Differentiation Of Adipose-derived Stem Cells Into Parathyroid-like Cells

Background: Hypoparathyroidism after thyroidectomy is a common and catastrophic iatrogenic injury with a high incidence.At present,the effect of existing drug replacement therapy is not satisfactory.We aimed to explore the method of establishing adipose-derived stem cells(ADSCs)to differentiate into parathyroid gland-like cells,so as to find sufficient seed cells for parathyroid autotransplantation,and open up a new way for clinical cure of hypoparathyroidism.Material/Methods: ADSCs were isolated,cultured and purified from inguinal adipose tissue of SD rats by enzyme digestion and adherent separation to induce adipogenesis and osteogenesis.The fifth generation of ADSCs was divided into control group and stimulated by Shh(100ng/ml)and three different concentrations of ActivinA(25ng / ml,50 ng / ml,75 ng / ml).On the 7th,14 th and 21 st day after induction,the content of parathyroid hormone(PTH)was detected by enzyme-linked immunosorbent assay(ELISA),the protein expression of CaSR,PTH and Gcm2 in cells was detected by western blotting(Western blot),and the mRNA expression of CaSR,PTH and Gcm2 in differentiated cells was detected by PCR.Results: Adipogenic induction and osteogenic induction and differentiation were successful,which was proved to be ADSCs.Under the stimulation of Shh and Activin A,PTH was detected in the culture medium in a concentration-and time-dependent manner.The best effect was shown in 100ng/ml Shh and 75ng/ml ActivinA group on 21 days in the experimental group.There was significant difference compared with the control group.The protein expression and mRNA expression of CaSR,PTH and Gcm2 showed the same trend as those of ELISA.Conclusion: ADSCs can be successfully isolated and purified from adipose tissue and further differentiated into parathyroid cells under the combined stimulation of Shh and Activin A.The best induction concentration was 100ng/ml Shh and 75ng/ml Activin A.The best action time is 21 days,which provides sufficient seed cells for parathyroid autotransplantation in the treatment of hypoparathyroidism,and lays a theoretical and practical foundation for the clinical application of this technique.