Angiotensin I converting enzyme and kinin–hydrolyzing enzymes along the rabbit nephron

Angiotensin I converting enzyme and kinin–hydrolyzing enzymes along the rabbit nephron. Angiotensin I converting enzyme (ACE) and kininase activities were measured in various segments of the rabbit nephron. ACE was determined with tritiated hippuryl-glycylglycine as substrate. Lysyl-bradykinin (LBK) hydrolysis (kininase activity) was measured by radioimmunoassay. ACE was only found in the glomerulus and in the two parts of proximal tubule: the convoluted proximal tubule and the pars recta (PR). It was distributed along a concentration gradient which increased from the glomerulus to PR. Kininase activity was found in both proximal and distal parts of the nephron. Besides intense LBK-hydrolyzing activity in the proximal tubule, a kininase activity was also found in the medullary collecting tubule (MCT). Kininase activity in the glomerulus and the proximal tubule was completely inhibited by chelating agents. Captopril inhibited this activity only in the PR and at high concentrations (above 10-7 M). These results indicate that several types of enzymes other than ACE hydrolyze kinins in the glomerulus and in the proximal tubule. The contribution of ACE to kinin hydrolysis appears only minimal. The kininase activity found in MCT was different from ACE and other proximal tubule kininases because it was not inhibited by chelating agents. This kininase may play a physiological role in inactivating the kinins formed by kallikrein at or beyond the connecting tubule

Novel splice site mutation in the PROS1 gene in a Polish patient with venous thromboembolism : c.602-2delA, splice acceptor site of exon 7

We identified a novel splice site mutation of the PROS1 gene in a Polish family with protein S (PS) deficiency and explored the molecular pathogenesis of this previously undescribed variant. A novel mutation was detected in a 26-year-old woman with a history of venous thromboembolism (VTE) provoked by oral contraceptives. Her family history of VTE was positive. The sequence analysis of the PROS1 gene was performed in the proband and the proband’s family. The proband and their asymptomatic father had lower free PS levels (45% and 50%, respectively) and PS activity (48% and 44%, respectively). Total PS levels were normal (65.6% and 62.4%, respectively). The sequence analysis of the PROS1 gene revealed the presence of heterozygous deletion at the nucleotide position c.602-2 in intron 6, just upstream of exon 7, detected in the proband and her father. This variant alters the splice acceptor site of exon 7, and, according to the in silico prediction, it is highly likely to cause in-frame exon 7 skipping. We also presented follow-up data of two other Polish patients with PS deficiency associated with splice site mutations in PROS1 gene

0052: Role of kinins in diabetic wound healing

The diabetic foot is associated with pain, decrease in patient's quality of life, considerable costs, and amputation. In this study, we determined the role of KKS, via activation of bradykinin receptors (B1R or B2R), in a mouse model of diabetic wound healing. Diabetic or nondiabetic mice are wounded with an 8-mm punch biopsy and then are treated or not with specific B1R or B2R agonists (720nmol/kg.d-1) and/or B2R antagonist (Icatibant, 500μg/kg.dg/d-1). The wound-healing surface was daily followed up. At 11 days, the scar were analysed by histology (Masson's trichrome staining) and B1R and B2R expression were assessed (RT-qPCR). Effects of the agonists on cells (fibroblasts and keratinocytes) migration and proliferation were also analysed. In diabetic condition, mRNA of B1R and B2R was increased in skin (p<0.01). B1R activation had no effect on wound closure in our model. In contrast, B2R activation dramatically delayed wound healing in diabetic (p<0.001) or nondiabetic (p<0.01) mice. Histological analysis of scar showed significant skin disorganization and epidermis thickening with B2R agonist (p<0.05). In vitro, B2R agonist induced an increase of keratinocyte proliferation (+46% after 48h, p<0.01) and a stimulation of keratinocyte migration (+30% after 24h, p<0.05). These effects was associated with ERK phosphorylation which occurs downstream of EGFR activation (p<0.05). B2R agonist had no effect on fibroblast migration but decreased fibroblast proliferation (–33% after 48h, p<0.05). Co-treatment with Icatibant abrogated in vivo and in vitro effects observed with B2R agonist. Moreover, Icatibant alone hastened wound healing and decrease the epidermis thickening induced by diabetes. In conclusion, KKS, through the B2R but not the B1R, plays a critical role in proliferation and remodelling phases of skin wound healing in mice. While more studies are needed, Icatibant could be used to correct the diabetic wound healing defect

Genetic characterization of antithrombin, protein C and protein S deficiencies in Polish patients

Inherited deficiencies of natural anticoagulants such as antithrombin (AT; gene: SERPINC1), protein C (PC; PROC), and protein S (PS; PROS1), with the prevalence in the general European population of 0.02% to 0.17%, 0.2% to 0.3%, and 0.5%, respectively, are associated with increased risk of thromboembolic events. Only a few case reports of Polish deficient patients with known causal mutations have been published so far. The aim of the study was to characterize the frequency of SERPINC1, PROC, and PROS1 mutations and their thromboembolic manifestations in patients with AT, PC, or PS deficiencies, inhabiting southern Poland. Ninety unrelated patients (mean [SD] age, 40.1 [13.2] years) with AT (n = 35), PC (n = 28), or PS (n = 27) deficiencies, with a history of venous 73 (81%) or arterial 17 (19%) thromboembolism, were screened for mutations using the Sanger sequencing or multiplex ligation‑dependent probe amplification. Twenty mutations (29%) described here were new, mostly in the SERPINC1 and PROC genes. Missense mutations accounted for 84% of all mutations in the PROC gene and approximately 50% of those in the SERPINC1 and PROS1 genes. In all 3 genes, the ratio of nonsense and splice-site mutations was 8% to 31% and 8% to 23%, respectively. The mutation detection rate was 90% for AT or PC when anticoagulant activity was below 70%, while for the PROS1 gene, the rate reached 80% at the free PS levels below 40%. To our knowledge, this is the largest cohort of Polish patients deficient in natural anticoagulants and evaluated for the causal genetic background. Several new Polish detrimental mutations were detected, mostly in AT- and PC‑deficient patients

Prognostic Value of the Insertion/Deletion Polymorphism of the ACE Gene in Type 2 Diabetic Subjects: Results from the Non-Insulin-Dependent Diabetes, Hypertension, Microalbuminuria or Proteinuria, Cardiovascular Events, and Ramipril (DIABHYCAR), Diabete de type 2, Nephropathie et Genetique (DIAB2NEPHROGENE), and Survie, Diabete de type 2 et Genetique (SURDIAGENE) studies

OBJECTIVE—We tested whether determination of the ACE insertion/deletion polymorphism is useful for renal and cardiovascular prognoses of type 2 diabetic subjects

Multilevel analysis of systolic blood pressure and ACE gene I/D polymorphism in 438 Swedish families – a public health perspective

BACKGROUND: Individuals belonging to the same family share a number of genetic as well as environmental circumstances that may condition a common SBP level. Among the genetic factors, the angiotensin converting enzyme (ACE) gene I/D polymorphism appears as a possible candidate as it might influence both SBP and the pharmacological effect of ACE inhibitors. We aimed to combine genetic epidemiology with public health ideas concerning life-course and multilevel epidemiology in order to understand the role of familial factors regarding individual SBP. METHODS: We applied multilevel regression analysis on 1926 individuals nested within 438 families from South Sweden. Modelling familial SBP variance as a function of age and use of ACE inhibitors we calculates a variance partition coefficient and the proportional change in familial SBP variance attributable to differences in ACE gene I/D polymorphism RESULTS: Our results suggest the existence of genetic or environmental circumstances that produce a considerable familial clustering of SBP, especially among individuals using ACE-inhibitors. However, ACE gene I/D polymorphism seems to play a minor role in this context. In addition, familial factors – genetic, environmental or their interaction – shape SBP among non-users of ACE inhibitors but their effect is expressed later in the life-course. CONCLUSION: Strategies directed to prevent hypertension should be launched in younger rather than in older ages and both prevention of hypertension and its treatment with ACE inhibitors should be focused on families rather than on individuals