Adiponectin up-regulates the decrease of myocardial autophagic flux induced by β1-adrenergic receptor autoantibody partly dependent on AMPK

Cardiomyocytes autophagy is essential for maintaining cardiac function. Our previous studies have found that β₁-adrenergic receptor autoantibody (β₁-AA) induced the decreased myocardial autophagic flux, which resulted in cardiomyocyte death and cardiac dysfunction. And other studies demonstrated that β₁-AA induced the decrease of AMPK phosphorylation, the key hub of autophagy pathway, while adiponectin up-regulated autophagic flux mediated by AMPK. However, it is not clear whether adiponectin improves the inhibition of myocardial autophagic flux induced by β₁-AA by up-regulating the level of AMPK phosphorylation. In this study, it has been confirmed that β₁-AA induced the decrease of AMPK phosphorylation level in both vivo and vitro. Moreover, pretreatment of cardiomyocytes with AMPK inhibitor Compound C could further reduce the autophagic flux induced by β₁-AA. Adiponectin deficiency could aggravate the decrease of myocardial AMPK phosphorylation level, autophagic flux and cardiac function induced by β₁-AA. Further, exogenous adiponectin could reverse the decline of AMPK phosphorylation level and autophagic flux induced by β₁-AA and even reduce cardiomyocyte death. While pretreated with the Compound C, the adiponectin treatment did not improve the decreased autophagosome formation, but still improved the decreased autophagosome clearance induced by β₁-AA in cardiomyocytes. This study is the first time to confirm that β₁-AA could inhibit myocardial autophagic flux by down-regulating AMPK phosphorylation level. Adiponectin could improve the inhibition of myocardial autophagic flux induced by β₁-AA partly dependent on AMPK, so as to provide an experimental basis for the treatment of patients with β₁-AA-positive cardiac dysfunction.     

Selective targeting of breast cancer cells through ROS-mediated mechanisms potentiates the lethality of paclitaxel by a novel diterpene, gelomulide K

Defects in apoptotic pathways confer resistance to tubulin-binding agents via downregulation of caspases or overexpression of antiapoptotic factors, urging the need for novel agents acting on an alternative pathway. The purpose of this study was to investigate whether induction of ROS can induce caspase-independent cell death in breast cancer cells and thereby enhance the activity of paclitaxel. Here, we report that gelomulide K acts as a caspase-independent cell death-inducing agent that synergizes with paclitaxel in breast cancer cells and has low toxicity in normal cells. Treatment with gelomulide K induced PARP-1 hyperactivation, AIF nuclear translocation, and cytoprotective autophagy. These effects were associated with increased ROS production and a decrease in cellular GSH levels in cancer cells. Furthermore, pretreatment with NAC, a precursor of intracellular GSH, effectively abrogated gelomulide K-induced caspase-independent cell death and autophagy, suggesting that ROS-mediated downstream signaling is essential to the anticancer effects of gelomulide K. Additionally, in a xenograft model, gelomulide K induced PARP-1 activation and reduced tumor growth. In terms of structure-activity relationships, analysis not only showed a correlation between ROS levels and drug activity but also highlighted the importance of the 8,14-epoxy group. Taken together, our results show that enhancement of paclitaxel activity can be achieved with gelomulide K and that the structurally relevant pharmacophore provides important insight into the development of new caspase-independent cell death-inducing agents.     

The vacuolated morphology of chordoma cells is dependent on cytokeratin intermediate filaments

Notochordal cells (NCs), characterized by their vacuolated morphology and coexpression of cytokeratin and vimentin intermediate filaments (IFs), form the immature nucleus pulposus (NP) of the intervertebral disc. As humans age, NCs give way to mature NP cells, which do not possess a vacuolated morphology and typically only express vimentin IFs. In light of their concomitant loss, we investigated the relationship between cytosolic vacuoles and cytokeratin IFs, specifically those containing cytokeratin-8 proteins, using a human chordoma cell line as a model for NCs. We demonstrate that the chemical disruption of IFs with acrylamide, F-actin with cytochalasin-D, and microtubules with nocodazole all result in a significant (p < 0.001) decrease in vacuolation. However, vacuole loss was the greatest in acrylamide-treated cells. Examination of the individual roles of vimentin and cytokeratin-8 IFs in the existence of vacuoles was accomplished using small interfering RNA–mediated RNA interference to knock down either vimentin or cytokeratin-8 expression. Reduction of cytokeratin-8 expression was associated with a less-vacuolated cell morphology. These data demonstrate that cytokeratin-8 IFs are involved in stabilizing vacuoles and that their diminished expression could play a role in the loss of vacuolation in NCs during aging. A better understanding of the NCs may assist in preservation of this cell type for NP maintenance and regeneration.     

The role of the Golgi complex in the isolation and digestion of organelles

The origin of the membranes and lytic enzymes involved in autophagy has been studied in metamorphosing insect fat body.The Golgi complex has two functions in the organelle destruction which takes place when fat body cells change their activities. (1) It gives rise to envelopes which externalize organelles scheduled for destruction. Microbodies, mitochondria and rough endoplasmic reticulum are sequentially removed from the cytoplasm by investment in isolation membranes. During the isolating phase, isolation membranes have the same osmiophilia as the outer saccular and microvesicular components of the Golgi complex, they do not contain lytic enzymes and they are specific in their adhesion to organelles scheduled for destruction. (2) The Golgi complex gives rise to lytic enzymes. Primary lysosomes which contain acid phosphatase fuse with the isolation bodies formed from invested organelles to become autophagic vacuoles. During this lytic phase, acid phosphatase is present in the inner saccules and microvesicular components of the Golgi complex, in the primary lysosomes seen fusing with isolation bodies and in autophagic vacuoles.     

Inactivation of the PPN1 gene exerts different effects on the metabolism of inorganic polyphosphates in the cytosol and the vacuoles of the yeast Saccharomyces cerevisiae

Inactivation of the PPN1 gene, encoding one of the enzymes involved in polyphosphate metabolism in the yeast Saccharomyces cerevisiae, was found to decrease exopolyphosphatase activity in the cytosol and vacuoles. This effect was more pronounced in the stationary growth phase than in the phase of active growth. The gene inactivation resulted in elimination of a 440-kDa exopolyphosphatase in the vacuoles but did not influence a previously unknown vacuolar exopolyphosphatase with a molecular mass of >1000 kDa, which differed from the former enzyme in the requirement for bivalent cations and sensitivity to heparin. Inactivation of the PPN1 gene did not influence the level of polyphosphates in the cytosol but increased it more than twofold in the vacuoles. In this case, the polyphosphate chain length in the cytosol increased from 10–15 to 130 phosphate residues both in the stationary and active growth phases. In the vacuoles, the polyphosphate length increased only in the stationary growth phase. A conclusion can be made that the PPN1 gene product has different effects on polyphosphate metabolism in the cytosol and the vacuoles.     

鱼腥藻7120三类液泡三种内含物的比较分析

分别从培养4d,24d和KCl诱导的材料分离液泡,对这3种液泡进行了蛋白质、还原糖和藻青蛋白测定,结果表明,3种物质含量呈现规律性变化。培养4d的材料液泡中各物质含量低,培养24d的材料液泡中物质含量升高,KCl诱导的液泡中含量下降,液泡中各种物质的相对含量在3种液泡中依次升高。这一结果说明,培养4d,液泡处于初期阶段,培养24d,液泡处于充分发育阶段,KCl诱导液泡为衰老阶段。随着细胞发育,液泡的生理作用提高。     

鱼腥藻7120细胞液泡内含物的初步测定

对鱼腥藻7120细胞液泡内含物中4种水溶性的物质进行了测定,液泡中4种物质占整个细胞中4种物质的比例分别为:蛋白质:14.1％;还原糖:34.4％;核酸:28.5％;藻青蛋白12.1％。     

Thymoquinone Suppresses the Proliferation,Migration and Invasiveness through Regulating ROS,Autophagic Flux and miR-877-5p in Human Bladder Carcinoma Cells

Bladder carcinoma is among the top 10 most frequently diagnosed cancer types in the world. As a phytochemical active metabolic, thymoquinone (TQ) is extracted from seeds of Nigella sativa, possessing various biological properties in a wide range of diseases. Moreover, the outstanding anti-cancer effect of TQ is attracting increasing attentions. In certain circumstances, moderate autophagy is regarded to facilitate the adaptation of malignant cells to different stressors. Conversely, closely linked with the mitochondrial membrane potential (MMP) loss, the upregulation of intracellular reactive oxygen species (ROS) is reported to activate the cell apoptosis in many cancer types. Furthermore, the vital effects of microRNAs in the pathological processes of cancer cells have also been confirmed by previous studies. The present research confirms that TQ restrains the viability, proliferation, migration and invasion through activating caspase-dependent apoptosis in bladder carcinoma cells, which is mediated by TQ induced ROS increase in bladder carcinoma cells. Furthermore, TQ is proved to block the fusion of autophagosomes and lysosomes, causing the accumulation of autophagosomes and subsequent cell apoptosis. In addition, TQ is also found to initiate the miR-877-5p/PD-L1 axis, which suppresses the epithelial mesenchymal transition (EMT) and invasion of bladder carcinoma cells. Taken together, TQ induces the apoptosis through upregulating ROS level and impairing autophagic flux, and inhibiting the EMT and cell invasion via activating the miR-877-5p/PD-L1 axis in bladder carcinoma cells.     

阔口尖毛虫形成包囊期间细胞超微结构的观察

阔口尖毛虫形成囊期间,细胞质内出现条带状或管产产的内质网和由不同大小的囊泡组成的包囊壁前体。并且,前体的产生与内质网有关;细胞质内发生自噬泡消化现象,这是细胞将原有结构和能量进行贮存,利用的一种重要形式;大核向细胞质突出形成阿米巴形结构,这与大核向细胞质排出部分核物质有关。