Reduction of Shp-2 expression by siRNA reduces murine embryonic stem cell-derived in vitro hematopoietic differentiation

¹ Herman B. Wells Center for Pediatric Research, Indiana University School of Medicine, Indianapolis, Indiana

^* To whom correspondence should be addressed. E-mail: myoder{at}iupui.edu.

	Abstract

Shp-2 is a member of a small family of cytoplasmic Src homology 2 (SH2) domaincontaining protein tyrosine phosphatases. Although Shp-2 has been shown to be necessary for hematopoiesis using a mouse model expressing a mutant residual protein (Shp-2^/), we used siRNA to reduce Shp-2 expression and examined the consequences on embryonic stem (ES) cell-derived hemangioblast, primitive, and definitive hematopoietic development. We found that Shp-2 siRNA at a concentration of 50 nM effectively diminished Shp-2 expression in differentiating embryoid bodies (EBs). Hemangioblast, primitive, and definitive hematopoietic progenitor formation was decreased significantly following transfection with Shp-2 siRNA, but not with scrambled siRNA. As Shp-2 is involved in signals emanating from the bFGF receptor, we asked whether Shp-2 functions in bFGF-mediated hemangioblast development. Reduction of Shp-2 expression using siRNA, but not scrambled siRNA, blocked the bFGF-induced increase in hemangioblast development. Using siRNA as an independent method of reducing Shp-2 function, in contrast to the mutant mouse model (Shp-2^/) previously utilized, we demonstrate that Shp-2 is required in hemangioblast, primitive, and definitive progenitor hematopoietic development and that Shp-2 is integrally necessary for bFGF-mediated hemangioblast production.

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