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Concept: Anoikis

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Whether responses of cells to extracellular environments affect the induction of apoptotic cell death is poorly understood. The current study aimed to unravel the different effects of culture media employed in vitro as extracellular environments on the susceptibility of cells to apoptosis. We found that apoptosis is stimulated to the higher levels by culturing human HeLa cells in Opti-MEM with unknown components, a medium that is specifically used for transfections, than by culturing cells in Dulbecco’s modified Eagle’s medium, a medium that is generally used for maintenance of cells. We showed that apoptosis is suppressed partially by culturing cells in heat-treated Opti-MEM, implicating a heat-sensitive component(s) in stimulating the apoptotic response of cells. Thus, different extracellular environments may contribute to different responses of cells to apoptosis, and this should be considered to evaluate the incidences of apoptotic cell death and could be applied to develop an efficient treatment for curing diseases such as cancer.

Concepts: Cancer, Death, Apoptosis, Cell culture, Programmed cell death, Vesicle, Caspase, Anoikis

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Apoptosis has been implicated in compensatory proliferation signaling (CPS), whereby dying cells induce proliferation in neighboring cells as a means to restore homeostasis. The nature of signaling between apoptotic cells and their neighboring cells remains largely unknown. Here we show that a fraction of apoptotic cells produce and release CrkI-containing microvesicles (distinct from exosomes and apoptotic bodies), which induce proliferation in neighboring cells upon contact. We provide visual evidence of CPS by videomicroscopy. We show that purified vesicles in vitro and in vivo are sufficient to stimulate proliferation in other cells. Our data demonstrate that CrkI inactivation by ExoT bacterial toxin or by mutagenesis blocks vesicle formation in apoptotic cells and inhibits CPS, thus uncoupling apoptosis from CPS. We further show that c-Jun amino-terminal kinase (JNK) plays a pivotal role in mediating vesicle-induced CPS in recipient cells. CPS could have important ramifications in diseases that involve apoptotic cell death.

Concepts: Immune system, Signal transduction, Death, Apoptosis, Programmed cell death, Vesicle, Caspase, Anoikis

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Synaptic activity increases the resistance of neurons to diverse apoptotic insults; however, the underlying mechanisms remain less well understood. Zinc promotes cell survival under varied conditions, but the role of synaptically released zinc in the activity-dependent anti-apoptotic effect is unknown. Using cultured hippocampal slices and primary neurons we show that a typical apoptosis inducer-staurosporine (STP) was able to cause concentration-dependent apoptotic cell death in brain slices; Enhanced synaptic activity by bicuculline (Bic)/4-Aminopyridine (AP) treatment effectively prevented neurons from STP-induced cell apoptosis, as indicated by increased cell survival and suppressed caspase-3 activity. Application of Ca-EDTA, a cell membrane-impermeable zinc chelator which can efficiently capture the synaptically released zinc, completely blocked the neuronal activity-dependent anti-apoptotic effect. Same results were also observed in cultured primary hippocampal neurons. Therefore, our results indicate that synaptic activity improves neuronal resistance to apoptosis via synaptically released zinc.

Concepts: Brain, Apoptosis, Membrane potential, Programmed cell death, Caspase, XIAP, Anoikis

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The aims of this study were to identify a robust apoptosis marker suitable for both quantification and back-to-back analyses of programmed cell death and to define specific upstream targets for apoptosis in corneal cells.

Concepts: DNA, Cell nucleus, Secretion, Apoptosis, Programmed cell death, Vesicle, Caspase, Anoikis

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The expression of microRNA‑206 (miR‑206) is aberrantly induced in steroid‑induced avascular necrosis of femoral head (SANFH). Therefore, investigating the function of miR‑206 in SANFH and uncovering the functional mechanism associated with the condition will promote the understanding and treatment of the disease. The purpose of the present study was to investigate the pro‑osteoclasteogenic effect of miR‑206 that occurs through regulation of programmed cell death 4 (PDCD4). The expression of miR‑206 and PDCD4 was analyzed in the clinical SANFH specimens. The level of miR‑206 and PDCD4 was regulated in human osteoblast lineage hFOB1.19 and the effect of different treatments on cell viability, proliferation, apoptosis and differentiation potential of osteoblasts were analyzed with a Cell Counting kit‑8, 5‑ethynyl‑2'‑deoxyuridine staining, flow cytometry and Hoechst staining. The expression of miR‑206 was upregulated while PDCD4 was downregulated in the SANFH specimens. Induced expression of miR‑206 decreased cell viability and proliferation, while apoptosis was induced. At the molecular level, overexpression of miR‑206 inhibited the expression of PDCD4, alkaline phosphatase (ALP) and B‑cell lymphoma 2 (Bcl‑2), and increased the expression of apoptosis regulator Bcl2‑X‑associated protein (Bax). Inhibiting the expression of miR‑206 increased cell viability and proliferation but had no effect on cell apoptosis, as detected by flow cytometry and Hoechst staining. However, at the molecular level, inhibiting the expression of miR‑206 induced expression of PDCD4, ALP and Bcl‑2, while it decreased the expression of Bax. Additionally, knockdown of PDCD4 blocked the effect of miR‑206 inhibition on hFOB1.19 cells, representing a PDCD4‑dependent manner of miR‑206 in inducing apoptosis of osteoblasts. Therefore, miR‑206 promoted the onset of SANFH by inducing apoptosis and suppressed the proliferation of osteoblasts, which was dependent on the inhibition of PDCD4.

Concepts: DNA, Gene expression, Apoptosis, Avascular necrosis, Necrosis, Programmed cell death, Caspase, Anoikis

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Apoptosis is a form of programmed cell death that is carried out by proteolytic enzymes called caspases. Executioner caspase activity causes cells to shrink, bleb, and disintegrate into apoptotic bodies and has been considered a point of no return for apoptotic cells. However, relatively recent work has shown that cells can survive transient apoptotic stimuli, even after executioner caspase activation. This process is called anastasis. In this Q&A, we answer common questions that arise regarding anastasis, including how it is defined, the origin of the name, the potential physiological consequences, molecular mechanisms, and open questions for this new field of study.

Concepts: Enzyme, Apoptosis, Programmed cell death, Caspase, Bcl-2, Caspase-9, Apoptosome, Anoikis

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Contrast medium (CM) is widely used in cardiac catheterization; however, it may induce acute kidney injury or renal failure, although the underlying mechanism remains to be elucidated. MicroRNA‑21 (miR‑21) is involved in renal disease and has been indicated to regulate cellular apoptosis and fibrosis, although its role in CM‑induced renal cell injury is unknown. The present study examined the expression and potential targets of miR‑21 in human renal proximal tubular epithelial (HK‑2) cells following CM treatment. CM induced renal cell apoptosis and decreased miR‑21 expression. The expression level of the apoptosis regulator protein, B‑cell lymphoma 2 (Bcl‑2) was upregulated, whereas that of the apoptosis regulator, Bcl‑2‑associated X protein (Bax) was downregulated upon transfection of miR‑21 mimics; miR‑21 overexpression additionally directly inhibited the expression of programmed cell death protein 4 (PDCD4), as determined by a dual luciferase reporter assay, and PDCD4 silencing reduced the rate of HK‑2 cell apoptosis. The results of the present study indicated that miR‑21 protected renal cells against CM‑induced apoptosis by regulating PDCD4 expression.

Concepts: DNA, Renal failure, Kidney, Gene expression, Apoptosis, Programmed cell death, Caspase, Anoikis

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Cartilage injury induced by acute excessive contact stress is common and mostly affects young adult. Although early detection of cartilage injury may prevent serious and lifelong arthritic complications, early detection and treatment is not possible due to the lack of a reliable detection method. Since chondrocyte injury and subsequent cell death are the early signs of cartilage injury, it is likely that cartilage cell apoptosis can be used to predict the extent of injury. To test this hypothesis, a near infrared probe was fabricated to have high affinity to apoptotic cells. In vitro tests show that this apoptosis probe has low toxicity, high specificity, and high affinity to apoptotic cells. In addition, there is a positive relationship between apoptotic cell numbers and fluorescence intensities. Using a mouse xiphoid injury model, we found significant accumulation of the apoptosis probes at the injured xiphoid cartilage site. There was also a positive correlation between probe accumulation and the number of apoptotic chondrocytes within the injured xiphoid cartilage, which was confirmed by TUNEL assay. The results support that the apoptosis probes may serve as a powerful tool to monitor the extent of mechanical force-induced cartilage injury in vivo.

Concepts: Apoptosis, Cartilage, In vitro, Programmed cell death, Vesicle, Caspase, Chondrocyte, Anoikis

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Far from being passive, apoptotic cells influence their environment. For instance, they promote tissue folding, myoblast fusion and modulate tumor growth. Understanding the role of apoptotic cells necessitates their efficient tracking within living tissues, a task which is currently challenging. In order to easily spot apoptotic cells in developing Drosophila tissues, we generated a series of fly lines expressing different fluorescent sensors of caspase activity. We show that three of these reporters (GFP, Cerulean and Venus derived molecules) are detected specifically in apoptotic cells and throughout the whole process of programmed cell death. These reporters allow the specific visualization of apoptotic cells directly within living tissues, without any post-acquisition processing. They overcome the limitations of other apoptosis detection methods developed so far and notably, they can be combined with any kind of fluorophore.

Concepts: Immune system, Cancer, Apoptosis, Programmed cell death, Vesicle, Caspase, Bcl-2, Anoikis