Nucleoporin 62-Like Protein is Required for the Development of Pharyngeal Arches through Regulation of Wnt/β-Catenin Signaling and Apoptotic Homeostasis in Zebrafish

We have previously observed the predominant expression of nucleoporin 62-like (Nup62l) mRNA in the pharyngeal region of zebrafish, which raises the question whether Nup62l has important implications in governing the morphogenesis of pharyngeal arches (PA) in zebrafish. Herein, we explored the functions of Nup62l in PA development. The disruption of Nup62l with a CRISPR/Cas9-dependent gene knockout approach led to defective PA, which was characterized by a thinned and shortened pharyngeal region and a significant loss of pharyngeal cartilages. During pharyngeal cartilage formation, prechondrogenic condensation and chondrogenic differentiation were disrupted in homozygous nup62l-mutants, while the specification and migration of cranial neural crest cells (CNCCs) were unaffected. Mechanistically, the impaired PA region of nup62l-mutants underwent extensive apoptosis, which was mainly dependent on activation of p53-dependent apoptotic pathway. Moreover, aberrant activation of a series of apoptotic pathways in nup62l-mutants is closely associated with the inactivation of Wnt/β-catenin signaling. Thus, these findings suggest that the regulation of Wnt/β-catenin activity by Nup62l is crucial for PA formation in zebrafish.

Download Full-text

Functions of SMC2 in the Development of Zebrafish Liver

Biomedicines ◽

10.3390/biomedicines9091240 ◽

2021 ◽

Vol 9 (9) ◽

pp. 1240

Author(s):

Xixi Li ◽

Guili Song ◽

Yasong Zhao ◽

Jing Ren ◽

Qing Li ◽

...

Keyword(s):

Dna Damage ◽

Chromosome Segregation ◽

Gene Knockout ◽

Chromosome Organization ◽

Liver Development ◽

Apoptotic Pathway ◽

The Core ◽

Apoptotic Pathways ◽

Structural Maintenance Of Chromosomes ◽

Zebrafish Liver

SMC2 (structural maintenance of chromosomes 2) is the core subunit of condensins, which play a central role in chromosome organization and segregation. However, the functions of SMC2 in embryonic development remain poorly understood, due to the embryonic lethality of homozygous SMC2−/− mice. Herein, we explored the roles of SMC2 in the liver development of zebrafish. The depletion of SMC2, with the CRISPR/Cas9-dependent gene knockout approach, led to a small liver phenotype. The specification of hepatoblasts was unaffected. Mechanistically, extensive apoptosis occurred in the liver of SMC2 mutants, which was mainly associated with the activation of the p53-dependent apoptotic pathway. Moreover, an aberrant activation of a series of apoptotic pathways in SMC2 mutants was involved in the defective chromosome segregation and subsequent DNA damage. Therefore, our findings demonstrate that SMC2 is necessary for zebrafish liver development.

Download Full-text

Development of a Decellularized Porcine Esophageal Matrix for Potential Applications in Cancer Modeling

Cells ◽

10.3390/cells10051055 ◽

2021 ◽

Vol 10 (5) ◽

pp. 1055

Author(s):

Hersh Chaitin ◽

Michael L. Lu ◽

Michael B. Wallace ◽

Yunqing Kang

Keyword(s):

Cell Growth ◽

Lymphatic Vessels ◽

Significant Loss ◽

Cancer Cell Growth ◽

Cell Growth And Migration ◽

Decellularized Extracellular Matrix ◽

Cancer Models ◽

The Matrix ◽

Potential Applications ◽

And Migration

Many decellularized extracellular matrix-derived whole organs have been widely used in studies of tissue engineering and cancer models. However, decellularizing porcine esophagus to obtain decellularized esophageal matrix (DEM) for potential biomedical applications has not been widely investigated. In this study a modified decellularization protocol was employed to prepare a porcine esophageal DEM for the study of cancer cell growth. The cellular removal and retention of matrix components in the porcine DEM were fully characterized. The microstructure of the DEM was observed using scanning electronic microscopy. Human esophageal squamous cell carcinoma (ESCC) and human primary esophageal fibroblast cells (FBCs) were seeded in the DEM to observe their growth. Results show that the decellularization process did not cause significant loss of mechanical properties and that blood ducts and lymphatic vessels in the submucosa layer were also preserved. ESCC and FBCs grew on the DEM well and the matrix did not show any toxicity to cells. When FBS and ESCC were cocultured on the matrix, they secreted more periostin, a protein that supports cell adhesion on matrix. This study shows that the modified decellularization protocol can effectively remove the cell materials and maintain the microstructure of the porcine esophageal matrix, which has the potential application of studying cell growth and migration for esophageal cancer models.

Download Full-text

Potential Biomarkers for Treatment Response to the BCL-2 Inhibitor Venetoclax: State of the Art and Future Directions

Cancers ◽

10.3390/cancers13122974 ◽

2021 ◽

Vol 13 (12) ◽

pp. 2974

Author(s):

Haneen T. Salah ◽

Courtney D. DiNardo ◽

Marina Konopleva ◽

Joseph D. Khoury

Keyword(s):

Treatment Response ◽

Gene Mutations ◽

Hematologic Malignancies ◽

Mechanisms Of Resistance ◽

Apoptotic Pathway ◽

Lymphoid Leukemia ◽

Optimal Drug ◽

Trisomy 12 ◽

Apoptotic Pathways ◽

Potential Biomarkers

Intrinsic apoptotic pathway dysregulation plays an essential role in all cancers, particularly hematologic malignancies. This role has led to the development of multiple therapeutic agents targeting this pathway. Venetoclax is a selective BCL-2 inhibitor that has been approved for the treatment of chronic lymphoid leukemia and acute myeloid leukemia. Given the reported resistance to venetoclax, understanding the mechanisms of resistance and the potential biomarkers of response is crucial to ensure optimal drug usage and improved patient outcomes. Mechanisms of resistance to venetoclax include alterations involving the BH3-binding groove, BCL2 gene mutations affecting venetoclax binding, and activation of alternative anti-apoptotic pathways. Moreover, various potential genetic biomarkers of venetoclax resistance have been proposed, including chromosome 17p deletion, trisomy 12, and TP53 loss or mutation. This manuscript provides an overview of biomarkers that could predict treatment response to venetoclax.

Download Full-text

Cadherin-6B proteolysis promotes the neural crest cell epithelial-to-mesenchymal transition through transcriptional regulation

The Journal of Cell Biology ◽

10.1083/jcb.201604006 ◽

2016 ◽

Vol 215 (5) ◽

pp. 735-747 ◽

Cited By ~ 21

Author(s):

Andrew T. Schiffmacher ◽

Vivien Xie ◽

Lisa A. Taneyhill

Keyword(s):

Neural Crest ◽

Epithelial To Mesenchymal Transition ◽

Neural Crest Cell ◽

Cranial Neural Crest ◽

Mesenchymal Transition ◽

Effector Genes ◽

A Disintegrin And Metalloproteinase ◽

And Migration ◽

Cranial Neural Crest Cells

During epithelial-to-mesenchymal transitions (EMTs), cells disassemble cadherin-based junctions to segregate from the epithelia. Chick premigratory cranial neural crest cells reduce Cadherin-6B (Cad6B) levels through several mechanisms, including proteolysis, to permit their EMT and migration. Serial processing of Cad6B by a disintegrin and metalloproteinase (ADAM) proteins and γ-secretase generates intracellular C-terminal fragments (CTF2s) that could acquire additional functions. Here we report that Cad6B CTF2 possesses a novel pro-EMT role by up-regulating EMT effector genes in vivo. After proteolysis, CTF2 remains associated with β-catenin, which stabilizes and redistributes both proteins to the cytosol and nucleus, leading to up-regulation of β-catenin, CyclinD1, Snail2, and Snail2 promoter-based GFP expression in vivo. A CTF2 β-catenin–binding mutant, however, fails to alter gene expression, indicating that CTF2 modulates β-catenin–responsive EMT effector genes. Notably, CTF2 association with the endogenous Snail2 promoter in the neural crest is β-catenin dependent. Collectively, our data reveal how Cad6B proteolysis orchestrates multiple pro-EMT regulatory inputs, including CTF2-mediated up-regulation of the Cad6B repressor Snail2, to ensure proper cranial neural crest EMT.

Download Full-text

Apoptosis mediated chemosensitization of tumor cells to 5-fluorouracil on supplementation of fish oil in experimental colon carcinoma

Tumor Biology ◽

10.1177/1010428317695019 ◽

2017 ◽

Vol 39 (3) ◽

pp. 101042831769501 ◽

Cited By ~ 10

Author(s):

Isha Rani ◽

Bhoomika Sharma ◽

Sandeep Kumar ◽

Satinder Kaur ◽

Navneet Agnihotri

Keyword(s):

Fatty Acids ◽

Dna Damage ◽

Polyunsaturated Fatty Acids ◽

Fish Oil ◽

Tumor Cells ◽

Apoptotic Index ◽

Apoptotic Pathway ◽

Apoptotic Effect ◽

Underlying Mechanisms ◽

Apoptotic Pathways

5-Fluorouracil has been considered as a cornerstone therapy for colorectal cancer; however, it suffers from low therapeutic response rate and severe side effects. Therefore, there is an urgent need to increase the clinical efficacy of 5-fluorouracil. Recently, fish oil rich in n-3 polyunsaturated fatty acids has been reported to chemosensitize tumor cells to anti-cancer drugs. This study is designed to understand the underlying mechanisms of synergistic effect of fish oil and 5-fluorouracil by evaluation of tumor cell–associated markers such as apoptosis and DNA damage. The colon cancer was developed by administration of N,N-dimethylhydrazine dihydrochloride and dextran sulfate sodium salt. Further these animals were treated with 5-fluorouracil, fish oil, or a combination of both. In carcinogen-treated animals, a decrease in DNA damage and apoptotic index was observed. There was also a decrease in the expression of Fas, FasL, caspase 8, and Bax, and an increase in Bcl-2. In contrast, administration of 5-fluorouracil and fish oil as an adjuvant increased both DNA damage and apoptotic index by activation of both extrinsic and intrinsic apoptotic pathways as compared to the other groups. The increased pro-apoptotic effect by synergism of 5-fluorouracil and fish oil may be attributed to the incorporation of n-3 polyunsaturated fatty acids in membrane, which alters membrane fluidity in cancer cells. In conclusion, this study highlights that the induction of apoptotic pathway by fish oil may increase the susceptibility of tumors to chemotherapeutic regimens.

Download Full-text

Mesenchymal stem cells attenuate liver fibrosis by targeting Ly6Chi/lo macrophages through activating the cytokine-paracrine and apoptotic pathways

Cell Death Discovery ◽

10.1038/s41420-021-00584-z ◽

2021 ◽

Vol 7 (1) ◽

Author(s):

Yuan-hui Li ◽

Shuang Shen ◽

Tong Shao ◽

Meng-ting Jin ◽

Dong-dong Fan ◽

...

Keyword(s):

Liver Fibrosis ◽

Molecular Mechanisms ◽

Stellate Cell ◽

Apoptotic Bodies ◽

Chronic Injury ◽

Apoptotic Pathway ◽

Fibrotic Liver ◽

Apoptotic Pathways ◽

Regulatory Functions ◽

Cell Inhibition

AbstractMesenchymal stem cell (MSC) therapy has become a promising treatment for liver fibrosis due to its predominant immunomodulatory performance in hepatic stellate cell inhibition and fibrosis resolution. However, the cellular and molecular mechanisms underlying these processes remain limited. In the present study, we provide insights into the functional role of bone marrow-derived MSCs (BM-MSCs) in alleviating liver fibrosis by targeting intrahepatic Ly6Chi and Ly6Clo macrophage subsets in a mouse model. Upon chronic injury, the Ly6Chi subset was significantly increased in the inflamed liver. Transplantation of BM-MSCs markedly promoted a phenotypic switch from pro-fibrotic Ly6Chi subset to restorative Ly6Clo subpopulation by secreting paracrine cytokines IL-4 and IL-10 from the BM-MSCs. The Ly6Chi/Ly6Clo subset switch significantly blocked the source of fibrogenic TGF-β, PDGF, TNF-α, and IL-1β cytokines from Ly6Chi macrophages. Unexpectedly, BM-MSCs experienced severe apoptosis and produced substantial apoptotic bodies in the fibrotic liver during the 72 h period of transplantation. Most apoptotic bodies were engulfed by Ly6Clo macrophages, and this engulfment robustly triggered MMP12 expression for fibrosis resolution through the PtdSer-MerTK-ERK signaling pathway. This paper is the first to show previously unrecognized dual regulatory functions of BM-MSCs in attenuating hepatic fibrosis by promoting Ly6Chi/Ly6Clo subset conversion and Ly6Clo macrophage restoration through secreting antifibrogenic-cytokines and activating the apoptotic pathway.

Download Full-text

Effects of Hsp70.1 Gene Knockout on the Mitochondrial Apoptotic Pathway After Focal Cerebral Ischemia

Stroke ◽

10.1161/01.str.0000136150.73891.14 ◽

2004 ◽

Vol 35 (9) ◽

pp. 2195-2199 ◽

Cited By ~ 53

Author(s):

Seung-Hoon Lee ◽

Hyung-Min Kwon ◽

Young-Ju Kim ◽

Kyung-Mi Lee ◽

Manho Kim ◽

...

Keyword(s):

Cerebral Ischemia ◽

Focal Cerebral Ischemia ◽

Gene Knockout ◽

Apoptotic Pathway ◽

Mitochondrial Apoptotic Pathway

Download Full-text

Ablation of the MiR-17-92 MicroRNA Cluster in Germ Cells Causes Subfertility in Female Mice

Cellular Physiology and Biochemistry ◽

10.1159/000487028 ◽

2018 ◽

Vol 45 (2) ◽

pp. 491-504 ◽

Cited By ~ 4

Author(s):

Jian Wang ◽

Bo Xu ◽

Geng G. Tian ◽

Tao Sun ◽

Ji Wu

Keyword(s):

Germ Cells ◽

Molecular Mechanisms ◽

Gene Knockout ◽

Conditional Knockout ◽

Luciferase Reporter ◽

Transcriptional Level ◽

Follicular Atresia ◽

Apoptotic Pathway ◽

Female Mice ◽

Important Regulator

Background/Aims: Oogenesis is a highly complex process that is intricately regulated by interactions of multiple genes and signaling molecules. However, the underlying molecular mechanisms are poorly understood. There is emerging evidence that microRNAs contribute to oogenesis. Here, we aimed to investigate the role of miR-17-92 cluster in regulating oogenesis. Methods: The miR-17-92 cluster was genetically ablated in germ cells of female mice by applying the Cre-loxp system for conditional gene knockout. Mating experiment, superovulation and histological analysis were used to assess the fertility of the model female mice. TUNEL assay was used to identify apoptotic cells in ovaries. The expression level of apoptosis- and follicular atresia- related genes was evaluated by qRT-PCR. Western blotting was performed to detect protein expression. Bioinformatics software and dual luciferase reporter assay were applied to predict and verify the target of miR-17-92 cluster. Results: Deletion of miR-17-92 cluster in germ cells of female mice caused increased oocyte degradation and follicular atresia, perturbed oogenesis, and ultimately led to subfertility. Genes involved in follicular atresia and the mitochondrial apoptotic pathway were obviously up-regulated. Furthermore, we verified that miR-19a regulated oogenesis at the post-transcriptional level by targeting Bmf in the ovaries of miR-17-92 cluster conditional knockout female mice. Conclusion: The miR-17-92 cluster is an important regulator of oogenesis. These findings will assist in better understanding the etiology of disorders in oogenesis and in developing new therapeutic targets for female infertility.

Download Full-text

L-securinine inhibits cell growth and metastasis of human androgen-independent prostate cancer DU145 cells via regulating mitochondrial and AGTR1/MEK/ERK/STAT3/PAX2 apoptotic pathways

Bioscience Reports ◽

10.1042/bsr20190469 ◽

2019 ◽

Vol 39 (5) ◽

Cited By ~ 5

Author(s):

Dongxu Zhang ◽

Houxian Liu ◽

Binbin Yang ◽

Jiasheng Hu ◽

Yue Cheng

Keyword(s):

Prostate Cancer ◽

Protein Expression ◽

Annexin V ◽

Inhibitory Effect ◽

Apoptotic Pathway ◽

Growth Inhibitory ◽

Du145 Cells ◽

Apoptotic Pathways ◽

Induced Apoptosis ◽

Androgen Independent

Abstract The present study aims to evaluate the anticancer effect of L-securinine on androgen-independent prostate cancer (AIPC) DU145 cells. L-securinine (2.5, 5, and 10 μM) treatment for 24, 48 and 72 h displayed strong growth inhibitory effect on DU145 cells in a concentration and time-dependent fashion but has less toxicity toward normal androgen-dependent LNCaP cells. Hoechst 332582 staining of DU145 cells and Annexin V-FITC/ PI dual-labeling followed by flow cytometry assay identified that this growth inhibition by L-securinine would be due to the induction of apoptosis. Moreover Transwell assay revealed that L-securinine significantly inhibited the cell migration/invasion ability of DU145 cells. Furthermore, results of western blotting showed that the involvement of mitochondrial apoptotic pathway in the L-securinine-induced apoptosis of DU145 cell, as evidenced by an increase in the protein expression of Bax, cleaved caspase-9, cleaved caspase-3, cytosolic cytochrome c, and cleaved PARP, together with a unchanged cleaved caspase-8 and decreased Bcl-2 protein expression. Also, L-securinine-induced antimetastatic activity in DU145 cells was associated with decreased protein expression of MMP-2 and MMP-9 and concurrent reduction of VEGF. In addition, further studies revealed that L-securinine may inhibit the protein expression of AGTR1, p-MEK1/2, p-ERK1/2, p-STAT3, PAX2, and p-PAX2, while the expression of ERK1/2, MEK1/2, and STAT3 protein retains intact. These findings suggest that L-securinine may be a promising chemopreventive agent against AIPC.

Download Full-text

Apaf-1 and caspase-9 are required for cytokine withdrawal-induced apoptosis of mast cells but dispensable for their functional and clonogenic death

Blood ◽

10.1182/blood-2005-05-2160 ◽

2006 ◽

Vol 107 (5) ◽

pp. 1872-1877 ◽

Cited By ~ 21

Author(s):

Vanessa S. Marsden ◽

Thomas Kaufmann ◽

Lorraine A. O'Reilly ◽

Jerry M. Adams ◽

Andreas Strasser

Keyword(s):

Mast Cells ◽

Fetal Liver ◽

Wild Type ◽

Caspase 9 ◽

Receptor Stimulation ◽

Apoptotic Pathway ◽

Apoptotic Pathways ◽

Induced Apoptosis ◽

Cytokine Deprivation

Cytokines promote survival of mast cells by inhibiting apoptotic pathways regulated by the Bcl-2 protein family. We previously showed that lymphocyte apoptosis can proceed via a Bcl-2-inhibitable pathway independent of the canonical initiator caspase, caspase-9, and its adaptor, Apaf-1. Here we report that mast cells lacking caspase-9 or Apaf-1 are refractory to apoptosis after cytotoxic insults but still lose effector function and ability to proliferate. In response to cytokine deprivation or DNA damage, fetal liver-derived mast cells lacking Apaf-1 or caspase-9 failed to undergo apoptosis. Nevertheless, the cytokine-starved cells were not functionally alive, because, unlike those overexpressing Bcl-2, they could not degranulate on Fcϵ receptor stimulation or resume proliferation on re-addition of cytokine. Furthermore, mast cells lacking Apaf-1 or caspase-9 had no survival advantage over wild-type counterparts in vivo. These results indicate that the Apaf-1/caspase-9-independent apoptotic pathway observed in lymphocytes is ineffective in cytokine-deprived mast cells. However, although Apaf-1 and caspase-9 are essential for mast cell apoptosis, neither is required for the functional or clonogenic death of the cells, which may be due to mitochondrial dysfunction.

Download Full-text