Effect Of Neuropeptide Y On Adipocyte Proliferation And Differentiation

Objective:1. To observe3T3-L1preadipocytes and human adipose-derived stem cells (hADSCs) morphological changes in the process of cultivation and differentiation, to product the growth curves.2. To investigate the influences of neuropeptide Y (NPY) with different concentrations on the proliferation and differentiation of3T3-L1preadipocytes and hADSCs, and to study the relationship between NPY and white adipose tissue formation preliminarily.Methods:1. Resurrecting, passage, cultivation, differentiation and producting the growth curve of3T3-L1preadipocytes and hADSCs; Cells were induced to differentiate by the classical hormonal cocktail and lipid droplets were observed by oil red O staining.2.3T3-L1preadipocytes and hADSCs were cultured in the existence of NPY with different concentrations(10-6M,10-7M,10-8M,10-9M,10-10M,10-11M,10-12M,10-13M,10-14M,10-15M) for24hours, MTT assay was used to determine the effects of NPY on adipocyte proliferation.3.3T3-L1preadipocytes and hADSCs were induced to differentiate by NPY with different concentrations(10-7M,10-9M,10-11M), oil red O staining was applied to observe lipid droplets during adipocyte differentiation.4. PPARy and C/EBPa protein expression in the process of adipocyte differentiation induced by NPY were detected by Western blot.5. Protein expression of Cidec and RIP140which were white adipose tissue marker were detected by Western blot when3T3-L1preadipocytes and hADSCs NPY-treated were induced to differentiate.Results:1.3T3-L1preadipocytes and hADSCs were fusiform cells and were induced into mature adipocytes by the classical hormonal cocktail successfully.Oil red O staining could observe the lipid droplets in orange.2. After3T3-L1preadipocytes and hADSCs were cultured in the presence of NPY with different concentrations(10-6M,10-7M,10-8M,10-9M,10-10M,10-11M,10-12M,10-13M,10-14M,10-15M) for24hours, compared with serum-free medium group(negative control group), we found10-6M～10-15M NPY could promote3T3-L1preadipocyte proliferation and10-11M～10-14M NPY could promote hADSCs proliferation, while10-7M、10-9M、10-10M NPY could inhibit hADSCs proliferation.3. In the process of3T3-L1preadipocytes and hADSCs differentiation induced by NPY, oil red O staining revealed that10-7M,10-9M,10-11M NPY could induce lipid droplet formation, lipid synthesis and adipocyte differentiation.4. Western-blot results showed that10-7M,10-9M,10-11M NPY could improve the expression of transcription factor PPARy and C/EBPa protein during adipose differentiation.5. Western-blot results suggested that10-7M,10-9M,10-11M NPY could promote the expression of Cidec(mouse as FSP-27) and RIP140protein which were white adipose tissue marker after adipocyte differentiation.Conclusions:1.10-6M～10-15M NPY promote3T3-L1preadipocyte proliferation;10-11M～10-14M NPY promote hADSCs proliferation while10-7M、10-9M、10-10M NPY inhibit hADSCs proliferation.2.10-7M,10-9M and10-11M NPY promote3T3-L1preadipocyte and hADSCs differentiation into mature adipocyte and may be associated with white adipose tissue formation.