QMPSF is sensitive and specific in the detection of NPHP1 heterozygous deletions.

BACKGROUND: Nephronophthisis, an autosomal recessive nephropathy, is responsible for 10% of childhood chronic renal failure. The deletion of its major gene, NPHP1, with a minor allele frequency of 0.24% in the general population, is the most common mutation leading to a monogenic form of childhood chronic renal failure. It is challenging to detect it in the heterozygous state. We aimed to evaluate the sensitivity and the specificity of the quantitative multiplex PCR of short fluorescent fragments (QMPSF) in its detection. METHODS: After setting up the protocol of QMPSF, we validated it on 39 individuals diagnosed by multiplex ligation-dependent probe amplification (MLPA) with normal NPHP1 copy number (n=17), with heterozygous deletion (n=13, seven parents and six patients), or with homozygous deletion (n=9). To assess the rate of the deletions that arise from independent events, deleted alleles were haplotyped. RESULTS: The results of QMPSF and MLPA correlated perfectly in the identification of 76 heterozygously deleted and 56 homozygously deleted exons. The inter-experimental variability of the dosage quotient obtained by QMPSF was low: control, 1.05 (median; range, 0.86-1.33, n = 102 exons); heterozygous deletion, 0.51 (0.42-0.67, n = 76 exons); homozygous deletion, 0 (0-0, n = 56 exons). All patients harboring a heterozygous deletion were found to carry a hemizygous mutation. At least 15 out of 18 deletions appeared on different haplotypes and one deletion appeared de novo. CONCLUSIONS: The cost- and time-effective QMPSF has a 100% sensitivity and specificity in the detection of NPHP1 deletion. The potential de novo appearance of NPHP1 deletions makes its segregation analysis highly recommended in clinical practice.

The 83,557insA variant of the gene coding 11ß-hydroxysteroid dehydrogenase type 1 enzyme associates with serum osteocalcin in patients with endogenous Cushing's syndrome.

OBJECTIVE: The type 1 and type 2 isoenzymes of the 11ß-hydroxysteroid dehydrogenase (HSD11B) play an important role in the prereceptor regulation of glucocorticoid bioavailability and action. The potential importance of gene variants coding HSD11B has not been previously evaluated in patients with endogenous hypercortisolism. The aim of the present study was to explore presumed associations between the 83,557insA variant of the HSD11B1 gene and circulating hormone concentrations, bone turnover and bone mineral density (BMD) in patients with endogenous Cushing's syndrome (CS). PATIENTS AND METHODS: Forty one patients with ACTH-producing pituitary adenomas (Cushing's disease-CD), 32 patients with cortisol-producing adrenal tumors (ACS) and 129 healthy control subjects were genotyped for the 83,557insA variant of the HSD11B1 gene using restriction fragment length analysis. BMD was measured by dual-energy X-ray absorptiometry. Serum cortisol, ACTH, osteocalcin (OC) and C-terminal crosslinks (CTX) of human collagen type I (C-telopeptide) were measured by electrochemiluminescence immunoassay. RESULTS: No statistically significant differences were found in the allelic frequencies of the 83,557insA polymorphism among patients with CD, ACS and healthy controls. Among all patients with CS, heterozygous carriers of the 83,557insA had significantly higher serum OC as compared to non-carriers. Patients with ACS carrying the 83,557insA variant had higher plasma ACTH concentrations compared to non-carriers. The 83,557insA variant failed to associate with BMD in patients and controls. CONCLUSIONS: Our present findings indicate that the 83,557insA variant of the HSD11B1 gene may influence serum markers of bone turnover, but not BMD in patients with endogenous Cushing's syndrome.

Effects of bone and mineral metabolism on arterial elasticity in chronic renal failure.

Arterial stiffness (Ast) individually predicts cardiovascular (CV) mortality. Ast increases via vascular calcification and can be characterized by pulse wave velocity (PWV). We assessed the influence of mineral and bone metabolism on Ast in dialyzed (D) and renal transplanted (Tx) children by measuring fetuin-A and bone markers [bone-specific alkaline phosphatase (BALP); beta-CrossLaps (beta)]. Normalized PWV/height (PWV/h) of 11 D and 17 Tx patients was measured by applanation tonometry. Levels of calcium (Ca), phosphate (P), fetuin-A, and bone markers were analyzed. Ca x P/fetuin-A ratio was calculated to characterize the balance of calcification and inhibition. Cumulative dose of calcitriol was also assessed. Fetuin-A was lower in D and Tx compared with healthy controls. Bone markers and Ca x P/ fetuin-A of D were significantly higher than those of Tx and controls. In D PWV/h correlated with Ca x P/fetuin-A and BALP (r=0.8; p=0.005, r=0.6, p=0.05, respectively); BALP correlated with Ca x P/fetuin-A (r=0.7, p=0.01). In Tx, there was a correlation between calcitriol administered before transplantation and PWV/h (r=0.5, p=0.04). Increased bone turnover was coupled with an increased potential of calcium-phosphate precipitation, as shown by the increased Ca x P/fetuin-A. It might explain the connection between disturbed mineral and bone metabolism and Ast. Tx might be beneficial on Ast, though follow-up studies are needed.