Identification and characterization of aquaporin-2 water channel mutations causing nephrogenic diabetes insipidus with partial vasopressin response

Congenital nephrogenic diabetes insipidus (NDI) is a rare disease caused most often by mutations in the vasopressin V2 receptor (AVPR2). We studied a family which included a female patient with NDI with symptoms dating from infancy. The patient responded to large doses of desmopressin (dDAVP) which decreased urine volume from 10 to 4 I/day. Neither the parents nor the three sisters were polyuric. The patient was found to be a compound heterozygote for two novel recessive point mutations in the aquaporin-2 (AQP2) gene: L22V in exon 1 and C181W in exon 3. Residue Cys181 in AQP2 is the site for inhibition of water permeation by mercurial compounds and is located near to the NPA motif conserved in all aquaporins. Osmotic water permeability (Pf) in Xenopus oocytes injected with cRNA encoding C181W-AQP2 was not increased over water control, while expression of L22V cRNA increased the Pf to approximately 60% of that for wild-type AQP2. Co-injection of the mutant cRNAs with the wild-type cRNA did not affect the function of the wild-type AQP2. Immunolocalization of AQP2-transfected CHO cells showed that the C181W mutant had an endoplasmic reticulum-like intracellular distribution, whereas L22V and wild-type AQP2 showed endosome and plasma membrane staining. Water permeability assays showed a high Pf in cells expressing wild-type and L22V AQP2. This study indicates that AQP2 mutations can confer partially responsive NDI.      

Reciprocal effect of Waardenburg syndrome mutations on DNA binding by the Pax-3 paired domain and homeodomain

The Pax-3 protein contains two DNA-binding domains, a paired domain and a homeodomain. Mutations in Pax-3 cause Waardenburg syndrome (WS) in humans and the mouse Splotch (Sp) phenotype. In the Sp-delayed mouse, a mutation in the Pax-3 paired domain (G9R) abrogates the DNA-binding activity of both the paired domain and the homeodomain, suggesting that they may functionally interact. To investigate this possibility further, we have analyzed the DNA-binding properties of additional point mutants in the Pax-3 paired domain and homeodomain that occur in WS patients (F12L, N14H, G15S, P17L, R23L, G48A, S51F and G66D in the paired domain, V47F and R53G in the homeodomain), the Pax-1 un mutation (G15A) and a substitution associated with Peters' anomaly in the PAX-6 gene (R23G). Within the paired domain, seven of 10 mutations were found to abrogate DNA-binding by the paired domain. Remarkably, these seven mutations also affected DNA binding by the homeodomain, causing either a complete loss (P17L and G66D), a reduction (R23G, R23L, G15S and G15A) or an increase in DNA-binding activity (N14H). In addition, the effect of paired domain mutations occurred at the level of monomer formation by the homeodomain, while the dimerization potential of this domain seemed unaffected in mutants where it could be analyzed. Furthermore, while both homeodomain mutations were found to abolish DNA binding by this domain, the R53G mutation also abrogated DNA binding by the paired domain. The important observation that independent mutations in either domain can affect DNA binding by the other in the intact Pax- 3 protein strongly suggests that the two domains are not functionally independent but bind DNA through cooperative interactions. Modeling the deleterlous mutations on the three-dimensional structure of the paired domain of Drosophila Prd shows that these mutations cluster at the DNA interface, thus suggesting that a series of DNA contacts are essential for DNA binding by both the paired domain and the homeodomain of Pax-3.      

Plasma protein binding of imipramine in patients with rheumatoid arthritis

Summary In 23 patients with rheumatoid arthritis the plasma protein binding of³H-imipramine and the plasma levels of 13 proteins were measured in order to examine the significance of the proteins for the binding of imipramine. The degree of³H-imipramine binding did not differ significantly from that in healthy controls. It was positively correlated with the concentrations of fibrinogen, ₁-acid-glycoprotein, ceruloplasmin, complement C3_c, haptoglobin and hemopexin. Erythrocyte sedimentation rate was also highly positively correlated with binding. The concentration of several of the proteins showed a significant covariation. The³H-imipramine binding was negatively correlated with the concentration of albumin and the latter was negatively correlated with some of the proteins mentioned-above. No correlation with the levels of apolipoproteins A and B was found. There appears to be more a qualitative than a quantitative change in³H-imipramine binding in patients with rheumatoid arthritis.     

PREVALENCE OF MICROALBUMINURIA IN NON-INSULIN-DEPENDENT DIABETES MELLITUS

The prevalence of microalbuminuria was assessed in 50 patients of non-insulin dependent diabetes mellitus. The mean age of patients was 52.1 ± 11.6 years and the duration of diabetes was 8.3 ± 6.8 years. Twenty (40%) patients had microalbuminuria. Microalbuminuria was more common in patients with a longer duration of diabetes (more than 5 years), a poor glycaemic control, and higher systolic blood pressure.KEY WORDS: Microalbuminuria, Diabetes mellitus, Diabetic nephropathy, Chronic renal failure     

Spinal gas collection demonstrated at CT.

In 234 consecutive CT examinations of the lumbar spine, gas collection was observed in 4 cases with disk herniation, and in 6 cases of disk protrusion. In 3 cases free gas was found in the epidural space, and one patient presented an intraspinal gas-filled "bleb". Gas collection in intervertebral disk spaces and facet joints was found in a total of 60 patients. The CT findings and surgical results were compared to determine whether gas collection contributes to clinical symptoms. In most cases the presence of gas was not clinically important, but in one patient it presented as a spinal mass, causing pain and radiculopathy.     

Impact of work environment on cardiovascular diseases in Denmark.

STUDY OBJECTIVE--The aim was to estimate the quantitative impact of working conditions on cardiovascular diseases in Denmark. DESIGN--The study was based on a review of recent epidemiological research publications in which relative risks were estimated and risk factor prevalences were determined. The impact of working conditions was quantified by means of aetiological fractions. SETTING--The aetiological fractions were estimated on the Danish population. MAIN RESULTS--16% of the premature cardiovascular mortality in men and 22% in occupationally active women is avoidable through interventions in the work environment. If "sedentary" work is included in the occupational risk factors, the aetiological fractions reach 51% for men and 55% for women. Taken separately, the major aetiological fractions for cardiovascular risk factors at work are respectively (women in parentheses) 6% (14%) for monotonous high paced work, 7% (7%) for shift work, and 2% (2%) for passive smoking. CONCLUSIONS--The aetiological fractions show that working conditions play a considerable role in cardiovascular diseases. Furthermore they might widen the focus of preventive cardiology from interventions directed only at individual and lifestyle risk factors to interventions directed also at working conditions.     

Portal recovery of short-chain fatty acids infused into the temporarily-isolated and washed reticulo-rumen of sheep

The present study was undertaken to study the metabolism of short-chain fatty acids (SCFA) by the reticulo-ruminal epithelium and the portal-drained viscera (PDV) under in vivo conditions with no interference from the metabolism of the rumen microbes. The technique of temporary isolation of the reticulo-rumen was applied to wethers implanted with catheters in a mesenteric artery, the hepatic portal vein and the right ruminal vein. Portal blood flow was measured by downstream dilution of p-aminohippuric acid; the PDV uptake of arterial acetate, as well as the whole-body irreversible loss rate (ILR) of acetate, was estimated by [2-(13)C]acetate infusion into the right ruminal vein. The sheep were maintained with a bicarbonate-buffered solution of SCFA in the reticulo-rumen along with continuous intraruminal infusion of SCFA for 4 h. The portal appearance of SCFA of non-reticulo-ruminal origin was estimated before and after the infusion protocol. Of the acetate absorbed by the sheep, 89 (SE 5), 109 (SE 7) and 101 (SE 7)% was recovered as portal net appearance of acetate, portal net appearance of acetate corrected for PDV uptake of arterial acetate and increase in the ILR of acetate respectively. Of the propionate, isobutyrate, butyrate, isovalerate and valerate absorbed by the sheep, 95 (SE 7), 102 (SE 9), 23 (SE 3), 48 (SE 5) and 32 (SE 4)% respectively was recovered as portal net appearance. In contrast to current concepts, the present study showed that the reticulo-ruminal epithelium metabolizes none (or only a small proportion) of the acetate and propionate absorbed from the rumen. This observation could lead to the more efficient use of results obtained with multi-catheterized animals to quantify the net metabolite output of the rumen microbes.