Filming a live cell by scanning electrochemical microscopy: label-free imaging of the dynamic morphology in real time

The morphology of a live cell reflects the organization of the cytoskeleton and the healthy status of the cell. We established a label-free platform for monitoring the changing morphology of live cells in real time based on scanning electrochemical microscopy (SECM). The dynamic morphology of a live human bladder cancer cell (T24) was revealed by time-lapse SECM with dissolved oxygen in the medium solution as the redox mediator. Detailed local movements of cell membrane were presented by time-lapse cross section lines extracted from time-lapse SECM. Vivid dynamic morphology is presented by a movie made of time-lapse SECM images. The morphological change of the T24 cell by non-physiological temperature is in consistence with the morphological feature of early apoptosis. To obtain dynamic cellular morphology with other methods is difficult. The non-invasive nature of SECM combined with high resolution realized filming the movements of live cells

Hedgehog Inhibition Promotes a Switch from Type II to Type I Cell Death Receptor Signaling in Cancer Cells

TRAIL is a promising therapeutic agent for human malignancies. TRAIL often requires mitochondrial dysfunction, referred to as the Type II death receptor pathway, to promote cytotoxicity. However, numerous malignant cells are TRAIL resistant due to inhibition of this mitochondrial pathway. Using cholangiocarcinoma cells as a model of TRAIL resistance, we found that Hedgehog signaling blockade sensitized these cancer cells to TRAIL cytotoxicity independent of mitochondrial dysfunction, referred to as Type I death receptor signaling. This switch in TRAIL requirement from Type II to Type I death receptor signaling was demonstrated by the lack of functional dependence on Bid/Bim and Bax/Bak, proapoptotic components of the mitochondrial pathway. Hedgehog signaling modulated expression of X-linked inhibitor of apoptosis (XIAP), which serves to repress the Type I death receptor pathway. siRNA targeted knockdown of XIAP mimics sensitization to mitochondria-independent TRAIL killing achieved by Hedgehog inhibition. Regulation of XIAP expression by Hedgehog signaling is mediated by the glioma-associated oncogene 2 (GLI2), a downstream transcription factor of Hedgehog. In conclusion, these data provide additional mechanisms modulating cell death by TRAIL and suggest Hedgehog inhibition as a therapeutic approach for TRAIL-resistant neoplasms

“Where, O Death, Is Thy Sting?” A Brief Review of Apoptosis Biology

Apoptosis was a term introduced in 1972 to distinguish a mode of cell death with characteristic morphology and apparently regulated, endogenously driven mechanisms. The effector processes responsible for apoptosis are now mostly well known, involving activation of caspases and Bcl2 family members in response to a wide variety of physiological and injury-induced signals. The factors that lead of the decision to activate apoptosis as opposed to adaptive responses to such signals (e.g. autophagy, cycle arrest, protein synthesis shutoff) are less well understood, but the intranuclear Promyelocytic Leukaemia Body (PML body) may create a local microenvironment in which the audit of DNA damage may occur, informed by the extent of the damage, the adequacy of its repair and other aspects of cell status

The inhibition of Bid expression by Akt leads to resistance to TRAIL-induced apoptosis in ovarian cancer cells

Epithelial ovarian cancer (EOC) cells often show increased activity of the PI3K/Akt pathway. In addition, we have previously shown that EOC ascites induce Akt activation in the tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-sensitive EOC cell line, CaOV3, leading to TRAIL-mediated apoptosis inhibition. In this study, we investigated the role of Akt in intrinsic resistance to TRAIL, which is common in EOC cells. We report that Akt activation reduces the sensitivity of EOC cells to TRAIL. TRAIL-resistant SKOV3ip1 and COV2 cells were sensitized to TRAIL-induced apoptosis by PI3K or Akt inhibitors although inhibition of PI3K/Akt signaling pathway did not interfere with the recruitment and processing of caspase-8 to the death-inducing signaling complex. Conversely, overexpression of Akt1 in TRAIL-sensitive cells promoted resistance to TRAIL. Although the fact that TRAIL-induced caspase-8 activation was observed in both sensitive and resistant cell lines, Bid cleavage occurred only in sensitive cells or in SKOV3ip1 cells treated with LY294002. Bid expression was low in resistant cells and Akt activation downregulated its expression. Depletion of Bid by siRNA in OVCAR3 cells was associated with a decrease in TRAIL-mediated apoptosis. Overexpression of Bid only in SKOV3ip1 cells enhanced TRAIL-induced apoptosis. Simultaneous blockade of Akt pathway further increased TRAIL-induced apoptosis. Thus, Akt acts upstream of mitochondria and inhibits TRAIL-induced apoptosis by decreasing Bid protein levels and possibly inhibiting its cleavage

TNFα Cooperates with IFN-γ to Repress Bcl-xL Expression to Sensitize Metastatic Colon Carcinoma Cells to TRAIL-mediated Apoptosis

BACKGROUND: TNF-related apoptosis-inducing ligand (TRAIL) is an immune effector molecule that functions as a selective anti-tumor agent. However, tumor cells, especially metastatic tumor cells often exhibit a TRAIL-resistant phenotype, which is currently a major impediment in TRAIL therapy. The aim of this study is to investigate the synergistic effect of TNFα and IFN-γ in sensitizing metastatic colon carcinoma cells to TRAIL-mediated apoptosis. METHODOLOGY/PRINCIPAL FINDINGS: The efficacy and underlying molecular mechanism of cooperation between TNFα and IFN-γ in sensitizing metastatic colon carcinoma cells to TRAIL-mediated apoptosis were examined. The functional significance of TNFα- and IFN-γ-producing T lymphocyte immunotherapy in combination with TRAIL therapy in suppression of colon carcinoma metastasis was determined in an experimental metastasis mouse model. We observed that TNFα or IFN-γ alone exhibits minimal sensitization effects, but effectively sensitized metastatic colon carcinoma cells to TRAIL-induced apoptosis when used in combination. TNFα and IFN-γ cooperate to repress Bcl-xL expression, whereas TNFα represses Survivin expression in the metastatic colon carcinoma cells. Silencing Bcl-xL expression significantly increased the metastatic colon carcinoma cell sensitivity to TRAIL-induced apoptosis. Conversely, overexpression of Bcl-xL significantly decreased the tumor cell sensitivity to TRAIL-induced apoptosis. Furthermore, TNFα and IFN-γ also synergistically enhanced TRAIL-induced caspase-8 activation. TNFα and IFN-γ was up-regulated in activated primary and tumor-specific T cells. TRAIL was expressed in tumor-infiltrating immune cells in vivo, and in tumor-specific cytotoxic T lymphocytes (CTL) ex vivo. Consequently, TRAIL therapy in combination with TNFα/IFN-γ-producing CTL adoptive transfer immunotherapy effectively suppressed colon carcinoma metastasis in vivo. CONCLUSIONS/SIGNIFICANCE: TNFα and IFN-γ cooperate to overcome TRAIL resistance at least partially through enhancing caspase 8 activation and repressing Bcl-xL expression. Combined CTL immunotherapy and TRAIL therapy hold great promise for further development for the treatment of metastatic colorectal cancer

Mécanisme moléculaires de l'acquisition d'une résistance à l'apoptose induite par TRAIL et à l'anoïkis par des cellules humaines coliques métastatiques

PARIS7-Bibliothèque centrale (751132105) / SudocSudocFranceF