Congenital bilateral absence of the vas deferens （CBAVD） is a frequent cause of obstructive azoospermia, and mutations of the cystic fibrosis transmembrane conductance regulator （CFTR） gene have also been frequent...Congenital bilateral absence of the vas deferens （CBAVD） is a frequent cause of obstructive azoospermia, and mutations of the cystic fibrosis transmembrane conductance regulator （CFTR） gene have also been frequently identified in patients with CBAVD. However, the distribution of the CFTR polymorphisms M470V, poly-T, TG-repeats and F508del mutation in the Chinese CBAVD population with presumed low cystic fibrosis （CF） frequency remains to be evaluated. Samples obtained from 109 Chinese infertile males with CBAVD and 104 normal controls were analyzed for the presence of CFTR （TG）m（T）n, M470V and F508del by PCR amplification followed by direct sequencing. Our study showed that the F5OSdel mutation was not found in our patients. The 5T mutation was present with high frequency in Chinese CBAVD patients and IVS8-5T linked to either 12 or 13 TG repeats was highly prevalent among CBAVD patients （97.22% of 72 cases and 96.91% of 97 alleles with IVS8-5T）. Moreover, a statistically significant relationship between TG 12-5T-V470 haplotype and CBAVD was detected. This study indicated that the CFTR polymorphisms poly-T, TG-repeats and M470V might affect the process of CBAVD in the Chinese population.展开更多
Mutations in the cystic fibrosis transmembrane conductance regulator （CFTR） gene have been implicated in the onset of cystic fibrosis and other clinical respiratory disorders. In the present study, we investigated t...Mutations in the cystic fibrosis transmembrane conductance regulator （CFTR） gene have been implicated in the onset of cystic fibrosis and other clinical respiratory disorders. In the present study, we investigated the role of CFTR variations, poly-T, TG-repeats, and M470V in susceptibility to bronchial asthma and chronic bronchitis in a Chinese population in Jiangsu province, China. A total of 72 bronchial asthma patients, 68 chronic bronchitis pa- tients, and 117 healthy subjects were included in this study. The Tn-TGm haplotype was sequenced and the CFTR variant M470V was detected using restriction fragment length polymorphism （RFLP）. We found that the fre- quency of TS-TG12-V470 in chronic bronchitis patients was 0.07%, which was notably higher than that in healthy subjects （0.01%） and bronchial asthma patients （0.04%）. Thus, the presence of the TS-TG12 haplotype of the CFTR gene is likely to play a role in the development and progression of respiratory conditions, such as chronic bronchitis.展开更多
Background: Genetically engineered animals are essential for gaining a proper understanding of the disease mechanisms of cystic fibrosis(CF). The rat is a relevant laboratory model for CF because of its zootechnical c...Background: Genetically engineered animals are essential for gaining a proper understanding of the disease mechanisms of cystic fibrosis(CF). The rat is a relevant laboratory model for CF because of its zootechnical capacity, size, and airway characteristics, including the presence of submucosal glands.Methods: We describe the generation of a CF rat model(F508 del) homozygous for the p.Phe508 del mutation in the transmembrane conductance regulator(Cftr) gene. This model was compared to new Cftr-/-rats(CFTR KO). Target organs in CF were examined by histological staining of tissue sections and tooth enamel was quantified by micro-computed tomography. The activity of CFTR was evaluated by nasal potential difference(NPD) and short-circuit current measurements. The effect of VX-809 and VX-770 was analyzed on nasal epithelial primary cell cultures from F508 del rats.Results: Both newborn F508 del and Knock out(KO) animals developed intestinal obstruction that could be partly compensated by special diet combined with an osmotic laxative. The two rat models exhibited CF phenotypic anomalies such as vas deferens agenesis and tooth enamel defects. Histology of the intestine, pancreas, liver, and lungs was normal. Absence of CFTR function in KO rats was confirmed ex vivo by short-circuit current measurements on colon mucosae and in vivo by NPD, whereas residual CFTR activity was observed in F508 del rats. Exposure of F508 del CFTR nasal primary cultures to a combination of VX-809 and VX-770 improved CFTR-mediated Cl-transport.Conclusions: The F508 del rats reproduce the phenotypes observed in CFTR KO animals and represent a novel resource to advance the development of CF therapeutics.展开更多
AIM: To investigate the three important cystic fibrosis transmembrane conductance regulator (CFTR) haplotypes poly-T, TG-repeats and the M470V polymorphisms in the Chinese population, and to compare their distribution...AIM: To investigate the three important cystic fibrosis transmembrane conductance regulator (CFTR) haplotypes poly-T, TG-repeats and the M470V polymorphisms in the Chinese population, and to compare their distribution with that in Caucasians and other Asian populations. METHODS: Genomic DNA was extracted from blood leukocytes. Exons 9 and 10 of the CFTR gene were obtained through polymerase chain reaction (PCR). Exon 9 DNA sequences were directly detected by an automated sequencer and poly-T and TG-repeats were identified by direct sequence analysis. Pure exon 10 PCR-amplified products were digested by HphⅠ restriction enzyme and the M470V mutation was detected by the AGE photos of digestion products. RESULTS: T7 was the most common (93.6%) haplotype and the (TG)11 frequency of 57.2% and (TG)12 frequency of 40.9% were dominant haplotypes in the junction of intron 8 (IVS-8) and exon 9. The frequency of T5 was 3.8% and all T5 allele tracts (10 alleles) were joined with (TG)12. Four new alleles of T6 (1.5%) were found in three healthy individuals. In exon 10, the V allele (56.1%) was slightly more frequent than the M allele (43.9%), and the M/V (45.5%) was the dominant genotype in these individuals. The three major haplotypes T7-(TG)11-V470, T7-(TG)12-M470 and T7-TG11-M470 were related to nearly 86.0% of the population. CONCLUSION: The polymorphisms of poly-T, TG-repeats, and M470V distribution were similar to those in other East Asians, but they had marked differences in frequency from those single haplotype polymorphisms or linkage haplotypes in Caucasians. Thus, they may be able to explain the low incidence of CF and CF-like diseases in Asians.展开更多
Congenital bilateral absence of the vas deferens(CBAVD)is observed in 1%–2%of males presenting with infertility and is clearly associated with cystic fibrosis transmembrane conductance regulator(CFTR)mutations.CFTR i...Congenital bilateral absence of the vas deferens(CBAVD)is observed in 1%–2%of males presenting with infertility and is clearly associated with cystic fibrosis transmembrane conductance regulator(CFTR)mutations.CFTR is one of the most well-known genes related to male fertility.The frequency of CFTR mutations or impaired CFTR expression is increased in men with nonobstructive azoospermia(NOA).CFTR mutations are highly polymorphic and have established ethnic specificity.Compared with F508Del in Caucasians,the p.G970D mutation is reported to be the most frequent CFTR mutation in Chinese patients with cystic fibrosis.However,whether p.G970D participates in male infertility remains unknown.Herein,a loss-of-function CFTR p.G970D missense mutation was identified in a patient with CBAVD and NOA.Subsequent retrospective analysis of 122 Chinese patients with CBAVD showed that the mutation is a common pathogenic mutation(4.1%,5/122),excluding polymorphic sites.Furthermore,we generated model cell lines derived from mouse testes harboring the homozygous Cftr p.G965D mutation equivalent to the CFTR variant in patients.The Cftr p.G965D mutation may be lethal in spermatogonial stem cells and spermatogonia and affect the proliferation of spermatocytes and Sertoli cells.In spermatocyte GC-2(spd)ts(GC2)Cftr p.G965D cells,RNA splicing variants were detected and CFTR expression decreased,which may contribute to the phenotypes associated with impaired spermatogenesis.Thus,this study indicated that the CFTR p.G970D missense mutation might be a pathogenic mutation for CBAVD in Chinese males and associated with impaired spermatogenesis by affecting the proliferation of germ cells.展开更多
文摘Congenital bilateral absence of the vas deferens （CBAVD） is a frequent cause of obstructive azoospermia, and mutations of the cystic fibrosis transmembrane conductance regulator （CFTR） gene have also been frequently identified in patients with CBAVD. However, the distribution of the CFTR polymorphisms M470V, poly-T, TG-repeats and F508del mutation in the Chinese CBAVD population with presumed low cystic fibrosis （CF） frequency remains to be evaluated. Samples obtained from 109 Chinese infertile males with CBAVD and 104 normal controls were analyzed for the presence of CFTR （TG）m（T）n, M470V and F508del by PCR amplification followed by direct sequencing. Our study showed that the F5OSdel mutation was not found in our patients. The 5T mutation was present with high frequency in Chinese CBAVD patients and IVS8-5T linked to either 12 or 13 TG repeats was highly prevalent among CBAVD patients （97.22% of 72 cases and 96.91% of 97 alleles with IVS8-5T）. Moreover, a statistically significant relationship between TG 12-5T-V470 haplotype and CBAVD was detected. This study indicated that the CFTR polymorphisms poly-T, TG-repeats and M470V might affect the process of CBAVD in the Chinese population.
基金supported by the Key Talent's Subsidy Project in Science and Education of the Jiangsu Province (No. 2007158)
文摘Mutations in the cystic fibrosis transmembrane conductance regulator （CFTR） gene have been implicated in the onset of cystic fibrosis and other clinical respiratory disorders. In the present study, we investigated the role of CFTR variations, poly-T, TG-repeats, and M470V in susceptibility to bronchial asthma and chronic bronchitis in a Chinese population in Jiangsu province, China. A total of 72 bronchial asthma patients, 68 chronic bronchitis pa- tients, and 117 healthy subjects were included in this study. The Tn-TGm haplotype was sequenced and the CFTR variant M470V was detected using restriction fragment length polymorphism （RFLP）. We found that the fre- quency of TS-TG12-V470 in chronic bronchitis patients was 0.07%, which was notably higher than that in healthy subjects （0.01%） and bronchial asthma patients （0.04%）. Thus, the presence of the TS-TG12 haplotype of the CFTR gene is likely to play a role in the development and progression of respiratory conditions, such as chronic bronchitis.
基金TEFOR,Grant/Award Number:RIIINSB-0014France Life Imaging,Grant/Award Number:ANR-11-INBS-0006+6 种基金Swiss Cystic Fibrosis Foundation CFCHFondation Maladies RaresInfrastructures Biologie-SantéIHU-CESTI,Grant/Award Number:ANR-10-IBHU-005IRSR Pays de la LoireVaincre la MucoviscidoseSwiss National Foundation,Grant/Award Number:310030_172909
文摘Background: Genetically engineered animals are essential for gaining a proper understanding of the disease mechanisms of cystic fibrosis(CF). The rat is a relevant laboratory model for CF because of its zootechnical capacity, size, and airway characteristics, including the presence of submucosal glands.Methods: We describe the generation of a CF rat model(F508 del) homozygous for the p.Phe508 del mutation in the transmembrane conductance regulator(Cftr) gene. This model was compared to new Cftr-/-rats(CFTR KO). Target organs in CF were examined by histological staining of tissue sections and tooth enamel was quantified by micro-computed tomography. The activity of CFTR was evaluated by nasal potential difference(NPD) and short-circuit current measurements. The effect of VX-809 and VX-770 was analyzed on nasal epithelial primary cell cultures from F508 del rats.Results: Both newborn F508 del and Knock out(KO) animals developed intestinal obstruction that could be partly compensated by special diet combined with an osmotic laxative. The two rat models exhibited CF phenotypic anomalies such as vas deferens agenesis and tooth enamel defects. Histology of the intestine, pancreas, liver, and lungs was normal. Absence of CFTR function in KO rats was confirmed ex vivo by short-circuit current measurements on colon mucosae and in vivo by NPD, whereas residual CFTR activity was observed in F508 del rats. Exposure of F508 del CFTR nasal primary cultures to a combination of VX-809 and VX-770 improved CFTR-mediated Cl-transport.Conclusions: The F508 del rats reproduce the phenotypes observed in CFTR KO animals and represent a novel resource to advance the development of CF therapeutics.
基金Supported by The Key Talent of Traditional Chinese Medicine, Foundation from the Government of Jiangsu Province, No. [2007] 158
文摘AIM: To investigate the three important cystic fibrosis transmembrane conductance regulator (CFTR) haplotypes poly-T, TG-repeats and the M470V polymorphisms in the Chinese population, and to compare their distribution with that in Caucasians and other Asian populations. METHODS: Genomic DNA was extracted from blood leukocytes. Exons 9 and 10 of the CFTR gene were obtained through polymerase chain reaction (PCR). Exon 9 DNA sequences were directly detected by an automated sequencer and poly-T and TG-repeats were identified by direct sequence analysis. Pure exon 10 PCR-amplified products were digested by HphⅠ restriction enzyme and the M470V mutation was detected by the AGE photos of digestion products. RESULTS: T7 was the most common (93.6%) haplotype and the (TG)11 frequency of 57.2% and (TG)12 frequency of 40.9% were dominant haplotypes in the junction of intron 8 (IVS-8) and exon 9. The frequency of T5 was 3.8% and all T5 allele tracts (10 alleles) were joined with (TG)12. Four new alleles of T6 (1.5%) were found in three healthy individuals. In exon 10, the V allele (56.1%) was slightly more frequent than the M allele (43.9%), and the M/V (45.5%) was the dominant genotype in these individuals. The three major haplotypes T7-(TG)11-V470, T7-(TG)12-M470 and T7-TG11-M470 were related to nearly 86.0% of the population. CONCLUSION: The polymorphisms of poly-T, TG-repeats, and M470V distribution were similar to those in other East Asians, but they had marked differences in frequency from those single haplotype polymorphisms or linkage haplotypes in Caucasians. Thus, they may be able to explain the low incidence of CF and CF-like diseases in Asians.
基金support from the National Key Research and Developmental Program of China (No.2018YFC1003603)the National Natural Science Foundation of China (No.81971445).
文摘Congenital bilateral absence of the vas deferens(CBAVD)is observed in 1%–2%of males presenting with infertility and is clearly associated with cystic fibrosis transmembrane conductance regulator(CFTR)mutations.CFTR is one of the most well-known genes related to male fertility.The frequency of CFTR mutations or impaired CFTR expression is increased in men with nonobstructive azoospermia(NOA).CFTR mutations are highly polymorphic and have established ethnic specificity.Compared with F508Del in Caucasians,the p.G970D mutation is reported to be the most frequent CFTR mutation in Chinese patients with cystic fibrosis.However,whether p.G970D participates in male infertility remains unknown.Herein,a loss-of-function CFTR p.G970D missense mutation was identified in a patient with CBAVD and NOA.Subsequent retrospective analysis of 122 Chinese patients with CBAVD showed that the mutation is a common pathogenic mutation(4.1%,5/122),excluding polymorphic sites.Furthermore,we generated model cell lines derived from mouse testes harboring the homozygous Cftr p.G965D mutation equivalent to the CFTR variant in patients.The Cftr p.G965D mutation may be lethal in spermatogonial stem cells and spermatogonia and affect the proliferation of spermatocytes and Sertoli cells.In spermatocyte GC-2(spd)ts(GC2)Cftr p.G965D cells,RNA splicing variants were detected and CFTR expression decreased,which may contribute to the phenotypes associated with impaired spermatogenesis.Thus,this study indicated that the CFTR p.G970D missense mutation might be a pathogenic mutation for CBAVD in Chinese males and associated with impaired spermatogenesis by affecting the proliferation of germ cells.
