Stellate Cells from Rat Pancreas Are Stem Cells and Can Contribute to Liver Regeneration
Figure 7
Contribution of transplanted eGFP+ PSC to liver regeneration in vivo.
(A) eGFP fluorescence (green) in PSC cultured for 7 days. (B) These culture-activated eGFP+ PSC were transplanted into wild type rats that underwent PHX in the presence of 2AAF. The eGFP+ PSC reached the host liver as determined by immunofluorescence staining of eGFP (green) 14 days after transplantation. (C, D) The eGFP+ PSC differentiated into hepatocytes in the host liver as determined by co-localization of eGFP fluorescence (green) and nuclear HNF4α immunofluorescence (red). (E) Their differentiation into hepatocytes was also confirmed by combined immunofluorescence staining of eGFP (green) and cytokeratin 18 (CK18; red). The fate of transplanted eGFP+ PSC was further analyzed by immunohistochemistry through Fast Red staining of eGFP-expressing cells (red). (F, G) The liver from wild type Wistar rats displayed no Fast Red staining after incubation with antibodies against eGFP, (H) but when eGFP+ PSC were transplanted, large areas of the host liver were colored red. This method labeled (I) hepatocytes (yellow arrows) and (J, K) bile duct cells (white arrows). (J) Other cell types in close proximity to bile ducts (black arrow) and liver sinusoids (yellow arrow) showed also eGFP expression as indicated by Fast Red staining. (L) The presence of eGFP-expressing cells in bile ducts of the host liver was further confirmed by combined immunofluorescence of eGFP (green) and CK19 (red). (M) Immunofluorescence staining of HNF4α (red) and FISH of chromosome Y (green) was used to identify hepatocytes in female liver tissue that derived from transplanted male PSC 14 days after PHX in the presence of 2AAF (white arrows). Also non-parenchymal cells with chromosome Y were detected (yellow arrows). (N) Combined FISH of chromosome Y (green) and immunofluorescence staining of panCK (red) revealed that the transplanted PSC had differentiated into duct-forming cholangiocytes. (O, P) Clonally expanded PSC of males were also transplanted into wild type female rats that underwent PHX in the presence of 2AAF. FISH of chromosome Y (green) and immunofluorescence staining of (O) HNF4α (red) or (P) panCK (red) indicated that single cell clones of PSC differentiated into hepatocytes and cholangiocytes (white arrows) as investigated after 14 days of regeneration. (P) Also PSC-derived cells that did not express cytokeratins were found close to bile ducts (yellow arrows). The cell nuclei were marked by DAPI (blue).