In vitro and in vivo functional and structural evaluation of cultivated fetal and adult hepatocytes
Adnan NARCI, Mevlit KORKMAZ, Tahsin YAKUT, Murat YAĞMURCA, Burcu BİLTEKİN, Barbaros YİĞİT, Evrim ÖZKARACA
Afyon Kocatepe Üniversitesi Tıp Fakültesi Çocuk Cerrahisi, Histoloji ve Embriyoloji Anabilim Dalları, Afyon, İstanbul Üniversitesi Tıp Fakültesi Histoloji ve Embriyoloji Anabilim Dalı, İstanbul, Uludağ Üniversitesi Tıp Fakültesi Tıbbi Genetik Anabilim Dalı, Bursa
Keywords: Hepatocyte, cell Culture
Abstract
Aim: In this study, we aimed to examine the in vitro physiological and morphological characteristics of fetal and adult hepatocytes and in situ tissue masses after implantation into the omentum.
Materials and Methods: Liver tissue samples were obtained from fetuses of twenty-day-gestation age and adult New Zealand rabbits. Hepatocytes were separated by collagenase digestion. Cell suspension was seeded onto collagen coated dishes and cultivation was maintained using hepatocyte medium (H 1777, Sigma) under standard culture condition. Growing cells were examined morphologically using light and electron microscopic images. After reaching confluence, flasks were treated with trypsin-EDTA and cell suspension was seeded onto the collagen coated unwoven poligliko-lactic acid (PLGA) polymers and cultivated for 10 days. Fetus and adult cell-polymer complexes were implanted (fixed with sutures) into the omentum of rabbits homologously and autologously respectively. After three weeks of implantation, tissue masses were resected and embedded into paraffin. Paraffin sections were stained with hemotoxilen-eosine, PAS, best carmine and examined histologically.
Results: Both fetal and adult hepatocytes were successfully isolated and cultivated. Morphologic analyzes have shown that all cell populations expressed typical spheroid shape with centrally located nucleus. Fetal cells reached confluence in two weeks whereas it took three weeks for the adult liver cells. Attached and proliferated hepatocytes on the polymer fibers were detected after 24 hours of seeding onto the scaffolds. Histological examination of artificial tissues harvested from omentum has shown well organized and vascularized liver tissues.
Conclusion: Although fetal liver cells seem proliferating rapidly than adults, there is no difference in histological examination of in situ tissue masses. Collagen coated PLGA polymers may be the preferred vehicle for cell transfer. We think that omentum is the most suitable tissue for adaptation and vascularization of hepatic cells. This model can be used to obtain artificial hepatic tissue and also for orthotopic transplantation.