Objective To investigate the disease-causing gene of Stargardt disease. Method Fifteen patients with Stargardt disease were analyzed with 11 primers of the 11 exons of ABCR gene by using PCR-SSCP and DNA direct sequencing techniques. Results Three newly detected disease-causing mutations were found. Among those mutations, one is a frameshift mutation and others are single base transition. Conclusion This research confirmed that ABCR gene is associated with Stargardt disease, and 3 new mutations of ABCR gene were found. (Chin J Ocul Fundus Dis,2000,16:240-243)
ObjectiveTo observe the clinical manifestation and gene mutation of a pedigree with Sorsby fundus dystrophy (SFD). Methods Ten members in 3 generations of a pedigree with SFD were included in this study. Four patients were observed in the pedigree, including 2 females and 2 males. All 10 members underwent comprehensive ophthalmic examinations, including best-corrected visual acuity, intraocular pressure, slit-lamp biomicroscopy, indirect ophthalmoscopy, fundus color photography and spectral domain optical coherence tomography. Genomic DNA was extracted from peripheral venous blood which was collected from all the members. Relevant exons of ocular diseases were detected by the next generation sequencing method from the proband. The other members underwent Sanger verification. Results Among the four patients, fading eyesight was appeared at their 44, 46, 47 and 40 year-old respectively. The two male patients had bilateral morbidity, and the two female patients had monocular symptoms. DNA sequencing results showed that the proband, other 3 patients and 2 members from the Ⅲ generation had heterozygous mutation of TIMP3 gene in exon 5. The amino acid encoded by TIMP3 gene No.204 codon changed from serine to cysteine (TIMP3:NM_000362:Exon5:c.A610T/p.S204C). CoclusionsThe invasion time of all the patients in this pedigree is after their 40 year-old. Heterozygous mutation at c.610A＞T (p.S204C) in TIMP3 gene is the causative gene of SFD in this pedigree.
ObjectiveTo analyze the clinical and genetic characteristics of ADGRV1 gene mutation epilepsy.MethodsA retrospective collection of 26 patients with epilepsy diagnosed and related gene sequencing was performed in the Affiliated Hospital of Jining Medical College from January 2018 to December 2018. Five epilepsy patients with ADGRV1 mutations were screened out, and their clinical characteristics and gene mutation characteristics were summarized.ResultsA total of 5 epilepsy patients with ADGRV1 mutation were collected, including 1 male and 4 females, with an average age of (7±5.83) years. Three patients had a family history of epilepsy, and the father of the other two patients had a history of febrile seizures. 2 cases showed generalized tonic-clonic seizures, and 3 cases showed partial seizures followed by generalized seizures. The results of genetic testing revealed 7 mutation sites in the ADGRV1 gene, of which one missense mutation site c.2039A>G has been reported in the literature. Two of the 5 patients underwent epilepsy surgery, and they were still treated with multiple anti-epileptic drugs for a long time after the operation, and the other 3 patients were treated with anti-epileptic drugs for a long time. At present, 4 out of 5 patients had seizures still not under effective control, and 1 case did not relapse after being followed up for nearly 1 year.ConclusionThe clinical features of epilepsy caused by ADGRV1 gene mutation are early onset, mainly manifested as general tonic-clonic seizures or partial seizures secondary to generalized seizures, accompanied by disturbance of consciousness during seizures. The combined treatment of anti-epileptic drugs and postoperative anti-epileptic drugs is less effective. Genetic testing can guide genetic counseling and assisted diagnosis.
ObjectiveTo observe the effects of penetrance, different time of onset and mutation sites on retinal nerve fiber layer (RNFL) and macular thickness in patients with Leber's hereditary optic neuropathy (LHON).MethodsThis was a cross-sectional observational study. A total of 88 patients with LHON and 1492 relatives of the maternal relatives (gene carriers) who received treatment in People’s Liberation Army General Hospital from 2015 to 2017 were included in the study. Among the 1492 family members, there were 694 males and 798 females. Peripheral venous blood was extracted from all subjects for mitochondrial DNA testing, and penetrance was calculated. A total of 117 patients underwent BCVA and SD-OCT examinations, including 82 patients and 35 gene carriers. The BCVA examination was performed using the Snellen visual acuity chart, which was converted into logMAR visual acuity. The thickness of RNFL, ganglion cell complex (GCC) and inner limiting membrane (ILM)-RPE were measured with OCT instrument. The mean follow-up was 50.02±86.27 months. The disease course was divided into 6 stages including ≤3 months, 4-6 months, 7-12 months and >12 months. The thickness of RNFL, GCC and ILM-RPE in patients with different time of onset and mutation sites were comparatively analyzed by covariance analysis. Categorical variables were expressed as a percentage, and the χ2 test was used for comparison among multiple groups.ResultsAmong the 1492 family members, 285 were diagnosed with LHON and highly suspected clinical manifestations (19.10%), including 190 males (21.98%) and 95 females (11.90%). The total penetrance rates of 11778, 14484 and rare mutation sites were 19.84% (228/1149), 20.50% (33/161), and 13.19% (24/182) respectively; male penetrance rates were 28.87% (153/530), 27.28% (20/72), and 18.48% (17/92) and female penetrance rates were 12.12% (75/619),14.61% (13/89) and 7.78% (7/90). There was no significant difference in total (χ2=4.732), male (χ2=4.263) and female (χ2=4.263) penetrance between different mutation sites (P=0.094, 0.110, 0.349). Compared with non-pathogenic carriers, the thickness of the RNFL, GCC and ILM-RPE were all different in the four stages ( ≤3months, 4-6 months, 7-12 months and >12 months). The thickness of RNFL, GCC and ILM-RPE decreased with the time of onset (P=0.000). There were significant differences in the thickness of each of the GCC and ILM-RPE layers in the macular area of LHON patients with different mutation sites (P＜0.05). Among them, the site 11778 and 3460 had the most severe damage in all quadrants of macular GCC and ILM-RPE layer, followed by 14484 site, and the rare site had the least damage in all quadrants.ConclusionsThe penetrance of LHON patients is 19.10%. With the extension of the onset time (within 1 year), the RNFL layer of the optic disc and all quadrants of the macular GCC and ILM-RPE layer gradually thinned. Compared with 11778 and rare site, 14484 site, and the rare site had the lighter damage on the thickness of RNFL, GCC and ILM-RPE.
Objective To analyze the BEST1 gene mutations and clinical features in patients with multifocal vitelliform retinopathy (MVR). Methods This is a retrospective case series study. Five MVR families with MVR, including 9 patients and 10 healthy family members were recruited. Clinical evaluations were performed in all MVR patients and their family members, including best-corrected visual acuity (BCVA), intraocular pressure (IOP), refraction, slit-lamp examination, 90 D preset lens examination, gonioscopy, color fundus photography, optical coherence tomography (OCT), fundus autofluorescence (AF), ultrasound biomicroscopy (UBM) and axial length measurement. Electro-oculogram (EOG) was performed in 12 eyes and visual field were performed in 13 eyes. Peripheral blood samples were collected in all subjects to extract genomic DNA. Coding exons and flanking intronic regions of BEST1 were amplified by polymerase chain reaction and analyzed by Sanger sequencing. Results Among the 5 MVR families, 3 probands from three families had family history, including 1 family had autosomal dominant inheritance pattern. Two patients from 2 families were sporadic cases. Screening of BEST1 gene identified four mutations, including three missense mutations (c.140G>T, p.R47L; c.232A>T, p.I78F; c.698C>T, p.P233L) and 1 deletion mutation (c.910_912del, p.D304del). Two mutations (p.R47L and p.I78F) were novel. The BCVA of affected eyes ranged from hand motion to 1.0. The mean IOP was (30.39±11.86) mmHg (1 mmHg=0.133 kPa). The mean refractive diopter was (-0.33±1.68) D. Twelve eyes had angle-closure glaucoma (ACG) and 4 eyes had angle closure (AC). EOG Arden ratio was below 1.55 in all patients. The mean anterior chamber depth was (2.17±0.29) mm. Visual field showed defects varied from paracentral scotoma to diffuse defects. The mean axial length was (21.87±0.63) mm. All MVR patients had multifocal vitelliform lesions in the posterior poles of retina. ACG eyes demonstrated pale optic disc with increased cup-to-disc ratio. OCT showed retinal edema, extensive serous retinal detachment and subretinal hyper-reflective deposits which had high autofluorescence in AF. The genetic testing and clinical examination were normal in 10 family members. Conclusions MVR patients harbored heterozygous mutation in the BEST1 gene. Two novel mutations (p.R47L and p.I78F) were identified. These patients had clinical features of multifocal vitelliform retinopathy and abnormal EOG. Most patients suffered from AC/ACG.
Objective To make the diagnosis of a pedigree of X-linked congenital stational night blindness(CSNB) and to identify the disease-causing gene. Methods Clinical examination and family analysis were made. Venous blood was drawn from 5 affected and 16 unaffected individuals from the family. Genomic DNA was extracted. The locus of the candidate gene was mapped by linkage study. Mutation was screened by polymerase chain reaction (PCR) of the candidate gene exons and flanked introns. The PCR products are directly sequenced. The healthy people in and out of the family who were selected according to certain standards were as the control. Results A Chinese family with X-linked complete congenital stationary night blindness (CSNB1) was diagnosed. A missense mutation A772C (T258P) in exon 2 of NYX gene was identified in all affected patients and all female carriers were heterozygous. This mutation was neither found in normal family members nor among 110 unrelated normal controls. Conclusion A novel mutation of NYX gene with threonine to proline change is responsible for this Chinese CSNB1 family. (Chin J Ocul Fundus Dis, 2007, 23: 184-188)
Objective To analyze the new primary mutation in Chinese people with Leberprime;s hereditary optic neuropathy (LHON). Methods Genomic DNA was collected from 260 suspected LHON patients and 100 normal healthy persons. The mitochondria DNA mutation at nucleotide position (NP) 15257 and the hot spot (14452-14601 bp) of ND6 gene which include the mutations at NP (14482, 14498, 14568, 14596, 14495, and 14459) were screened by using polymerase chain reaction (PCR), heteroduplex-single strand conformation polymorphism (HA-SSCP) and restriction fragment length polymorphism (RFLP) analysis and sequencing. Primary mutation spectrum of Chinese race was analyzed. Results Eight kinds of polymorphism of mitochondria DNA were found in 260 suspected LHON patients and 100 normal healthy persons, including NP 14488C, 14518G, and 14617G which hadnrsquo;t been reported (http://www.mitomap.org/). No mutation at NP 15257, 14482, 14498, 14568, 14596, 14495, and 14459 was found. Conclusion The NP 15257A may not be the primary mutation in Chinese. Because of the race difference, 14452-14601 bp in ND6 gene may not be the hot spot in Chinese patients with LHON, and other hot spots may exist. (Chin J Ocul Fundus Dis, 2006, 22: 82-85)
ObjectiveTo reveal the pathogenic mutation in a three-generation Chinese family with autosomal dominant familial exudative vitreoretinopathy (FEVR). MethodsThree patients and a healthy spouse from the index family with FEVR were recruited. The proband was a 5 years old boy. His mother and grandpa were presented with typical FEVR presentations, while his father with normal ocular fundus. DNA was extracted from peripheral blood samples taken from all four participants. All coding and exon-intron boundary regions of five targeted genes, including NDP, FZD4, LRP5, TSPAN12 and ZNF408 were amplified with polymerase chain reaction and sequenced using direct sequencing. In silico analyses were applied to determine the conservation of the mutation site, pathogenic effect and the potential protein crystal structural changes caused by the mutation. ResultsFZD4 c.478G > A, a susceptible mutation was found after four high frequency mutation sites which MAF values were higher than 0.001 was filtered among 5 single nucleotide variations detected in four participants, leading to the residue 160 changing from glutamate to lysine (p.E160K). Co-segregation analysis between genotypes and phenotypes revealed FZD4 p.E160K as the disease-causing mutation for this family. Conservational analysis suggested that this mutation site was highly conserved among all tested species. Functional analysis predicated that this mutation may be a damaging mutation. Crystal structural analysis also indicated that this mutation could lead to the elimination of the hydrogen bond between residue 160 and asparagine at residue 152, thus altering the tertiary structure of the protein and further impairing the protein function. ConclusionOur study demonstrates FZD4 p.E160K as a novel pathogenic mutation for FEVR.
ObjectiveTo explore distribution characteristics of drug-resistant mutations and analyze drug-resistant genotypes in Mycobacterium tuberculosis in Deyang district, Sichuan. MethodsA total of 257 patients infected with Mycobacterium tuberculosis and positive for mycobacterium tuberculosis DNA who were detected from February 2010 to March 2013 were included in our research. Drug-resistance mutations were detected and analyzed using gene chip technology combining by polymerase chain reaction (PCR) and reverse dot hybridization (RDB). ResultsIn these 257 pulmonary tuberculosis patients, drug-resistance mutations were detected in 49 with pulmonary tuberculosis. Drug-resistance mutation rate at katG 315, rpsL 43, embB 306 and rpoB 531 (S531L) was 11.67% (30/257), 7.00% (18/257), 4.28% (11/257) and 3.89% (10/257), respectively. In 234 initially treated pulmonary tuberculosis patients, the rate of isoniazid-resistant genotype, rifampicin-resistant genotype, ethambutol-resistant genotype, streptomycin-resistant genotype and multi-drug resistant genotype was 9.83%, 4.27%, 3.42%, 5.13% and 2.99%, respectively. In 23 retreated pulmonary tuberculosis patients, these rates was 52.17%, 26.09%, 13.04%, 43.48% and 13.04%, respectively. ConclusionIn Deyang district, Sichuan, drug-resistant genotypes for isoniazid, rifampicin, ethambutol and streptomycin are detected in Mycobacterium tuberculosis. Most of the drug-resistant mutations occur at katG 315, rpsL 43, embB 306 and rpoB 531. The rates of drug-resistant genotypes and multi-drug resistance in initially treated pulmonary tuberculosis patients are lower than those in retreated patients. Multi-drug resistant rate is relatively low in our research.
Objective To investigate the feasibility of detection of epidermal growth factor receptor ( EGFR) exon 19 deletions and exon 21 L858R mutations in pleural effusion fromnon-small-cell lung cancer ( NSCLC) patients by mutant enriched PCR assay. Methods The mutations of exon 19 and 21 of EGFR gene in pleural samples fromthirty NSCLC patients were analyzed using both the mutant-enriched PCR assay and the non-enriched PCR assay. Results Ten ( 33. 3% , 10/ 30) exon 19 deletions and five ( 16. 7% , 5/30) exon 21 L858R mutation were detected by the mutant-enriched PCR assay, while only 6 cases ( 20. 0% ) and 1 case ( 3. 3% ) were detected by the non-enriched PCR assay respectively. The difference of mutation detection rate of EGFR gene between the two methods was statistically significant ( P = 0. 032) . Mutations were detected in all of partial responders ( 2 /4) among the four patients who received gefitinib therapy. Conclusions Mutant-enriched PCR assay can detect EGFR exon 19 deletions and exon 21 L858R mutation in pleural effusion from NSCLC patients effectively, economically and accurately. It may be a valuable biomarker for gefitinib therapy in advanced NSCLC.