Biochemical mechanisms of apoptosis: ordering of the biochemical events in chemical-induced apoptosis.

Apoptosis is an important form of cell death induced by anti-cancer drugs and chemicals. The biochemical mechanisms of chemical-induced apoptosis remain, however, largely unknown, and the major biochemical events in this model of apoptosis are poorly characterised. In this study, apoptosis, induced in human monocytic THP.l cells by etoposide and N-tosyl-L-phenylalanyl chloromethyl ketone and in leukaemic U937 cells by etoposide, was accompanied by the release of mitochondrial cytochrome c, activation of caspases, proteolysis of poly(ADP-ribose) polymerase (PARP), DNA fragmentation, externalisation of phosphatidylserine (PS), and reduction in mitochondrial membrane potential (ɅѰm). Time course studies demonstrated that activation of caspases occurred prior to both PS externalisation and reduction in A\|/m. This was further supported by the observation that the caspase inhibitor, benzyloxycarbonyl-Val-Ala-Asp (OMe) fluoromethyl ketone (Z-VAD.FMK) inhibited all the ultrastructural and biochemical characteristics of apoptosis except the release of cytochrome c. The execution phase of apoptosis may thus be initiated by the translocation of cytochrome c to the cytosol, where it interacts with dATP and apoptotic-protease activating factor-1, resulting in activation of the initiator caspase-9, which in turn activates effector caspases, such as -3 and -7. Cytochrome c release was a later event in non-apoptotic cell death occurring after commitment to cell death and without caspase activation. Thus its release may represent a commitment to cell death in chemical-induced apoptosis. This study also showed that in cells triggered into apoptosis the plasma membrane changes leading to recognition and phagocytosis may be uncoupled from other features of apoptosis by using the mitochondrial inhibitors, antimycin A and oligomycin. These inhibitors blocked increased plasma membrane permeability, PS externalisation and recognition by two classes of phagocytes but not activation of caspases-3 and -7, PAR? cleavage or DNA fragmentation in THP.l cells. PS externalisation was also dissociated from caspase activation in U937 cells. Thus cell surface changes governing safe clearance of apoptotic cells may be regulated by an independent pathway to those mediated by caspases. This finding may have important consequences for attempts to manipulate cell death for therapeutic gain in vivo. Finally, this study demonstrated that in chemical-induced apoptosis caspases functioned solely as executioners and activation of caspases occurred after commitment to cell death, through post-mitochondrial activation pathway(s). Z- VAD.FMK inhibited apoptosis at a stage after this commitment by blocking the post- mitochondrial activation cascade of caspases. This contrasted to its effect in death receptor-mediated apoptosis in U937 cells, where Z.VAD.FMK inhibited apoptosis prior to commitment to cell death induced by tumour necrosis factor-ɑ

Passive Limb Movement Augments Ventilatory Response to CO\u3csub\u3e2\u3c/sub\u3e via Sciatic Inputs in Anesthetized Rats

Passive limb movement (PLM) in humans induces a phasic hyperpnea, but the underlying physiological mechanisms remain unclear. We asked whether PLM in anesthetized rats would produce a similar phasic hyperpnea associated with an augmented ventilatory (V̇E) response to CO2 that is dependent on sciatic afferents. The animals underwent 5 min threshold PLM, 3 min hypercapnia (5% CO2), and their combination (CO2 exposure at the end of 2nd min of 5-min PLM) before and after bilateral transection of the sciatic nerves. We found that a threshold PLM evoked a phasic hyperpnea, similar to that denoted in humans, and an augmented (V̇E) response to CO2. Both responses were greatly diminished by sciatic nerve transection. Moreover, similar responses were also evoked by electrically stimulating the central end of the transected sciatic nerve. Our findings suggest an ability of the sciatic afferents to augment the (V̇E) response to CO2 that likely contributes to the PLM-induced hyperpnea

Class Attention to Regions of Lesion for Imbalanced Medical Image Recognition

Automated medical image classification is the key component in intelligent diagnosis systems. However, most medical image datasets contain plenty of samples of common diseases and just a handful of rare ones, leading to major class imbalances. Currently, it is an open problem in intelligent diagnosis to effectively learn from imbalanced training data. In this paper, we propose a simple yet effective framework, named \textbf{C}lass \textbf{A}ttention to \textbf{RE}gions of the lesion (CARE), to handle data imbalance issues by embedding attention into the training process of \textbf{C}onvolutional \textbf{N}eural \textbf{N}etworks (CNNs). The proposed attention module helps CNNs attend to lesion regions of rare diseases, therefore helping CNNs to learn their characteristics more effectively. In addition, this attention module works only during the training phase and does not change the architecture of the original network, so it can be directly combined with any existing CNN architecture. The CARE framework needs bounding boxes to represent the lesion regions of rare diseases. To alleviate the need for manual annotation, we further developed variants of CARE by leveraging the traditional saliency methods or a pretrained segmentation model for bounding box generation. Results show that the CARE variants with automated bounding box generation are comparable to the original CARE framework with \textit{manual} bounding box annotations. A series of experiments on an imbalanced skin image dataset and a pneumonia dataset indicates that our method can effectively help the network focus on the lesion regions of rare diseases and remarkably improves the classification performance of rare diseases.Comment: Accepted by Neurocomputing on July 2023. 37 page

Myocyte-Specific Overexpressing HDAC4 Promotes Myocardial Ischemia/Reperfusion Injury

Background: Histone deacetylases (HDACs) play a critical role in modulating myocardial protection and cardiomyocyte survivals. However, Specific HDAC isoforms in mediating myocardial ischemia/reperfusion injury remain currently unknown. We used cardiomyocyte-specific overexpression of active HDAC4 to determine the functional role of activated HDAC4 in regulating myocardial ischemia and reperfusion in isovolumetric perfused hearts. Methods: In this study, we created myocyte-specific active HDAC4 transgenic mice to examine the functional role of active HDAC4 in mediating myocardial I/R injury. Ventricular function was determined in the isovolumetric heart, and infarct size was determined using tetrazolium chloride staining. Results: Myocyte-specific overexpressing activated HDAC4 in mice promoted myocardial I/R injury, as indicated by the increases in infarct size and reduction of ventricular functional recovery following I/R injury. Notably, active HDAC4 overexpression led to an increase in LC-3 and active caspase 3 and decrease in SOD-1 in myocardium. Delivery of chemical HDAC inhibitor attenuated the detrimental effects of active HDAC4 on I/R injury, revealing the pivotal role of active HDAC4 in response to myocardial I/R injury. Conclusions: Taken together, these findings are the first to define that activated HDAC4 as a crucial regulator for myocardial ischemia and reperfusion injury

Development of a cell-line model to mimic the pro-survival effect of nurse-like cells in chronic lymphocytic leukemia

The interaction between Chronic lymphocytic leukemia (CLL) cells and monocyte-derived nurse-like cells (NLCs) is fundamentally important to CLL biology. However, studies of how CLL cells and NLCs interact have been hampered by the need for freshly obtained CLL blood samples, coupled with wide variation in the number of monocytes present in the blood of individual patients. Here, we report the development and validation of a cell-line model of NLCs which overcomes these difficulties. Co-culture of primary CLL cells with THP-1 cells induced to differentiate into macrophages by phorbol 12-myristate 13-acetate (PMA) significantly reduced both spontaneous and fludarabine-induced cell death of leukemic cells. Furthermore, compared with their M1-polarized counterparts, M2-polarized macrophages derived from PMA-differentiated THP-1 cells conferred to CLL cells greater protection from spontaneous and fludarabine-induced apoptosis. Since NLCs resemble M2 tumor-associated macrophages, this cell-line model could be useful for investigating the mechanisms through which NLCs protect CLL cells from spontaneous and drug-induced apoptosis

LEPREL1 Expression in Human Hepatocellular Carcinoma and Its Suppressor Role on Cell Proliferation

Background. Hepatocellular carcinoma (HCC) is one of the most aggressive malignancies worldwide. It is characterized by its high invasive and metastatic potential. Leprecan-like 1 (LEPREL1) has been demonstrated to be downregulated in the HCC tissues in previous proteomics studies. The present study is aimed at a new understanding of LEPREL1 function in HCC. Methods. Quantitative RT-PCR, immunohistochemical analysis, and western blot analysis were used to evaluate the expression of LEPREL1 between the paired HCC tumor and nontumorous tissues. The biology function of LEPREL1 was investigated by Cell Counting Kit-8 (CCK8) assay and colony formation assay in HepG2 and Bel-7402 cells. Results. The levels of LEPREL1 mRNA and protein were significantly lower in the HCC tissues as compared to those of the nontumorous tissues. Reduced LEPREL1 expression was not associated with conventional clinical parameters of HCC. Overexpression of LEPREL1 in HepG2 and Bel-7402 cells inhibited cell proliferation (P<0.01) and colony formation (P<0.05). LEPREL1 suppressed tumor cell proliferation through regulation of the cell cycle by downregulation of cyclins. Conclusions. Clinical parameters analysis suggested that LEPREL1 was an independent factor in the development of HCC. The biology function experiments showed that LEPREL1 might serve as a potential tumor suppressor gene by inhibiting the HCC cell proliferation