Leishmania major MAPK4 intercepts and redirects CD40 signaling promoting infection.

The parasite Leishmania resides as amastigotes within the macrophage parasitophorous vacuoles inflicting the disease Leishmaniasis. Leishmania selectively modulates mitogen-activated protein kinase (MAPK) phosphorylation subverting CD40-triggered anti-leishmanial functions of macrophages. The mechanism of any pathogen-derived molecule induced host MAPK modulation remains poorly understood. Herein, we show that of the fifteen MAPKs, LmjMAPK4 expression is higher in virulent L. major. LmjMAPK4- detected in parasitophorous vacuoles and cytoplasm- binds MEK-1/2, but not MKK-3/6. Lentivirally-overexpressed LmjMAPK4 augments CD40-activated MEK-1/2-ERK-1/2-MKP-1, but inhibits MKK3/6-p38MAPK-MKP-3, phosphorylation. A rationally-identified LmjMAPK4 inhibitor reinstates CD40-activated host-protective anti-leishmanial functions in L. major-infected susceptible BALB/c mice. These results identify LmjMAPK4 as a MAPK modulator at the host-pathogen interface and establish a pathogen-intercepted host receptor signaling as a scientific rationale for identifying drug targets.

TLR induced IL-27 plays host-protective role against B16BL6 melanoma in C57BL/6 mice.

Elicitation of the tumor-eliminating immune response is a major challenge, as macrophages- constituting a major component of solid tumor mass- play important roles in development, maintenance and tumor regression. The macrophage-expressed Toll-Like Receptors (TLRs) enhance macrophage function and their ability to activate T cells via secretion of cytokines, which may help in tumor regression. IL-27, a member of the IL-12 family of cytokines, is shown to exhibit anti-tumor and anti-angiogenic activities. Herein, we developed B16BL6 melanoma model in C75BL/6 mouse to dissect the crosstalk between TLRs and IL-27 in tumors. We report existence of a novel TLR- IL-27 feed-forward loop, whereby TLRs and IL-27 up-regulated each other's expression, which we found perturbed during melanoma tumorigenesis. Intra-tumoral injection of Imiquimod, a TLR7/8 ligand, reduced the tumor burden; the anti-tumor effect was reversed upon IL-27 and IL-27R silencing by intra-tumorally administered, lentivirally expressed IL-27 and IL-27R shRNA. The reduced tumor growth was accompanied by significantly fewer Treg cells but increased IFN-γ and granzyme B expression by CD8+ T cells. These data indicate the preventative role for TLR-induced IL-27 in aggressive and highly invasive melanoma.

Comparison of clinical methods to diagnose pediatric endobronchial intubation-A randomized controlled trial.

BACKGROUND AND AIMS: Diagnosing accurate placement of the tip of the endotracheal tube is crucial in pediatric practice. This study was conducted to find out the efficacy of five clinical methods to ascertain the tube position by a resident anesthesiologist. MATERIAL AND METHODS: This was a randomized crossover study conducted in a research institute. Fifty pediatric patients were enrolled. All patients were randomly allocated to tracheal (group T) or bronchial group (group B). The five clinical methods which were evaluated include the auscultation, observation of chest movements, bag compliance, tube depth, and capnography. In group T, the tube was placed in the trachea and later positioned in bronchus (assisted by fiberoptic bronchoscopy). The vice versa was done in group B. In each position, a single test followed by all tests was performed and after the change of position, the same single test followed by all tests was performed. Correct and incorrect diagnoses by tests in detecting tube positions were made and their sensitivity and odds ratio were estimated. RESULTS: The tube depth and combination of all tests detected endobronchial intubation with a sensitivity of 88% and 97%, respectively, which is more than that of auscultation (70%) and observation (55%). Evaluation of the difference in agreement level of tube depth to detect tube-position showed the odds ratio of 2.28 (0.17-30.95) for detecting endobronchial intubation. CONCLUSION: We observed that the tube-depth was better than the other individual tests in diagnosing endobronchial intubation in pediatric patients. However, its efficacy is lesser than that of performing all clinical tests together.

Emergency Pancreaticoduodenectomy for Exsanguinating Ampullary Malignancy.

How to cite this article: Subbarayan R, Anand S, Selvaraj S. Emergency Pancreaticoduodenectomy for Exsanguinating Ampullary Malignancy. Indian J Crit Care Med 2020;24(12):1279-1280.

DAMP-TLR-cytokine axis dictates the fate of tumor.

Random mutations leading to loss of cell cycle control is not a rare occurrence in an organism but the mutated cells are recognized and eliminated preventing the development of a tumor. These potentially tumorigenic cells release damage-associated molecular patterns (DAMPs), which are recognized by toll-like receptors (TLRs) on macrophages and dendritic cells. The initial TLR-DAMP interactions lead to different responses such as altered antigen presentation and cytokine release that directly affect T cell activation and removal of the tumorigenic cells. The indirect effects of TLR-DAMP interaction include chemokine-directed altered T cell trafficking, angiogenesis for both T cell infiltration and tumor cell metastasis, and alteration of intra-tumoral milieu contributing to the development of tumor cells heterogeneity. Thus, the initial TLR-DAMP interaction has a set of local effects that modulate tumor cell growth and heterogeneity and a disseminating set of central effects that dynamically affect T cell trafficking and functions. Herein, we argue that the DAMP-TLR-cytokine axis in the tumor microenvironment serves as the mainstay that orchestrates and regulates the pro- and anti-tumor elements which dynamically interact between themselves eventuating in tumor regression or growth. The knowledge of this TLR-based immuno-surveillance framework is a key to developing a novel immunotherapy against cancer.

Pegylated bisacycloxypropylcysteine, a diacylated lipopeptide ligand of TLR6, plays a host-protective role against experimental Leishmania major infection.

TLRs recognize pathogen-expressed Ags and elicit host-protective immune response. Although TLR2 forms heterodimers with TLR1 or TLR6, recognizing different ligands, differences in the functions of these heterodimers remain unknown. In this study, we report that in Leishmania major-infected macrophages, the expression of TLR1 and TLR2, but not TLR6, increased; TLR2-TLR2 association increased, but TLR2-TLR6 association diminished. Lentivirus-expressed TLR1-short hairpin RNA (shRNA) or TLR2-shRNA administration reduced, but TLR6-shRNA increased L. major infection in BALB/c mice. Corroboratively, Pam3CSK4 (TLR1-TLR2 ligand) and peptidoglycan (TLR2 ligand) increased L. major infection but reduced TLR9 expression, whereas pegylated bisacycloxypropylcysteine (BPPcysMPEG; TLR2-TLR6 ligand) reduced L. major number in L. major-infected macrophages, accompanied by increased TLR9 expression, higher IL-12 production, and inducible NO synthase expression. Whereas MyD88, Toll/IL-1R adaptor protein, and TNFR-α-associated factor 6 recruitments to TLR2 were not different in Pam3CSK4-, peptidoglycan-, or BPPcysMPEG-treated macrophages, only BPPcysMPEG enhanced p38MAPK and activating transcription factor 2 activation. BPPcysMPEG conferred antileishmanial functions to L. major-infected BALB/c-derived T cells in a macrophage-T cell coculture and in BALB/c mice; the protection was TLR6 dependent and IL-12 dependent, and it was accompanied by reduced regulatory T cell number. BPPcysMPEG administration during the priming with fixed L. major protected BALB/c mice against challenge L. major infection; the protection was accompanied by low IL-4 and IL-10, but high IFN-γ productions and reduced regulatory T cells. Thus, BPPcysMPEG, a novel diacylated lipopeptide ligand for TLR2-TLR6 heterodimer, induces IL-12-dependent, inducible NO synthase-dependent, T-reg-sensitive antileishmanial protection. The data reveal a novel dimerization partner-dependent duality in TLR2 function.

Anti-VEGF antibody enhances the antitumor effect of CD40.

As its central immunomodulatory effects, CD40 induces interleukin (IL)-12-dependent antitumor immune responses; as its local protumor effects, CD40 induces the expression of vascular endothelial growth factor (VEGF) that promotes tumor angiogenesis and growth. Therefore, using a previously established tumor model in mouse, we examined if the antitumor functions of CD40 are self-limited by VEGF induction. We observed that as the tumor mass grew during day 6 to day 18, VEGF expression in the tumor peaked with concomitant decrease in expressions of CD40 and IL-12 but not of IL-10. Among the angiogenic factors, VEGF-B, VEGFR-1, VEGFR-2, angiopoietin-1 and Tie2 expressions decreased, whereas the expressions of angiopoietin-2 and angiopoietin-3 increased with tumor growth. As significant changes in the expressions of these factors were observed on day 6, we treated the tumor-bearing mice with the agonistic anti-CD40 antibody or neutralizing anti-VEGF antibody-alone or in combination-from the fifth day after the injection of tumor cells. The anti-VEGF antibody significantly enhanced the antitumor effects of the anti-CD40 antibody, as observed through increased survival of the mice, accompanied by reduced angiogenesis and angiopoietin-2 expression but higher T-cell proliferation in response to tumor antigens, increased interferon-γ production and tumor cell cytotoxicity and higher levels of tumor antigen-specific serum IgM, IgG1 and IgG2a, indicating B-cell activation. Thus, our data show for the first time that the combined treatment with an agonistic anti-CD40 antibody and a neutralizing anti-VEGF antibody, which increases antitumor immune response or reduces local angiogenesis, respectively, is a novel antitumor strategy.