Characterization of the Osteogenesis Imperfecta mutation pattern and in vitro evaluation of the effects of resveratrol, ascorbic acid and curcumin antioxidants on mesenchymal stem cells

Name: Maira Trancozo
Type: PhD thesis
Publication date: 03/02/2022
Advisor:

Namesort descending Role
Flavia de Paula (M/D) Advisor *

Examining board:

Namesort descending Role
Agnes Lumi Nishimura External Examiner *
Carlos Magno da Costa Maranduba External Examiner *
Debora Dummer Meira Internal Examiner *
Flavia de Paula (M/D) Advisor *
Maria do Carmo Pimentel Batitucci Internal Examiner *

Summary: Osteogenesis Imperfecta (OI) is an inherited disorder of connective tissues that contain collagen in their formation. More than 20 OI-related genes have been reported in the last years. Most cases happen due mutations inherited with autossomic dominant inheritance in the COL1A1 or COL1A2 genes that codifing the collagen fibers type I proteins. Mutations with recessive inheritance were also related to the disease. There is no cure for OI, but some treatments can significantly improve the quality of life of affected individuals. Therefore, this work aimed to characterize the pattern of mutations in OI-related genes in patients from Espírito Santo state/Brazil as well as search for new OI treatments. The techniques of next generation sequencing (NGS), Sanger sequencing and/or Screening of mutations by single-stranded conformational polymorphism (SSCP) were used for the molecular analysis of the mutations. We focused on evaluating whether antioxidants are able to improve the cellular proliferation in vitro experiments in the search for new OI treatments. The samples of this research were obtained from patients treated at the Nossa Senhora da Glória Children`s Hospital in Vitória/ES, Brazil and at the Dório Silva Hospital/ES, Brazil. The COL1A1, COL1A2, P3H1, CRTAP, PPIB, SERPINH1, SERPINF1, FKBP10, SP7, WNT1 and IFITM5 genes were studied in 29 patients for the analyze the mutations. As expected, most individuals (23/29) carried mutations in COL1A1 or COL1A2 genes. Approximately 10% of the patients (3/29) had mutations in the FKBP10 gene. We found one mutation in the IFITM5 gene in one patient and another change in the P3H1 gene in other patient. The results suggest that the study of these five genes is able to identify 95% of OI mutations in Brazilian patients. Stem cell cultures were exposed to the antioxidants Resveratrol, Ascorbic Acid and Curcumin to search for compounds that improve cellular proliferation. Samples treated with Curcumin had their cell viability compromised by almost 80. This result suggests that Curcumin causes cell death in culture. By the other hand, increased cell proliferation was observed in cells treated with resveratrol and ascorbic acid. This result suggests that the compounds Resveratrol, Ascorbic acid are potential targets in the treatment of diseases and could be analyzed how much they can improve the bone mineralization. The results of this research can help in the planning of more efficient molecular diagnostic strategies, as well as in the identification of new compounds for the treatment of OI, contributing, in the future, for genetic counseling and improving the life quality of patients.

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