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111:Fate of Epithelial Palatal Seam With Smad2 Over-expression[AAOM2021]
Poster Title: 111:Fate of Epithelial Palatal Seam With Smad2 Over-expression[AAOM2021]
Submitted on 17 Apr 2021
Author(s): Dr. Husein Al-Omer¹ʼ³, Dr. Gihan E-H Gawish²ʼ³
Affiliations: Prince Abdulrahman Advanced Dental Institute,, Saudi Arabia
This poster was presented at 2021 American Academy of Oral Medicine Virtual Conference
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Poster Information
Abstract: OBJECTIVES In the present study, we investigated the mechanism of palatal fusion in TGF-β3 -/- /K14Smad2 compared to normal fusion in wild type mice.

Methods Samples: Pregnant C57BL/6 female were killed at day 14.5 , fetal heads were dissected under microscope, and genomic PCR performed on tissue from each embryo to determine genotypes. Heads were fixed and embedded in paraffin according to standard procedure. Serial coronal sections (5um) were placed on lysine-treated slides.

Immunofluorescence: After deparaffinization, rehydration and antigen retrieval using EDTA buffer, sections were blocked with 3% BSA/PBS for 30 min at room temperature to reduce background staining and incubated with the primary antibodies at 4oC overnight,. Fluorescence-labeled secondary antibodies were added for 1h at room temperature. After washing with PBS, cover slips were mounted with mounting medium including DAPI. To confirm the specificity of antibodies, additional slides were incubated without the primary antibodies. All sections were examined by using a Nikon Laser Scanning Confocal microscope. Quantitative counting for the MES cells were performed for sections of the anterior, middle, and posterior parts of the palate.

Apoptosis marker: antibody for cleaved caspase3 was used with E-cadherin and DAPI.

EMT marker: Snail antibody was used with DAPI


The targeted over-expression of Smad2 protein in the MEE rescued the cleft in TGF-β3 null mice palate, and the mechanism of palatal fusion in the presence of extra amounts of phosphorylated Smad2 was different than events happening in wild type. The MES in Smad2 over-expression mice have a higher ratio of apoptotic cells compared to wild type mice.

Of the three types of mice examined, the TGF-β3 -/- /K14Smad2 mice show the highest ratio of apoptotic cells in the midline seam where the two halves of the palate are fusing.

Sections from the middle part of the secondary palate show a higher apoptosis ratio than sections from more anterior and posterior along the seam.

Phospho-Smad2 expression in MEE of Smad2 over-expressed and TGF-β3 -/- /KSmad2 mice was related to the high apoptosis ratio.

Secondary palate in TGF-β3 -/- /K14Smad2 fused and the MES disintegrated mainly by apoptosis with high expression of Phospho-Smad2.
Summary: In the present study, we investigated the mechanism of palatal fusion in TGF-β3 -/- /K14Smad2 compared to normal fusion in wild type mice.

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References: Anderson JG, Peralta S, Kol A, Kass PH, Murphy B. Clinical and Histopathologic Characterization of Canine Chronic Ulcerative Stomatitis. Vet Pathol. 2017;54(3):511-9.

Anderson JG, Kol A, Bizikova P, Stapelton BP, Ford K, Villarreal A, et al. Immunopathogenesis of canine chronic ulcerative stomatitis. PLoS One. 2020;15(1):e0227386.

Chen T, Yu WH, Izard J, Baranova OV, Lakshmanan A, Dewhirst FE. The Human Oral Microbiome Database: a web accessible resource for investigating oral microbe taxonomic and genomic information. Database (Oxford). 2010;2010:baq013.

Dewhirst FE, Klein EA, Thompson EC, Blanton JM, Chen T, Milella L, et al. The canine oral microbiome. PLoS One. 2012;7(4):e36067.

Hajishengallis G, Darveau RP, Curtis MA. The keystone pathogen hypothesis. Nat Rev Microbiol. 2012;10(10):717-25.
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