Exploring caspase functions in mouse models.

Caspases are enzymes with protease activity. Despite being known for more than three decades, caspase investigation still yields surprising and fascinating information. Initially associated with cell death and inflammation, their functions have gradually been revealed to extend beyond, targeting pathways such as cell proliferation, migration, and differentiation. These processes are also associated with disease mechanisms, positioning caspases as potential targets for numerous pathologies including inflammatory, neurological, metabolic, or oncological conditions. While in vitro studies play a crucial role in elucidating molecular pathways, they lack the context of the body's complexity. Therefore, laboratory animals are an indispensable part of successfully understanding and applying caspase networks. This paper aims to summarize and discuss recent knowledge, understanding, and challenges in caspase knock-out mice.

Disease-associated astrocyte epigenetic memory promotes CNS pathology.

Disease-associated astrocyte subsets contribute to the pathology of neurologic diseases, including multiple sclerosis and experimental autoimmune encephalomyelitis1-8 (EAE), an experimental model for multiple sclerosis. However, little is known about the stability of these astrocyte subsets and their ability to integrate past stimulation events. Here we report the identification of an epigenetically controlled memory astrocyte subset that exhibits exacerbated pro-inflammatory responses upon rechallenge. Specifically, using a combination of single-cell RNA sequencing, assay for transposase-accessible chromatin with sequencing, chromatin immunoprecipitation with sequencing, focused interrogation of cells by nucleic acid detection and sequencing, and cell-specific in vivo CRISPR-Cas9-based genetic perturbation studies we established that astrocyte memory is controlled by the metabolic enzyme ATP-citrate lyase (ACLY), which produces acetyl coenzyme A (acetyl-CoA) that is used by histone acetyltransferase p300 to control chromatin accessibility. The number of ACLY+p300+ memory astrocytes is increased in acute and chronic EAE models, and their genetic inactivation ameliorated EAE. We also detected the pro-inflammatory memory phenotype in human astrocytes in vitro; single-cell RNA sequencing and immunohistochemistry studies detected increased numbers of ACLY+p300+ astrocytes in chronic multiple sclerosis lesions. In summary, these studies define an epigenetically controlled memory astrocyte subset that promotes CNS pathology in EAE and, potentially, multiple sclerosis. These findings may guide novel therapeutic approaches for multiple sclerosis and other neurologic diseases.

YKL-40 as a biomarker in various inflammatory diseases: A review.

YKL-40 or Chitinase-3-Like Protein 1 (CHI3L1) is a highly conserved glycoprotein that binds heparin and chitin in a non-enzymatic manner. It is a member of the chitinase protein family 18, subfamily A, and unlike true chitinases, YKL-40 is a chitinase-like protein without enzymatic activity for chitin. Although its accurate function is yet unknown, the pattern of its expression in the normal and disease states suggests its possible engagement in apoptosis, inflammation and remodeling or degradation of the extracellular matrix. During an inflammatory response, YKL-40 is involved in a complicated interaction between host and bacteria, both promoting and attenuating immune response and potentially being served as an autoantigen in a vicious circle of autoimmunity. Based on its pathophysiology and mechanism of action, the aim of this review was to summarize research on the growing role of YKL-40 as a persuasive biomarker for inflammatory diseases' early diagnosis, prediction and follow-up (e.g., cardiovascular, gastrointestinal, endocrinological, immunological, musculoskeletal, neurological, respiratory, urinary, infectious) with detailed structural and functional background of YKL-40.

Eicosanoids and cancer

Eicosanoids are 20-carbon bioactive lipids derived from the metabolism of polyunsaturated fatty acids, which can modulate various biological processes including cell proliferation, adhesion and migration, angiogenesis, vascular permeability and inflammatory responses. In recent years, studies have shown the importance of eicosanoids in the control of physiological and pathological processes associated with several diseases, including cancer. The polyunsaturated fatty acid predominantly metabolized to generate 2-series eicosanoids is arachidonic acid, which is the major n-6 polyunsaturated fatty acid found in animal fat and in the occidental diet. The three main pathways responsible for metabolizing arachidonic acid and other polyunsaturated fatty acids to generate eicosanoids are the cyclooxygenase, lipoxygenase and P450 epoxygenase pathways. Inflammation plays a decisive role in various stages of tumor development including initiation, promotion, invasion and metastasis. This review will focus on studies that have investigated the role of prostanoids and lipoxygenase-derived eicosanoids in the development and progression of different tumors, highlighting the findings that may provide insights into how these eicosanoids can influence cell proliferation, cell migration and the inflammatory process. A better understanding of the complex role played by eicosanoids in both tumor cells and the tumor microenvironment may provide new markers for diagnostic and prognostic purposes and identify new therapeutic strategies in cancer treatment.

Los inhibidores de las proteínas-cinasas en enfermedades autoinmunes e inflamatorias: presente y futuro de nuevas dianas terapéuticas / Protein-kinase inhibitors: A new treatment pathway for autoimmune and inflammatory diseases?

Pese a los avances terapéuticos en las enfermedades autoinmunes e inflamatorias, muchos pacientes no logran un control adecuado de la enfermedad. De ahí la necesidad de optimizar el uso de las terapias biológicas y de explorar nuevas opciones terapéuticas. La disponibilidad de fármacos que inhiben proteínas-cinasas ya es una realidad en especialidades como oncología y hematología, donde los resultados asociados a la evolución clínica de la enfermedad han sido prometedores. La principal ventaja de estos fármacos es la administración oral, que podría favorecer la adherencia del paciente y reducir los costes asociados al tratamiento. Tofacitinib, inhibidor de tirosinas-cinasas, actualmente es el único fármaco de esta categoría aprobado para el tratamiento de la artritis reumatoide por la FDA. Estas dianas terapéuticas son evaluadas actualmente en diversas enfermedades autoinmunes e inflamatorias. Sin embargo, el conocimiento y la comprensión de las vías de señalización intracelular siguen siendo limitados, persistiendo dudas en cuanto al mecanismo de acción, la eficacia y los posibles efectos secundarios asociados al uso de estos nuevos fármacos (AU)

Although advances in biological medicine have seen significant progress in the treatment of autoimmune and inflammatory disease, many patients do not experience a satisfactory response. Hence, there are two challenges facing the medical research community. The first is to continue development in the field of existing biological therapies, such as monoclonal antibodies. The second is to open new frontiers of research and explore treatment alternatives for non-responders to other therapies. Attention has increasingly turned to the therapeutic potential of small molecule weight kinase inhibitors (SMKIs), currently used extensively in oncology and haematology. Initial research into the therapeutic value of SMKIs for autoimmune and inflammatory diseases has been encouraging. SMKIs are taken orally, which reduces cost for the health provider, and could increase compliance for the patient. This is why research is now focusing increasingly on SMKIs as a new generation line of treatment in these diseases. Tofacitinib, an inhibitor of Janus-kinase, is currently the only drug approved for the treatment of rheumatoid arthritis by FDA. However, much more needs to be done to understand the intracellular signalling pathways and how these might affect disease progression before solid conclusions can be drawn (AU)

Modulación farmacológica de la cicatriz glial para la reparación de lesiones del SNC / Pharmacological modulation of the glial scar for CNS injury repair

Objetivo: Estudiar la actividad de la neurostatina obtenida enzimáticamente y purificada mediante un nuevo método, en cultivos de células implicadas en la formación de la cicatriz glial. Material y métodos: La neurostatina se obtuvo mediante una reacción enzimática a partir del gangliósido GD1b comercial y se purificó con un método nuevo simplificado. La actividad de la neurostatina se probó en ensayos de proliferación con MTT en pericitos y microglía de rata. Resultados: La actividad de la neurostatina purificada fue similar a la obtenida mediante purificación a partir de cerebro de mamífero. La neurostatina inhibió la proliferación de los pericitos inducida por el factor de crecimiento PDGF-B y la proliferación de la microglía de rata inducida por la toxina bacteriana LPS a concentraciones nanomolar. Conclusión: La nueva metodología de obtención y purificación de la neurostatina y su actividad justifican su ensayo en modelos de lesión del SNC en animales, para evaluar su posible uso como terapia en pacientes con lesiones del SNC (AU)

Objective: To study the activity of neurostatin obtained enzymatically and purified using a new method, in cultures of cells involved in the formation of the glial scar. Material and methods: Neurostatin was obtained from the commercial ganglioside GD1b, using enzymatic Oacetylation, and was purified by a new simplified method. The activity of neurostatin was tested by an MTT proliferation assay in pericytes and rat microglial cells. Results: The activity of neurostatin obtained and purified by this new method was similar to the neurostatin obtained by the purification from mammalian brain. Neurostatin inhibited the PDGF-B growth factor induced proliferation of pericytes and LPS bacteria toxin induced proliferation of rat microglial cells at nanomolar concentrations. Conclusion: This new methodology to synthesize and purify neurostatin and its activity justify further studies to test its effect in animal models of CNS injuries and to evaluate its possible use as a therapy in patients with CNS injuries (AU)

Angiotensin II induces NF-κB, JNK and p38 MAPK activation in monocytic cells and increases matrix metalloproteinase-9 expression in a PKC- andRho kinase-dependent manner

Angiotensin II (ANG II), the main effector of the renin-angiotensin system, is implicated in endothelial permeability, recruitment and activation of the immune cells, and also vascular remodeling through induction of inflammatory genes. Matrix metalloproteinases (MMPs) are considered to be important inflammatory factors. Elucidation of ANG II signaling pathways and of possible cross-talks between their components is essential for the development of efficient inhibitory medications. The current study investigates the inflammatory signaling pathways activated by ANG II in cultures of human monocytic U-937 cells, and the effects of specific pharmacological inhibitors of signaling intermediates on MMP-9 gene (MMP-9) expression and activity. MMP-9 expression was determined by real-time PCR and supernatants were analyzed for MMP-9 activity by ELISA and zymography methods. A multi-target ELISA kit was employed to evaluate IκB, NF-κB, JNK, p38, and STAT3 activation following treatments. Stimulation with ANG II (100 nM) significantly increased MMP-9 expression and activity, and also activated NF-κB, JNK, and p38 by 3.8-, 2.8- and 2.2-fold, respectively (P < 0.01). ANG II-induced MMP-9 expression was significantly reduced by 75 and 67 percent, respectively, by co-incubation of the cells with a selective inhibitor of protein kinase C (GF109203X, 5 µM) or of rho kinase (Y-27632, 15 µM), but not with inhibitors of phosphoinositide 3-kinase (wortmannin, 200 nM), tyrosine kinases (genistein, 100 µM) or of reactive oxygen species (α-tocopherol, 100 µM). Thus, protein kinase C and Rho kinase are important components of the inflammatory signaling pathways activated by ANG II to increase MMP-9 expression in monocytic cells. Both signaling molecules may constitute potential targets for effective management of inflammation.

Fetal programming and cardiometabolic diseases: the role of angiotensin II and inflammation

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Rebamipide-induced downregulation of phospholipase D inhibits inflammation and proliferation in gastric cancer cells

Rebamipide a gastroprotective drug, is clinically used for the treatment of gastric ulcers and gastritis, but its actions on gastric cancer are not clearly understood. Phospholipase D (PLD) is overexpressed in various types of cancer tissues and has been implicated as a critical factor in inflammation and carcinogenesis. However, whether rebamipide is involved in the regulation of PLD in gastric cancer cells is not known. In this study, we showed that rebamipide significantly suppressed the expression of both PLD1 and PLD2 at a transcriptional level in AGS and MKN-1 gastric cancer cells. Downregulation of PLD expression by rebamipide inhibited its enzymatic activity. In addition, rebamipide inhibited the transactivation of nuclear factor kappa B (NFkappaB), which increased PLD1 expression. Rebamipide or PLD knockdown significantly suppressed the expression of genes involved in inflammation and proliferation and inhibited the proliferation of gastric cancer cells. In conclusion, rebamipide-induced downregulation of PLD may contribute to the inhibition of inflammation and proliferation in gastric cancer.

Associação entre fatores de risco cardiovascular e proteína C-reativa em mulheres idosas / Association between cardiovascular risk factors and C-reactive protein in aged women

INTRODUÇÃO: O processo inflamatório desempenha um importante papel na etiologia das doenças cardiovasculares. Muitos estudos demonstram que níveis elevados de proteína C-reativa (PCR), uma proteína hepática de fase aguda, estão associados ao risco de tais eventos. Objetivos: Investigar a existência de associação entre PCR e fatores de risco cardiovascular em mulheres idosas MATERIAL E MÉTODO: Foram realizadas avaliações antropométricas, impedanciometria, verificação de pressão arterial, mensurações de perfil lipídico, glicemia em jejum e PCR. RESULTADOS: Observou-se que a PCR esteve relacionada com índice de massa corporal (p = 0,001) e com percentual de gordura corporal (p = 0,015), não apresentando relação significativa com nenhuma outra variável estudada. DISCUSSÃO: A associação entre PCR e marcadores de obesidade é consenso na literatura, podendo, no entanto, não significar verdadeira progressão da aterosclerose ou de um estado inflamatório. Em relação à inexistência de associação com os demais fatores de risco cardiovascular observada neste estudo, os dados encontrados são conflitantes. Há autores que indicam a correlação entre PCR e tais fatores; outros apontam sua inexistência. CONCLUSÕES: Este trabalho demonstra a associação da PCR a marcadores de obesidade, mas não a outros fatores de risco cardiovascular.

BACKGROUND: The inflammatory process plays an important role in the etiology of cardiovascular diseases. Several studies have shown that high levels of C-reactive protein (CRP), a hepatic acute phase protein, are associated with the risk of such diseases. OBJECTIVES: In this study we investigated the existence of association between CRP and cardiovascular risk factors in elderly women. MATERIAL AND METHOD: Anthropometric data, body impedance, blood pressure, lipid profiles, fasting glucose and CRP levels were evaluated. RESULTS: We observed that CRP was linked with body mass index (p = 0.001) and body fat percentage (p = 0.015) and there was no significant connection with any other studied variable. DISCUSSION: The association between CRP and measures of obesity is a consensus in literature. However, it may not show a true progression of atherosclerosis or an inflammatory state. Regarding the inexistence of association with other cardiovascular risk factors observed in this study, the gathered data are conflicting. Some authors indicate correlation between PCR and such factors, whereas others point out its inexistence. CONCLUSIONS: This study demonstrates the association of CRP with obesity, but not with other cardiovascular risk factors.

Inducción de citocinas por efecto de la LDL electronegativa en monocitos y linfocitos / Cytokine induction by the effect of electronegative LDL on monocytes and lymphocytes

Introducción y objetivo. La lipoproteína de baja densidad (LDL) electronegativa (LDL[­]) es una fracción minoritaria modificada de la LDL presente en circulación plasmática con propiedades aterogénicas e inflamatorias. Se ha descrito que la LDL(­) induce, en cultivos de células endoteliales, la producción de diversos mediadores de la inflamación, así como apoptosis y/o citotoxicidad. Sin embargo, no se ha evaluado previamente su efecto sobre otros tipos celulares, como células en circulación, sobre las que es más posible su interacción durante la circulación plasmática. Por ello, el objetivo fue analizar las citocinas, los factores de crecimiento y otras moléculas proinflamatorias implicadas en el proceso arteriosclerótico que son inducidos por la LDL(­) en monocitos y linfocitos aislados de sangre periférica. Material y métodos. La LDL total fue aislada mediante ultracentrifugación y se separaron las fracciones electropositiva (LDL[+] o LDL nativa) y electronegativa por cromatografía de intercambio aniónico. Se incubaron monocitos y linfocitos aislados de voluntarios normolipémicos con las LDL durante 20 h; se utilizó lipopolisacárido (LPS) como control positivo. Se valoró en los sobrenadantes celulares la producción de 42 mediadores inflamatorios relacionados con la arteriosclerosis mediante un array de proteínas. Se cuantificó por ELISA las proteínas inducidas y mediante PCR a tiempo real se evaluó si su inducción era transcripcional. Resultados y conclusión. La LDL(­) indujo una mayor liberación y expresión, comparada con la LDL(+), de interleucina (IL) 6, IL-8, IL-10, MCP-1, growth-related oncogene (GRO) (GROß y GRO*, tanto en monocitos como en linfocitos. Así pues, la LDL(­) es capaz de inducir en células mononucleares la producción de factores implicados en el proceso inflamatorio que actúan en diferentes estadios de la lesión arteriosclerótica (AU)

Introduction and objective. Electronegative low-density lipoprotein (LDL(­)) is a minor modified LDL fraction present in plasma with atherogenic and inflammatory properties. In cultured endothelial cells, LDL(­) has been reported to induce production of several mediators of inflammation, as well as apoptosis and/or cytotoxicity. However, the effect of LDL(­) on other cell types, such as white blood cells ­with which its interaction is more feasible during plasma circulation­ has not previously been evaluated. Therefore, the objective of this study was to analyze the cytokines, growth factors and other proinflammatory molecules involved in the atherosclerotic process that are induced by LDL(­) in monocytes and lymphocytes isolated from peripheral blood. Material and methods. Total LDL was isolated by ultracentrifugation and electropositive (LDL(+) or native LDL) and electronegative fractions were separated by anion-exchange chromatography. Monocytes and lymphocytes isolated from normolipemic volunteers were incubated with LDLs for 20 h; lipopolysaccharide was used as positive control. The production of 42 inflammatory mediators related to atherosclerosis was determined in cell supernatants by protein array. Induced proteins were quantified by ELISA assays. The question of whether induction was transcriptional was determined by real time-polymerase chain reaction. Results and conclusion. LDL(­) induced greater release and expression of interleukin (IL)-6, IL-8, IL-10, MCP-1, GROß and GRO* than did LDL(+) in monocytes and lymphocytes. Therefore, in mononuclear cells, LDL(­) is able to induce production of several factors involved in the inflammatory process that act on different stages of the atherosclerotic lesion (AU)

The Expression of Thymidine Phosphorylase in Cancer-infiltrating Inflammatory Cells in Stomach Cancer

Thymidine phosphorylase (TP) has shown to be up-regulated in several cancers and to play a role in angiogenesis and invasion. Most studies regarding TP have focused on cancer cells. Recently, evidences suggest that TP in cancer-infiltrating inflammatory cells (CIICs) also affect the cancer cell behavior. To evaluate the significance of TP expression of CIICs in gastric cancer, we assessed TP expression of cancer cells and CIICs separately using immunohistochemical assay on 116 paraffin-embedded tissue samples from stomach cancer patients and investigated their clinical significance. When subjects were divided into 4 groups according to the TP expression: cancer/matrix (+/+), C/M (+/-), C/M (-/+), and C/M (-/-), intratumoral microvessel density scores were higher in the C/M (+/-) group than in the C/M (-/-) group (p=0.02). For lymph node metastasis and survival, there were no significant differences among the 4 groups. However, there were significant differences in survival (p=0.035) and LN metastasis (p=0.023) between the two groups divided by TP expression of CIICs alone irrespective of TP expression of cancer cells. Taken together, this study suggested the TP expression in CIICs could affect lymph node metastasis and patients' survival in gastric cancer.

Role of matrix metalloproteinases in the development of airway inflammation and remodeling

Matrix metalloproteinases (MMPs) are a major group of proteases known to regulate extracellular matrix (ECM) turnover and so they have been suggested to be important in the process of lung disease associated with tissue remodeling. This has led to the concept that modulation of airway remodeling including excessive proteolysis damage to the tissue may be of interest for future treatment. Within the MMP family, macrophage elastase (MMP-12) is able to degrade ECM components such as elastin and is involved in tissue remodeling processes in chronic obstructive pulmonary disease including emphysema. Pulmonary fibrosis has an aggressive course and is usually fatal within an average of 3 to 6 years after the onset of symptoms. Pulmonary fibrosis is associated with deposition of ECM components in the lung interstitium. The excessive airway remodeling as a result of an imbalance in the equilibrium of the normal processes of synthesis and degradation of ECM components could justify anti-protease treatments. Indeed, the correlation of the differences in hydroxyproline levels in the lungs of bleomycin-treated mice strongly suggests that a reduced molar pro-MMP-9/TIMP-1 ratio in bronchoalveolar lavage fluid is associated with collagen deposition, beginning as early as the inflammatory events at day 1 after bleomycin administration. Finally, these observations emphasize that effective treatment of these disorders must be started early during the natural history of the disease, prior to the development of extensive lung destruction and fibrosis.

Macrophage elastase (MMP-12): a pro-inflammatory mediator?

As many metalloproteinases (MMPs), macrophage elastase (MMP-12) is able to degrade extracellular matrix components such as elastin and is involved in tissue remodeling processes. Studies using animal models of acute and chronic pulmonary inflammatory diseases, such as pulmonary fibrosis and chronic obstrutive pulmonary disease (COPD), have given evidences that MMP-12 is an important mediator of the pathogenesis of these diseases. However, as very few data regarding the direct involvement of MMP-12 in inflammatory process in the airways were available, we have instilled a recombinant form of human MMP-12 (rhMMP-12) in mouse airways. Hence, we have demonstrated that this instillation induced a severe inflammatory cell recruitment characterized by an early accumulation of neutrophils correlated with an increase in proinflammatory cytokines and in gelatinases and then by a relatively stable recruitment of macrophages in the lungs over a period of ten days. Another recent study suggests that resident alveolar macrophages and recruited neutrophils are not involved in the delayed macrophage recruitment. However, epithelial cells could be one of the main targets of rhMMP-12 in our model. We have also reported that a corticoid, dexamethasone, phosphodiesterase 4 inhibitor, rolipram and a non-selective MMP inhibitor, marimastat could reverse some of these inflammatory events. These data indicate that our rhMMP-12 model could mimic some of the inflammatory features observed in COPD patients and could be used for the pharmacological evaluation of new anti-inflammatory treatment. In this review, data demonstrating the involvement of MMP-12 in the pathogenesis of pulmonary fibrosis and COPD as well as our data showing a pro-inflammatory role for MMP-12 in mouse airways will be summarized.

Protease-activated receptors and inflammatory hyperalgesia

Recent advances in basic science pointed to a role for proteinases, through the activation of proteinase-activated receptors (PARs) in nociceptive mechanisms. Activation of PAR1, PAR2 and PAR4 either by proteinases or by selective agonists causes inflammation inducing most of the cardinal signs of inflammation: swelling, redness, and pain. Sub-inflammatory doses of PAR2 agonist still induced hyperalgesia and allodynia while PAR2 has been shown to be implicated in the generation of hyperalgesia in different inflammatory models. In contrast, sub-inflammatory doses of PAR1 increases nociceptive threshold, inhibiting inflammatory hyperalgesia, thereby acting as an analgesic agent. PARs are present and functional on sensory neurons, where they participate either directly or indirectly to the transmission and/or inhibition of nociceptive messages. Taken together, the results discussed in this review highlight proteinases as signaling molecules to sensory nerves. We need to consider proteinases and the receptors that are activated by proteinases as important potential targets for the development of analgesic drugs in the treatment of inflammatory pain.

Role of protease-activated receptor-2 in inflammation, and its possible implications as a putative mediator of periodontitis

Proteinase-activated receptor-2 (PAR2) belongs to a novel subfamily of G-protein-coupled receptors with seven-transmembrane domains. This receptor is widely distributed throughout the body and seems to be importantly involved in inflammatory processes. PAR2 can be activated by serine proteases such as trypsin, mast cell tryptase, and bacterial proteases, such as gingipain produced by Porphyromonas gingivalis. This review describes the current stage of knowledge of the possible mechanisms that link PAR2 activation with periodontal disease, and proposes future therapeutic strategies to modulate the host response in the treatment of periodontitis.

Topical antiinflammatory activity of phytosterols isolated from Eryngium foetidum on chronic and acute inflammation models

Eryngium foetidum L. (Apiaceae) is a Caribbean endemic plant used in folk medicine for the treatment of several antiinflammatory disorders. A preliminary phytochemical study showed that the hexane extract is rich in terpenic compounds. Chromatographic fractionation of this extract yielded: alpha-cholesterol, brassicasterol, campesterol, stigmasterol (as the main component, 95 percent) clerosterol, beta-sitosterol, delta 5-avenasterol, delta (5) 24-stigmastadienol and delta 7-avenasterol. The topical antiinflammatory activity of the hexane extract and of stigmasterol was evaluated by auricular oedema, induced by 12-0-tetradecanoylphorbol acetate (TPA), in the mouse, using single and multiple applications of the phlogistic agent. Both reduced the oedema in a similar proportion in the two model assays (acute and chronic). Meloperoxidase activity was strongly reduced by both the extract and the compound, in the acute but not the chronic model. These results indicate that the leaves of Eryngium foetidum L may be effective against topical inflammation processes. Stigmasterol also exerts a significant topical antiinflammatory activity although it cannot be considered to be a major antiinflammatory agent, therefore other bioactive components are probably involved in the activity of the hexane extract.(AU)