DNA hypermethylation of Fgf16 and Tbx22 associated with cleft palate during palatal fusion

We previously reported that mutation of the transforming growth factor-beta3 (TGF-beta3) gene caused cleft palate in homozygous null (−/−) mice. TGF-beta3 is normally expressed in the medial edge epithelial (MEE) cells of the palatal shelf. In the present study, we investigated the mechanisms by which TGF-beta3 deletions caused cleft palate in 129 × CF-1 mice. For organ culture, palatal shelves were dissected from embryonic day 13.5 (E13.5) mouse embryos. Palatal shelves were placed singly or in pairs on Millipore filters and cultured in DMEM/F12 medium. Shelves were placed in homologous (+/+ vs +/+, −/− vs −/−, +/− vs +/−) or heterologous (+/+ vs −/−, +/− vs −/−, +/+ vs +/−) paired combinations and examined by macroscopy and histology. Pairs of −/− and −/− shelves failed to fuse over 72 hours of culture whereas pairs of +/+ (wild-type) and +/+ or +/− (heterozygote) and +/−, as well as +/+ and −/− shelves, fused within the first 48 hour period. Histological examination of the fused +/+ and +/+ shelves showed complete disappearance of the midline epithelial seam whereas −/− and +/+ shelves still had some seam remnants. In order to investigate the ability of TGF-beta family members to rescue the fusion between −/− and −/− palatal shelves in vitro, either recombinant human (rh) TGF-beta1, porcine (p) TGF-beta2, rh TGF-beta3, rh activin, or p inhibin was added to the medium in different concentrations at specific times and for various periods during the culture. In untreated organ culture −/− palate pairs completely failed to fuse, treatment with TGF-beta3 induced complete palatal fusion, TGF-beta1 or TGF-beta2 near normal fusion, but activin and inhibin had no effect. We investigated ultrastructural features of the surface of the MEE cells using SEM to compare TGF-beta3-null embryos (E 12. 5-E 16.5) with +/+ and +/− embryos in vivo and in vitro. Up to E13.5 and after E15.5, structures resembling short rods were observed in both +/+ and −/− embryos. Just before fusion, at E14.5, a lot of filopodia-like structures appeared on the surface of the MEE cells in +/+ embryos, however, none were observed in −/− embryos, either in vivo or in vitro. With TEM these filopodia are coated with material resembling proteoglycan. Interestingly, addition of TGF-beta3 to the culture medium which caused fusion between the −/− palatal shelves also induced the appearance of these filopodia on their MEE surfaces. TGF-beta1 and TGF-beta2 also induced filopodia on the −/− MEE but to a lesser extent than TGF-beta3 and additionally induced lamellipodia on their cell surfaces. These results suggest that TGF-beta3 may regulate palatal fusion by inducing filopodia on the outer cell membrane of the palatal medial edge epithelia prior to shelf contact. Exogenous recombinant TGF-beta3 can rescue fusion in −/− palatal shelves by inducing such filopodia, illustrating that the effects of TGF-beta3 are transduced by cell surface receptors which raises interesting potential therapeutic strategies to prevent and treat embryonic cleft palate.

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Cleft palate formation after palatal fusion occurs due to the rupture of epithelial basement membranes

Journal of Cranio-Maxillofacial Surgery ◽

10.1016/j.jcms.2018.09.016 ◽

2018 ◽

Vol 46 (12) ◽

pp. 2027-2031 ◽

Cited By ~ 6

Author(s):

Chisato Sakuma ◽

Hideto Imura ◽

Tomohiro Yamada ◽

Toshio Sugahara ◽

Azumi Hirata ◽

...

Keyword(s):

Cleft Palate ◽

Basement Membranes ◽

Palatal Fusion ◽

Epithelial Basement

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Ras signaling and its effector RREB1 are required for the dissociation of MEE cells in palatogenesis

Disease Models & Mechanisms ◽

10.1242/dmm.049093 ◽

2021 ◽

Author(s):

Toshihiro Inubushi ◽

Ayaka Fujiwara ◽

Takumi Hirose ◽

Gozo Aoyama ◽

Toshihiro Uchihashi ◽

...

Keyword(s):

Embryonic Development ◽

Organ Culture ◽

Cleft Palate ◽

Birth Defects ◽

Crucial Role ◽

Initial Step ◽

Ras Signaling ◽

Palatal Fusion ◽

Element Binding Protein ◽

Responsive Element

Cleft palate is one of the major congenital craniofacial birth defects. The etiology underlying the pathogenesis of cleft palate has largely remained unelucidated. Dissociation of the medial edge epithelium (MEE) at the contacting region of palatal shelves and subsequent migration or apoptosis of MEE cells is required for the proper MEE removal. Ras Responsive Element Binding Protein 1 (RREB1), a RAS transcriptional effector, has recently been shown to play a crucial role in developmental EMT, in which loss of epithelial characteristics is an initial step, during mid-gastrulation of embryonic development. Interestingly, the involvement of RREB1 in cleft palate has been indicated in humans. Here, we demonstrated that pan-Ras inhibitor prevents the dissociation of MEE during palatal fusion. Rreb1 is expressed in the palatal epithelium during palatal fusion, and knockdown of Rreb1 in palatal organ culture resulted in palatal fusion defects by inhibiting the dissociation of MEE cells. Our present findings provide evidence that RREB1-mediated Ras signaling is required during palatal fusion. Aberrant RREB1-mediated Ras signaling might be involved in the pathogenesis of cleft palate.

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In utero gene delivery of adenovirus encoded TGF-beta3 restores physiologic palatal fusion and rescues cleft palate in a TGF-beta3 knockout mouse

Journal of the American College of Surgeons ◽

10.1016/j.jamcollsurg.2007.06.229 ◽

2007 ◽

Vol 205 (3) ◽

pp. S92 ◽

Cited By ~ 2

Author(s):

Ryan M. Spivak ◽

Masayuki Endo ◽

Allison Zajac ◽

Philip Zoltick ◽

Brian Ang ◽

...

Keyword(s):

Gene Delivery ◽

Cleft Palate ◽

Knockout Mouse ◽

In Utero ◽

Palatal Fusion

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Anterior Cleft Palate due to Cbfb deficiency and its rescue by folic acid

10.1101/515650 ◽

2019 ◽

Author(s):

Safiye E. Sarper ◽

Toshihiro Inubushi ◽

Hiroshi Kurosaka ◽

Hitomi Ono Minagi ◽

Yuka Murata ◽

...

Keyword(s):

Transcription Factors ◽

Folic Acid ◽

Cleft Palate ◽

Anterior Portion ◽

Pharmaceutical Application ◽

Palatal Fusion ◽

Stat3 Phosphorylation ◽

Signaling Axis ◽

Summary Statement

AbstractCore binding factor β (Cbfb) is a cofactor of Runx transcription factors. Among Runx transcription factors, Runx1 is a prerequisite for anterior-specific palatal fusion. However, whether Cbfb serves as a modulator or obligatory factor in Runx signaling that regulates palatogenesis is unclear. We herein report that Cbfb is essential and indispensable in anterior palatogenesis. Palatal fusion in Cbfb mutants is disturbed due to failed disintegration of the fusing epithelium specifically at the anterior portion, as is observed in Runx1 mutants. In this mutants, the Tgfb3 expression is disturbed at the corresponding area of the failed palatal fusion, where phosphorylation of Stat3 is also disturbed. TGFB3 protein rescues the palatal fusion in vitro. Strikingly, the anterior cleft palate in Cbfb mutants is further rescued by pharmaceutical application of folic acid that activates suppressed Stat3 phosphorylation and Tgfb3 expression in vitro. With these findings, we provide the first evidence that Cbfb is a prerequisite for anterior palatogenesis as an obligatory cofactor in the Runx1/Cbfb-Stat3-Tgfb3 signaling axis. Furthermore, the rescue of the mutant cleft palate using folic acid may elucidate potential therapeutic targets by Stat3 modification for the prevention and pharmaceutical intervention of cleft palate.Summary StatementEpithelial deletion of Cbfb results in an anterior cleft palate with impaired fusion of the palatal process and folic acid application rescues the mutant phenotype with Stat3 activation in vitro.

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Occurrence of Cleft-Palate and Alteration of Tgf-β3 Expression and the Mechanisms Leading to Palatal Fusion in Mice following Dietary Folic-Acid Deficiency

Cells Tissues Organs ◽

10.1159/000323213 ◽

2011 ◽

Vol 194 (5) ◽

pp. 406-420 ◽

Cited By ~ 17

Author(s):

Estela Maldonado ◽

Jorge Murillo ◽

Carmen Barrio ◽

Aurora del Río ◽

Juliana Pérez-Miguelsanz ◽

...

Keyword(s):

Folic Acid ◽

Cleft Palate ◽

Folic Acid Deficiency ◽

Palatal Fusion ◽

Acid Deficiency

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Classroom Teacher and Speech Clinician Severity Ratings of Different Speech Disorders

Language Speech and Hearing Services in Schools ◽

10.1044/0161-1461.0603.119 ◽

1975 ◽

Vol 6 (3) ◽

pp. 119-124 ◽

Cited By ~ 3

Author(s):

Robert T. Wertz ◽

Michael D. Mead

Keyword(s):

Public School ◽

Cleft Palate ◽

Classroom Teacher ◽

First Grade ◽

Speech Disorders ◽

Third Grade ◽

Second Grade ◽

Classroom Teachers

Typical examples of four different speech disorders—voice, cleft palate, articulation, and stuttering—were ranked for severity by kindergarten, first-grade, second-grade, and third-grade teachers and by public school speech clinicians. Results indicated that classroom teachers, as a group, moderately agreed with speech clinicians regarding the severity of different speech disorders, and classroom teachers displayed significantly more agreement among themselves than did the speech clinicians.

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Holistic Communication Assessment for Young Children With Cleft Palate Using the International Classification of Functioning, Disability and Health:Children and Youth

Language Speech and Hearing Services in Schools ◽

10.1044/2020_lshss-19-00122 ◽

2020 ◽

Vol 51 (4) ◽

pp. 914-938

Author(s):

Anna Cronin ◽

Sharynne McLeod ◽

Sarah Verdon

Keyword(s):

Young Children ◽

Cleft Palate ◽

Case History ◽

Significant Others ◽

International Classification ◽

Assessment Tools ◽

International Classification Of Functioning ◽

Speech Language Pathologists ◽

Holistic Assessment

Purpose Children with a cleft palate (± cleft lip; CP±L) can have difficulties communicating and participating in daily life, yet speech-language pathologists typically focus on speech production during routine assessments. The International Classification of Functioning, Disability and Health: Children and Youth Version (ICF-CY; World Health Organization, 2007 ) provides a framework for holistic assessment. This tutorial describes holistic assessment of children with CP±L illustrated by data collected from a nonclinical sample of seven 2- to 3-year-old children, 13 parents, and 12 significant others (e.g., educators and grandparents). Method Data were collected during visits to participants' homes and early childhood education and care centers. Assessment tools applicable to domains of the ICF-CY were used to collect and analyze data. Child participants' Body Functions including speech, language, and cognitive development were assessed using screening and standardized assessments. Participants' Body Structures were assessed via oral motor examination, case history questionnaires, and observation. Participants' Activities and Participation as well as Environmental and Personal Factors were examined through case history questionnaires, interviews with significant others, parent report measures, and observations. Results Valuable insights can be gained from undertaking holistic speech-language pathology assessments with children with CP±L. Using multiple tools allowed for triangulation of data and privileging different viewpoints, to better understand the children and their contexts. Several children demonstrated speech error patterns outside of what are considered cleft speech characteristics, which underscores the importance of a broader assessment. Conclusion Speech-language pathologists can consider incorporating evaluation of all components and contextual factors of the ICF-CY when assessing and working with young children with CP±L to inform intervention and management practices.

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