Preparation Of Lignin-carbohydrate Complexes Based Scaffold And Application In Culturing Human Hepatocytes

Liver is one of the important organs in humam body. However, there are a lot ofpeople died of liver cancer and acute liver failure every year. Therefore, livertransplantation is the best healing method. Looking for a suitable scaffold is a basicresearch in the culture of liver organs from human liver cells. As lignin-carbohydratecomplexes (LCCs) have good amphilic, biocompatibal, non-toxic and biodegradblity,they are proming biomedical polymer material. In this article, the application of LCCsin tissue engineering is investigated.Lignin-carbohydrate complexes were extracted from Gymnosperm wood (gingkobiloba L.), hard wood (aspen tomentosa Carr.) and non-wood material（wheat triticumaestivum Linn.）. Then, water-soluble fraction and water-insoluble fraction were isolatedand applied in the culture of human hepatocytes in this article.The water-soluble fraction of the LCCs was characterized by Fourier transforminfrared spectrometer（FT-IR） and High Performance Liquid Chromatograph(HPLC).Then soluble LCCs were used in human hepatocytes culture L-02with contious monitorof cell growth. The proliferation and morphology of hepatocytes were also observed bycell counting and inverted biological microscope，and hepatocytes’ metabolic activity,was detected. The results show in gingko water-soluble LCC, the components ofgalactose and mannose are2.05%and28.31%respectively; in aspen water-soluble LCC,the components of galactose and mannose are5.59%and24.5%respectively; in wheatwater-soluble LCC, the component of mannose is10.62%, the component of galactoseis little. Cell counting results show that in fourth day, hepatocytes proliferate fastest andhepatocytes number is the most, hepatocytes proliferation rate is as follow aspenwater-soluble LCC> gingko water-soluble LCC> control group> wheat water-solubleLCC.Spherical biological carriers were prepared with gingko and aspen water-insolubleLCC by the method of freeze-drying. Fourier transform infrared spectrometer（FT-IR）,High Performance Liquid Chromatograph(HPLC) and scanning electron microscope(SEM) were applied to elucidate the composition, chemical structure and morphologyof spherical biological carriers. Then, carriers were applied in human liver cell cultureL-02with contious monitor of cell growth. The proliferation and morphology ofhepatocytes were observed by cell counting, inverted biological microscope and SEM.meanwhile, hepatocytes metabolic activity was analysized. The results show that Theresults show in gingko spherical biological carriers, the components of galactose andmannose are3.30%and21.94%respectively; in aspen spherical biological carriers, thecomponents of galactose and mannose are3.82%and25.25%respectively, gingko andaspen water-insoluble LCC spherical biological carriers’ specific suiface area are17m2/g and9m2/g, both of spherical biological carriers’ dry-diameter is1.8mm~2.0mm.In fifth day, hepatocytes proliferate fastest and hepatocytes number is the most. From inverted biological microscope and SEM, we can find that hepatocytes grew on aspenwateer-insoluble LCC spherical biological carriers group is more than hepatocytes grewon gingko wateer-insoluble LCC spheriacal biological carriers. Hepatocytesproliferation rate and ALB, BUN and glucose consumption are all as follow aspenwater-soluble LCC> gingko water-soluble LCC> control group. According to invertedbiological microscope and SEM, we can find that a lot of cells were successfullyadhered on spherical biological carriers. LCCs are proming biomedical polymermaterial in the tissue engineering.