Nonsyndromic Cleft Lip Palate Research Articles

Association of methylenetetrahydrofolate reductase gene variant C677T and folate levels in non-syndromic cleft lip/palate among Sindhi, Pakistani population.

The object ives of this study were to determine the association of methylenetetrahydrofolate reduc tase (MTHFR) gene variant C67 7 T with non -syndromic cl eft lip/palate (NSCLP) in Pakistani population and compare the m aternal serum foli c acid levels in NSCLP-affected and healthy group. A c om parative cross sec ti onal study was conducted between 2017 and 2019 at Liaquat U niversity of Medi cal and Health Science s, Jamshoro. Sixty motherinfant dy ads were recruited (n=120), inc luding NSCLP-affected and healthy infants alo ng with t heir mother s. The MTHFR C677T vari ant exhibited si gnificant association with NSCLP in dominant and over-domi nant models. No differences in maternal serum folic acid levels were obse rved between both th e groups; however, the folic acid intake during pre-conception period was associated w ith decreased risk for NSC LP. Our stu dy suggested that MTHFR 677 CT genotype was related with decreased risk for NSCLP in Sindhi, Pakistani, population. Pre -conception folic acid may decrease the ri sk for ora l clefts.

The impact of passive alveolar molding vs. nasoalveolar molding on cleft width and other parameters of maxillary growth in unilateral cleft lip palate

ObjectivePassive alveolar molding (PAM) and nasoalveolar molding (NAM) are established presurgical infant orthodontic (PSIO) therapies for cleft lip palate (CLP) patients. PAM guides maxillary growth with a modified Hotz appliance, while NAM also uses extraoral taping and includes nasal stents. The effects of these techniques on alveolar arch growth have rarely been compared.Material and methodsWe retrospectively compared 3D-scanned maxillary models obtained before and after PSIO from infants with unilateral, non-syndromic CLP treated with PAM (n = 16) versus NAM (n = 13). Nine anatomical points were set digitally by four raters and transversal/sagittal distances and rotations of the maxilla were measured.ResultsBoth appliances reduced the anterior cleft, but NAM percentage wise more. NAM decreased the anterior and medial transversal width compared to PAM, which led to no change. With both appliances, the posterior width increased. The alveolar arch length of the great and small segments and the sagittal length of the maxilla increased with PAM but only partially with NAM. However, NAM induced a significant greater medial rotation of the larger and smaller segment compared to PAM with respect to the lateral angle.ConclusionsNAM and PAM presented some significant differences regarding maxillary growth. While NAM reduced the anterior cleft and effectively rotated the segments medially, PAM allowed more transversal and sagittal growth.Clinical relevanceThe results of this study should be taken into consideration when to decide whether to use PAM or NAM, since they show a different outcome within the first few months. Further studies are necessary regarding long-term differences.

Association of Single-Nucleotide Polymorphisms of MAFB Gene with Nonsyndromic Cleft Lip with or without Cleft Palate in Kinh Vietnamese Patients.

Background Cleft lip with or without palate (CL/P) is the most common orofacial birth defect. Single-nucleotide polymorphisms (SNPs) in MAFB gene (V-Maf avian musculoaponeurotic fibrosarcoma oncogene homolog B) were identified as susceptible to this defect in a genome-wide association study. To further evaluate its role in this birth defect, we conducted this study with the aim of identifying allele frequencies, genotype frequencies, and association of SNPs rs13041247, rs6065259, and rs6072081 of MAFB gene with nonsyndromic cleft lip/palate (NCL/P) in Kinh Vietnamese patients. Methods We performed case–control study involved 79 patients with NCL/P and 77 healthy controls. DNAs were extracted from participants' saliva and tetra-amplification refractory mutation system polymerase chain reaction (tetra-ARMS PCR) was used for genotyping SNPs. Results SNPs of MAFB gene were genotyped using the Tetra-ARMS PCR method. We found that genotype CT of rs13041247 was associated with an increased risk of NCL/P in Kinh Vietnamese (odds ratio TCTT [OR TC/TT ] = 1.63, 95% confidence interval [CI] = 0.83–3.19, p = 0.17). The G allele genotypes of SNP rs6072081 increase high risk for the malformation, statistically significant result (OR GG/AA = 7.06, 95% CI = 2.13–23.42, p < 0.001). There is no clear association between rs6065259 and CL/P (OR AA/GG = 0.75, 95% CI = 0.22–2.50, p = 0.32; OR AG/GG = 1.53, 95% CI = 0.79–2.97, p = 0.32). When the patients were divided into the phenotypic subgroups, there was a similar significant trend between the patients and controls for all SNPs. Conclusions Our study provides further evidence of role of MAFB gene variations with NCL/P defect in Kinh Vietnamese.

Parental transmission effects of the PAX7 Polymorphisms among non-syndromic cleft lip palate: A case-parent trio study

Background: Orofacial clefts (OFCs) are the most common congenital deformities in the human face, with a prevalence of around 1:800 to 1:1000 among the Indian population. Aim: To assess the Paired box 7 (PAX7) gene polymorphisms in the etiology of non-syndromic cleft lip and cleft palate (NSCL/P) in the Indian population using a case-parent trio design and to contemplate parent-of-origin effects. Materials and Methods: The study comprised forty case-parent trios of NSCL/P from the Indian population. Genomic deoxyribonucleic acid (DNA) was isolated from the cases and their parents. The polymorphisms rs6659735, rs553934, rs624761, rs609959, and rs4075768 of the PAX7 gene were genotyped using Agena Bio MassARRAY analysis. The transmission disequilibrium test (TDT) was performed using the PLINK software and the pairwise linkage disequilibrium analysis by the Haploview software. Results: The genotyping of the polymorphisms rs6659735, rs553934, rs624761, rs609959, and rs4075768 was done. We found a significant association of the rs6659735 (P-value = 0.03, Odds ratio = 4.5) with NSCL/P in the allelic frequencies of the Indian case-parent trios. In addition, the parent-of-origin effects were observed as the allelic TDT analysis showed a significant paternal transmission of the rs6659735 (P-value = 0.04), whereas the rs553934 (P-value = 0.04) showed an excess maternal transmission. Conclusions: The parent-of-origin effects were observed as the two polymorphisms demonstrated a significant parental transmission. The rs6659735 showed a significant paternal transmission, whereas the rs553934 showed an excess maternal transmission, suggesting that the PAX7 gene may influence the risk of NSCL/P in the Indian population.

Linkage and association of PAX7 polymorphisms (rs742071, rs766325, and rs4920520) with the risk of non-syndromic cleft lip with/without cleft palate: A systematic review and meta-analysis

The studies have reported several additional non-syndromic cleft lip/palate (NSCL/P) susceptibility loci. This systematic review and meta-analysis aimed to evaluate the linkage and association of PAX7 polymorphisms with the NSCL/P risk. A comprehensive search was conducted in the PubMed, Cochrane Library, Web of Science, and Scopus databases until May 15, 2021. The association between PAX7 polymorphisms and NSCL/P susceptibility was analyzed by calculation of the odds ratios (ORs) and 95% confidence intervals (CIs), and the linkage was assessed by the allelic transmission disequilibrium test. Some articles included more than one study (reporting more than one polymorphism). Therefore, nine articles including 13 studies were entered into the meta-analysis. With regard to the association of rs742071 polymorphism with the risk of NSCL/P, the pooled OR was 1.33 for the allelic (P < 0.0001), 1.98 for the homozygous (P = 0.0007), 1.41 for the heterozygous (P = 0.0267), and 1.15 for the dominant (P = 0.2542) models. For the association of PAX7 rs766325 and rs4920520 polymorphisms with the risk of NSCL/P, the pooled OR was 0.96 (P = 0.5571) and 1.14 (P = 0.1020), respectively. In addition, the pooled allelic transmission disequilibrium test for s742071 and rs766325 polymorphisms did not show any allelic linkage between these polymorphisms and susceptibility to NSCL/P. The main results of the present systematic review and meta-analysis showed an association between PAX7 rs742071 polymorphism and NSCL/P susceptibility; but there was no linkage or association with rs766325 and rs4920520 polymorphisms.

Analysis of Single Nucleotide Polymorphisms on Locus 13q33.1-34 in Multigenerational Families of Cleft Lip Palate using MassArray

BACKGROUND: Cleft lip palate is a common congenital anomaly with multifactorial etiology. Many high-risk markers at different loci were reported to be involved in its etiology. Advanced genetic research led to the discovery of evidence of a new linkage on 13q33.1-34 region at marker rs1830756 in two multigenerational Indian families. However, no further study was reported to confirm or validate this linkage in other families. Hence, the present study was designed.METHODS: Twenty multigenerational families affected by non-syndromic cleft lip palate were selected for the study. Polymorphisms, rs1830756, rs1323672, rs1935135 of FAM155A gene; rs1961495, rs953386, rs1411040 of COL4A1 gene; and rs726449, rs984300 of MYO16 gene were selected. Genomic DNA was isolated and sent for genetic analysis by single nucleotide polymorphism (SNP) genotyping using the MassArray method. Statistical analysis of the genomic data was done by PLINK. Bonferroni correction was applied and haplotype analysis was done using Haploview software.RESULTS: Polymorphisms followed the Hardy Weinberg Equilibrium. In the allelic association, all the polymorphisms analysed showed no statistical significance. Hence, there was no significant difference in the allelic frequencies between non-syndromic cleft lip palate patients and healthy controls. The odds ratio was not more than 1.6 for all the SNPs. Haplotype analysis showed that haplotypes were not significantly higher in non-syndromic cleft patients than in control subjects.CONCLUSION: There is no association between SNPs analysed in the locus 13q33.1-34 with cleft lip palate.KEYWORDS: cleft lip palate, chromosome, polymorphism

Association of Single Nucleotide Polymorphisms on Locus 18q21.1 in the Etiology of Nonsyndromic Cleft Lip Palate (NSCLP) in Indian Multiplex Families.

Background Cleft lip palate (CLP) is a common congenital anomaly with multifactorial etiology. Many polymorphisms at different loci on multiple chromosomes were reported to be involved in its etiology. Genetic research on a single multigenerational American family reported 18q21.1 locus as a high-risk locus for nonsyndromic CLP (NSCLP). However, its association in multiple multiplex families and Indian population is not analyzed for its association in NSCLP. Aim This study was aimed to evaluate whether high-risk single nucleotide polymorphisms (SNPs) on chromosome 18q21.1 are involved in the etiology of NSCLP in multiplex Indian families. Materials and Methods Twenty multigenerational families affected by NSCLP were selected for the study after following inclusion and exclusion criteria. Genomic DNA was isolated from the affected and unaffected members of these 20 multiplex families and sent for genetic analysis. High-risk polymorphisms, such as rs6507872 and rs8091995 of CTIF , rs17715416, rs17713847 and rs183559995 of MYO5B , rs78950893 of SMAD7 , rs1450425 of LOXHD1 , and rs6507992 of SKA1 candidate genes on the 18q21.1 locus, were analyzed. SNP genotyping was done using the MassARRAY method. Statistical analysis of the genomic data was done by PLINK. Results Polymorphisms followed the Hardy–Weinberg equilibrium. In the allelic association, all the polymorphisms had a p -value more than 0.05. The odds ratio was not more than 1.6 for all the SNPs. Conclusion High-risk polymorphisms, such as rs6507872 and rs8091995 of CTIF , rs17715416, rs17713847 and rs183559995 of MYO5B , rs78950893 of SMAD7 , rs1450425 of LOXHD1 , and rs6507992 of SKA1 in the locus 18q21.1, are not associated with NSCLP in Indian multiplex families.

Polymorphisms of ATP-Binding Cassette, Sub-Family A, Member 4 (rs560426 and rs481931) and Non-Syndromic Cleft Lip/Palate: A Meta-Analysis.

Background: A number of genes are associated with the incidence of non-syndromic cleft lip/palate (NSCL/P). Studies have shown a significant association between polymorphisms of ATP-binding cassette, sub-family A, member 4 (ABCA4) with the risk of NSCL/P. The present meta-analysis assessed the association between ABCA4 polymorphisms (rs560426 and rs481931) and the NSCL/P risk by reviewing case-control studies. Methods: Four databases (Scopus; Cochrane Library; Web of Science; and PubMed) were searched for articles published up to June 2020. The Review Manager 5.3 software was used to calculate the crude odds ratio (OR) and 95% confidence interval (CI). Both subgroup analyses for ethnicity and source of controls and a meta-regression related to publication year were conducted. Results: Of 94 retrieved studies, 12 were analyzed in this meta-analysis (2859 NSCL/P patients and 3792 controls for ABCA4 rs560426 polymorphism and 1333 NSCL/P patients and 1884 controls for ABCA4 rs481931 polymorphism). Overall, there was no significant association between both polymorphisms and the risk of NSCL/P. However, subgroup analysis demonstrated that there was a higher risk of NSCL/P for specific models: the allelic model (OR = 1.13; p = 0.03), the homozygote model (OR = 1.53; p = 0.04), and the recessive model (OR = 1.30; p = 0.03) in the Asian ethnicity for the rs560426 polymorphism. Conclusion: The findings confirmed that the NSCL/P risk was significantly associated with the G allele and GG genotype of rs560426 polymorphism but not for rs481931 polymorphism. There were no associations between both polymorphisms (rs560426 and rs481931) and the NSCL/P risk in those of European descent and the mixed ethnicities.

Screening of IRF6 Variants in Patients Subjected to Genetic Association Studies for Nonsyndromic Cleft Lip/Palate

To screen for interferon regulatory factor 6 (IRF6) pathogenic variants in patients clinically diagnosed with nonsyndromic cleft lip palate (NSCL/P) and establish the proportion of misdiagnosed Van der Woude syndrome (VWS) cases, which could have biased previous NSCL/P case-control association studies. Retrospective case series. Tertiary care children's hospital. One hundred seventy-two unrelated Mexican patients with NSCL/P, 128 of whom had previously been included in a NSCL/P case-control association study. Sanger sequencing of the 9 IRF6 exons were performed, all variants respect with sequence reference were reported and classified for their pathogenic significance according to the American College of Medical Genetics and Genomics guidelines. Seven percent of cases were familial. No pathogenic variant was identified in IRF6. We identified 12 previously reported benign variants; their frequencies did not significantly differ from those reported for individuals of Mexican ancestry. Three of them were uncommon intronic variants not reported in ClinVar. The rs2235371 and rs2235375 variants, which were previously analyzed in a NSCL/P case-control association study (containing 132 patients, 128 of whom were analyzed herein) did not show discordant association results comparing to the 370 controls from the previous study. The misdiagnosis of IRF6-related VWS as NSCL/P appears to be infrequent in our sample, suggesting that mutational screening of IRF6 would have a low diagnostic yield in patients with NSCL/P. The absence of IRF6 pathogenic alleles could be related to the application of an exhaustive clinical evaluation that discarded the syndromic forms and/or the low proportion of familial cases included.

CRISPLD2 Gene Polymorphisms with Nonsyndromic Cleft Lip Palate in Indian Population

Cleft lip palate (CLP) is one of the common congenital anomalies with multifactorial etiology. Many genes are associated with its etiology. In one of the studies CRISPLD2 gene polymorphisms rs1546124, rs4783099, and rs16974880 were reported in the Chinese population. However, its role in the Indian population is not yet studied. Hence, this research was conducted on the Indian population to know the role of these high-risk polymorphisms in patients with nonsyndromic CLP. Following an inclusion and exclusion criteria, 20 multiplex CLP families were selected from a high volume cleft center in India. Genomic DNA was isolated from these families. Single nucleotide polymorphism (SNP) rs1546124, rs4783099, and rs16974880 were analyzed for their association using MassARRAY method. A whole-genome association analysis toolset, PLINK was used for statistical analysis. The polymorphisms followed Hardy–Weinberg equilibrium. None of the polymorphisms showed any significance. Hence the high-risk polymorphisms rs1546124, rs4783099, and rs16974880 are not associated with nonsyndromic CLP in Indian population.

Methylenetetrahydrofolate reductase C677T polymorphism is not associated with the risk of nonsyndromic cleft lip/palate: An updated meta-analysis

Both genetic and environmental factors affect the risk of orofacial clefts. The present meta-analysis aimed to evaluate the association between methylenetetrahydrofolate reductase (MTHFR) C677T polymorphism and risk of nonsyndromic cleft lip/palate (NSCL/P) in cases-control studies. The PubMed/Medline, Scopus, Web of Science, and Cochrane Library databases were searched up to April 2019 with no restrictions. The odds ratios (ORs) and 95% confidence intervals (CIs) in all analyses were calculated by Review Manager 5.3 software. The funnel plot analysis was carried out by the Comprehensive Meta-Analysis version 2.0 software. Subgroup analysis, meta-regression, and sensitivity analysis were performed for the pooled analyses. Thirty-one studies reviewed in this meta-analysis included 4710 NSCL/P patients and 7271 controls. There was no significant association between MTHFR C677T polymorphism and NSCL/P susceptibility related to allelic model (OR = 1.04; P = 0.49), homozygote model (OR = 1.11; P = 0.35), heterozygote model (OR = 0.99; P = 0.91), dominant model (OR = 1.00; P = 0.96), or recessive model (OR = 1.08; P = 0.23). There was no significant association between MTHFR C677T polymorphism and NSCL/P susceptibility based on the ethnicity or the source of cases. There was a significant linear relationship between the year of publication and log ORs for the allele model. The results of the present meta-analysis failed to show an association between MTHFR C677T polymorphism and NSCL/P susceptibility. The subgroup analyses based on the ethnicity and the source of cases further confirmed this result.

Association between cystathionine beta-synthase c.844ins68 polymorphism and risk of non-syndromic cleft lip/palate: A meta-analysis of family-based and case-control studies

Both genetics and environmental factors play a role in the occurrence of non-syndromic cleft lip/palate (NSCL/P). This meta-analysis evaluated the association between cystathionine beta-synthase (CBS) c.844ins68 polymorphism and risk of NSCL/P in family-based and case-control studies. The PubMed, Web of Science, Scopus, and Cochrane Library databases were searched for articles published until September 2018. RevMan 5.3 software was used to calculate the odds ratio (OR) for the association between CBS c.844ins68 polymorphism and risk of NSCL/P by using five genetic models in the studies. The transmission disequilibrium test (TDT) was conducted for family-based studies. Three case-control and three family-based studies were evaluated. Based on the analysis of five genetic models, risk of NSCL/P was not related to CBS c.844ins68 polymorphism in case-control studies. The results of family-based studies did not show any association between the CBS c.844ins68 allele and NSCL/P either. The results of this meta-analysis showed that there was no association between CBS c.844ins68 polymorphism and risk of NSCL/P; therefore, this polymorphism does not play a role in susceptibility to NSCL/P.

Association of Betaine-Homocysteine S-Methyl Transferase (rs3797546 and rs3733890) polymorphisms with non-syndromic cleft lip/palate: A meta-analysis

Non-syndromic cleft lip/palate (NSCL/P) has a multifactorial and polygenic aetiology. The role of genetics in its occurrence has not been fully clarified. The present meta-analysis aimed to evaluate the association of betaine-homocysteine S-methyltransferase (BHMT) polymorphisms (rs3797546 and rs3733890) with the risk of NSCL/P. PubMed/Medline, Scopus, Cochrane Library, and Web of Science databases were systematically searched for articles published up until December 2018 with no language restriction. Quality evaluation of each study was performed by the Newcastle-Ottawa Scale (NOS). The crude odds ratio (OR) and 95% confidence interval (CI) were calculated for each study by RevMan 5.3 software, and a funnel plot analysis was performed by the CMA 2.0 software using the Egger's and Begg's tests. Review of the four selected studies revealed that the CC genotype of rs3797546 polymorphism significantly increased the risk of NSCL/P. No association was noted between NSCL/P risk and rs3733890 polymorphism except in Chinese (elevated risk of NSCL/P) and Polish (decreased risk of NSCL/P) populations. According to the present meta-analysis, rs3733890 polymorphism does not play a role in susceptibility to NSCL/P; whereas, rs3797546 polymorphism may play a role in susceptibility to NSCL/P. Future studies are required to examine the association between BHMT polymorphisms and the NSCL/P risk in different ethnicities with a larger sample size.

Polymorphic Variants of V-Maf Musculoaponeurotic Fibrosarcoma Oncogene Homolog B (rs13041247 and rs11696257) and Risk of Non-Syndromic Cleft Lip/Palate: Systematic Review and Meta-Analysis

Background: Non-syndromic cleft lip/palate (NSCL/P) has an etiology, including both genetic and environmental factors. Herein, we evaluated the association of rs13041247 and rs11696257 v-maf musculoaponeurotic fibrosarcoma oncogene homolog B (MAFB) polymorphisms with the risk of NSCL/P in a meta-analysis. Methods: The PubMed/Medline, Scopus, Cochrane Library, Web of Science, and HuGE Navigator databases were systematically searched to retrieve relevant articles published up to January 2019. The Newcastle–Ottawa scale was applied for quality evaluation of retrieved articles. The 95% confidence interval (CI) and crude odds ratio (OR) were calculated for each study using the Review Manager 5.3 software to show the association between MAFB polymorphisms and risk of NSCL/P. The comprehensive meta-analysis 2.0 software was used to calculate the publication bias. In addition, sensitivity analysis was carried out to show the stability of results. Results: Of 102 articles retrieved from the databases, 10 articles were analyzed in this meta-analysis. Ten articles, including eleven studies reporting rs13041247 MAFB polymorphism, included 3082 NSCL/P patients and 4104 controls. Three studies that reported rs11696257 MAFB polymorphism involved 845 NSCL/P patients and 927 controls. The rs11696257 MAFB polymorphism was not associated with the risk of NSCL/P, but the CC and TC genotypes of rs13041247 polymorphism were associated with the risk of NSCL/P. Nevertheless, the C allele and CC and TC genotypes were associated with a significant decline in the risk of NSCL/P in population-based studies. Conclusions: The results of this meta-analysis demonstrated that the risk of NSCL/P was related to rs13041247 polymorphism, not rs11696257 MAFB polymorphism. Well-designed studies are required to assess the interaction of MAFB and other genes with environmental factors in different ethnic groups.

A systematic review and meta-analysis on protective role of forkhead box E1 (FOXE1) polymorphisms in susceptibility to non-syndromic cleft lip/palate

Several environmental and genetic factors have a role in the aetiology of non-syndromic cleft lip/palate (NSCL/P). This meta-analysis evaluated the association of rs3758249 and rs4460498 forkhead box E1 (FOXE1) polymorphisms with the NSCL/P risk. The Scopus, Cochrane Library, Web of Science, and PubMed databases were searched for articles published until March 2019. The analyses were performed by Review Manager 5.3 using the crude odds ratio (OR) and 95% confidence interval (CI) for a strong association between FOXE1 polymorphisms and the risk of NSCL/P. Out of 161 articles retrieved from the databases, four case-control articles were involved in the meta-analysis. The pooled ORs of rs4460498 polymorphism based on allelic, homozygous, heterozygous, dominant, and recessive models were 0.74 (95% CI: 0.69, 0.80; P<0.00001), 0.43 (95% CI: 0.30, 0.61; P<0.00001), 0.66 (95% CI: 0.55, 0.80; P<0.0001), 0.66 (95% CI: 0.59, 0.73; P<0.00001), and 0.70 (95% CI: 0.60, 0.82; P<0.0001), respectively; whereas, the pooled OR of rs3758249 polymorphism were 0.86 (95% CI: 0.71, 1.04; P=0.12), 0.68 (95% CI: 0.57, 0.82; P<0.0001), 0.79 (95% CI: 0.57, 1.09; P=0.15), 0.79 (95% CI: 0.58, 1.08; P=0.14), and 0.80 (95% CI: 0.68, 0.95; P=0.010) for the afore-mentioned models, respectively. The results showed that the T allele, TT, and CT genotypes of rs4460498 polymorphism were significantly associated with a decreased risk of NSCL/P; whereas, for rs3758249 polymorphism, only the AA genotype had a significant protective role in NSCL/P. Thus, FOXE1 is strongly associated with NSCL/P in the populations.

Study on DNA methylation profiles in non-syndromic cleft lip/palate based on bioinformatics

To study DNA methylation patterns of non-syndromic cleft lip/palate(NSCL/P) using bioinformatic methods, including methylated positions and regions. Whole blood DNA methylation data of NSCL/P samples was download from Gene Expression Omnibus(GEO) database, including 67 NSCL/P cases and 59 controls without birth defects. Data analysis included ①data cleaning, such as probes filtering, quality control and normalization; ②differential methylation analysis, including methylated positions and regions; ③Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) analysis on differential methylated genes. R 3.4.3 software was used for data cleaning, differential methylated positions and regions analysis. DAVID6.8 tool was used for GO and KEGG analysis. 814 differential methylated positions were detected (adjusted P＜0.001，|Δβ|＞0.125）, of which 178 were hypermethylated in NSCL/P patients, and 636 were hypomethylated. In addition, 386 differential methylated regions were identified (P＜0.05), of which 204 were hypermethylation regions and 182 were hypomethylation regions. GO analysis showed that 38 differential methylated genes were involved in 7 kinds of biological processes, 163 differential methylated genes were involved in 3 kinds of molecular functions, and 114 differential methylated genes were involved in 3 kinds of cellular components (P＜0.01). KEGG analysis showed that 59 differential methylated genes were involved in 9 kinds of signal pathways. Abnormal DNA methylation patterns of NSCL/P might be an important epigenetic mechanism affecting the development of NSCL/P. This study might contribute to the identification of identification of biomarkers and targeted interventions of NSCL/P.

Association Studies Between Regulatory Regions of IRF6/TP63 Genes and Nonsyndromic Oral Clefts.

To evaluate genetic variants within the regulatory regions of interferon regulatory factor 6 (IRF6) and TP63 for the etiology of nonsyndromic oral clefts risk factors. We performed allelic transmission disequilibrium test analysis on 5 eligible single-nucleotide polymorphisms (SNPs) and SNP haplotypes using the Family-Based Association Test. The study sample consisted of 334 case-parent trios of nonsyndromic oral clefts from Taiwanese population, separated into nonsyndromic cleft lip/palate (NSCL/P) and nonsyndromic cleft palate only (NSCPO) groups. We found all 3 selected SNPs of the IRF6 gene show significant association with nonsyndromic oral clefts (rs2235371, P = 5.10E-07; rs642961, P = .00194; and rs77542756, P = 9.08E-07). Haplotype analyses identified 3 possible SNP combination haplotypes in the IRF6 gene and found that C-G-G showed significant undertransmission (P = .058), whereas 2 other haplotypes, T-G-A and C-A-G (P = 2.71E-06 and P = 5.00E-04, respectively), were significantly overtransmitted to the NSCL/P children but not to the NSCPO children. For the TP63 gene, we failed to detect evidence of nonsyndromic oral cleft association in the 2 SNPs within the TP63 large intron 1 region. We used a family-based analysis in 334 Taiwanese case-parent trios to evaluate selected SNPs of IRF6 genes and TP63 genes for a risk of orofacial clefting. This study provides additional evidence for an association between IRF6 and NSCL/P, including the genetic variants within the 5'-noncoding region of the gene. We also confirmed that NSCL/P and NSCPO individuals belong to different groups. For the TP63, our data did not favor the direct involvement of TAp63 isoforms during orofacial development.

Novel TBX22 mutations in Chinese nonsyndromic cleft lip/palate families

TBX22 is a gene which contribute to cleft lip/palate, and many mutation sites of TBX22 have been reported. However, the exact role of TBX22 mutation in Chinese nonsyndromic cleft lip/palate (NSCL/P) family was not clearly explored. In this study, we tried to investigate the profiles and effects of TBX22 mutation in Chinese NSCL/P family. Members of two Chinese NSCL/P families and 200 normal controls were enrolled in this study. Further, DNA sequence and bioinformatic analysis for TBX22 were performed. The results showed that a novel and essential splicing site mutation, IVS6-1G>C , was detected in a family with cleft palate. The bioinformatic analysis results showed that this mutation would lead to abnormal transcription or translation, followed by a loss of function of TBX22. In addition, a hemizygous missense mutation, c.874G>A (p.D292N), was first reported in another Chinese family, which may exhibit aggravated effects on the phenotypes of CL/P. Taking these findings together, this study provides a profile of TBX22 mutation in Chinese NSCL/P families, and further confirmed the important role of TBX22 in familial cases with X-linked cleft palate.

Updating Genetics Polymorphisms of Non-Syndromic Clefts Lip-Palates

Introduction: Non-Syndromic Clefts Lip-Palates (NSCLP/CP) are most common congenital malformation in the world, with very important psychic and social impact. Formation of NSCLP/CP arises from the interaction of environmental and genetic factors. This paper provides a review of recent progress in defining the genetic causes of NSCLP. Methods: A literature review was conducted on the Medline data by searching for the following keywords: genes, non-syndromic cleft lip-palate, and genetics of clefts lip-palates, until January 2018. Results: Various genes are identified in different population and country, with the study using case parent’s trio. The aim of this study contributes to review relative gene which has been identify in non-syndromic cleft lip and palate, and to help to have a better understanding of the inheritance pattern of this pathology and the prevention of genetic disease. Conclusion: Although three major genes have been confirmed, the genetic research is necessary to provide an understanding of the pathophysiology of the clefts lip-palates.

Disrupted IRF6-NME1/2 Complexes as a Cause of Cleft Lip/Palate

Mutations and common polymorphisms in interferon regulatory factor 6 (IRF6) are associated with both syndromic and nonsyndromic forms of cleft lip/palate (CLP). To date, much of the focus on this transcription factor has been on identifying its direct targets and the gene regulatory network in which it operates. Notably, however, IRF6 is found predominantly in the cytoplasm, with its import into the nucleus tightly regulated like other members of the IRF family. To provide further insight into the role of IRF6 in the pathogenesis of CLP, we sought to identify direct IRF6 protein interactors using a combination of yeast 2-hybrid screens and co-immunoprecipitation assays. Using this approach, we identified NME1 and NME2, well-known regulators of Rho-type GTPases, E-cadherin endocytosis, and epithelial junctional remodeling, as bona fide IRF6 partner proteins. The NME proteins co-localize with IRF6 in the cytoplasm of primary palatal epithelial cells in vivo, and their interaction with IRF6 is significantly enhanced by phosphorylation of key serine residues in the IRF6 C-terminus. Furthermore, CLP associated IRF6 missense mutations disrupt the ability of IRF6 to bind the NME proteins and result in elevated activation of Rac1 and RhoA, compared to wild-type IRF6, when ectopically expressed in 293T epithelial cells. Significantly, we also report the identification of 2 unique missense mutations in the NME proteins in patients with CLP (NME1 R18Q in an IRF6 and GRHL3 mutation-negative patient with van der Woude syndrome and NME2 G71V in a patient with nonsyndromic CLP). Both variants disrupted the ability of the respective proteins to interact with IRF6. The data presented suggest an important role for cytoplasmic IRF6 in regulating the availability or localization of the NME1/2 complex and thus the dynamic behavior of epithelia during lip/palate development.