Endotoxin-induced silencing of mesoderm induction and functional differentiation: role of HMGB1 in pluripotency and infection

Sivasubramaniyan, Kavitha and Atluri, Rajesh Reddy and Sarda, Kanchan and Arvind, Milan (2008) Endotoxin-induced silencing of mesoderm induction and functional differentiation: role of HMGB1 in pluripotency and infection. Regenerative Medicine, 3 (1). pp. 23-31. ISSN 1746-0751

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Objectives: Mechanisms underpinning Gram-negative bacterial vaginosis-induced birth anomalies are obscure. Ethical issues limit such studies on peri-implantation-stage human embryos. Here we have used embryoid bodies (EBs) as an in vitro model to examine the effect of Gram-negative bacterial endotoxins/lipopolysaccharides (LPS) on the faithful induction of germ lineages during embryogenesis. The role of LPS-inducible cytokine and pluripotency-related DNA-binding protein HMGB1 was also studied in these EBs. Methods: EBs derived from the human embryonic stem cell line HUES9 were exposed to 12.5 pg/ml of LPS for 48 h. The expression profile of the ectoderm, endoderm, mesoderm and trophectoderm lineage markers, such as βIII-tubulin, GATA4, BMP2, Brachury and β-hCG, were studied, by RT-PCR and immunofluorescence. Inhibition of mesoderm induction was confirmed by RT-PCR analysis for hANP, cTnT, ABCG2, GATA2, BMP4 and HAND1. Osteoblast differentiation was induced in the EBs, and confirmed by von Kosa and Alizarin red staining. A comet assay was also carried out to assess the degree of apoptosis in these EBs. Results and conclusions: We found that the LPS-treated EBs were selectively silenced for mesoderm markers and failed to differentiate into functional osteoblasts. HMGB1 expression was absent in the normal EBs and was found to be localized in the cytoplasm of the LPS-treated EBs. Overall, our data indicate that endotoxin-induced HMGB1 expression in the peri-implantation-stage embryos can bring about severe birth defects of, for example, the bone and heart. This study also indicates that HMGB1 could be involved in maintenance of pluripotency in the human embryonic stem cells by impeding their differentiation.

Item Type: Article
Additional Information: Copyright of this article belongs to Future Medicine.
Subjects: Regenerative Medicine > MIRM Bangalore
Depositing User: MCON Library
Date Deposited: 20 Mar 2012 06:49
Last Modified: 20 Mar 2012 06:49
URI: http://eprints.manipal.edu/id/eprint/3606

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