Investigation of Genes and Gene Proteins in Cleft Affected Tissue. Summary of the Doctoral Thesis

Abstract

Craniofacial clefts are congenital defects of the facial region which occur when the growing facial folds do not fuse fully together. Clefts can be unilateral or bilateral and they can affect the oral cavity mucosal tissue in different locations, forming cleft lip and cleft palate. Facial cleft morphopathogenesis is associated with cleft candidate genes which encode proteins needed for the regulation of facial tissue growth and differentiation. The involvement of cleft candidate genes during facial cleft morphopathogenesis in human tissue is unclear because the interactions between said genes is complicated, these genes have been studied more in animals and the available information about their interaction in human tissue is very limited. The aim of this research was to determine the distribution and the relative number of cleft candidate gene coded protein containing cells in different cleft affected tissue types. In our research cleft tissue samples were divided into three groups depending on the cleft tissue type – unilateral cleft lip, bilateral cleft lip, and cleft palate tissue. Unilateral cleft lip tissue was gathered from 36 patients during cleft lip plastic surgery, bilateral cleft lip tissue was obtained from 13 patients during cleft lip plastic surgery, but cleft palate tissue was gathered from 26 patients during soft palate plastic surgery. Cleft tissue samples were obtained from patients aged before or during the age of primary dentition. Healthy oral cavity tissue needed for the control group was obtained during upper lip frenulum plastic surgery from seven patients (from five to fourteen years old) and from five newborn patients whose tissue were gathered from the historical collection of the Institute of Anatomy and Anthropology of Rīga Stradiņš University. Immunohistochemistry was used to determine the relative number of factor positive cells for BarH-like Homeobox 1 (BARX1), Distal-less Homeobox 4 (DLX4), Forkhead Box E1 (FOXE1), Homeobox B3 (HOXB3), Muscle Segment Homeobox 2 (MSX2), Paired Box Transcription Factor 7 (PAX7), Paired Box Transcription Factor 9 (PAX9), Receptor-like Tyrosine Kinase (RYK), Sonic Hedgehog (SHH), SRY Box Transcription Factor 3 (SOX3), Wingless-type MMTV Integration Site Protein 3A (WNT3A) and Wingless-type MMTV Integration Site Protein 9B (WNT9B). Healthy oral cavity tissue is characterized by the presence of DLX4, FOXE1, HOXB3, PAX7, RYK, SHH, SOX3, WNT3A and WNT9B gene proteins. In healthy oral cavity tissue WNT9B, WNT3A and SOX3 were found the most, SHH, PAX7, HOXB3, FOXE1 were found in moderate number, PAX9, DLX4, RYK were found a bit less, but practically no BARX1 and MSX2 containing cells were found. Cleft affected tissue is characterized by the increase of BARX1 and MSX2 and the decrease of SOX3, WNT3A and WNT9B. Unilateral cleft is characterized by the intensive production of BARX1, FOXE1, HOXB3 and PAX7 which indicates the increase of cell regeneration and proliferation potential intensity. Bilateral cleft is characterized by changes in DLX4, PAX9 and SHH expression – DLX4 and PAX9 increase, while SHH decreases. This reflects the compensatory functions of these factors within bilateral cleft tissue – DLX4 affects the vascularization of connective tissue, PAX9 regulates tissue growth and SHH could affect the formation of more severe cleft phenotypes. All cleft phenotypes are characterized by the increase of MSX2 and RYK and by the decrease of SOX3, WNT3A and WNT9B in tissue.