Piotr Wójcicki

Associations of folate and choline metabolism gene polymorphisms with orofacial clefts

Background Non-syndromic isolated cleft lip with or without cleft palate (NCL/P) is a common congenital anomaly in humans, the aetiology of which is complex and associated with both genetic and environmental factors. It has been reported that maternal nutritional factors are likely to play a major role in development of NCL/P in the embryo. Objective As the mechanism by which folic acid and choline supplementation prevents NCL/P is poorly understood, the relationship between 16 polymorphic variants of 12 genes encoding enzymes involved in the metabolism of these two nutrients and the risk of facial clefts was investigated. Results It was found that individuals with the AA genotype of the BHMT rs3733890 polymorphism have a significantly lower risk of orofacial clefts (OR 0.1450, 95% CI 0.0420 to 0.4995; p=0.0005; pcorr=0.008). It was also demonstrated that the rs7639752 polymorphism of the PCYT1A gene increases the risk of NCL/P nearly twofold in the Polish population (OR 1.891, 95% CI 1.151 to 3.107; p=0.011), but this association would not withstand correction for multiple testing (pcorr=0.176). The genetic variations in CBS, MTHFD1, MTHFR, MTR, MTRR, TCN2, BHMT2, CHDH, CHKA, and PEMT were not separately correlated with NCL/P risk. However, the Multifactor Dimensionality Reduction (MDR) analysis showed a significant epistatic interaction between MTHFR (rs1801133), MTR (rs1805087), and PEMT (rs4646406) in NCL/P susceptibility. Conclusion This study demonstrates that choline metabolism may play an important role in the aetiology of NCL/P. Polymorphic variants of BHMT and PCYT1A and interactions between genes of choline and folate metabolism might influence the risk of NCL/P in the Polish population.

Association between genetic variants of reported candidate genes or regions and risk of cleft lip with or without cleft palate in the polish population

BACKGROUND Cleft lip with or without cleft palate (CL/P) is one of the most common craniofacial malformations, with a complex and multifactorial etiology. Because of the genetic heterogeneity of facial clefts, the aim of this study was to investigate the contribution of previously reported candidate genes and chromosomal loci to the risk of CL/P in the Polish population. METHODS We performed an analysis of 18 polymorphisms of FOXE1, IRF6, MSX1, PAX9, TBX10, FGF10, FGFR1, TGFalpha, TGFbeta3, SUMO1, and the chromosomal region 8q24 in a group of 175 patients with CL/P and a properly matched control group. RESULTS Highly significant results were observed for the IRF6 rs642961 variant and the 8q24 regions rs987525 (odds ratio [OR](AG+AAvsGG), 1.635; 95% confidence interval [CI], 1.153-2.319; p = 0.005; and OR(AC+AAvsCC), 1.962; 95% CI, 1.382-2.785; p = 1.4 x 10(-4), respectively). For rs987525, the results were also significant after correction for multiple comparisons. Borderline association with an increased risk of CL/P was also identified for the SUMO1 locus (rs2350350; OR(CGvsGG), 1.580; 95% CI, 1.056-2.363; p = 0.025). CONCLUSIONS Our findings confirmed that genetic variants of IRF6 and the polymorphism located in the 8q24 gene desert are strongly involved in the etiology of facial clefts in the Polish population sample.

Genotype and haplotype analysis of WNT genes in non-syndromic cleft lip with or without cleft palate

The wingless-type MMTV integration site family (Wnt) signalling pathway plays a crucial role in craniofacial development. Recently, nucleotide variants in WNT genes have been shown to be associated with oral congenital anomalies, including facial clefts. Therefore, in the current study we decided to assay the association of nucleotide variants in selected WNT genes with the risk of non-syndromic cleft lip with or without cleft palate (NCL/P) in the Polish population. Fourteen polymorphisms in WNT3, WNT3A, WNT5A, WNT8A, WNT9B, and WNT11 were tested in a group of 210 patients with NCL/P and in a properly matched control group. The most significant results were found for the WNT3 rs3809857 variant, which, under the assumption of a recessive model, was associated with a two-fold decrease in the risk of NCL/P (OR(TT vs. GT + GG) = 0.492, 95% CI: 0.276-0.879, P = 0.015). Moreover, haplotype analysis revealed that WNT3 is significantly correlated with NCL/P. The global P-values for haplotypes of rs12452064_rs7207916 and rs3809857_rs12452064_rs7207916 were 0.0034 and 0.0014, respectively, and these results were statistically significant, even after the permutation test correction. In conclusion, our study confirmed the involvement of polymorphisms in the WNT3 gene in NCL/P aetiology in the tested population.

Surgical treatment of Treacher Collins syndrome.

Morphology, genetic conditioning, terminology, and the principles of treatment of Treacher Collins syndrome have been presented on the basis of our own material, as well as review of literature. Fifty patients (27 males, 23 females) were operated on at the Hospital of Plastic Surgery in Polanica Zdrój from 1976 to 2005. The patients were first seen when they were from 1 to 32 years of age (mean, 7). The disease was hereditary in 17 patients, while the remaining subjects did not reveal any genetic conditioning. The lifesaving surgical treatment was undertaken in 4 children with the most severe form of the disease who presented with dyspnea and paroxysmal apnea. In those cases, the treatment aimed at improving the respiratory function by restoring patency of the nostrils and distraction of the mandible. Twelve patients were operated on for cleft palate between 1 and 2 years of age. Twelve patients had eyelid correction with the use of musculocutaneous flap transposition from the upper eyelid (Z-plasty). The zygomatic bone and lateral wall of the orbit were reconstructed by means of iliac bone grafts in 26 patients. The auricular reconstruction was usually undertaken after 10 years of age. Nine patients underwent bilateral auricular reconstruction by means of a modified Brent method. Fifteen patients aged 12–14 had chin osteotomy according to the Obwegeser method. Nasal osteotomy was performed in 10 patients with characteristic broad, long, and hooked noses, who were operated on after 16 years of age and after completion of orthognathic treatment. In total, 258 surgical procedures were performed in 50 patients, an average of 5.2 operations per every patient. Apart from a multistage surgical treatment, the patients required a combined multidisciplinary approach, mainly due to hearing impairment and occlusal disturbances. The obtained outcome of treatment, although far from being perfect, but still beneficial, confirmed the correctness of applied approach but at the same time pointed to the necessity of introducing new methods of treatment aiming, among others, at compensating for the tissue deficiency by means of tissue engineering.

A mutation in mouse Pak1ip1 causes orofacial clefting while human PAK1IP1 maps to 6p24 translocation breaking points associated with orofacial clefting.

Orofacial clefts are among the most common birth defects and result in an improper formation of the mouth or the roof of the mouth. Monosomy of the distal aspect of human chromosome 6p has been recognized as causative in congenital malformations affecting the brain and cranial skeleton including orofacial clefts. Among the genes located in this region is PAK1IP1, which encodes a nucleolar factor involved in ribosomal stress response. Here, we report the identification of a novel mouse line that carries a point mutation in the Pak1ip1 gene. Homozygous mutants show severe developmental defects of the brain and craniofacial skeleton, including a median orofacial cleft. We recovered this line of mice in a forward genetic screen and named the allele manta-ray (mray). Our findings prompted us to examine human cases of orofacial clefting for mutations in the PAK1IP1 gene or association with the locus. No deleterious variants in the PAK1IP1 gene coding region were recognized, however, we identified a borderline association effect for SNP rs494723 suggesting a possible role for the PAK1IP1 gene in human orofacial clefting.

Novel mutation in the 5' splice site of exon 4 of the TCOF1 gene in the patient with treacher Collins syndrome

Treacher Collins syndrome (TCS) is caused by mutations in the TCOF1 gene. This gene encodes a serine/alanine‐rich protein called treacle. The structure of the entire TCOF1 gene was investigated in a patient with TCS. We detected a novel deletion (376delAAGGTGAGTGGGACTGCC) spanning 3 bp of exon 4 and 15 bp of the adjacent intronic sequence. This mutation causes premature termination of translation, resulting in a truncated protein devoid of nucleolar localization signal, and potential phosphorylation sites. Real‐time PCR analysis showed different melting temperatures of the amplified fragment containing normal allele and that harboring the 18 bp deletion, thus providing a rapid screening assay for this and other deletions of the TCOF1 gene.

Bone graft healing in alveolar osteoplasty in patients with unilateral lip, alveolar process, and palate clefts.

Abstract Secondary osteoplasty by means of autogenic spongy bone grafting is the most common procedure used in the reconstruction of the continuity of the maxillary alveolar process. The aim of the study was to analyze retrospectively the effect of certain factors on the course of the bone graft healing process in patients with unilateral complete clefts of the lip, alveolar process, and palate. The investigations involved 62 children aged 8 to 14 years (mean age, 11 years) with unilateral complete cleft of the lip, alveolar process, and palate operated on at the Clinic of Plastic Surgery in Polanica Zdrój from November 2007 to April 2009. All the procedures consisted in the reconstruction of the maxillary alveolar process by means of autogenic spongy bone grafting from the iliac bone. The analysis was performed on the basis of computed tomography scans presenting maxillary alveolar processes in the horizontal cross-sectional planes performed on the second or third postoperative day and after 6 months. They were used as the basis for the measurement of the volume and density (condensation) of the bone graft, the surface of its adhesion to the maxillary alveolar bone, and the volume and density of the healed bone. The following correlation coefficients were determined: between the adhesion surface of the bone to the alveolar bone and the volume of the healed bone, between the adhesion surface of the bone to the alveolar bone and the density of the healed bone, and between the density of the graft and the volume of the healed bone. Increasing the surface of the graft adhesion to the bone ridges of the alveolar cleft contributes to increased volume of the healed bone and slows down the increase in its density (on 6-month follow-up). Crushing of the bone graft increases its resorption and reduces volume of the healed bone.

Association of DVL2 and AXIN2 gene polymorphisms with cleft lip with or without cleft palate in a Polish population.

BACKGROUND Nonsyndromic cleft lip with or without cleft palate (NSCL/P) is one of the most common congenital anomalies, with a complex and still not fully understood etiology. Given the important role of the Wnt/β-catenin pathway during craniofacial development, we decided to test the hypothesis that common polymorphic variants of the genes encoding crucial components of this signaling pathway might contribute to the risk of NSCL/P in the Polish population. METHODS A set of 19 single nucleotide polymorphisms (SNPs) in the APC, AXIN1, AXIN2, CTNNB1, DVL2, and GSK-3β genes were analyzed using restriction fragment length polymorphism and high-resolution melting curve methods in a group of 280 patients with NSCL/P and a properly matched control group (n = 330). RESULTS Both single-marker and haplotype analyses showed an association between SNPs in the DVL2 gene and the risk for NSCL/P. The strongest association was found under an overdominant model for the rs35594616 variant located in the exonic sequence of DVL2 (odds ratio [OR], 1.90; 95% confidence interval [CI], 1.37-2.62; p < 0.0001). Moreover, the gene-gene interaction analysis revealed a significant epistatic interaction between DVL2 gene SNPs in the susceptibility to orofacial clefts. Borderline association with a decreased risk of NSCL/P was also observed for the AXIN2 rs3923087 variant (dominant model OR, 0.69; 95% CI, 0.50-0.95; p = 0.03). CONCLUSION This study suggests that polymorphic variants of the Wnt/β-catenin pathway genes have a role in the susceptibility to orofacial clefts. The DVL2 and AXIN2 genes might be candidate genes for this craniofacial anomaly in the Polish population. Birth Defects Research (Part A), 2012.

In utero surgery – current state of the art – part II

Summary Background Fetal surgery, also referred to as in utero, prenatal or intrauterine surgery, consists of treatment of congenital malformations during the fetal period. The idea of treating malformations diagnosed in the course of intrauterine life dates back to 1963, when Lilly performed the first blood transfusion in a fetus. Since then it has been introduced as a treatment option in a series of lethal malformations. Efforts are being made to treat nonlethal malformations by means of fetal surgery. Material/Methods A comprehensive search of the literature using MEDLINE and PubMed between 1925 and February 2009 was performed. Search terms for MEDLINE and PubMed were: fetal surgery, foetal surgery, in utero surgery, prenatal surgery, and in utero treatment. In addition, information was obtained at Web sites of the International Medicine and Surgery Society and the University of California Fetal Treatment Centre. Results Authors’ attention focused on the survey of indications to intrauterine operations. We outline potential directions of its development, quoting the groundwork of the most experienced researchers and clinicians. Moreover, owing to the authors’ interest in plastic surgery, some remarks on the role of intrauterine medicine and surgery in this branch of medicine are made. Conclusions In utero surgery may be regarded as an efficient tool of preventive medicine. It offers some advantages that no other branch of medicine may offer. However, its implementation is more troublesome than in the past, therefore only selected cases may benefit from its advantages. Nevertheless, current tendencies are to include, after thorough evaluation of benefit-risk ratio, some new indications for fetal surgery.

Surgical treatment of patients with Parry-Romberg syndrome.

Introduction:Parry–Romberg syndrome is an extremely rare disease characterized by the slow atrophy of the face involving the skin, subcutaneous tissue, fatty tissue, muscles, and even bones. The cause of the disease is unknown. The onset is slow and it begins usually during the first 2 decades of life, more often between the ages of 5 and 15. The purpose of the study was to present the principles of management and treatment outcomes in patients with Parry–Romberg syndrome. Material and Method:A total of 14 patients (12 females and 2 males) aged between 14 and 36 years (mean, 21.5 years), were treated at the Hospital of Plastic Surgery in Polanica Zdrój between 1970 and 2005. Hemifacial atrophy affected the left side in 7 patients, and the right side in the remaining 7. Altogether 45 operations were performed. Less severe changes were reconstructed by means of dermal grafts, galeal flaps; patients with more severe facial deformities were treated with free tissue transfer: parascapular and latissimus dorsi flaps (6 cases). The treatment was completed with facial modeling, dermal regrafting, and scars correction. Results:The appearance was improved in all the patients. There was only 1 free flap loss reported during postoperative management. Conclusions:Dermal grafts are used in the treatment of milder forms of Romberg disease to improve facial symmetry. In severe cases of facial atrophy, the surgical management is based on free-tissue transfer.