Avinash Kharat1, Avinash Sanap1, Supriya Kheur1, Madhura Shekatkar1, Ramesh Bhonde2. 1. Regenerative Medicine Laboratory, Dr. D. Y. Patil Dental College and Hospital, Dr. D. Y. Patil Vidyapeeth, Pimpri, Pune, India. 2. Regenerative Medicine Laboratory, Dr. D. Y. Patil Dental College and Hospital, Dr. D. Y. Patil Vidyapeeth, Pimpri, Pune, India. rrbhonde@gmail.com.
Abstract
BACKGROUND: The human gingiva-derived mesenchymal stem cells (hGMSCs) possess a great potential to develop the cell-based therapy for diabetes due to its unscarred healing capacity and reparative potential. In this current study, we isolated, cultured and characterised the GMSCs and explored their potential to differentiate into Insulin Producing Cell Clusters (IPCCs). METHODS: The cells derived from gingival tissues exhibited fibroblast-like morphology. The flow cytometric analysis revealed positive expression of CD73(97.43%), CD90(95.05%), and CD105(93.17%) and negative expression of CD34(0.05%), CD45(0.09%), and HLA-DR (0.025) surface markers. We then converted this adherent fibroblast-like GMSCs into floating IPCCs using a sequential three-step protocol containing a different combination of differentiating agents. Initially, the presence of insulin in IPCCs was confirmed by dithizone staining. Glucose-stimulated insulin secretion (GSIS) assay confirmed that IPCCs secrete insulin in response to glucose. RESULTS: Generated IPCCs express pancreatic markers such as insulin, pdx1, glucagon, GLUT4 and GLUT2 as evidenced by RT-PCR analysis. Our results unequivocally showed that IPCCs can be generated from gingiva which is a potential source of postnatal MSCs. Our results offer the IPCCs generated from hGMSCs a platform for screening anti-diabetic drugs and a new autologous source of tissue for islet transplantation for the treatment of diabetes. CONCLUSIONS: Our results unequivocally demonstrate for the first time that hGMSCs can be used as an attractive non-invasive tissue source for generating IPCCs, which can be employed in diabetes research for screening antidiabetic agents and also for transplantation in type 1 diabetic patients as autologous source without the need of immunosuppression.
BACKGROUND: The human gingiva-derived mesenchymal stem cells (hGMSCs) possess a great potential to develop the cell-based therapy for diabetes due to its unscarred healing capacity and reparative potential. In this current study, we isolated, cultured and characterised the GMSCs and explored their potential to differentiate into Insulin Producing Cell Clusters (IPCCs). METHODS: The cells derived from gingival tissues exhibited fibroblast-like morphology. The flow cytometric analysis revealed positive expression of CD73(97.43%), CD90(95.05%), and CD105(93.17%) and negative expression of CD34(0.05%), CD45(0.09%), and HLA-DR (0.025) surface markers. We then converted this adherent fibroblast-like GMSCs into floating IPCCs using a sequential three-step protocol containing a different combination of differentiating agents. Initially, the presence of insulin in IPCCs was confirmed by dithizone staining. Glucose-stimulated insulin secretion (GSIS) assay confirmed that IPCCs secrete insulin in response to glucose. RESULTS: Generated IPCCs express pancreatic markers such as insulin, pdx1, glucagon, GLUT4 and GLUT2 as evidenced by RT-PCR analysis. Our results unequivocally showed that IPCCs can be generated from gingiva which is a potential source of postnatal MSCs. Our results offer the IPCCs generated from hGMSCs a platform for screening anti-diabetic drugs and a new autologous source of tissue for islet transplantation for the treatment of diabetes. CONCLUSIONS: Our results unequivocally demonstrate for the first time that hGMSCs can be used as an attractive non-invasive tissue source for generating IPCCs, which can be employed in diabetes research for screening antidiabetic agents and also for transplantation in type 1 diabetic patients as autologous source without the need of immunosuppression.
Authors: V Govindasamy; V S Ronald; A N Abdullah; K R Ganesan Nathan; Z A C Ab Aziz; M Abdullah; S Musa; N H Abu Kasim; R R Bhonde Journal: J Dent Res Date: 2011-02-18 Impact factor: 6.116
Authors: Vikrant R Patil; Avinash H Kharat; Deepak G Kulkarni; Supriya M Kheur; Ramesh R Bhonde Journal: Cell Biol Int Date: 2018-12 Impact factor: 3.612