Single Strand Conformation Polymorphism and Sequencing of HS6ST2 Gene in Patients of Idiopathic Premature Ovarian Failure GC01-GC08
Dr. Mona Sharma,
Room No. 2087, Teaching Block, 2nd Floor, Department of Reproductive Biology,
AIIMS, New Delhi-110029, India.
Introduction: Premature Ovarian Failure (POF) is a condition characterised by hypergonadotropic hypoestrogenic amenorrhea in women under the age of 40 years. Most cases of POF remain idiopathic despite of various aetiologies described. X-chromosome aberrations remain the major underlying cause of idiopathic POF. Heparan Sulfate 6-O Sulfotransferase 2 (HS6ST2) gene is one of the gene of X chromosome implicated in POF.
Aim: To detect HS6ST2 gene mutations in patients of idiopathic POF by Single Strand Conformation Polymorphism (SSCP) and to confirm the results by Sanger sequencing.
Materials and Methods: A cross-sectional study was planned from february 2017 to february 2018 by including total of 25 women with secondary amenorrhea and age less than 40 years as cases. Out of those 25 cases, 10 idiopathic cases of POF were selected by excluding the other causes of POF. The study also included 10 healthy fertile women with normal hormonal profile as the control group. Study was done in two parts. In the first part, idiopathic cases of POF were identified based on clinical and laboratory parameters. In second part, SSCP was standardised and was applied to identify mutations in HS6ST2 gene and results were confirmed by Sanger sequencing. Results were compared statistically by using two sample t-test and two sample Wilcoxon rank sum (Mann-Whitney) test. The values obtained were expressed as mean±SD along with the p-value.
Results: Idiopathic cases of POF were selected based on clinical and laboratory parameters. There were no mutations found in the selected HS6ST2 exons and results were confirmed by Sanger sequencing.
Conclusion: As the present work was planned in smaller number of cases, involvement of HS6ST2 gene cannot be completely ruled out. The study shall be planned in larger cohorts to confirm validity of results. In case if genomic variants are found in more number of studies planned, it will help in developing functional studies in both human cell lines and in vivo mouse models to validate the association of genomic variants in POF. SSCP is a rapid, easy technique and it is as sensitive as Sanger sequencing for identifying gene mutations.