Effects of biomimetic network membrane prepared by chitosan/gelatin/pectin on proliferation and mineralization of mesenehymal stem cells / 吉林大学学报(医学版)

ABSTRACT

Objective:To explore the effects of the biomimetic network membrane prepared by chitosan/gelatin/pectin on the proliferation and mineralization of mesenchymal stem cells (MSCs) , and to evaluate its feasibility of constructing tissue engineering bone.Methods:Chitosan, gelatin and pectin were made into a new biomimetic network membrane in a certain ratio by biomimetics.The experiment was divided into control group (MSCs+conventional medium) , material group (MSCs+network membrane+conventional medium) and material+OS group (MSCs+network membrane+OS medium) .The cell morphology was observed by inverted phase contrast microscope;the growth and secretion of extracellular matrix of the MSCs were observed under scanning electron microscope (SEM) .The proliferation of cells was determined by MTT assay (The MSCs were divided into negative control group and material group, and they were cultivated with blank medium and medium including materials) .The expression of calcium in MSCs was detected by Alizarin Red staining.Real-time polymerase chain reaction (RT-PCR) was used to determine the expression levels of osteocalcin (OC) mRNA and osteopontin (OPN) mRNA in the MSCs.Results:The network membrane was semitransparent thin film.The MSCs were short shuttle and clustered under inverted phase contrast microscope.After cultured for 7d, the MSCs were shuttle;after cultured for 14d, the number of MSCs was increased, with pseudo feet on the membrane;after cultured for21d, the MSCs clustered with a lot of neo-formed extracellular matrix.The MTT results showed that there was no significant difference in the proliferation level of MSCs between material group and negative control group (P＞0.05) .The Alizarin Red staining results showed that the MSCs in the network membrane were dyed orange red.The RT-PCR results showed that the expression levels of OC mRNA in the MSCs in material group and material+OS group were lower on the 7th and 14th days, but on the 21th day, the expression levels were significantly increased and reached the peak;the expression level of OC mRNA in the MSCs in material group was significantly increased on the 7th day, and the expression level reached the peak on the 14th day, then fell slightly on the 21th day;compared with control group, the expression levels of OC mRNA and OPN mRNA in the cells in material group and material+OS group at different time points were significantly increased (P＜0.01) , but there were no significant differences between material group and material+OS group (P＞0.05) .Conclusion:Chitosan/gelatin/pectin biomimetic network membrane has good biocompatibility, and MSCs can grow and proliferate well on the membrane.The membrane can induce the MSCs to express mineralization-related genes and proteins without inducers.