Molecular characterization of the cystic fibrosis transmembrane conductance regulator gene in congenital absence of the vas deferens

Abstract

Purpose: Approximately 20% of patients with congenital absence of the vas deferens remain without two mutations identified. We applied a strategy of serial screening steps to 45 patients with congenital absence of the vas deferens and characterized cystic fibrosis transmembrane conductance regulator gene mutations in all cases.Methods: DNA samples of 45 patients with congenital absence of the vas deferens were screened by successive different molecular genetics approaches. Results: Initial screening for the 31 most frequent cystic fibrosis mutations, IVS8 poly(TG)m, poly(T)n, and M470V polymorphisms, identified 8 different mutations in 40 patients (88.9%). Extensive cystic fibrosis transmembrane conductance regulator gene analysis by denaturing gradient gel electrophoresis, denaturing high-performance liquid chromatography, and DNA sequencing detected 17 further mutations, of which three were novel. Cystic fibrosis transmembrane conductance regulator gene rearrangements were searched by semiquantitative fluorescent multiplex polymerase chain reaction, which detected a CFTRdele2,3 (21 kb) large deletion and confirmed two homozygous mutations. Overall, 42 patients (93.3%) had two mutations and 3 patients (6.7%) had one mutation detected. Conclusions: The present screening strategy allowed a higher mutation detection rate than previous studies, with at least one cystic fibrosis transmembrane conductance regulator gene mutation found in all patients with congenital absence of the vas deferens. Genet Med 2007:9(3):163-172. Key Words: CAVD, CBAVD, CUAVD, CFTR mutations, CFTR rearrangements, cystic fibrosis, male infertilityCongenital absence of the vas deferens (CAVD) in otherwise healthy males is responsible for 1% to 2% of male infertility and 6% of obstructive azoospermia cases. The disease was classified as a primary genital form of cystic fibrosis (CF) because 67% to 85% of patients with CAVD have mutations and/or splicing variants in the cystic fibrosis transmembrane conductance regulator (CFTR) gene. Patients with CAVD with upper urinary tract malformations (20% of cases) have a normal frequency of CFTR mutations and are thus considered a distinct clinical entity with a different cause. [1][2][3][4][5][6][7][8] CFTR genotypes in patients with CAVD are markedly different from those found in patients with CF. Patients with CAVD are usually heterozygous for a mild mutation on one allele and a severe mutation on the second allele (88%), or for mild mutations on both CFTR alleles (12%). On the other hand, homozygosity or compound heterozygosity for two severe mutations is responsible for the classic CF phenotype. The most common CFTR mutations found in CAVD are T5 allele, deltaF508, and R117H, with the combination between the T5 allele and a severe CF mutation in the other allele being the main cause of CAVD. 9 The polymorphic polythymidine locus (Tn) is located within the 3= splice acceptor site of intron 8 (IVS8 poly[T]n). The length of the IVS8 poly(T)n tract (T5/T7/T9) is inversely correlated with the degree of ...

Molecular and Functional Characterization of CBAVD-Causing Mutations Located in CFTR Nucleotide-Binding Domains

“…Thus CBAVD may represent an atypical incomplete (genital) form of CF [20]. The genetic link between CF and CBAVD has been confirmed by several studies that identified CFTR mutations in 80-100% of isolated CBAVD patients [24][25][26]. Clinical symptoms of CBAVD are characterized by azoospermia with low semen plasma volume (<1.5 ml) and low pH (<7.5), unilateral or bilateral non-palpable vas deferens, absence of the distal part of the epididymis and hypoplasia of the seminal vesicles [21].…”

“…Mutations in CFTR were found to have an impact on human reproduction and consequently about 98% of male CF patients are infertile due to obstructive azoospermia, which is associated with congenital absence of the vas deferens (CBAVD; MIM 277180) [17,18]. Mutations in CFTR gene have also been identified in patients with isolated CBAVD and in the absence of other clinical CF symptoms [19][20][21][22][23][24]. Thus CBAVD may represent an atypical incomplete (genital) form of CF [20].…”

“…CBAVD patients are usually heterozygous for a mild mutation in one and a severe mutation in the other allele (88%), or carry two mild mutations in both CFTR alleles (12%) [26]. In a previous study we described a spectrum of CFTR mutations causing CBAVD [24]. Here we analyzed the impact of three novel missense mutations on the CFTR processing, intracellular transport and Cl -channel activity using a heterologous expression system.…”

Immunohystochemical analysis of CFTR in normal and disrupted spermatogenesis

“…CFTR mutations may cause CBAVD in the absence of CF. Patients with CBAVD but without CF, present one severe CFTR mutation in one allele and one mild CFTR mutation in the other, or two mild CFTR mutations in the two alleles [Grangeia et al 2004;Grangeia et al 2007]. Functional sperm are produced, since testicular sperm can be recovered for intracytoplasmic sperm injection (ICSI) treatment cycles by testicular sperm extraction, with very satisfactory results [Larriba et al 1998;Wong et al 1998;Sousa et al 2002;Grangeia et al 2004;Grangeia et al 2007].…”

Spectrum of CFTR gene sequence variants in a northern Portugal population

“…15 In addition to CF, CFTR variants are also responsible for other diseases, usually monosymptomatic and classified as CFTR-related disorders (RD). 16 Many of these patients may carry a CF-pathogenic variant on one allele and a likely pathogenic variant on the other allele, or even in both alleles.…”