rat Hsp65. The AIA-protective effect of mycobacterial Hsp65-induced tolerance was associated with a significantly reduced T cell proliferative response to mycobacterial Hsp65, which was reversed by interleukin-2 (IL-2), indicating anergy induction. The production of interferon-gamma (but not IL-4/IL-10) was increased, with concurrent down-regulation of IL-17 expression by mycobacterial Hsp65-primed T cells. Neither the frequency nor the suppressive activity of CD4+FoxP3+ T cells changed following tolerization, but the serum level of anti-mycobacterial Hsp65 antibodies was increased. However, no evidence was observed for a role of IDO or cross-tolerance to mycobacterial Hsp70, mycobacterial Hsp10, or rat Hsp65. Tolerization with soluble mycobacterial Hsp65 leads to suppression of IL-17, anergy induction, and enhanced production of anti-mycobacterial Hsp65 antibodies, which play a role in protection against AIA. These results are relevant to the development of effective immunotherapeutic approaches for autoimmune arthritis.

Extract of the Chinese herbal formula Huo Luo Xiao LingDan inhibited adjuvant arthritis in rats

Ethnopharmacological relevance The herbal formula Huo Luo Xiao Ling Dan (HLXL) and its modifications have been used in traditional Chinese medicine for about one hundred years to alleviate pain and inflammation. Aim To investigate the effects of HLXL on complete Freund's adjuvant (CFA)-induced multiple-joint arthritis in rats. Materials and methods Male Lewis rats, 190–210 g, were immunized subcutaneously at the base of the tail with 200 μl of heat-killed Mycobacterium tuberculosis in mineral oil (5 mg/ml). HLXL (2.30 and 4.60 g/kg) or vehicle control ( n = 8 per group) was administered orally (i.g.) once a day between days 16 and 25 post-CFA injection. The rats were observed for signs of arthritis with arthritic changes (erythema, edema, induration) being scored on a scale of 0–4 of increasing severity using a standard scoring system. The maximum arthritis score per rat was 16. A plethysmometer was used to measure edema volume in each paw. Adverse effects of HLXL were monitored by closely observing the animals for unusual behavioral changes. Levels of tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1β) in local tissue were measured by enzyme-linked immunosorbent assay on day 25 post-CFA. Results HLXL significantly decreased arthritis scores between days 23–25 in the 2.30 g/kg group and 21–25 in the 4.60 g/kg group ( p < 0.05). It reduced paw edema on days 22 and 24 in the 2.30 g/kg group and on days 20, 22 and 24 in the 4.60 g/kg group compared to control ( p < 0.05). Local tissue TNF-α and IL-1β levels on day 25 post-CFA injection were significantly ( p < 0.05) lower in rats treated with HLXL than in control rats. No observable adverse effects were found. Conclusion The data suggest that HLXL produces significant anti-arthritic effects that may be mediated by suppressing pro-inflammatory cytokines, and it appears to be safe.

Green Tea Protects Rats against Autoimmune Arthritis by Modulating Disease-Related Immune Events

Green tea, a product of the dried leaves of Camellia sinensis, is the most widely consumed beverage in the world. The polyphenolic compounds from green tea (PGT) possess antiinflammatory properties. We investigated whether PGT can afford protection against autoimmune arthritis and also examined the immunological basis of this effect using the rat adjuvant arthritis (AA) model of human rheumatoid arthritis (RA). AA can be induced in Lewis rats (RT.1 l) by immunization with heat-killed Mycobacterium tuberculosis H37Ra (Mtb), and arthritic rats raise a T cell response to the mycobacterial heat-shock protein 65 (Bhsp65). Rats consumed green tea (2–12 g/L) in drinking water for 1–3 wk and then were injected with Mtb to induce disease. Thereafter, they were observed regularly and graded for signs of arthritis. Subgroups of these rats were killed at defined time points and their draining lymph node cells were harvested and tested for T cell proliferative and cytokine responses. Furthermore, the sera collected from these rats were tested for anti-Bhsp65 antibodies. Feeding 8 g/L PGT to Lewis rats for 9 d significantly reduced the severity of arthritis compared with the water-fed controls. Interestingly, PGT-fed rats had a lower concentration of the proinflammatory cytokine interleukin (IL)-17 but a greater concentration of the immunoregulatory cytokine IL-10 than controls. PGT feeding also suppressed the anti-Bhsp65 antibody response. Thus, green tea induced changes in arthritis-related immune responses. We suggest further systematic exploration of dietary supplementation with PGT as an adjunct nutritional strategy for the management of RA.

Antibody-mediated demyelination in experimental allergic encephalomyelitis is independent of complement membrane attack complex formation

The effects of decomplementation by cobra venom factor (CVF) on the pathogenesis of inflammation and demyelination in experimental allergic encephalomyelitis (EAE) and acute antibody-mediated demyelinating EAE (ADEAE) have been quantified histologically and immunocytochemically. In rats immunized with 50 micrograms of myelin basic protein in Freund's complete adjuvant containing 100 micrograms heat-killed Mycobacterium tuberculosis H37Ra, clinical signs of EAE were completely suppressed by two injections of CVF given 9 and 12 days post-immunization. Suppression of clinical disease was associated with a dramatic reduction in peri-vascular inflammation in the CNS, although immunohistochemical staining identified small numbers of infiltrating T cells and macrophages. In contrast, CVF treatment had no significant effect on the clinical severity of ADEAE and although C9 deposition within the CNS was virtually abolished, there was no statistically significant decrease in the extent of demyelination or inflammation. These observations indicate that in the absence of complement components C3 and C5 an antibody-dependent cell-mediated cytotoxic response plays an important role in the pathogenesis of antibody-mediated demyelination. The major role of the complement cascade in EAE appears to be the generation of pro-inflammatory factors that enhance the inflammatory response within the CNS in animals facing a mild encephalitogenic challenge.

The Dynamics of Articular Leukocyte Trafficking and the Immune Response to Self Heat-Shock Protein 65 Influence Arthritis Susceptibility

Adjuvant arthritis (AA) shares several features with human rheumatoid arthritis, and it can be induced in the Lewis (LEW) rat but not the Wistar Kyoto (WKY) rat (both RT.1(l)) by immunization with heat-killed Mycobacterium tuberculosis (Mtb). We set out to unravel the mechanisms underlying the differential susceptibility to AA of these MHC-compatible rat strains. We compared the levels of T-cell proliferative and cytokine response to the immunoregulatory self (rat) hsp65 (Rhsp65) after an arthritogenic (Mtb) challenge and the kinetics of migration of adoptively transferred, (111)Indium-labeled, Mtb-primed leukocytes into the hind paw joints of recipient rats. The WKY rats raised a significantly higher level of T-cell proliferative response coupled with a temporally opposite cytokine profile against the disease-regulating Rhsp65 compared to that of LEW rats. Moreover, the arthritogenic leukocytes accumulated into the joints of WKY rats at significantly lower numbers than that in LEW rats. These results offer novel insights into the immune events influencing the pathogenesis of autoimmune arthritis.

Exogenous tumour necrosis factor α induces suppression of autoimmune arthritis

IntroductionOur previous studies showed that arthritic Lewis (LEW) rats produced the highest levels of tumour necrosis factor (TNF)α in the recovery phase of adjuvant arthritis (AA), suggesting a correlation between high TNFα levels and reduced severity of arthritis. To further explore this correlation, we compared the TNFα secretion profile of the AA-resistant Wistar Kyoto (WKY) rats with that of LEW rats, determined the effect of exogenous TNFα on the course of AA in LEW rats, and examined various mechanisms involved in TNFα-induced disease modulation.MethodsA cohort each of LEW and WKY rats was immunised subcutaneously with heat-killed Mycobacterium tuberculosis H37Ra (Mtb). At different time points thereafter, subgroups of rats were killed and their draining lymph node cells were tested for cytokine production. Another group of LEW rats was injected with TNFα intraperitoneally daily for a total of 10 injections, 3 before and 6 after Mtb challenge, and then observed for signs of AA. In parallel, TNFα-treated rats were examined for changes in other cytokines, in CD4+CD25+ T cell frequency, and in indoleamine 2,3-dioxygenase (IDO) mRNA expression levels.ResultsLEW rats displayed a TNFα secretion profile that was opposite to that of the WKY rats. Furthermore, TNFα treatment significantly downmodulated the severity of AA in LEW rats, and decreased the interferon (IFN)-γ secretion in response to the pathogenic determinant of the disease-related antigen. No significant alterations were observed in other parameters tested.ConclusionThe role of endogenous TNFα in the induction and propagation of arthritis is well established. However, exogenous TNFα can downmodulate the course of AA, displaying an immunoregulatory functional attribute of this cytokine.

Differential activation of alveolar and peritoneal macrophages from BCG-vaccinated guinea pigs

We compared the effect of BCG vaccination on the mRNA expression of two prototypic cytokines, IL-12 (Type 1) and IL-10 (Type 2), in guinea pig resident alveolar macrophages (AM) or resident peritoneal macrophages (PM). Cells were stimulated with live or heat-killed Mycobacterium tuberculosis, and/or with recombinant guinea pig (rgp) TNF-alpha and/or rgp IFN-gamma. AM from BCG-vaccinated guinea pigs expressed significantly less IL-10 mRNA and more IL-12p40 mRNA compared to AM from naive animals following stimulation with heat-killed mycobacteria. In PM from BCG-vaccinated guinea pigs, IL-12p40 mRNA was significantly up-regulated; however, the level of IL-10 mRNA was not affected by prior vaccination. rgp TNF-alpha or rgp IFN-gamma, both alone and together, induced a significant increase of H(2)O(2) production in PM from BCG-vaccinated animals. MHC class II expression was dramatically up-regulated in PM from BCG-vaccinated animals stimulated with both rgp TNF-alpha and rgp IFN-gamma. The levels of IL-10 and IL-12p40 mRNA were significantly enhanced in PM stimulated with combinations of rgp TNF-alpha and rgp IFN-gamma, and those cells suppressed the intracellular accumulation of viable, virulent M. tuberculosis. BCG vaccination results in the differential activation of guinea pig AM and PM to promote a Type 1 cytokine milieu and control intracellular mycobacteria.

Low frequency and low intensity pulsed electromagnetic field exerts its antiinflammatory effect through restoration of plasma membrane calcium ATPase activity

Rheumatoid arthritis (RA) is a chronic inflammatory disorder affecting 1% of the population worldwide. Pulsed electromagnetic field (PEMF) has a number of well-documented physiological effects on cells and tissues including antiinflammatory effect. This study aims to explore the antiinflammatory effect of PEMF and its possible mechanism of action in amelioration of adjuvant induced arthritis (AIA). Arthritis was induced by a single intradermal injection of heat killed Mycobacterium tuberculosis at a concentration of 500 μg in 0.1 ml of paraffin oil into the right hind paw of rats. The arthritic animals showed a biphasic response regarding changes in the paw edema volume. During the chronic phase of the disease, arthritic animals showed an elevated level of lipid peroxides and depletion of antioxidant enzymes with significant radiological and histological changes. Besides, plasma membrane Ca 2+ ATPase (PMCA) activity was inhibited while intracellular Ca 2+ level as well as prostaglandin E 2 levels was noticed to be elevated in blood lymphocytes of arthritic rats. Exposure of arthritic rats to PEMF at 5 Hz × 4 μT × 90 min, produced significant antiexudative effect resulting in the restoration of the altered parameters. The antiinflammatory effect could be partially mediated through the stabilizing action of PEMF on membranes as reflected by the restoration of PMCA and intracellular Ca 2+ levels in blood lymphocytes subsequently inhibiting PGE 2 biosynthesis. The results of this study indicated that PEMF could be developed as a potential therapy for RA in human beings.