The physiological response of protease inhibition in dystrophic muscle

Duchenne muscular dystrophy (DMD) is caused by the production of a non‐functional dystrophin gene product and a failure to accumulate functional dystrophin protein in muscle cells. This leads to membrane instability, loss of Ca²⁺ homoeostasis and widespread cellular injury. Associated with these changes are increased protease activities in a variety of proteolytic systems. As such, there have been numerous investigations directed towards determining the therapeutic potential of protease inhibition. In this review, evidence from genetic and/or pharmacological inhibition of proteases as a treatment strategy for DMD is systematically evaluated. Specifically, we review the potential roles of calpain, proteasome, caspase, matrix metalloproteinase and serine protease inhibition as therapeutic approaches for DMD. We conclude that despite early results to the contrary, inhibition of calpain proteases is unlikely to be successful. Conversely, evidence suggests that inhibition of proteasome, matrix metalloproteinases and serine proteases does appear to decrease disease severity. An important caveat to these conclusions, however, is that the fundamental cause of DMD, dystrophin deficiency, is not corrected by this strategy. Hence, this should not be viewed as a cure, but rather, protease inhibitors should be considered for inclusion in a therapeutic cocktail. Physiological Relevance. Selective modulation of protease activity has the potential to profoundly change intracellular physiology resulting in a possible treatment for DMD. However, alteration of protease activities could also lead to worsening of disease progression by promoting the accumulation of substrates in the cell. The balance of benefit and potential damage caused by protease inhibition in human DMD patients is largely unexplored.     

Melatonin modulates neonatal brain inflammation through endoplasmic reticulum stress,autophagy, and miR‐34a/silent information regulator 1 pathway

Maternal infection/inflammation represents one of the most important factors involved in the etiology of brain injury in newborns. We investigated the modulating effect of prenatal melatonin on the neonatal brain inflammation process resulting from maternal intraperitoneal (i.p.) lipopolysaccharide (LPS) injections. LPS (300 μg/kg) was administered to pregnant rats at gestational days 19 and 20. Melatonin (5 mg/kg) was administered i.p. at the same time as LPS. Melatonin counteracted the LPS sensitization to a second ibotenate‐induced excitotoxic insult performed on postnatal day (PND) 4. As melatonin succeeded in reducing microglial activation in neonatal brain at PND1, pathways previously implicated in brain inflammation regulation, such as endoplasmic reticulum (ER) stress, autophagy and silent information regulator 1 (SIRT1), a melatonin target, were assessed at the same time‐point in our experimental groups. Results showed that maternal LPS administrations resulted in an increase in CHOP and Hsp70 protein expression and eIF2α phosphorylation, indicative of activation of the unfolded protein response consequent to ER stress, and a slighter decrease in the autophagy process, determined by reduced lipidated LC3 and increased p62 expression. LPS‐induced inflammation also reduced brain SIRT1 expression and affected the expression of miR‐34a, miR146a, and miR‐126. All these effects were blocked by melatonin. Cleaved‐caspase‐3 apoptosis pathway did not seem to be implicated in the noxious effect of LPS on the PND1 brain. We conclude that melatonin is effective in reducing maternal LPS‐induced neonatal inflammation and related brain injury. Its role as a prophylactic/therapeutic drug deserves to be investigated by clinical studies.     

Tephrosin-induced autophagic cell death in A549 non-small cell lung cancer cells

Anticancer effect of tephrosin (1) has been documented; however, the molecular mechanisms underlying the cytotoxicity of tephrosin in cancer cells remain unclear. In the present paper, the proliferation inhibition rate of several cancer cells was tested using the MTT assay; cell cycle, reactive oxygen species (ROS), and mitochondrial membrane potential (MMP) were determined by flow cytometry; poly(ADP-ribose) polymerase (PARP) cleavage and heat shock protein 90 (Hsp90) expression were evaluated by Western blotting; autophagy was examined by confocal microscopy and light chain 3 (LC3) conversion assay. The results showed that exposure of the cells to tephrosin induced significant proliferation inhibition in a dose-dependent manner, especially on A549 with G₂/M being arrested. Tephrosin was not found to induce cell apoptosis as PARP cleavage was not detected after 24 h treatment, but the formation of acidic vesicular organelle of autophagy character was found, and autophagy was further confirmed by the increase in the ratio of LC3-II to LC3-I. It was observed that tephrosin induced ROS generation and Hsp90 expression inhibition. These results indicate that tephrosin induces A549 cancer cell death via the autophagy pathway, and the roles of ROS generation and Hsp90 expression inhibition in this process need further study in the future.     

自噬在心血管疾病中的影响及中药的干预作用

<正>自噬是一种通过促进溶酶体消化分解受损或衰老的蛋白质、大分子和细胞器,从而调节细胞内的平衡,避免细胞凋亡和损伤的自体机制。在心血管疾病如心肌缺血、心衰和动脉粥样硬化等疾病的进程中,自噬均产生一定的影响。结合现代医学研究,较多学者发现,自噬与中医也存在着相关性。中医学不仅可以从理论上阐述自噬,中药提取物及方剂也能调节自噬,从而改善心血管功能。本文将近年来的相关研究做一综述。现报告如下:     

Autophagy mediates neutrophil responses to bacterial infection

Neutrophils constitute the first line of cellular defense against pathogens and autophagy is a fundamental cellular homeostasis pathway that operates with the intracellular degradation/recycling system. Induction of the autophagic process in neutrophils, in response to invading pathogens, constitutes a crucial mechanism in innate immunity. Exploration of autophagy has greatly progressed and diverse strategies have been reported for studying this molecular process in different biological systems; especially in infectious and inflammatory diseases. Furthermore, the role of autophagy in neutrophils, during pathogenic infection, continues to be of interest, due to the role of the cell in immunity function, its recruitment to the site of infection and its implication in inflammatory diseases. This review focuses on the known role of autophagy in neutrophils defence against pathogenic infections. A more detailed discussion will concern the recent findings highlighting the role of autophagy in inflammation and cell death in infected neutrophils.     

[pemetrexed + sildenafil], via autophagy-dependent HDAC downregulation,enhances the immunotherapy response of NSCLC cells

Pemetrexed is an approved therapeutic in NSCLC and ovarian cancer. Our studies focused on the ability of [pemetrexed + sildenafil] exposure to alter the immunogenicity of lung and ovarian cancer cells. Treatment of lung and ovarian cancer cells with [pemetrexed + sildenafil] in vitro rapidly reduced the expression of PD-L1, PD-L2 and ornithine decarboxylase (ODC), and increased the expression of class I MHCA. In a cell-specific fashion, some cells also released the immunogenic nuclear protein HMGB1 into the extracellular environment. [Pemetrexed + sildenafil] reduced the expression of multiple histone deacetylases that was blocked by knock down of autophagy regulatory proteins. [Pemetrexed + sildenafil] lethality was enhanced by the histone deacetylase inhibitors AR42 and sodium valproate; AR42 and valproate as single agents also rapidly reduced the expression of PD-L1, PD-L2 and ODC, and increased expression of MHCA and CerS6. Nitric oxide and CerS6 signaling was required for drug-induced death receptor activation and tumor cell killing. In vivo, [pemetrexed + sildenafil] lethality against lung cancer cells was enhanced by sodium valproate. Using syngeneic mouse lung cancer cells [pemetrexed + sildenafil] enhanced the anti-tumor effects of antibodies directed to inhibit PD-1 or CTLA4. [Pemetrexed + sildenafil] interacted with the anti-PD-1 antibody to strongly enhance tumor infiltration by M1 macrophages; activated NK cells and activated T cells. Our data demonstrate that treatment of tumor cells with [pemetrexed + sildenafil] results in tumor cell killing and via autophagy-dependent downregulation of HDACs, it opsonizes the remaining tumor cells to anti-tumor immunotherapy antibodies.     

基于“脾主运化”理论探讨肝细胞自噬与脂质代谢的关系＊

脾位于中焦,主运化,为气血生化之源。膏脂由脾运化水谷所生,中土得运,纳运有常,则膏脂可随血循环以濡养五脏六腑及四肢百骸;肝脏是脂质代谢重要的器官,其参与脂质消化、吸收、转运等功能,脾气健运,肝失疏泄,肝脏脂质代谢功能与"脾"运化功能相关。肝细胞自噬是维持肝脏脂质代谢稳态的关键因素之一,自噬功能正常则肝脏脂质代谢趋于平衡。本文基于脾与脂质相关性,从"脾主运化"探讨肝细胞自噬对脂质代谢的影响及机制。     

Autophagy,mitochondria and 3-nitropropionic acid joined in the same model

Huntington''s disease (HD) is a neurodegenerative disorder caused by a mutation in the gene encoding the huntingtin protein. Although the precise mechanism by which neuronal degeneration occurs is still unclear, several elements are important to its development: (1) altered gene expression and protein synthesis, (2) mitochondrial damage and (3) improper regulation of the autophagy programme. In this issue of British Journal of Pharmacology, Galindo and co-workers provide the first evidence for a role of the mitochondrial permeability transition pore (mPTP) in mitochondrial fragmentation and autophagy activation. In a model of cell death induced by 3-nitropropionic acid (3-NP) in human neural cells, the authors describe clear functions for mPTP and Bax, but not the mitochondrial fusion/fission machinery, mitochondrial fragmentation and autophagy (mitophagy). This commentary summarises the significance of this relationship and suggests several points for future development.