The Association of TOX3 Copy Number Variation with Gene Expression and Susceptibility to Nonsyndromic Cleft Lip and/or Palate in a Malay Cohort

IntroductionNon-syndromic cleft lip with or without cleft palate is a common worldwide birth defect due to a combination of environmental and genetic factors. Genome-wide association studies reported the rs7078160 of Vax1 is closely related to non-syndromic cleft lip with or without cleft palate in European populations. The following studies showed the same results in Mongolian, Japanese, Filipino, Vietnamese populations etc. However, conflicting research had been reported in Chinese population, ObjectiveThe aim of this study was to investigate the association between the rs7078160 polymorphism and non-syndromic cleft lip with or without cleft palate in Southern Chinese patients. MethodsIn this study, we investigated the polymorphism distribution of rs7078160 in 100 complete patient trios (39 patients with non-syndromic cleft lip and palate; 36 patients with non-syndromic cleft lip only; 25 had non-syndromic cleft palate only; and their parents) from Southern ethnic Han Chinese. 60 healthy trios were selected as control. Polymerase chain reaction and Sanger sequencing were used to genotype rs7078160 in Vax1; both case–control and family-based associations were analyzed. ResultsThe case–control analyses revealed the rs7078160 polymorphism was significant, associated with non-syndromic cleft lip with or without cleft palate (p=0.04) and non-syndromic cleft lip and palate (p=0.01), but not associated with non-syndromic cleft lip only and non-syndromic cleft palate only patients. The genotype composition of rs7078160 comprises mutated homozygous AA, heterozygous AG and wild homozygous GG. Cases with AG+AA genotypes compared with GG homozygotes showed an increased risk of non-syndromic cleft lip with or without cleft palate (p=0.04, OR=2.05, 95% CI: 1.01–4.16) and non-syndromic cleft lip and palate (p=0.01, OR=3.94, 95% CI: 1.34–11.54). In addition, we did not detect any transmission-disequilibrium in rs7078160 (p=0.68). ConclusionThis study suggests that rs7078160 polymorphism is a risk factor of non-syndromic cleft lip with or without cleft palate, and Vax1 is strongly associated with non-syndromic cleft lip with or without cleft palate in Southern Chinese Han populations.

Among complications associated with non-syndromic cleft lip and/or palate (NSCL/P) in school age children, conductive hearing loss has been thoroughly investigated because the symptoms are noticeable and the treatment is often easy to access. Research on central auditory processing disorder [(C)APD] has been rarely explored in this clinical population. However, children with NSCL/P have been reported to have delayed speech and language development, as well as poor academic performance in general compared to craniofacially normal peers, despite their peripheral hearing problems typically resolving with age. In order to investigate suspected (C)APD in children with NSCL/P, the present research program was initiated.
\nThe study aimed to utilize a comprehensive test battery to assess auditory status in children with NSCL/P, and to investigate whether they have a greater frequency of (C)APD compared to craniofacially normal children. 147 children with NSCL/P and 60 normal children were recruited. They were Mandarin speakers and attending regular schools. The children with NSCL/P did not have peripheral hearing problems at time of assessment. There were three types of assessment tools used in the present research program. Firstly, hearing health tests were conducted to evaluate peripheral hearing function. Case history and auditory questionnaire reports were completed by caregivers to obtain basic medical and developmental information (Chapter 3). Secondly, auditory behavioral tests of (C)APD functioning were administered, including a gap detection test and a speech in noise recognition test (Chapter 4). Thirdly, an auditory evoked potential (AEP) assessment was conducted as an objective evaluation of the central auditory nervous system (Chapter 5).
\nBased on the results of this research program, it was concluded that, firstly, children with NSCL/P showed behaviors typically found in children with (C)APD when questionnaire results were considered. Children with cleft palate (CP) showed the most negative outcomes, and cleft lip (CL) group children showed results equivalent to craniofacially normal children. Secondly, behavioral assessment results showed that compared to control group children, maturation for temporal resolution abilities was delayed in children with NSCL/P. Also, the ability to use interaural timing and intensity cues for speech recognition in a noisy environment was poorer in children with CP and CLP. Finally, abnormal AEP findings in children with cleft suggested longer neural transmission times and delayed development of the auditory nervous system may occur in this population.
\nIn summary, the research program found that children with NSCL/P are at higher risk of auditory processing difficulties compared to craniofacially normal children. In addition, a comprehensive test battery is more appropriate for making an accurate diagnosis of (C)APD in this population than a single assessment protocol. The present research program has contributed to an enhanced awareness of potential (C)APD in children with NSCL/P, which had not been investigated using a comprehensive test battery for a large sample of children with cleft disorders in any previous studies. Since Chinese language specific assessment tools are limited, further studies to develop an appropriate, comprehensive test battery for the diagnosis of auditory processing disorder in Chinese children with oral cleft and to explore effective management of this disorder are required.

The aim of this study was to evaluate the association of the polymorphisms in TCN2 (rs1801198) gene and in MTRR (rs1801394) gene with nonsyndromic cleft lip and/or palate (NSCL/P) in a Brazilian population. Genomic DNA was extracted from buccal cells. The polymorphisms in TCN2 (rs1801198) and MTRR (rs1801394) genes were genotyped by carrying out real-time PCR and Taqman assay. Chi-square test was used to determine the association between genotype and allele frequencies with NSCL/P and NSCL/P subgroups (cleft lip only, cleft lip and palate, and cleft palate only). Eight hundred and sixty seven unrelated individuals (401 cases with NSCL/P and 466 individuals without cleft) were evaluated. Genotype distributions of TCN2 and MTRR polymorphisms were in Hardy-Weinberg equilibrium. The TCN2 polymorphic genotype GG was identified in 16.7% of the NSCL/P group and in 14.1% of the non-cleft group (p>0.05). Similarly, the frequency of MTRR genotype (GG) was similar in NSCL/P group (15.5%) and control group (17.8%) (p>0.05). Multivariate analysis showed an association between MTRR and the subgroup that the mother smoked during pregnancy (p=0.039). Our findings did not demonstrate an association between TCN2 polymorphisms and NSCL/P, however suggests an association between MTRR and NSCL/P etiology.

Background Cleft lip with or without cleft palate (CL/P) is the most common facial birth defect, with a worldwide incidence of 1 in 700-1000 live births. CL/P can be divided into syndromic CL/P (SCL/P) and nonsyndromic CL/P (NSCL/P). Genetic factors are an important component to the etiology of NSCL/P. ARHGAP29, one of the NSCL/P disease-causing genes, mediates the cyclical regulation of small GTP binding proteins such as RhoA and plays an essential role in cellular shape, proliferation, and craniofacial development. Methods The present study investigated a Chinese family with NSCL/P and explored potential pathogenic variants using whole-exome sequencing (WES). Variants were screened and filtered through bioinformatic analysis and prediction of variant pathogenicity. Cosegregation was subsequently conducted. Results We identified a novel heterozygous missense variant of ARHGAP29 (c.2615C > T, p.A872V) in a Chinese pedigree with NSCL/P. Conclusion We detected the disease-causing variant in this NSCL/P family. Our identification expands the genetic spectrum of ARHGAP29 and contributes to novel approaches to the genetic diagnosis and counseling of CL/P families.

PURPOSE: Nonsyndromic cleft lip and/or palate (NSCLP) is the most common congenital craniofacial anomaly. Early recognition of any associated developmental delay is critical to counseling families and developing individualized treatment plans. Here we sought to identify developmental delays associated with NSCLP in a large population of children in order to begin identifying etiology and improve multidisciplinary management. METHODS: This is an IRB-approved, single-center retrospective analysis of all patients with a diagnosis of cleft lip and/or cleft palate between 5 and 21 years of age. Demographic and clinical variables were collected from this patient population as well as from children comprising the 2018 National Survey of Children’s Health database. RESULTS: All children with an identified or suspected genetic syndrome were excluded (160 in our cohort and 1383 in the National Survey of Children’s Health database). Subsequently, 619 children in our cohort and 29,147 in the National Survey of Children’s Health database were identified with NSCLP and included in our analysis. The mean birth weight amongst NSCLP children was lower than that in the national cohort (108.5 ± 24.8 oz versus 117.8 ± 19.1 oz; P < 0.0001). Nearly one-fourth (25.8%) of children with NSCLP were admitted to the NICU at birth. The distribution of cleft lip/palate diagnoses in the NSCLP cohort is shown. Compared with the national cohort, children with isolated cleft palate had significantly higher rates of intellectual disability (3.2% versus 0.5%, P < 0.00001), speech delay (70.8% versus 7.1%, P < 0.00001), global developmental delay (15.7% versus 5.8%, P < 0.00001), cerebral palsy (2.2% versus 0.3%, P < 0.00001), and hearing loss (25.9% versus 1.0%, P < 0.00001). Rates of learning disability (7.0% versus 5.9%, P = 0.529), behavioral delay (7.6% versus 11.4%, P = 0.1038), ADD/ADHD (2.7% versus 2.3%, P = 0.7032), autism (4.3% versus 5.5%, P = 0.5005), and vision loss (1.6% versus 1.2%, P = 0.5764) were comparable between those with isolated cleft palate and the national cohort. Children with cleft lip (with or without cleft palate) had significantly higher rates of ADD/ADHD compared with the normative national cohort: isolated cleft lip (7.7% versus 2.3%, P = 0.0092), unilateral cleft lip and palate (4.6% versus 2.3%, P = 0.0088), bilateral cleft lip & palate (5.9% versus 2.3%, P = 0.0153). CONCLUSIONS: Our study demonstrates, for the first time, higher rates of various developmental delays in children with NSCLP compared with the general pediatric population. This includes increased rates of intellectual disability, global delay, and cerebral palsy in children with nonsyndromic isolated cleft palate and increased ADD/ADHD in children with cleft lip (with or without cleft palate). The association of NSCLP diagnoses with developmental delays highlights the importance of proper risk assessment of patients, appropriate family counseling, and multi-disciplinary team management.

Hypermethylation of WNT3A gene and non-syndromic cleft lip and/or palate in association with in utero exposure to lead: A mediation analysis

Non-syndromic cleft lip and/or palate (NCL/P) is a congenital craniofacial anomaly with significant psychosocial and economic impacts. In Ethiopia, the prevalence of NCL/P and access to specialized care are not well documented. This study aimed to evaluate the knowledge and awareness of NCL/P among families of affected children and dental school students in Ethiopia and explore their perceptions and attitudes toward NCL/P. From 2009 to 2018, we conducted questionnaire surveys involving 86 patients with NCL/P and their families in the towns of Addis Ababa and Butajira, including 161 students from the School of Dentistry, Addis Ababa University. The surveys assessed the knowledge, perceptions, and attitudes toward NCL/P and their social implications. The majority of patients with NCL/P were born at home and in low-income families. Concerns about the future social life of patients were prominent, with stigma and discrimination reported by 32% of the patients' families. Among the dental school students, 66% had some knowledge of NCL/P, primarily from media sources. Students perceived higher levels of societal blame towards mothers of patients with NCL/P than reported by the patients' families. This study revealed significant gaps in the knowledge and awareness regarding NCL/P among dental school students. It also revealed the substantial social stigma that the patients affected by NCL/P and their families faced in Ethiopia. Enhancing public education and providing comprehensive multidisciplinary care is crucial for improving the quality of life of patients with NCL/P in Ethiopia.

Behavioral assessment of auditory processing disorder in children with non-syndromic cleft lip and/or palate

Mutation at Paired box gene 9 is associated with non-syndromic cleft lip only from Western Han Chinese population

To determine the association of single-nucleotide polymorphisms (SNPs) in genes related to craniofacial development, which were previously identified as susceptibility signals for nonsyndromic oral clefts, in Brazilians with nonsyndromic cleft lip and/or palate (NSCL/P). The SNPs rs748044 (TNP1), rs1106514 (MSX1), rs28372960, rs15251 and rs2569062 (TCOF1), rs7829058 (FGFR1), rs1793949 (COL2A1), rs11653738 (WNT3), and rs242082 (TIMP3) were assessed in a family-based transmission disequilibrium test (TDT) and a structured case-control analysis based on the individual ancestry proportions. The SNPs were initially analyzed by TDT, and polymorphisms showing a trend toward excess transmission were subsequently studied in an independent case-control sample. The study sample consisted of 189 case-parent trios of nonsyndromic cleft lip with or without cleft palate (NSCL±P), 107 case-parent trios of nonsyndromic cleft palate (NSCP), 318 isolated samples of NSCL±P, 189 isolated samples of NSCP, and 599 healthy controls. Association of alleles with NSCL/P pathogenesis. Preferential transmission of SNPs rs28372960 and rs7829058 in NSCL±P trios and rs11653738 in NSCP trios (P = .04) were observed, although the structured case-control analysis did not confirm these associations. The haplotype T-C-C formed by TCOF1 SNPs rs28372960, rs15251, and rs2569062 was more frequently transmitted from healthy parents to NSCL±P offspring, but the P value (P = .01) did not withstand Bonferroni correction for multiple tests. With the modest associations, our results do not support the hypothesis that TNP1, MSX1, TCOF1, FGFR1, COL2A1, WNT3, and TIMP3 variants are risk factors for nonsyndromic oral clefts in the Brazilian population.

To study the relationship between Sonic hedgehog (Shh) associated single-nucleotide polymorphism (SNP) and non-syndromic cleft lip and/or palate (NSCL/P), and to explore the risk factors of cleft lip and/or palate. Many studies suggest that the pathogenesis of NSCL/P could be related to genes that control early development, in which the Shh signaling pathway plays an important role. Peripheral blood was collected from 197 individuals (100 patients with NSCL/P and 97 healthy controls). Haploview software was used for haplotype analysis and Tag SNP were selected, based on the population data of Han Chinese in Beijing of the international human genome haplotype mapping project. A total of 27 SNP were selected for the 4 candidate genes of SHH, PTCH1, SMO and GLI2 in the Shh signaling pathway. The genotypes of 27 SNP were detected and analyzed by Sequenom mass spectrometry. The data were analyzed by chi-squared test and an unconditional Logistic regression model. The selected SNP basically covered the potential functional SNP of the target genes, and its minimum allele frequency (MAF) was >0.05: GLI2 73.5%, PTCH1 91.0%, SMO 100.0%, and SHH 75.0%. It was found that the genotype frequency of SNP (rs12674259) located in SMO gene and SNP (rs2066836) located in PTCH1 gene were significantly different between the NSCL/P group and the control group. Linkage disequilibrium was also found on 3 chromosomes (chromosomes 2, 7 and 9) where the 4 candidate genes were located. However, in the analysis of linkage imbalance haplotype, there was no significant difference between the disease group and the control group. In China, NSCL/P is the most common congenital disease in orofacial region. However, as it is a multigenic disease and could be affected by multiple factors, such as the external environment, the etiology of NSCL/P has not been clearly defined. This study indicates that Shh signaling pathway is involved in the occurrence of NSCL/P, and some special SNP of key genes in this pathway are related to cleft lip and/or palate, which provides a new direction for the etiology research of NSCL/P and may provide help for the early screening and risk prediction of NSCL/P.

ObjectiveThe aim of the study was to conduct a meta-analysis of research examining the early speech and language functioning of young children, birth to age 8;11 (years;months), with nonsyndromic cleft lip and/or palate (NSCL/P) compared to their peers without NSCL/P.MethodWe conducted a random-effects metaregression using 241 effect sizes from 31 studies comparing 955 young children with NSCL/P to 938 typically developing peers on measures of speech and language functioning. Moderators were sample characteristics (i.e., age, cleft type, publication year, and study location) and measurement characteristics (i.e., speech sample material, language modality and domain, and assessment type).ResultsYoung children with NSCL/P scored significantly lower on measures of speech and language compared to children without NSCL/P. Children with NSCL/P had smaller consonant inventories (standardized mean difference effect size [ESg] = −1.24), less accurate articulation (ESg = −1.13), and more speech errors (ESg = 0.93) than their peers. Additionally, children with NSCL/P had poorer expressive (ESg = −0.57) and receptive (ESg = −0.59) language skills than their peers. Age and assessment type moderated effect sizes for expressive language. As children with NSCL/P aged, their expressive language performance became more similar to their peers. Expressive language effect sizes from parent reports and observational language measures (estimated effect size = −0.74) were significantly lower than those from standardized norm-referenced tests (estimated effect size = −0.45).ConclusionsThese findings suggest that young children with NSCL/P experience delays relative to their peers across multiple speech and language constructs. Differences between children with NSCL/P and their typically developing peers appear to decrease with age.Supplemental Material https://doi.org/10.23641/asha.11356904

Objective: The aim of the study was to conduct a meta-analysis of research examining the early speech and language functioning of young children, birth to age 8;11 (years;months), with nonsyndromic cleft lip and/or palate (NSCL/P) compared to their peers without NSCL/P.Method: We conducted a random-effects metaregression using 241 effect sizes from 31 studies comparing 955 young children with NSCL/P to 938 typically developing peers on measures of speech and language functioning. Moderators were sample characteristics (i.e., age, cleft type, publication year, and study location) and measurement characteristics (i.e., speech sample material, language modality and domain, and assessment type).Results: Young children with NSCL/P scored significantly lower on measures of speech and language compared to children without NSCL/P. Children with NSCL/P had smaller consonant inventories (standardized mean difference effect size [ESg] = −1.24), less accurate articulation (ESg = −1.13), and more speech errors (ESg = 0.93) than their peers. Additionally, children with NSCL/P had poorer expressive (ESg = −0.57) and receptive (ESg = −0.59) language skills than their peers. Age and assessment type moderated effect sizes for expressive language. As children with NSCL/P aged, their expressive language performance became more similar to their peers. Expressive language effect sizes from parent reports and observational language measures (estimated effect size = −0.74) were significantly lower than those from standardized norm-referenced tests (estimated effect size = −0.45).Conclusions: These findings suggest that young children with NSCL/P experience delays relative to their peers across multiple speech and language constructs. Differences between children with NSCL/P and their typically developing peers appear to decrease with age.Supplemental Material S1. PRISMA 2009 Checklist.Supplemental Material S2. Code for meta-analysis. Supplemental Material S3. List of citations and study ID number for included studies.Lancaster, H. P., Lien, K. M., Chow, J. C., Frey, J. R., Scherer, N. J., & Kaiser, A. P. (2019). Early speech and language development in children with nonsyndromic cleft lip and/or palate: A meta-analysis. Journal of Speech, Language, and Hearing Research. Advance online publication. https://doi.org/10.1044/2019_JSLHR-19-00162

Association between FOXE1 and non-syndromic orofacial clefts in a northeastern Chinese population

To analyse the sex ratio of non-syndromic cleft lip and/or palate (NSCL/P) in Hong Kong Chinese. All cases of NSCL/P delivered in a teaching hospital in Hong Kong from 2001–2014 were reviewed. The sex ratio for different kinds of NSCL/P was compared using chi square test. During the study period, 87,989 (45,562 male and 42,427 female) live births were delivered. There were 96 cases of NSCL/P, 50 male and 46 female. The incidence of NSCL/P was 1:916 live births (1:911 male and 1:922 female). There were 54 (56%) cleft lip and palate, 24 (25%) cleft lip only and 18 (19%) cleft palate only. Male was more frequently affected than female, with male: female (M:F) ratio of 1.1:1. This feature of male predominance was observed in cleft lip only, cleft lip and palate and cleft lip with or without cleft palate, with M:F ratio of 1.7:1, 1.3:1 and 1.4:1 respectively. However, the sex ratio was reversed in cleft palate only, with M:F ratio of 0.3:1. The difference is statistically significant (p = 0.0175). Non-syndromic cleft lip with or without palate is more common in male while non-syndromic cleft palate only is more common in female. This may indicate that they represent two different entities with totally different genetic basis and environmental trigger. Researchers working on NSCL/P may need to make a distinction between the two entities.