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Treacher Collins syndrome is an autosomal dominant craniofacial malformation mainly caused by mutations in the TCOF1 gene. Few cases have been observed in the Chinese population. Herein, the authors report the mutational analysis of TCOF1, GSC, and HOXA2 to determine the mutational features of the 3 genes in Chinese patients with Treacher Collins syndrome. Genomic DNA of the patients and their parents was extracted from peripheral blood following a standard protocol. DNA sequencing analysis was performed on all exons and the exon-intron borders of TCOF1, GSC, and HOXA2 in addition to the 1200-bp upstream of TCOF1. Four novel single nucleotide polymorphisms were detected in TCOF1, one of which was in the promoter region. Mutations in GSC and HOXA2 were not found in the 3 patients. Our results suggest the possibility of genetic heterogeneity or different mechanisms leading to the disease. Further functional study of the alteration is necessary to obtain more definitive information.

Microtia is a complicated congenital anomaly with a genetic and environmental predisposition, and the molecular events underlying this disease are not fully understood. MicroRNAs (miRNAs) are a class of 20-22 nucleotide non-coding RNAs that function to control post-transcriptional gene expression. We want to find the miRNA expression profiling of microtia by using Affymetrix GeneChip(®) miRNA 2.0 Arrays.We selected 9 microtia cartilages and 3 normal controls for GeneChip(®) miRNA 2.0 Arrays analysis. The altered miRNA were analyzed by poly (A) RT-PCR from 58 microtia samples and 16 normal controls. We predicted the target genes of miRNAs by bioinformatics and RT-PCT was used to confirm the target genes.We found 11 miRNAs with significantly altered expression in the microtic group compared to the normal controls, which included 6 up-regulated miRNAs and 5 down-regulated miRNAs. These miRNAs were further examined using poly (A) RT-PCR analysis, we found that miR-451 and miR-486-5p were significantly up-regulated and miR-200c was significantly down-regulated in the microtic group compared to the normal controls (p0.05). Several complementary target messenger RNAs (mRNAs) had been predicted. OSR1, the target gene of miR-451 and miR-200c, was significantly up-regulated (p0.01); TRPS1, the target gene of miR-200c, was significantly down-regulated in the microtic group compared to the controls (p0.0001).The reduction in miR-200c expression and the accretion of miR-451 and miR-486-5p expression in microtic samples could be possible causes of the abnormal development of the external ear. OSR1 and TRPS1, as the complementary target mRNAs, may play important roles during the development of the external ear. Further studies are still needed to identify the miRNA target genes and to determine their function in the pathogenesis of microtia. This is the first report of a relationship between miRNAs and microtia.

Objective Microtia is defined as a developmental malformation characterized by a small, abnormal shaped auricle, with atresia or stenosis of the auditory canal. Genes responsible for nonsyndromic microtia have remained elusive. We therefore report a mutational analysis of GSC, HOXA2 and PRKRA in 106 congenital microtia patients without any combined malformation to explore the relationship between GSC, HOXA2, PRKRA and nonsyndromic microtia. Methods A total of 106 patients with a clinical diagnosis of congenital microtia and a control group (100 unaffected controls) were recruited through the Eye and ENT Hospital of Fudan University in China. Genomic DNA was extracted following a standard protocol. DNA sequencing analysis was performed in all exons and the exon-intron borders of GSC, HOXA2 and PRKRA. Results We identified 5 genomic variants in GSC, HOXA2 and PRKRA. As to the GSC, we obtained a reported variant g.994C > T in exon 2, which resulted in no change of protein. Our results revealed that g.994C > T was also detected in 10 control cases. We also detected 2 novel variants, g.90G > A and g.114A > C, in the 5′UTR of HOXA2. No class 5 or 4 genomic variant of PRKRA was identified in our microtia patients. Additionally, two previously reported SNVs in GSC and PRKRA were also presented. Conclusions We suggest that g.994C > T is a new SNV, which is different from the previous report. Further study is needed to prove the function of 2 novel variants in the 5′UTR of HOXA2, and to explore the possible mechanism of these variants in the occurrence of microtia.

Velocardiofacial syndrome (VCFS) is a disease in human with an expansive phenotypic spectrum and diverse genetic mechanisms mainly associated with copy number variations (CNVs) on 22q11.2 or other chromosomes. However, the correlations between CNVs and phenotypes remain ambiguous. This study aims to analyze the types and sizes of CNVs in VCFS patients, to define whether correlations exist between CNVs and clinical manifestations in Chinese VCFS patients. In total, 55 clinically suspected Chinese VCFS patients and 100 normal controls were detected by multiplex ligation-dependent probe amplification (MLPA). The data from MLPA and all the detailed clinical features of the objects were documented and analyzed. A total of 44 patients (80.0%) were diagnosed with CNVs on 22q11.2. Among them, 43 (78.2%) presented with 22q11.2 heterozygous deletions, of whom 40 (93.0%) had typical 3-Mb deletion, and 3 (7.0%) exhibited proximal 1.5-Mb deletion; no patient was found with atypical deletion on 22q11.2. One patient (1.8%) presented with a 3-Mb duplication mapping to the typical 3-Mb region on 22q11.2, while none of the chromosomal abnormalities in the MLPA kit were found in the other 11 patients and 100 normal controls. All the 43 patients with 22q11.2 deletions displayed characteristic face and palatal anomalies; 37 of them (86.0%) had cognitive or behavioral disorders, and 23 (53.5%) suffered from immune deficiencies; 10 patients (23.3%) manifested congenital heart diseases. Interestingly, all patients with the characteristic face had 22q11.2 heterozygous deletions, but no difference in phenotypic spectrum was observed between 3-Mb and 1.5-Mb deletions. Our data suggest that the characteristic face can be used as a key indicator for direct diagnosis of 22q11.2 deletions in Chinese VCFS patients.

Background miRNAs are a group of small RNA molecules regulating target genes by inducing mRNA degradation or translational repression. Aberrant expression of miRNAs correlates with various cancers. Although miR-135a has been implicated in several other cancers, its role in breast cancer is unknown. HOXA10 however, is associated with multiple cancer types and was recently shown to induce p53 expression in breast cancer cells and reduce their invasive ability. Because HOXA10 is a confirmed miR-135a target in more than one tissue, we examined miR-135a levels in relation to breast cancer phenotypes to determine if miR-135a plays role in this cancer type. Methods Expression levels of miR-135a in tissues and cells were determined by poly (A)-RT PCR. The effect of miR-135a on proliferation was evaluated by CCK8 assay, cell migration and invasion were evaluated by transwell migration and invasion assays, and target protein expression was determined by western blotting. GFP and luciferase reporter plasmids were constructed to confirm the action of miR-135a on downstream target genes including HOXA10. Results are reported as means ± S.D. and differences were tested for significance using 2-sided Student"s t-test. Results Here we report that miR-135a was highly expressed in metastatic breast tumors. We found that the expression of miR-135a was required for the migration and invasion of breast cancer cells, but not their proliferation. HOXA10, which encodes a transcription factor required for embryonic development and is a metastasis suppressor in breast cancer, was shown to be a direct target of miR-135a in breast cancer cells. Our analysis showed that miR-135a suppressed the expression of HOXA10 both at the mRNA and protein level, and its ability to promote cellular migration and invasion was partially reversed by overexpression of HOXA10. Conclusions In summary, our results indicate that miR-135a is an onco-miRNA that can promote breast cancer cell migration and invasion. HOXA10 is a target gene for miR-135a in breast cancer cells and overexpression of HOXA10 can partially reverse the miR-135a invasive phenotype.

Background The purpose of this study is to evaluate the prognostic value of TFPI-2 expression in breast cancer patients through examining the correlation between TFPI-2 expression and breast cancer clinicopathologic features. Methods Immunohistochemical staining combined with digital image analysis was used to quantify the expression of TFPI-2 protein in breast tumor tissues. For evaluation of the prognostic value of TFPI-2 expression to each clinicopathologic factor, Kaplan-Meier method and COX’s Proportional Hazard Model were employed. Results TFPI-2 expression was significantly correlated with tumor size, lymph node metastasis, histologic grade, clinical stage, and vessel invasion. More importantly, TFPI-2 expression was also associated with disease-free survival (DFS) of breast cancer patients. We found that patients with high TFPI-2 expression had longer DFS compared with those with low or negative expression of TFPI-2 (P P Conclusions Low or negative expression of TFPI-2 is associated with breast cancer progression, recurrence and poor survival outcome after breast cancer surgery. TFPI-2 expression in breast tumors is a potential prognostic tool for breast cancer patients.