The Zebrafish Antiapoptotic Protein BIRC2 Promotes Edwardsiella piscicida Infection by Inhibiting Caspases and Accumulating p53 in a p53 Transcription-Dependent and -Independent Manner

IAPs (inhibitors of apoptosis) are endogenous caspase inhibitors with multiple biological activities. In the present study, we show functional characteristics of antiapoptotic protein BIRC2 (cIAP1) in response to Edwardsiella piscicida infection. Overexpression of BIRC2 in zebrafish larvae promoted the proliferation of E. piscicida, leading to a decreased larvae survival. The expression levels of caspases including casp3, casp8, and casp9 were significantly inhibited by BIRC2 overexpression in the case of E. piscicida infection. Treatment of zebrafish larvae microinjected with BIRC2 with the caspase activator PAC-1 completely blocked the negative regulation of BIRC2 on the E. piscicida infection, with the reduced inhibition on the casp3 and without inhibition on casp8 and casp9. In contrast to the regulation of BIRC2 on the caspases, BIRC2 overexpression significantly induced the expression of p53, especially at 24 hpi. In addition to the cytoplasmic p53 expression, BIRC2 overexpression also induced the expression of the nuclear p53 protein. Further analysis demonstrated that BIRC2 could interact and colocalize with p53 in the cytoplasm. The numbers of E. piscicida in larvae overexpressed with BIRC2 and treated with pifithrin-μ (an inhibitor of mitochondrial p53) or pifithrin-α (an inhibitor of p53 transactivation) were lower than those of larvae without pifithrin-μ or pifithrin-α treatment. Critically, the p53 inactivators pifithrin-μ and pifithrin-α had no significant effect on larval survival, but completely rescued larval survival for zebrafish microinjected with BIRC2 in the case of E. piscicida infection. Collectively, the present study suggest that piscine BIRC2 is a negative regulator for antibacterial immune response in response to the E. piscicida infection via inhibiting caspases, and accumulating p53 in a p53 transcription-dependent and -independent manner.

BCL-XL is crucial for progression through the adenoma-to-carcinoma sequence of colorectal cancer

Evasion of apoptosis is a hallmark of cancer, which is frequently mediated by upregulation of the antiapoptotic BCL-2 family proteins. In colorectal cancer (CRC), previous work has highlighted differential antiapoptotic protein dependencies determined by the stage of the disease. While intestinal stem cells (ISCs) require BCL-2 for adenoma outgrowth and survival during transformation, ISC-specific MCL1 deletion results in disturbed intestinal homeostasis, eventually contributing to tumorigenesis. Colon cancer stem cells (CSCs), however, no longer require BCL-2 and depend mainly on BCL-XL for their survival. We therefore hypothesized that a shift in antiapoptotic protein reliance occurs in ISCs as the disease progresses from normal to adenoma to carcinoma. By targeting antiapoptotic proteins with specific BH3 mimetics in organoid models of CRC progression, we found that BCL-2 is essential only during ISC transformation while MCL1 inhibition did not affect adenoma outgrowth. BCL-XL, on the other hand, was crucial for stem cell survival throughout the adenoma-to-carcinoma sequence. Furthermore, we identified that the limited window of BCL-2 reliance is a result of its downregulation by miR-17-5p, a microRNA that is upregulated upon APC-mutation driven transformation. Here we show that BCL-XL inhibition effectively impairs adenoma outgrowth in vivo and enhances the efficacy of chemotherapy. In line with this dependency, expression of BCL-XL, but not BCL-2 or MCL1, directly correlated to the outcome of chemotherapy-treated CRC patients. Our results provide insights to enable the rational use of BH3 mimetics in CRC management, particularly underlining the therapeutic potential of BCL-XL targeting mimetics in both early and late-stage disease.

Modern aspects of the etiology and pathogenesis of hyperplastic endometrial processes

The article presents modern approaches to the etiology, classification and pathogenesis of endometrial hyperplastic processes. Hyperplastic processes of the endometrium, representing an urgent medical and social problem, which is due to the peculiarities of the clinical manifestation of the disease (uterine bleeding), reduced reproductive potential, high recurrence rate, limited conservative treatment of this pathology in pre- and postmenopausal women, due to concomitant extragenital diseases and a high risk of malignancy.On the basis of new criteria of pathomorphological diagnostics, drawing parallels with the accumulated experience and clinical data, the immunohistochemical profile for each histotype of hyperplastic process of endometrium is created, certain diagnostic determinants are defined. Namely, simple and complex endometrial hyperplasia without atypia are characterized by imbalance between proliferation and apoptosis, manifested in moderate activation of the intratissue APUD system, lack of activation of proapoptotic protein p53, increased levels of antiapoptotic protein 2. Atypical forms of endometrial hyperplasia (simple and complex) are characterized by expression in the epithelial component of the proapoptotic protein p53, an increase in the level of antiapoptotic protein Bcl-2 in 2-2.5 times, expressed by activation of the intratissue APUD system.

Targeting MCL-1 in cancer: current status and perspectives

Myeloid leukemia 1 (MCL-1) is an antiapoptotic protein of the BCL-2 family that prevents apoptosis by binding to the pro-apoptotic BCL-2 proteins. Overexpression of MCL-1 is frequently observed in many tumor types and is closely associated with tumorigenesis, poor prognosis and drug resistance. The central role of MCL-1 in regulating the mitochondrial apoptotic pathway makes it an attractive target for cancer therapy. Significant progress has been made with regard to MCL-1 inhibitors, some of which have entered clinical trials. Here, we discuss the mechanism by which MCL-1 regulates cancer cell apoptosis and review the progress related to MCL-1 small molecule inhibitors and their role in cancer therapy.

Early Lipid Raft-Related Changes: Interplay between Unilateral Denervation and Hindlimb Suspension

Muscle disuse and denervation leads to muscle atrophy, but underlying mechanisms can be different. Previously, we have found ceramide (Cer) accumulation and lipid raft disruption after acute hindlimb suspension (HS), a model of muscle disuse. Herein, using biochemical and fluorescent approaches the influence of unilateral denervation itself and in combination with short-term HS on membrane-related parameters of rat soleus muscle was studied. Denervation increased immunoexpression of sphingomyelinase and Cer in plasmalemmal regions, but decreased Cer content in the raft fraction and enhanced lipid raft integrity. Preliminary denervation suppressed (1) HS-induced Cer accumulation in plasmalemmal regions, shown for both nonraft and raft-fractions; (2) HS-mediated decrease in lipid raft integrity. Similar to denervation, inhibition of the sciatic nerve afferents with capsaicin itself increased Cer plasmalemmal immunoexpression, but attenuated the membrane-related effects of HS. Finally, both denervation and capsaicin treatment increased immunoexpression of proapoptotic protein Bax and inhibited HS-driven increase in antiapoptotic protein Bcl-2. Thus, denervation can increase lipid raft formation and attenuate HS-induced alterations probably due to decrease of Cer levels in the raft fraction. The effects of denervation could be at least partially caused by the loss of afferentation. The study points to the importance of motor and afferent inputs in control of Cer distribution and thereby stability of lipid rafts in the junctional and extrajunctional membranes of the muscle.

TOM20-mediated transfer of Bcl2 from ER to MAM and mitochondria upon induction of apoptosis

In this work, we have explored the subcellular localization of Bcl2, a major antiapoptotic protein. In U251 glioma cells, we found that Bcl2 is localized mainly in the ER and is translocated to MAM and mitochondria upon induction of apoptosis; this mitochondrial transfer was not restricted to the demonstrator cell line, even if cell-specific modulations exist. We found that the Bcl2/mitochondria interaction is controlled by TOM20, a protein that belongs to the protein import machinery of the mitochondrial outer membrane. The expression of a small domain of interaction of TOM20 with Bcl2 potentiates its anti-apoptotic properties, which suggests that the Bcl2–TOM20 interaction is proapoptotic. The role of MAM and TOM20 in Bcl2 apoptotic mitochondrial localization and function has been confirmed in a yeast model in which the ER–mitochondria encounter structure (ERMES) complex (required for MAM stability in yeast) has been disrupted. Bcl2–TOM20 interaction is thus an additional player in the control of apoptosis.

Therapeutic Effect of Mongolian Medicine RuXian-I on Hyperplasia of Mammary Gland Induced by Estrogen/Progesterone through CRYAB-Promoted Apoptosis

The traditional Mongolian medicine (TMM) RuXian-I is an empirical formula specifically used for treating the hyperplasia of mammary gland (HMG) in clinic based on the principles of traditional Mongolian medicine, but the treatment mechanism is not completely clear. In this paper, we elaborated the mechanism of RuXian-I in the treatment of HMG induced by estrogen and progestogen from its toxicity and activity. Firstly, RuXian-I exhibited no toxic effect on HMG rats through no changes of body weight and food intake measurement and no pathologic changes of the organs (heart, liver, spleen, lung, and kidney) detected. Secondly, RuXian-I could decrease the increased nipple height and diameter and remarkably relieve the pathologic changes of HMG rats and also alleviate serum sex hormone levels (estradiol (E2), luteinizing hormone (LH), progesterone (P), and testosterone (T)) of HMG rats. Finally, RuXian-I could obviously inhibit the upregulation level of antiapoptotic protein CRYAB of HMG rats and promote mammary gland cell apoptosis of HMG rats via increases of promoting apoptosis protein caspases-3, 8, and 9 and Bax and tumor suppressor protein p53, decreases of antiapoptosis protein Bcl-2, and release of cytochrome c. These results suggested that RuXian-I has protective and therapeutic effects on HMG rats induced by estrogen and progestogen possibly via promoting apoptotic pathway regulated by CRYAB and is a promising agent for treating HMG.

Epigenetic Upregulation of Chicken MicroRNA-16-5p Expression in DF-1 Cells following Infection with Infectious Bursal Disease Virus (IBDV) Enhances IBDV-Induced Apoptosis and Viral Replication

ABSTRACT MicroRNAs (miRNAs) are small noncoding RNAs that regulate gene expression posttranscriptionally by silencing or degrading their targets and play important roles in the host response to pathogenic infection. Although infectious bursal disease virus (IBDV)-induced apoptosis in host cells has been established, the underlying molecular mechanism is not completely unraveled. Here, we show that infection of DF-1 cells by IBDV induced gga-miR-16-5p (chicken miR-16-5p) expression via demethylation of the pre-miR-16-2 (gga-miR-16-5p precursor) promoter. We found that ectopic expression of gga-miR-16-5p in DF-1 cells enhanced IBDV-induced apoptosis by directly targeting the cellular antiapoptotic protein B-cell lymphoma 2 (Bcl-2), facilitating IBDV replication in DF-1 cells. In contrast, inhibition of endogenous miR-16-5p markedly suppressed apoptosis associated with enhanced Bcl-2 expression, arresting viral replication in DF-1 cells. Furthermore, infection of DF-1 cells with IBDV reduced Bcl-2 expression, and this reduction could be abolished by inhibition of gga-miR-16-5p expression. Moreover, transfection of DF-1 cells with gga-miR-16-5p mimics enhanced IBDV-induced apoptosis associated with increased cytochrome c release and caspase-9 and -3 activation, and inhibition of caspase-3 decreased IBDV growth in DF-1 cells. Thus, epigenetic upregulation of gga-miR-16-5p expression by IBDV infection enhances IBDV-induced apoptosis by targeting the cellular antiapoptotic protein Bcl-2, facilitating IBDV replication in host cells. IMPORTANCE Infectious bursal disease (IBD) is an acute, highly contagious, and immunosuppressive disease in young chickens, causing severe economic losses to stakeholders across the globe. Although IBD virus (IBDV)-induced apoptosis in the host has been established, the underlying mechanism is not very clear. Here, we show that infection of DF-1 cells by IBDV upregulated gga-miR-16-5p expression via demethylation of the pre-miR-16-2 promoter. Overexpression of gga-miR-16-5p enhanced IBDV-induced apoptosis associated with increased cytochrome c release and caspase-9 and -3 activation. Importantly, we found that IBDV infection induced expression of gga-miR-16-5p that triggered apoptosis by targeting Bcl-2, favoring IBDV replication, while inhibition of gga-miR-16-5p in IBDV-infected cells restored Bcl-2 expression, slowing down viral growth, indicating that IBDV induces apoptosis by epigenetic upregulation of gga-miR-16-5p expression. These findings uncover a novel mechanism employed by IBDV for its own benefit, which may be used as a potential target for intervening IBDV infection.