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BACKGROUND: Excessive hepatic glycogen accumulation benefits tumorigenesis and cancer cell survival. We previously reported that BMP4 has the strongest ability to promote glycogenesis among the 14 BMPs in hepatocytes and augmented hepatocellular carcinoma (HCC) cell survival under hypoxia and hypoglycemia conditions by promoting the glycolysis pathway. However, the mechanism underlying BMP4's effect on glycogenesis in HCC remains elusive. METHODS: The expression of BMP4 and SLC2A1 were acquired by analyzing the TCGA-LIHC dataset, as well as by immunohistochemical analysis of the 40 pairs of human HCC samples and para-tumor tissues. Gene expressions were detected by qPCR, immunoflurorescence staining, and Western blotting. Overexpression and silencing of BMP4 were accomplished through adenoviruses Ad-B4 and Ad-siB4 infection. Hepatic glycogen was detected by PAS staining. SLC2A1 (GLUT1) function was blocked by the inhibitor BAY-876. ChIP assay was used to determine the binding of SMADs to the promoter region of SLC2A1 in HCC cells. Lastly, the in vivo effect of BMP4-regulated SLC2A1 on HCC tumor growth was assessed in a xenograft model of HCC. RESULTS: The elevated expression of BMP4 in HCC tumor tissues was highly correlated with hepatic glycogen accumulation in clinical samples. SLC2A1 was highly expressed in HCC tumor tissue and correlated with clinical stage and prognosis. Exogenous BMP4 augmented glycogen accumulation and upregulated the expression of glycogen synthesis-related genes in Huh7 and HepG2 cells, both of which were effectively blunted by SLC2A1inhibitor BAY-876. In mechanism, BMP4 activated SMAD5 to regulate the promoter of SLC2A1to enhance its expression. The in vivo xenograft experiments revealed that BMP4 promoted glycogen accumulation and tumor growth, which were effectively diminished by BAY-876. CONCLUSION: These results demonstrate that BMP4 upregulates glycogen synthesis through the SMAD/SLC2A1 (GLUT1) signaling axis in HCC cells, which may be exploited as novel therapeutic targets for HCC treatment.
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[This corrects the article on p. 793 in vol. 11, PMID: 33791154.].
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Effective and safe liver-directed gene therapy has great promise in treating a broad range of liver diseases. While adenoviral (Ad) vectors have been widely used for efficacious in vivo gene delivery, their translational utilities are severely limited due to the short duration of transgene expression and solicitation of host immune response. Used as a promising polymeric vehicle for drug release and nucleic acid delivery, carboxymethyl chitosan (CMC) is biocompatible, biodegradable, anti-microbial, inexpensive, and easy accessible. Here, by exploiting its biocompatibility, controlled release capability and anti-inflammatory activity, we investigated whether CMC can overcome the shortcomings of Ad-mediated gene delivery, hence improving the prospect of Ad applications in gene therapy. We demonstrated that in the presence of optimal concentrations of CMC, Ad-mediated transgene expression lasted up to 50 days after subcutaneous injection, and at least 7 days after intrahepatic injection. Histologic evaluation and immunohistochemical analysis revealed that CMC effectively alleviated Ad-induced host immune response. In our proof-of-principle experiment using the CCl4-induced experimental mouse model of chronic liver damage, we demonstrated that repeated intrahepatic administrations of Ad-IL10 mixed with CMC effectively mitigated the development of hepatic fibrosis. Collectively, these results indicate that CMC can improve the prospect of Ad-mediated gene therapy by diminishing the host immune response while allowing readministration and sustained transgene expression.
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Skin injury is repaired through a multi-phase wound healing process of tissue granulation and re-epithelialization. Any failure in the healing process may lead to chronic non-healing wounds or abnormal scar formation. Although significant progress has been made in developing novel scaffolds and/or cell-based therapeutic strategies to promote wound healing, effective management of large chronic skin wounds remains a clinical challenge. Keratinocytes are critical to re-epithelialization and wound healing. Here, we investigated whether exogenous keratinocytes, in combination with a citrate-based scaffold, enhanced skin wound healing. We first established reversibly immortalized mouse keratinocytes (iKera), and confirmed that the iKera cells expressed keratinocyte markers, and were responsive to UVB treatment, and were non-tumorigenic. In a proof-of-principle experiment, we demonstrated that iKera cells embedded in citrate-based scaffold PPCN provided more effective re-epithelialization and cutaneous wound healing than that of either PPCN or iKera cells alone, in a mouse skin wound model. Thus, these results demonstrate that iKera cells may serve as a valuable skin epithelial source when, combining with appropriate biocompatible scaffolds, to investigate cutaneous wound healing and skin regeneration.
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Hepatocellular carcinoma (HCC) is one of the leading causes of cancer death worldwide although its pathogenic mechanism remains to be fully understood. Unlike normal cells, most cancer cells rely on aerobic glycolysis and are more adaptable to the microenvironment of hypoxia and hypoglycemia. Bone Morphogenetic Protein 4 (BMP4) plays important roles in regulating proliferation, differentiation, invasion and migration of HCC cells. We have recently shown that BMP4 plays an important role in regulating glucose metabolism although the effect of BMP4 on glucose metabolic reprogramming of HCC is poorly understood. In this study, we found that BMP4 was highly expressed in HCC tumor tissues, as well as HCC cell lines that were tolerant to hypoxia and hypoglycemia. Mechanistically, we demonstrated that BMP4 protected HCC cells from hypoxia and hypoglycemia by promoting glycolysis since BMP4 up-regulated glucose uptake, the lactic acid production, the ATP level, and the activities of rate limiting enzymes of glycolysis (including HK2, PFK and PK). Furthermore, we demonstrated that BMP4 up-regulated HK2, PFKFB3 and PKM2 through the canonical Smad signal pathway as SMAD5 directly bound to the promoter of PKM. Collectively, our findings shown that BMP4 may play an important role in regulating glycolysis of HCC cells under hypoxia and hypoglycemia condition, indicating that novel therapeutics may be developed to target BMP4-regulated glucose metabolic reprogramming in HCC.