Mycobacterium tuberculosis lipoarabinomannan-mediated IRAK-M induction negatively regulates toll-like receptor-dependent interleukin-12 p40 production in macrophages

Mannose-capped lipoarabinomannans (Man-LAMs) are members of the repertoire of Mycobacterium tuberculosis modulins that the bacillus uses to subvert the host innate immune response. Interleukin-12 (IL-12) production is critical for mounting an effective immune response by the host against M. tuberculosis. We demonstrate that Man-LAM inhibits IL-12 p40 production mediated by subsequent challenge with lipopolysaccharide (LPS). Man-LAM inhibits LPS-induced IL-12 p40 expression in an IL-10-independent manner. It attenuates LPS-induced NF-κB-driven luciferase gene expression, suggesting that its effects are likely directly related to inhibition of NF-κB. This is probably because of dampening of the Toll-like receptor signaling. Man-LAM inhibits IL-1 receptor-associated kinase (IRAK)-TRAF6 interaction as well as IκB-α phosphorylation. It directly attenuates nuclear translocation and DNA binding of c-Rel and p50. Man-LAM exerts these effects by inducing the expression of Irak-M, a negative regulator of TLR signaling. Knockdown of Irak-M expression by RNA interference reinstates LPS-induced IL-12 production in Man-LAM-pretreated cells. The fact that Irak-M expression could be elicited by yeast mannan suggested that ligation of the mannose receptor by the mannooligosaccharide caps of LAM was the probable trigger for IRAK-M induction

Helicobacter pylori protein HPO175 transactivates epidermal growth factor receptor through TLR4 in gastric epithelial cells

The pathophysiology of Helicobacter pylori-associated gastroduodenal diseases, ulcerogenesis, and carcinogenesis is intimately linked to activation of epidermal growth factor receptor (EGFR) and production of vascular endothelial growth factor (VEGF). Extracellular virulence factors, such as CagA and VacA, have been proposed to regulate EGFR activation and VEGF production in gastric epithelial cells. We demonstrate that the H. pylori secretory protein, HP0175, by virtue of its ability to bind TLR4, transactivates EGFR and stimulates EGFR-dependent VEGF production in the gastric cancer cell line AGS. Knock-out of the hp0175 gene attenuates the ability of the resultant H. pylori strain to activate EGFR or to induce VEGF production. HP0175-induced activation of EGFR is preceded by translocation of TLR4 into lipid rafts. In lipid rafts, the Src kinase family member Lyn interacts with TLR4, leading to tyrosine phosphorylation of TLR4. Knockdown of Lyn prevents HP0175-induced activation of EGFR and VEGF production. Tyrosine-phosphorylated TLR4 interacts with EGFR. This interaction is necessary for the activation of EGFR. Disruption of lipid rafts with methyl β-cyclodextrin prevents HP0175-induced tyrosine phosphorylation of TLR4 and activation of EGFR. This mechanism of transactivation of EGFR is novel and distinct from that of metalloprotease-dependent shedding of EGF-like ligands, leading to autocrine activation of EGFR. It provides new insight into our understanding of the receptor cross-talk network

Toll-like receptor 2 and mitogen- and stress-activated kinase 1 are effectors of Mycobacterium avium-induced cyclooxygenase-2 expression in macrophages

Understanding how pathogenic mycobacteria subvert the protective immune response is crucial to the development of strategies aimed at controlling mycobacterial infections. Prostaglandin E2 exerts an immunosuppressive function in the context of mycobacterial infection. Because cyclooxygenase-2 (COX-2) is a rate-limiting enzyme in prostaglandin biosynthesis, there is a need to delineate the mechanisms through which pathogenic mycobacteria regulate COX-2 expression in macrophages. Our studies demonstrate that the NF-κB and CRE elements of the COX-2 promoter are critical to Mycobacterium avium-induced COX-2 gene expression. M. avium-triggered signaling originates at the Toll-like receptor 2 (TLR2). Ras associates with TLR2 and activates the mitogen-activated protein kinase (MAPK) extracellular signal-regulated kinase (ERK), whereas tumor necrosis factor receptor-associated factor 6 (TRAF6)/transforming growth factor β -activated kinase 1 (TAK1)-dependent signaling activates p38 MAPK. Both ERK and p38 MAPK activation converge to regulate the activation of mitogen- and stress-activated kinase 1 (MSK1). MSK1 mediates the phosphorylation of the transcription factor CREB accounting for its stimulatory effect on CRE-dependent gene expression. M. avium-triggered cytoplasmic NF-κB activation following IκB phosphorylation is necessary but not sufficient for COX-2 promoter-driven gene expression. MSK1 activation is also essential for M. avium-triggered NF-κB-dependent gene expression, presumably mediating nucleosomal modifications. These studies demonstrate that the nuclear kinase MSK1 is necessary in regulating the pathogen-driven expression of a gene by controlling two transcription factors. The attenuation of MSK1 may therefore have potential benefit in restricting survival of pathogenic mycobacteria in macrophages

Execution of macrophage apoptosis by PE_PGRS33 of Mycobacterium tuberculosis is mediated by toll-like receptor 2-dependent release of tumor necrosis factor-α

Combating tuberculosis requires a detailed understanding of how mycobacterial effectors modulate the host immune response. The role of the multigene PE family of proteins unique to mycobacteria in the pathogenesis of tuberculosis is still poorly understood, although certain PE_PGRS genes have been linked to virulence. Tumor necrosis factor-α (TNF-α ) is essential for successfully combating tuberculosis. In this study we provide evidence that PE_PGRS33, a surface exposed protein, elicits TNF-α release from macrophages in a TLR2 (Toll-like receptor 2)-dependent manner. ASK1 (apoptosis signal-regulating kinase 1) is activated downstream of TLR2. ASK1 activates the MAPKs p38 and JNK. PE_PGRS33-induced signaling leads to enhanced expression of TNF-α and TNF receptor I (TNFRI) genes. Mycobacterium smegmatis expressing PE_ PGRS33 elicits the same effects as purified PE_PGRS33. TNF-α release occurs even when internalization of the bacteria is blocked by cytochalasin D, suggesting that interaction of PE_ PGRS33 with TLR2 is sufficient to trigger the effects described. Release of TNF-α plays the determining role in triggering apoptosis in macrophages challenged with PE_PGRS33. The death receptor-dependent signals are amplified through classical caspase 8-dependent mitochondrial release of cytochrome c, leading to the activation of caspases 9 and 3. An important aspect of our findings is that deletions within the PGRS domain (simulating those occurring in clinical strains) attenuate the TNF-α-inducing ability of PE_PGRS33. These results provide the first evidence that variations in the polymorphic repeats of the PGRS domain modulate the innate immune response

EFFECT OF ETHANOLIC EXTRACT OF CYANOTIS CRISTATA LEAVES APPLIED TOPICALLY ON WOUND HEALING IN WISTAR RATS

  Objective: The objective of this study was to assess the effect of ethanolic extract of Cyanotis cristata leaves when applied topically on healing of wound in Wistar rats.Methods: C. cristata leaves were evaluated for its effect on healing of wound in three models, namely, incision, excision, and burn wound. Each model included five groups, each consisting of 6 rats. The five groups were as follows: Group I rats (control and ointment base), Group II rats (standard and silver sulfadiazine), and Group III, Group IV, Group V (treated with C. cristata extract ointment of 0.5%, 1%, and 2%, respectively). Variables evaluated included breaking strength in incision wound while it was contraction rate and epithelialization in excision and burn wound. One-way analysis of variance and Tukey's post hoc test was used to analyze data.Result: In incision wound, breaking strength in test group was significantly increased (p<0.001) as compared to control. In excision and burn wound of test group, time to epithelialization and contraction rate was significantly decreased (p<0.001). The granulation tissue from excision and burn wound showed increased collagen and less inflammatory cells in test groups in comparison to the control.Conclusion: The ethanolic extract of leaves of C. cristata, when applied topically, enhanced wound strength and accelerated healing of incision, excision, and burn wounds in rats

Exposure to Apoptotic Activated CD4+ T Cells Induces Maturation and APOBEC3G- Mediated Inhibition of HIV-1 Infection in Dendritic Cells

Dendritic cells (DCs) are activated by signaling via pathogen-specific receptors or exposure to inflammatory mediators. Here we show that co-culturing DCs with apoptotic HIV-infected activated CD4+ T cells (ApoInf) or apoptotic uninfected activated CD4+ T cells (ApoAct) induced expression of co-stimulatory molecules and cytokine release. In addition, we measured a reduced HIV infection rate in DCs after co-culture with ApoAct. A prerequisite for reduced HIV infection in DCs was activation of CD4+ T cells before apoptosis induction. DCs exposed to ApoAct or ApoInf secreted MIP-1α, MIP-1β, MCP-1, and TNF-α; this effect was retained in the presence of exogenous HIV. The ApoAct-mediated induction of co-stimulatory CD86 molecules and reduction of HIV infection in DCs were partially abrogated after blocking TNF-α using monoclonal antibodies. APOBEC3G expression in DCs was increased in co-cultures of DCs and ApoAct but not by apoptotic resting CD4+ T cells (ApoRest). Silencing of APOBEC3G in DC abrogated the HIV inhibitory effect mediated by ApoAct. Sequence analyses of an env region revealed significant induction of G-to-A hypermutations in the context of GG or GA dinucleotides in DNA isolated from DCs exposed to HIV and ApoAct. Thus, ApoAct-mediated DC maturation resulted in induction of APOBEC3G that was important for inhibition of HIV-infection in DCs. These findings underscore the complexity of differential DC responses evoked upon interaction with resting as compared with activated dying cells during HIV infection

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

rJHP0290 modulates cell cycle progression in gastric epithelial cells.

<p>AGS cells were incubated either with protein storage buffer (Ctrl) or rJHP0290 (100 ng/ml) for 24 h. Cells were washed with PBS, fixed in ice-cold 70% ethanol at 4°C and stained with PI and RNAse A in PBS for 30 min. The DNA profile was generated by FACS. One representative profile (A) and mean ± SD of six independent experiments (B) are shown. Statistically significant differences are indicated by * (p < 0.05).</p