Zhihe Liu

Apolipoprotein L1, a Novel Bcl-2 Homology Domain 3-only Lipid-binding Protein, Induces Autophagic Cell Death

The Bcl-2 family proteins are important regulators of type I programmed cell death apoptosis; however, their role in autophagic cell death (AuCD) or type II programmed cell death is still largely unknown. Here we report the cloning and characterization of a novel Bcl-2 homology domain 3 (BH3)-only protein, apolipoprotein L1 (apoL1), that, when overexpressed and accumulated intracellularly, induces AuCD in cells as characterized by the increasing formation of autophagic vacuoles and activating the translocation of LC3-II from the cytosol to the autophagic vacuoles. Wortmannin and 3-methyladenine, inhibitors of class III phosphatidylinostol 3-kinase and, subsequently, autophagy, blocked apoL1-induced AuCD. In addition, apoL1 failed to induce AuCD in autophagy-deficient ATG5-/- and ATG7-/- mouse embryonic fibroblast cells, suggesting that apoL1-induced cell death is indeed autophagy-dependent. Furthermore, a BH3 domain deletion construct of apoL1 failed to induce AuCD, demonstrating that apoL1 is a bona fide BH3-only pro-death protein. Moreover, we showed that apoL1 is inducible by p53 in p53-induced cell death and is a lipid-binding protein with high affinity for phosphatidic acid (PA) and cardiolipin (CL). Previously, it has been shown that PA directly interacted with mammalian target of rapamycin and positively regulated the ability of mammalian target of rapamycin to activate downstream effectors. In addition, CL has been shown to activate mitochondria-mediated apoptosis. Sequestering of PA and CL with apoL1 may alter the homeostasis between survival and death leading to AuCD. To our knowledge, this is the first BH3-only protein with lipid binding activity that, when overproduced intracellularly, induces AuCD.

Epidermal stem cells cultured on collagen-modified chitin membrane induce in situ tissue regeneration of full-thickness skin defects in mice.

A Large scale of full-thickness skin defects is lack of auto-grafts and which requires the engineered skin substitutes for repair and regeneration. One major obstacle in skin tissue engineering is to expand epidermal stem cells (ESCs) and develop functional substitutes. The other one is the scaffold of the ESCs. Here, we applied type I collagen-modified chitin membrane to form collagen-chitin biomimetic membrane (C-CBM), which has been proved to have a great biocompatibility and degraded totally when it was subcutaneously transplanted into rat skin. ESCs were cultured, and the resulting biofilm was used to cover full-thickness skin defects in nude mice. The transplantation of ESCs- collagen- chitn biomimetic membrane (ESCs-C-CBM) has achieved in situ skin regeneration. In nude mice, compared to controls with collagen-chitin biomimetic membrane (C-CBM) only, the ESCs-C-CBM group had significantly more dermatoglyphs on the skin wound 10 w after surgery, and the new skin was relatively thick, red and elastic. In vivo experiments showed obvious hair follicle cell proliferation in the full-thickness skin defect. Stem cell markers examination showed active ESCs in repair and regeneration of skin. The results indicate that the collagen-modified chitin membrane carry with ESCs has successfully regenerated the whole skin with all the skin appendages and function.

Amentoflavone inhibits angiogenesis of endothelial cells and stimulates apoptosis in hypertrophic scar fibroblasts.

Amentoflavone (8-[5-(5,7-dihydroxy-4-oxo-chromen-2-yl)-2-hydroxy-phenyl]-5,7-dihydroxy-2-(4-hydroxyphenyl) chromen-4-one; AF) is a biflavonoid derived from the extracts of Selaginella tamariscina. It has been shown that AF has diverse biological effects such as antitumour, etc. It is well known that high cell proliferation, viability, angiogenesis and low apoptosis are key factors in hypertrophic scar formation. In this study, we report that AF inhibited viability and stimulated apoptosis in hypertrophic scar fibroblasts (HSFBs). Incubation of HSFBs with AF showed its inhibitory effect on cell viability and the exhibition of a series of cellular changes that were consistent with apoptosis. By Western-blot analysis, our data indicated significant increases in the amounts of cleaved caspases 3, 8, 9 and Bax, several apoptotic promoters and a significant decrease in translationally controlled tumour protein (TCTP), an apoptotic inhibitor, in HSFBs treated with AF. Furthermore, AF showed significant inhibitions on the viability, migration and tube formation of endothelial cells, which are associated with angiogenesis. In conclusion, this study suggests that AF stimulates apoptosis in HSFBs and inhibits angiogenesis of endothelial cells. Therefore, AF is a promising molecule that can be used in hypertrophic scar treatment.

Knockdown of FUSE binding protein 1 enhances the sensitivity of epithelial ovarian cancer cells to carboplatin

Epithelial ovarian cancer (EOC) affects almost 25,000 women annually and is the fifth most common malignancy in women in North America. A combination of surgery and cytotoxic chemotherapy may produce a favorable clinical response. The platinum-paclitaxel combination regimen is the chemotherapy gold-standard for advanced ovarian cancer, and carboplatin is one of the agents in this combination therapy. However, the majority of patients eventually experience a relapse due to the development of platinum resistance. FUSE binding protein 1 (FBP1) has been identified as an anti-apoptotic and pro-proliferative oncoprotein that is overexpressed in hepatocellular carcinoma. Its high expression is also associated with carboplatin resistance. In the present study, it was identified that the expression of FBP1 was significantly higher in EOC tissues than in normal epithelial ovarian or in epithelial ovarian adenoma tissue. FBP1 expression was significantly correlated with the grade of epithelial ovarian cancer. Carboplatin inhibited the expression of FBP1 in epithelial ovarian cancer cells and the knockdown of FBP1 enhanced the inhibition of cell viability and migration by carboplatin. In addition to FBP1, carboplatin also inhibited the expression of β-catenin and matrix metalloproteinase (MMP)-9. Furthermore, the expression of β-catenin and MMP-9 were lower in FBP1 knockdown cells compared with control EOC cells. FBP1 may thus serve a role in the regulation of the expression of β-catenin and MMP-9; the inhibition of β-catenin and MMP-9 by carboplatin may be mediated through the inhibition of FBP1. The inhibition of FBP1 expression by carboplatin may be a mechanism in the treatment of EOC by carboplatin.

FBP1 promotes ovarian cancer development through the acceleration of cell cycle transition and metastasis

Epithelial ovarian cancer (EOC) is the fifth most common malignancy in women, with a 5-year mortality of >70% in North America. As the symptoms are often not observed until the cancer has spread extensively, few women are diagnosed at an early stage of disease. Large-scale gene expression analyses have identified molecular subtypes within high-grade ovarian cancer with variable survival rates and drug resistance. The understanding of gene expression, the mechanisms underlying cancer processes and drug resistances have facilitated the development of targeted therapies. The far-upstream element (Fuse)-binding protein 1 (FBP1) is overexpressed in a number of malignancies such as hepatocellular carcinoma, and has been identified as an oncoprotein. In our early studies, FBP1 was demonstrated to physically interact with p53 and suppresses p53 transcription activity. In the present study, FBP1 expression increased as ovarian cancer developed. Among ovarian normal, adenoma and carcinoma tissues, the highest FBP1 expression was identified in carcinoma tissues. Furthermore FBP1 did not influence the apoptosis of ovarian carcinoma cells, yet enhanced cell cycle transition and metastasis. Therefore, it was hypothesized that FBP1 promotes ovarian cancer development through the acceleration of cell cycle transition and metastasis, and FBP1 is a novel potential biological marker for epithelial ovarian cancer diagnosis.

Protective effect of p53 on the viability of intervertebral disc nucleus pulposus cells under low glucose condition

P53 is a famous cancer suppressor and plays key roles in metabolism. Intervertebral disc (IVD) is the largest avascular cartilaginous structure in humans and its degeneration is a common cause of spine diseases initiated from damaged nucleus pulposus (NP) cells. The potential cause of disc degeneration has been attributed to aging, genetic factors, mechanical factors and nutrition. In this study, we found that p53 decreased and leaked to the cytoplasm in NP cells as the glucose level decreases, in contrast to cancer cells in which p53 increases and concentrates to the nuclei. Comparing with in p53 knockdown NP cells, relative high p53 expression in normal control NP cells inhibited autophagy and the pentose phosphate pathway. Furthermore, the expression of Sox 9 and type II collagen were higher in p53 normal control than p53 knockdown NP cells. Based on these results, we believe that relative high p53 facilitates NP cell viability and integrity.

Apoptosis and Autophagy: The Yin–Yang of Homeostasis in Cell Death in Cancer

Apoptosis and autophagy are physiologically necessary pathways that are vital for cell homeostasis. Apoptosis facilitates type I programmed cell death, while the autophagic survival mechanism counteracts apoptosis. Dysregulation in the homeostatic balance between these two essential cellular pathways has been linked to various diseases. We review relevant Janus molecules and their interactomes, as well as lysosomes which play multiple roles in apoptosis and autophagy, and discuss how targeted interventions can be used in cancer prevention and/or therapy.

Abstract B35: Amentoflavone inhibits angiogenesis and stimulates apoptosis in cells

Amentoflavone (8-[5-(5,7-dihydroxy-4-oxo-chromen-2-yl)-2-hydroxy-phenyl]-5,7-dihydroxy -2-(4-hydroxyphenyl) chromen-4-one, AF) is a biflavonoid derived from the extracts of Selaginella tamariscina. It has been shown that AF has diverse biological effects including anti-inflammatory, anti-viral, inhibition of fatty acid synthesis, and anti-tumor. It is well known that high cell proliferation and angiogenesis are two key factors in tumorigenesis. In this study, we report that AF inhibited proliferation, and stimulated apoptosis in fibroblasts. Incubation of fibroblast with AF showed inhibitory effect on cell viability and exhibited a series of cellular changes that were consistent with apoptosis. By western blot analysis, our date indicated significant increases in the amounts of cleaved-caspase 3, 8, 9 and Bax, several apoptotic promoters, and a significant decrease in TCTP, an apoptotic inhibitor, in fibroblast treated with AF. Furthermore, AF showed significant inhibitions on the proliferation, migration, and tube formation of endothelial cells, which are associated with angiogenesis. In conclusion, this study suggests that AF stimulates apoptosis in fibroblasts and inhibits angiogenesis of endothelial cells. Therefore, AF is a promising molecule which can be used in cancer treatment. Note: This abstract was not presented at the conference. Citation Format: Zhihe Liu, Jinli Zhang, Wenjuan Cao. Amentoflavone inhibits angiogenesis and stimulates apoptosis in cells. [abstract]. In: Proceedings of the Third AACR International Conference on Frontiers in Basic Cancer Research; Sep 18-22, 2013; National Harbor, MD. Philadelphia (PA): AACR; Cancer Res 2013;73(19 Suppl):Abstract nr B35.