Altered expression of caspases-4 and -5 during inflammatory bowel disease and colorectal cancer : diagnostic and therapeutic potential

Caspases are a group of proteolytic enzymes involved in the co-ordination of cellular processes, including cellular homeostasis, inflammation and apoptosis. Altered activity of caspases, particularly caspase-1, has been implicated in the development of intestinal diseases, such as inflammatory bowel disease (IBD) and colorectal cancer (CRC). However, the involvement of two related inflammatory caspase members, caspases-4 and -5, during intestinal homeostasis and disease has not yet been established. This study demonstrates that caspases-4 and -5 are involved in IBD-associated intestinal inflammation. Furthermore, we found a clear correlation between stromal caspase-4 and -5 expression levels, inflammation and disease activity in ulcerative colitis patients. Deregulated intestinal inflammation in IBD patients is associated with an increased risk of developing CRC. We found robust expression of caspases-4 and -5 within intestinal epithelial cells, exclusively within neoplastic tissue, of colorectal tumours. An examination of adjacent normal, inflamed and tumour tissue from patients with colitis-associated CRC confirmed that stromal expression of caspases-4 and -5 is increased in inflamed and dysplastic tissue, while epithelial expression is restricted to neoplastic tissue. In addition to identifying caspases-4 and -5 as potential targets for limiting intestinal inflammation, this study has identified epithelial-expressed caspases-4 and -5 as biomarkers with diagnostic and therapeutic potential in CRC

Low frequency of the TIRAP S180L polymorphism in Africa, and its potential role in malaria, sepsis, and leprosy

<p>Abstract</p> <p>Background</p> <p>The Toll-like receptors (TLRs) mediate innate immunity to various pathogens. A mutation (S180L) in the TLR downstream signal transducer <it>TIRAP </it>has recently been reported to be common in Europeans and Africans and to roughly half the risks of heterogeneous infectious diseases including malaria, tuberculosis, bacteremia, and invasive pneumococal disease in heterozygous mutation carriers.</p> <p>Methods</p> <p>We assessed the <it>TIRAP </it>S180L variant by melting curve and RFLP analysis in 1095 delivering women from malaria-endemic Ghana, as well as in a further 1114 individuals participating in case control studies on sepsis and leprosy in Germany, Turkey and Bangladesh.</p> <p>Results</p> <p>In Ghana, the <it>TIRAP </it>S180L polymorphism was virtually absent. In contrast, the mutation was observed among 26.6%, 32.9% and 12% of German, Bangladesh and Turkish controls, respectively. No significant association of the heterozygous genotype with sepsis or leprosy was observed. Remarkably, homozygous <it>TIRAP </it>180L tend to increase the risk of sepsis in the German study (<it>P </it>= 0.04).</p> <p>Conclusion</p> <p>A broad protective effect of <it>TIRAP </it>S180L against infectious diseases <it>per se </it>is not discernible.</p

NLRP6 negatively regulates innate immunity and host defence against bacterial pathogens

Members of the intracellular nucleotide-binding and oligomerization domain (NOD)-like receptor (NLR) family contribute to immune responses through activation of nuclear factor-kappa B (NF-kappa B), type I interferon and inflammasome signalling(1). Mice lacking the NLR family member NLRP6 were recently shown to be susceptible to colitis and colorectal tumorigenesis(2-4), but the role of NLRP6 in microbial infections and the nature of the inflammatory signalling pathways regulated by NLRP6 remain unclear. Here we show that Nlrp6-deficient mice are highly resistant to infection with the bacterial pathogens Listeria monocytogenes, Salmonella typhimurium and Escherichia coli. Infected Nlrp6-deficient mice had increased numbers of monocytes and neutrophils in circulation, and NLRP6 signalling in both haematopoietic and radioresistant cells contributed to increased susceptibility. Nlrp6 deficiency enhanced activation of mitogen-activated protein kinase (MAPK) and the canonical NF-kappa B pathway after Toll-like receptor ligation, but not cytosolic NOD1/2 ligation, in vitro. Consequently, infected Nlrp6-deficient cells produced increased levels of NF-kappa B-and MAPK-dependent cytokines and chemokines. Thus, our results reveal NLRP6 as a negative regulator of inflammatory signalling, and demonstrate a role for this NLR in impeding clearance of both Gram-positive and -negative bacterial pathogens

Recent evolution of the NF-κB and inflammasome regulating protein POP2 in primates

<p>Abstract</p> <p>Background</p> <p>Pyrin-only protein 2 (POP2) is a small human protein comprised solely of a pyrin domain that inhibits NF-κB p65/RelA and blocks the formation of functional IL-1β processing inflammasomes. Pyrin proteins are abundant in mammals and several, like POP2, have been linked to activation or regulation of inflammatory processes. Because <it>POP2 </it>knockout mice would help probe the biological role of inflammatory regulation, we thus considered whether <it>POP2 </it>is common in the mammalian lineage.</p> <p>Results</p> <p>BLAST searches revealed that <it>POP2 </it>is absent from the available genomes of not only mice and rats, but those of other domestic mammals and New World monkeys as well. <it>POP2 </it>is however present in the genome of the primate species most closely related to humans including <it>Pan troglodytes </it>(chimpanzees), <it>Macaca mulatta </it>(rhesus macaques) and others. Interestingly, chimpanzee POP2 is identical to human POP2 (huPOP2) at both the DNA and protein level. Macaque POP2 (mqPOP2), although highly conserved is not identical to the human sequence; however, both functions of the human protein are retained. Further, <it>POP2 </it>appears to have arisen in the mammalian genome relatively recently (~25 mya) and likely derived from retrogene insertion of <it>NLRP2</it>.</p> <p>Conclusion</p> <p>Our findings support the hypothesis that the NLR loci of mammals, encoding proteins involved in innate and adaptive immunity as well as mammalian development, have been subject to recent and strong selective pressures. Since POP2 is capable of regulating signaling events and processes linked to innate immunity and inflammation, its presence in the genomes of hominids and Old World primates further suggests that additional regulation of these signals is important in these species.</p

Combination of RGD Compound and Low-Dose Paclitaxel Induces Apoptosis in Human Glioblastoma Cells

) peptide, to human glioblastoma U87MG cells with combination of low dose Paclitaxel (PTX) pre-treatment to augment therapeutic activity for RGD peptide-induced apoptosis. peptide induced U87MG programmed cell death. The increased expression of PTX-induced integrin-αvβ3 was correlated with the enhanced apoptosis in U87MG cells.This study provides a novel concept of targeting integrin-αvβ3 with RGD peptides in combination with low-dose PTX pre-treatment to improve efficiency in human glioblastoma treatment

Role of stromal cell-mediated Notch signaling in CLL resistance to chemotherapy

Stromal cells are essential components of the bone marrow (BM) microenvironment that regulate and support the survival of different tumors, including chronic lymphocytic leukemia (CLL). In this study, we investigated the role of Notch signaling in the promotion of survival and chemoresistance of human CLL cells in coculture with human BM-mesenchymal stromal cells (hBM-MSCs) of both autologous and allogeneic origin. The presence of BM-MSCs rescued CLL cells from apoptosis both spontaneously and following induction with various drugs, including Fludarabine, Cyclophosphamide, Bendamustine, Prednisone and Hydrocortisone. The treatment with a combination of anti-Notch-1, Notch-2 and Notch-4 antibodies or γ-secretase inhibitor XII (GSI XII) reverted this protective effect by day 3, even in presence of the above-mentioned drugs. Overall, our findings show that stromal cell-mediated Notch-1, Notch-2 and Notch-4 signaling has a role in CLL survival and resistance to chemotherapy. Therefore, its blocking could be an additional tool to overcome drug resistance and improve the therapeutic strategies for CLL

An Accessory to the ‘Trinity’: SR-As Are Essential Pathogen Sensors of Extracellular dsRNA, Mediating Entry and Leading to Subsequent Type I IFN Responses

Extracellular RNA is becoming increasingly recognized as a signaling molecule. Virally derived double stranded (ds)RNA released into the extracellular space during virus induced cell lysis acts as a powerful inducer of classical type I interferon (IFN) responses; however, the receptor that mediates this response has not been identified. Class A scavenger receptors (SR-As) are likely candidates due to their cell surface expression and ability to bind nucleic acids. In this study, we investigated a possible role for SR-As in mediating type I IFN responses induced by extracellular dsRNA in fibroblasts, a predominant producer of IFNβ. Fibroblasts were found to express functional SR-As, even SR-A species thought to be macrophage specific. SR-A specific competitive ligands significantly blocked extracellular dsRNA binding, entry and subsequent interferon stimulated gene (ISG) induction. Candidate SR-As were systematically investigated using RNAi and the most dramatic inhibition in responses was observed when all candidate SR-As were knocked down in unison. Partial inhibition of dsRNA induced antiviral responses was observed in vivo in SR-AI/II-/- mice compared with WT controls. The role of SR-As in mediating extracellular dsRNA entry and subsequent induced antiviral responses was observed in both murine and human fibroblasts. SR-As appear to function as ‘carriers’, facilitating dsRNA entry and delivery to the established dsRNA sensing receptors, specifically TLR3, RIGI and MDA-5. Identifying SR-As as gatekeepers of the cell, mediating innate antiviral responses, represents a novel function for this receptor family and provides insight into how cells recognize danger signals associated with lytic virus infections. Furthermore, the implications of a cell surface receptor capable of recognizing extracellular RNA may exceed beyond viral immunity to mediating other important innate immune functions

Antileukemic activity of the HSP70 inhibitor pifithrin-μ in acute leukemia

Heat shock protein (HSP) 70 is aberrantly expressed in different malignancies and has emerged as a promising new target for anticancer therapy. Here, we analyzed the in vitro antileukemic effects of pifithrin-μ (PFT-μ), an inhibitor of inducible HSP70, in acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) cell lines, as well as in primary AML blasts. PFT-μ significantly inhibited cell viability at low micromolar concentrations in all cell lines tested, with IC50 values ranging from 2.5 to 12.7 μ, and was highly active in primary AML blasts with a median IC50 of 8.9 μ (range 5.7–37.2). Importantly, higher IC50 values were seen in normal hematopoietic cells. In AML and ALL, PFT-μ induced apoptosis and cell cycle arrest in a dose-dependent fashion. PFT-μ also led to an increase of the active form of caspase-3 and reduced the intracellular concentrations of AKT and ERK1/2 in NALM-6 cells. Moreover, PFT-μ enhanced cytotoxicity of cytarabine, 17-(allylamino)-17-desmethoxygeldanamycin, suberoylanilide hydroxamic acid, and sorafenib in NALM-6, TOM-1 and KG-1a cells. This is the first study demonstrating significant antileukemic effects of the HSP70 inhibitor PFT-μ, alone and in combination with different antineoplastic drugs in both AML and ALL. Our results suggest a potential therapeutic role for PFT-μ in acute leukemias

Sertoli cells have a functional NALP3 inflammasome that can modulate autophagy and cytokine production

Sertoli cells, can function as non-professional tolerogenic antigen-presenting cells, and sustain the blood-testis barrier formed by their tight junctions. The NOD-like receptor family members and the NALP3 inflammasome play a key role in pro-inflammatory innate immunity signalling pathways. Limited data exist on NOD1 and NOD2 expression in human and mouse Sertoli cells. Currently, there is no data on inflammasome expression or function in Sertoli cells. We found that in primary pre-pubertal Sertoli cells and in adult Sertoli line, TLR4\NOD1 and NOD2 crosstalk converged in NF?B activation and elicited a NALP3 activation, leading to de novo synthesis and inflammasome priming. This led to caspase-1 activation and IL-1? secretion. We demonstrated this process was controlled by mechanisms linked to autophagy. NOD1 promoted pro-IL-1? restriction and autophagosome maturation arrest, while NOD2 promoted caspase-1 activation, IL-1? secretion and autophagy maturation. NALP3 modulated NOD1 and pro-IL-1? expression, while NOD2 inversely promoted IL-1?. This study is proof of concept that Sertoli cells, upon specific stimulation, could participate in male infertility pathogenesis via inflammatory cytokine induction

A comprehensive characterization of the caspase gene family in insects from the order Lepidoptera

<p>Abstract</p> <p>Background</p> <p>The cell suicide pathway of apoptosis is a necessary event in the life of multicellular organisms. It is involved in many biological processes ranging from development to the immune response. Evolutionarily conserved proteases, called caspases, play a central role in regulating apoptosis. Reception of death stimuli triggers the activation of initiator caspases, which in turn activate the effector caspases. In Lepidoptera, apoptosis is crucial in processes such as metamorphosis or defending against baculovirus infection. The discovery of p35, a baculovirus protein inhibiting caspase activity, has led to the characterization of the first lepidopteran caspase, Sf-Caspase-1. Studies on Sf-Caspase-1 mode of activation suggested that apoptosis in Lepidoptera requires a cascade of caspase activation, as demonstrated in many other species.</p> <p>Results</p> <p>In order to get insights into this gene family in Lepidoptera, we performed an extensive survey of lepidopteran-derived EST datasets. We identified 66 sequences distributed among 27 species encoding putative caspases. Phylogenetic analyses showed that Lepidoptera possess at least 5 caspases, for which we propose a unified nomenclature. According to homology to their <it>Drosophila </it>counterparts and their primary structure, we determined that Lep-Caspase-1, -2 and -3 are putative effector caspases, whereas Lep-Caspase-5 and -6 are putative initiators. The likely function of Lep-Caspase-4 remains unclear. Lep-Caspase-2 is absent from the silkworm genome and appears to be noctuid-specific, and to have arisen from a tandem duplication of the Caspase-1 gene. In the tobacco hawkmoth, 3 distinct transcripts encoding putative Caspase-4 were identified, suggesting at least 2 duplication events in this species.</p> <p>Conclusions</p> <p>The basic repertoire of five major types of caspases shared among Lepidoptera seems to be smaller than for most other groups studied to date, but gene duplication still plays a role in lineage-specific increases in diversity, just as in Diptera and mammals.</p