Reduced heparan sulfate accumulation in enterocytes contributes to protein-losing enteropathy in a congenital disorder of glycosylation

Intestinal biopsy in a boy with gastroenteritis-induced protein-losing enteropathy (PLE) showed loss of heparan sulfate (HS) and syndecan-1 core protein from the basolateral surface of the enterocytes, which improved after PLE subsided. Isoelectric focusing analysis of serum transferrin indicated a congenital disorder of glycosylation (CDG) and subsequent analysis showed three point mutations in the ALG6 gene encoding an alpha1,3-glucosyltransferase needed for the addition of the first glucose to the dolichol-linked oligosaccharide. The maternal mutation, C998T, causing an A333V substitution, has been shown to cause CDG-Ic, whereas the two paternal mutations, T391C (Y131H) and C924A (S308R) have not previously been reported. The mutations were tested for their ability to rescue faulty N:-linked glycosylation of carboxypeptidase Y in an ALG6-deficient Saccharomyces cerevisiae strain. Normal human ALG6 rescues glycosylation and A333V partially rescues, whereas the combined paternal mutations (Y131H and S308R) are ineffective. Underglycosylation resulting from each of these mutations is much more severe in rapidly dividing yeast. Similarly, incomplete protein glycosylation in the patient is most severe in rapidly dividing enterocytes during gastroenteritis-induced stress. Incomplete N:-linked glycosylation of an HS core protein and/or other biosynthetic enzymes may explain the selective localized loss of HS and PLE.     

Novel variants and clinical symptoms in four new ALG3‐CDG patients,review of the literature,and identification of AAGRP‐ALG3 as a novel ALG3 variant with alanine and glycine‐rich N‐terminus

ALG3‐CDG is one of the very rare types of congenital disorder of glycosylation (CDG) caused by variants in the ER‐mannosyltransferase ALG3. Here, we summarize the clinical, biochemical, and genetic data of four new ALG3‐CDG patients, who were identified by a type I pattern of serum transferrin and the accumulation of Man₅GlcNAc₂‐PP‐dolichol in LLO analysis. Additional clinical symptoms observed in our patients comprise sensorineural hearing loss, right‐descending aorta, obstructive cardiomyopathy, macroglossia, and muscular hypertonia. We add four new biochemically confirmed variants to the list of ALG3‐CDG inducing variants: c.350G>C (p.R117P), c.1263G>A (p.W421*), c.1037A>G (p.N346S), and the intron variant c.296+4A>G. Furthermore, in Patient 1 an additional open‐reading frame of 141 bp (AAGRP) in the coding region of ALG3 was identified. Additionally, we show that control cells synthesize, to a minor degree, a hybrid protein composed of the polypeptide AAGRP and ALG3 (AAGRP‐ALG3), while in Patient 1 expression of this hybrid protein is significantly increased due to the homozygous variant c.160_196del (g.165C>T). By reviewing the literature and combining our findings with previously published data, we further expand the knowledge of this rare glycosylation defect.     

CDG-Id caused by homozygosity for an ALG3 mutation due to segmental maternal isodisomy UPD3(q21.3-qter)

We report on a patient with a congenital disorder of glycosylation type Id (CDG-Id) caused by a homozygous mutation in the ALG3 gene, which results from a de novo mutation in combination with a segmental maternal uniparental isodisomy (UPD). The patient presented with severe psychomotor delay, primary microcephaly, and opticus atrophy, compatible with a severe form of CDG. Isoelectric focusing of transferrin showed a type I pattern and lipid-linked oligosaccharide analysis showed an accumulation of dol-PP-GlcNAc2Man5 in patient's fibroblasts suggesting a defect in the ALG3 gene. A homozygous ALG3 missense mutation p.R266C (c.796C > T) was identified. Further evaluation revealed that neither the mother nor the father were carrier of the p.R266C mutation. Marker analysis revealed a segmental maternal isodisomy for the chromosomal region 3q21.3-3qter. UPD for this region has not been described before. More important, the combination of UPD with a de novo mutation is an exceptional coincidence and an extraordinary observation.     

CDG-Id in two siblings with partially different phenotypes

We present two sibs with congenital disorder of glycosylation (CDG) type Id. Each shows severe global delay, failure to thrive, seizures, microcephaly, axial hypotonia, and disaccharidase deficiency. One sib has more severe digestive issues, while the other is more neurologically impaired. Each is compound heterozygous for a novel point mutation and an already known mutation in the ALG3 gene that leads to the synthesis of a severely truncated oligosaccharide precursor for N-glycans. The defect is corrected by introduction of a normal ALG3 cDNA. CDG should be ruled out in all patients with severe seizures and failure to thrive. (c) 2007 Wiley-Liss, Inc.     

Acid sphingomyelinase: identification of nine novel mutations among Italian Niemann Pick type B patients and characterization of in vivo functional in-frame start codon

Niemann Pick disease (NPD) is an autosomal recessive disorder due to the deficit of lysosomal acid sphingomyelinase, which results in intracellular accumulation of sphingomyelin. In the present work we studied 18 patients with NPD type B, including five individuals who presented an intermediate phenotype characterised by different levels of neurological involvement. We identified nine novel mutations in the SMPD1 gene including six single base changes c.2T>G, c.96G>A, c.308T>C, c.674T>C, c.732G>C, c.841G>A (p.M1_W32del, p.W32X, p.L103P, p.L225P, p.W244C, p.A281T) and three frameshift mutations c.100delC, c.565dupC, c.575dupC (p.G34fsX42, p.P189fsX1 and p.P192fsX14). The novel c.2T>G (p.M1_W32del) mutation inactivates the first in-frame translation start site of the SMPD1 gene and in the homozygous status causes NPD type B indicating that in'vivo translation of wild type SMPD1 initiates from the first in-frame ATG. Moreover, the new c.96G>A (p.W32X) introduces a premature stop codon before the second in-frame ATG. As a consequence of either c.2T>G (p.M1_W32del) or c.96G>A (p.W32X), impaired translation from the first in-frame ATG results in a mild NPD-B phenotype instead of the severe phenotype expected for a complete deficiency of the enzyme, suggesting that when the first ATG is not functional, the second initiation codon (ATG33) still produces a fairly functional sphingomyelinase. Analysis of the patients'clinical and molecular data demonstrated that all five patients with the intermediate phenotype carried at least one severe mutation. No association between the onset of pulmonary symptoms and genotype was observed. Finally, the presence of c.96G>A (p.W32X), the most frequent allele among Italian NPD type B population, and c.1799G>C (p.R600P) as compound heterozygotes in association with severe mutations suggested a beneficial effect for both mutations.     

Ten novel mutations in VMD2 associated with Best macular dystrophy (BMD)

Mutations in the vitelliform macular dystrophy 2 (VMD2) gene encoding besrtophin are responsible for Best macular dystrophy (BMD), a juvenile-onset autosomal dominant disorder of the central retina. Here, we report ten novel VMD2 mutations identified in clinically diagnosed BMD patients. The heterozygous alterations include nine missense mutations (c.32A>T, c.76G>C, c.85T>C, c.122T>C, c.122T>C, c.310G>C, c.722C>A, c.880C>G, c.893T>C) resulting in amino acid changes (respectively: Asn11Ile, Gly26Arg, Tyr29His, Leu41Pro, Trp102Arg, Asp104His, Thr241Asn, Leu294Val and Phe298Ser) located within four previously defined hotspot regions of the gene. In addition, a silent exonic mutation (c.624G>A) was identified in a two generation BMD pedigree. To determine a possible pathogenic effect of this variant, the consequences on splicing behaviour and potential exonic splice enhancer (ESE) motifs were analyzed. Finally, a 1-bp deletion (c.779delC) resulting in a frameshift mutation (Pro260fsX288) was found in exon 7, representing the first case of a potential frameshift mutation that affects the N-terminal half of the VMD2 protein. Besides a dominant negative effect which is likely attributable to the identified missense mutations, the deletion mutation suggests haploinsufficiency as an infrequent disease-causing mechanism in BMD.     

Clinical and molecular characterization of the first adult congenital disorder of glycosylation (CDG) type Ic patient

Congenital disorder of glycosylation (CDG) type Ic, the second largest subtype of CDG, is caused by mutations in human ALG6 (hALG6). This gene encodes the alpha1,3-glucosyltransferase that catalyzes transfer of the first glucose residue to the lipid-linked oligosaccharide precursor for N-linked glycosylation. In this report, we describe the first adult patient diagnosed with CDG-Ic, carrying two previously unknown mutations. The first is a three base deletion (897-899delAAT) leading to the loss of I299, the second is an intronic mutation (IVS7 + 2T > G) that causes aberrant splicing. Wildtype hALG6, delivered by a lentiviral vector into patient's fibroblasts, clearly improves the biochemical phenotype, which confirms that the mutations are disease-causing. Striking clinical findings include limb deficiencies in the fingers, resembling brachydactyly type B, a deep vein thrombosis, pseudotumor cerebri, and endocrine disturbances with pronounced hyperandrogenism and virilization. However, even in adulthood, this patient shows normal magnetic resonance imaging of the brain.     

Congenital disorder of glycosylation Ic in patients of Indian origin

Congenital disorder of glycosylation type Ic (CDG-Ic) is caused by mutations in ALG6, encoding an alpha 1,3-glucosyltransferase. The most frequent mutation found in this gene (C998T resulting in an A333V substitution) has until now been found only in patients of European origin. Here we describe the first occurrence of this CDG-Ic mutation in patients of Indian origin. Of three Indian patients described in this study, patient 1 was homozygous and patient 2 heterozygous for the A333V mutation. In patient 2 we also found a new mutation, IVS3+2_3insT, just 3bp away from the previously described IVS3+5G>A substitution; both mutations resulted in exon 3 skipping. We screened a panel of >350 genomic DNA samples from an ethnically diverse American population to determine the frequency of the A333V mutation. None of the samples carried this mutation, indicating the frequency of patients carrying this homozygous mutation should be <1 in 5x10(5). The discovery of the common CDG-Ic mutation A333V in an Indian population raises questions as to its ethnic origin.     

Mutation screening of TRPM1, GRM6, NYX and CACNA1F genes in patients with congenital stationary night blindness

The aim of this study was to identify mutations in the TRPM1, GRM6, NYX and CACNA1F genes in patients with congenital stationary night blindness (CSNB). Twenty-four unrelated patients with CSNB were ascertained. Sanger sequencing was used to analyze the coding exons and adjacent intronic regions of TRPM1, GRM6, NYX and CACNA1F. Six mutations were identified in six unrelated patients, including five novel and one known. Of the six, three novel hemizygous mutations, c.92G>A (p.Cys31Tyr), c.149G>C (p.Ary50Pro), and c.[272T>A;1429G>C] (p.[Leu91Gln;Gly477Arg]), were found in NYX in three patients, respectively. A novel c.[1984_1986delCTC;3001G>A] (p.[Leu662del;Gly1001Arg]) mutation was detected in CACNA1F in one patient. One novel and one known heterozygous variation, c.1267T>C (p.Cys423Arg) and c.1537G>A (p.Val513Met), were detected in GRM6 in two patients, respectively. No variations were found in TRPM1. The results expand the mutation spectrum of NYX, CACNA1F and GRM6. They also suggest that NYX mutations are a common cause of CSNB.     

Ethnic variation in AMD-associated complement factor H polymorphism p.Tyr402His

Age-related macular degeneration (AMD) is the most common cause of irreversible visual loss in the developed world. Previous studies have demonstrated that the c.1204T>C, p.Tyr402His allelic variant in the complement factor H (CFH) gene is associated with an approximately three-fold increased risk for AMD in Caucasians of predominantly European descent. Both the prevalence as well as the phenotypic spectrum of AMD varies widely among persons of different ethnicities. We hypothesized that populations with a lower prevalence of AMD might also have a lower prevalence of the CFH risk allele. In this study we sought to determine the frequency of this sequence variant in control populations of Caucasians, African Americans, Hispanics, Somalis, and Japanese. Normal control populations were assembled for each ethnic group: Caucasian (n=148), Somali (n=128), African American (n=75), Hispanic (n=81), and Japanese (n=82). Individuals were genotyped using a restriction digest assay and the frequency of the C allele at nucleotide position 1204 of the CFH gene was determined. A bioinformatic approach was used to identify SNPs in linkage disequilibrium with rs1061170 (c.1204T>C, p.Tyr402His) from the human haplotype map project database (HapMap) in order to validate the findings. We found widely discordant frequencies of the risk allele between some of the different ethnic groups: Japanese 0.07+/-0.02, Hispanics 0.17+/-0.03, African-Americans 0.35+/-0.04, Caucasians 0.34+/-0.03, and Somalis 0.34+/-0.03. Allele frequencies generated by analysis of the HapMap database were consistent with these findings. This study suggests that there are other yet unidentified genetic factors important in the pathogenesis of AMD that may mitigate the effects of c.1204T>C, p.Tyr402His variant.     

Functional analysis of 13 GBA mutant alleles identified in Gaucher disease patients: Pathogenic changes and "modifier" polymorphisms

Gaucher disease, the most prevalent sphingolipidosis, is caused by the deficient activity of acid beta-glucosidase, mainly due to mutations in the GBA gene. Over 200 mutations have been identified worldwide, more than 25 of which were in Spanish patients. In order to demonstrate causality for Gaucher disease, some of them: c.662C>T (p.P182L), c.680A>G (p.N188S), c.886C>T (p.R257X), c.1054T>C (p.Y313H), c.1093G>A (p.E326K), c.1289C>T (p.P391L), c.1292A>T (p.N392I), c.1322T>C (p.I402T), and the double mutants [c.680A>G; c.1093G>A] ([p.N188S; p.E326K]) and [c.1448T>C; c.1093G>A] ([p.L444P; p.E326K]), were expressed in Sf9 cells using a baculovirus expression system. Other well-established Gaucher disease mutations, namely c.1226A>G (p.N370S), c.1342G>C (p.D409H), and c.1448T>C (p.L444P), were also expressed for comparison. The levels of residual acid beta-glucosidase activity of the mutant enzymes produced by the cDNAs carrying alleles c.662C>T (p.P182L), c.886C>T (p.R257X), c.1054T>C (p.Y313H), c.1289C>T (p.P391L), and c.1292A>T (p.N392I) were negligible. The c.1226A>G (p.N370S), c.1322T>C (p.I402T), c.1342G>C (p.D409H), c.1448T>C (p.L444P), and [c.1448T>C; c.1093G>A] ([p.L444P; p.E326K]) alleles produced enzymes with levels ranging from 6 to 14% of the wild-type. The three remaining alleles, c.680A>G (p.N188S), c.1093G>A (p.E326K), and [c.680A>G; c.1093G>A] ([p.N188S; p.E326K]), showed higher activity (66.6, 42.7, and 23.2%, respectively). Expression studies revealed that the c.1093G>A (p.E326K) change, which was never found alone in a Gaucher disease-causing allele, when found in a double mutant such as [c.680A>G; c.1093G>A] ([p.N188S; p.E326K]) and [c.1448T>C; c.1093G>A] ([p.L444P; p.E326K]), decreases activity compared to the activity found for the other mutation alone. These results suggest that c.1093G>A (p.E326K) should be considered a "modifier variant" rather than a neutral polymorphism, as previously considered. Mutation c.680A>G (p.N188S), which produces a mutant enzyme with the highest level of activity, is probably a very mild mutation or another "modifier variant."     

PMM2 mutation spectrum, including 10 novel mutations, in a large CDG type 1A family material with a focus on Scandinavian families

Carbohydrate-deficient glycoprotein syndrome type IA (CDG IA) is an autosomal recessive disease characterized clinically by severe involvement of the central and peripheral nervous system, and biochemically by complex defects in carbohydrate residues in a number of serum glycoproteins. CDG IA is caused by mutations in the PMM2 gene located in chromosome region 16p13. In this study, 61 CDG type IA patients (122 chromosomes) were screened for mutations in the PMM2 gene using a combination of SSCP and sequence analysis. More than 95% of the mutations could be detected. All of them were missense mutations. Mutations 422G>A and 357C>A were strikingly more common in the material and comprised 58% of mutations detected. Of the 20 mutations found, 10 were not reported previously. Seven mutations, e.g. 26G>A (five alleles) and 548T>C (seven alleles), were found only in Scandinavian families. The most common genotype was 357C>A/422G>A (36%). Three patients were homozygous, 357C>A/357C>A (two cases), and 548T>C/548T>C (one case). No patients homozygous for the most common mutation 422G>A were detected. The different mutations were clustered e.g., in that most were located in exon 5 (five) and exon 8 (six), while no mutation was detected in exon 2. When the frequencies of each mutation were included, exon 5 comprised 61% (65 chromosomes) of the mutations; in Scandinavian patients the frequency of these mutations was 72%. Thus, analysis of exon five in these patients enables both reliable and time-saving first screening in prenatal diagnostic cases. This could be followed by a second step of additional strategies for the detection of other mutations.     

Novel RS1 mutations associated with X-linked juvenile retinoschisis

To identify mutations in the retinoschisin (RS1) gene in families with X-linked retinoschisis (XLRS). Twenty families with XLRS were enrolled in this study. All six coding exons and adjacent intronic regions of RS1 were amplified by polymerase chain reaction (PCR). The nucleotide sequences of the amplicons were determined by Sanger sequencing. Ten hemizygous mutations in RS1 were detected in patients from 14 of the 20 families. Four of the ten mutations were novel, including c:176G>A (p:Cys59Tyr) in exon?3, c:531T>G (p:Tyr177X), c:607C>G (p:Pro203Ala) and c:668G>A (p:Cys223Tyr) in exon?6. These four novel mutations were not present in 176 normal individuals. The remaining six were recurrent mutations, including c:214G>A (p:Glu72Lys), c:304C>T (p:Arg102Trp), c:436G>A (p:Glu146Lys), c:544C>T (p:Arg182Cys), c:599G>A (p:Arg200His) and c:644A>T (p:Glu215Val). Our study expanded the mutation spectrum of RS1 and enriches our understanding of the molecular basis of XLRS.     

Novel TP53 gene mutations in tumors of Russian patients with breast cancer detected using a new solid phase chemical cleavage of mismatch method and identified by sequencing

Mutations in the tumor-suppressor p53 gene TP53 are frequent in most human cancers including breast cancer. A new solid phase chemical cleavage of mismatch method (CCM) allowed rapid and efficient screening and analysis of the TP53 gene in DNA samples extracted from tumors of 89 breast cancer patients. The novel CCM technique utilized silica beads and the potassium permanganate/tetraethylammonium chloride (KMnO(4)/TEAC) and hydroxylamine (NH(2)OH) reactions were performed sequentially in a single tube. Mutation analysis involved amplification of five different fragments of the TP53 gene using DNA from the 89 tumor samples, then pairing of the 391 labeled PCR products and forming heteroduplexes. A total of 41 unique signals were revealed in the analysis of TP53 exons 5-9 and eight were identified by direct sequencing. The three novel mutations detected are c.600T>G (p.Asn200Lys), c.601T>G (p.Leu201Val), and c.766-768delACA (p.Thr256del). The detected mutations c.638G>T (p.Arg213Leu), c.730G>T (p.Gly244Cys), and c.758C>T (p.Thr253Ile) have not been reported in breast cancer but have been recorded in tumors of other organs. A previously reported mutation c.535C>T (p.His179Tyr) and a heterozygous polymorphism c.639A>G were also detected. Of the 41 unique signals, 36 were not identified as a sequence change. As direct sequencing requires the mutant allele concentration to be greater than 30% when the mutant allele is present in a mixture with the wild-type allele, the CCM method represents a more sensitive technique requiring a lower mutant allele concentration in the wild-type mixture compared with direct sequencing. This reveals the advantage of CCM for unknown point mutation detection in DNA samples of cancer patients.     

RFT1 deficiency in three novel CDG patients

The medical significance of N‐glycosylation is underlined by a group of inherited human disorders called Congenital Disorders of Glycosylation (CDG). One key step in the biosynthesis of the Glc₃Man₉GlcNAc₂‐PP‐dolichol precursor, essential for N‐glycosylation, is the translocation of Man₅GlcNAc₂‐PP‐dolichol across the endoplasmic reticulum membrane. This step is facilitated by the RFT1 protein. Recently, the first RFT1‐deficient CDG (RFT1‐CDG) patient was identified and presented a severe N‐glycosylation disorder. In the present study, we describe three novel CDG patients with an RFT1 deficiency. The first patient was homozygous for the earlier reported RFT1 missense mutation (c.199C>T; p.R67C), whereas the two other patients were homozygous for the missense mutation c.454A>G (p.K152E) and c.892G>A (p.E298 K), respectively. The pathogenic character of the novel mutations was illustrated by the accumulation of Man₅GlcNAc₂‐PP‐dolichol and by reduced recombinant DNase 1 secretion. Both the glycosylation pattern and recombinant DNase 1 secretion could be normalized by expression of normal RFT1 cDNA in the patients' fibroblasts. The clinical phenotype of these patients comprised typical CDG symptoms in addition to sensorineural deafness, rarely reported in CDG patients. The identification of additional RFT1‐deficient patients allowed to delineate the main clinical picture of RFT1‐CDG and confirmed the crucial role of RFT1 in Man₅GlcNAc₂‐PP‐dolichol translocation. Hum Mutat 30:1–7, 2009. © 2009 Wiley‐Liss, Inc.     

An unusual case of hemochromatosis due to a new compound heterozygosity in HFE (p.[Gly43Asp;His63Asp]+[Cys282Tyr]): structural implications with respect to binding with transferrin receptor 1

Most adults affected with HFE hereditary hemochromatosis (HH type 1, MIMmusical sharp 235200) are homozygous for the p.Cys282Tyr mutation in HFE (NC_000006.10, region 26195427 to 26205038). The aim of this study was to investigate the molecular basis of iron overload in a patient presenting with severe clinical HH with one c.845G>A (p.Cys282Tyr) allele only. Molecular and pedigree studies demonstrated the presence of the c.845G>A (p.Cys282Tyr) mutation in one allele whereas the other carried the c.187C>G (p.His63Asp) mutation plus a new c.128G>A (p.Gly43Asp) substitution in cis. A molecular modeling study of the p.[Gly43Asp;His63Asp] and p.His63Asp variants versus the wild type was carried out using molecular dynamics (MD) simulation in presence of implicit solvent. We found that the c.187C>G (p.His63Asp) mutation does not introduce any major change in the 1- domains of HFE whereas the c.128G>A (p.Gly43Asp) substitution is responsible for a modification of the dynamics and the structure of the Gln40-Ser45 loop, a critical region for HFE-TfR1 interaction thus impairing HFE-TfR1 normal contact. We conclude that the occurrence of complex alleles may be an alternative explanation for the variability of the phenotype in individuals who are compound heterozygous for c.[187C>G]+[845G>A] (p.[His63Asp]+[Cys282Tyr]).     

ALG11‐CDG syndrome: Expanding the phenotype

ALG11‐Congenital Disorder of Glycosylation (ALG11‐CDG, also known as congenital disorder of glycosylation type Ip) is an inherited inborn error of metabolism due to abnormal protein and lipid glycosylation. We describe two unrelated patients with ALG11‐CDG due to novel mutations, review the literature of previously described affected individuals, and further expand the clinical phenotype. Both affected individuals reported here had severe psychomotor disabilities and epilepsy. Their fibroblasts synthesized truncated precursor glycan structures, consistent with ALG11‐CDG, while also showing hypoglycosylation of a novel biomarker, GP130. Surprisingly, one patient presented with normal transferrin glycosylation profile, a feature that has not been reported previously in patients with ALG11‐CDG. Together, our data expand the clinical and mutational spectrum of ALG11‐CDG.     

Molecular and clinical description of the first US patients with congenital disorder of glycosylation Ig

In this report we describe the first two US patients with congenital disorder of glycosylation type Ig (CDG-Ig). Both patients presented with symptoms indicating CDG, including developmental delay, hypotonia and failure to thrive, and tested positive for deficient glycosylation of transferrin. Labeling of the patients' lipid-linked oligosaccharides suggested mutations in the hALG12 gene, encoding a mannosyltransferase. Both patients were shown to carry previously unpublished hALG12-mutations. Patient 1 has one allele with a deletion of G29, resulting in a premature stop codon, and another allele with an 824G>A mutation yielding an S275N amino acid change. Patient 2 carries two heterozygous mutations (688T>G and 931C>T), resulting in two amino acid exchanges, Y230D and R311C. An adenoviral vector expressing wild type hALG12 corrects the abnormal lipid-linked oligosaccharide pattern of the patients' cells. In addition to common CDG symptoms, these patients also presented with low IgG and genital hypoplasia, symptoms previously described in CDG-Ig patients. We therefore conclude that a combination of developmental delay, low IgG, and genital hypoplasia should prompt CDG testing.     

FBN1新生突变引起的马凡综合征及其基因型-表型的关联研究

 目的: 本研究对2个不同马凡综合征(Marfan syndrome)的小家系进行致病基因FBN1的编码区和剪切位点突变检测,以寻找致病的突变,并初步探索马凡综合征基因型-表型的关联。方法: 通过临床检查、实验室检查及心脏超声检查确诊2个无血缘关系的家庭中原疑似为马凡综合征的3例患者。运用新一代测序对家系1的疑似患者行FBN1基因的全外显子组测序,并对检出的致病性遗传变异进行Sanger验证及在所有家系成员中验证;对于家系2的存活成员,本研究直接进行PCR扩增FBN1基因的所有编码区及剪切位点,对产物进行直接Sanger测序。另外在50个正常对照中对新发现的突变位点进行基于PCR产物的测序分析,以排除多态性;并对实验结果行生物信息学分析。结果: 所有存活的疑似患者均确诊为马凡综合征。在家系1中,我们检测到了一个FBN1基因数据库中尚未报道的新突变c.4685G>A(p.Cys1562Tyr),并且患者父母和同胞姐姐均未检测到此变异,故此突变为一个新生突变。该错义突变使第1562位上极性中性的含硫的半胱氨酸被极性中性的含羟苯基的酪氨酸所替代,影响了fibrillin-1蛋白一个TGF-β结合结构域,导致蛋白质的二级结构发生改变。家系2含父母及一对同卵双胎患者,其中一患者已去世。我们在存活患者检测到1个FBN1基因的已报道致病突变c.3706T>C(p.Cys1236Arg),该突变在患者父母中不存在,故也为新生突变。结论: 本文报道了一例FBN1基因的新突变及另一例由FBN1基因已知突变引起的马凡综合征,二者皆为新生突变,并在家系中进行了基因型-表型的比较,表明家系1的新突变可能与经典马凡综合征的表型相关,而家系2的已知突变确和新生儿重症马凡综合征表型相关。