Inactivation of endothelial proprotein convertase 5/6 decreases collagen deposition in the cardiovascular system: role of fibroblast autophagy

Proprotein convertase (PC) 5/6 belongs to a family of secretory proteases involved in proprotein proteolysis. Several studies suggest a role for PC5/6 in cardiovascular disease. Because lethality at birth of mice lacking PC5/6 precluded elucidation of its function in the adult, we generated mice in which the gene of PC5/6 (pcsk5) is specifically inactivated in endothelial cells (ecKO), which are viable and do not exhibit overt abnormalities. In order to uncover the function of PC5/6 in the cardiovascular system, the effect of ecKO was studied in aging mice. In 16 to 18-month-old ecKO mice, the left ventricle (LV) mass, media cross-sectional area of aorta and coronary arteries, and media-to-lumen ratio of mesenteric arteries were decreased. The LV presented decreased diastolic function, and mesenteric arteries showed decreased stiffness. Collagen was decreased in the LV myocardial interstitium and perivascularly in coronary arteries and aorta. Cardiovascular hypotrophy likely develops with aging, since no significant changes were observed in 2-month-old ecKO mice. Fibroblasts, as a source of collagen in myocardium and vasculature, may play a role in the decrease in collagen deposition. Fibroblasts co-cultured with ecKO endothelial cells showed decreased collagen production, decreased insulin-like growth factor (IGF)-1/Akt/mTOR signaling, and enhanced autophagic activation. PC5/6 inactivation in endothelial cells results in cardiovascular hypotrophy associated with decreased collagen deposition, decreased LV diastolic function, and vascular stiffness, suggesting a trophic role of endothelial PC5/6 in the cardiovascular system, likely mediated by IGF-1/Akt/mTOR signaling and control of autophagy.     

Arenavirus envelope glycoproteins mimic autoprocessing sites of the cellular proprotein convertase subtilisin kexin isozyme-1/site-1 protease

A crucial step in the arenavirus life cycle is the proteolytic processing of the viral envelope glycoprotein precursor (GPC) by the cellular proprotein convertase (PC) subtilisin kexin isozyme-1 (SKI-1)/site-1 protease (S1P). Here we conducted a systematic and quantitative analysis of SKI-1/S1P processing of peptides derived from the recognition sites of GPCs of different Old World and New World arenaviruses. We found that SKI-1/S1P showed a strong preference for arenaviral sequences resembling its autoprocessing sites, which are recurrent motifs in arenaviral GPCs. The African arenaviruses Lassa, Mobala, and Mopeia resemble the SKI-1/S1P autoprocessing C-site, whereas sequences derived from Clade B New World viruses Junin and Tacaribe have similarities to the autoprocessing B-site. In contrast, analogous peptides derived from cellular SKI-1/S1P substrates were remarkably poor substrates. The data suggest that arenavirus GPCs evolved to mimic SKI-1/S1P autoprocessing sites, likely ensuring efficient cleavage and perhaps avoiding competition with SKI-1/S1P's cellular substrates.     

人类枯草溶菌素转化酶9基因影响胆固醇代谢的研究进展

家族性高胆固醇血症(FH)是一种由低密度脂蛋白受体(LDLR)缺陷所导致的、以血总胆固醇升高为特征的常染色体显性遗传病.目前普遍认为FH是复杂的多基因病.近年研究发现,人类枯草溶菌素转化酶9(PCSK9)在血胆固醇代谢中起重要作用.一些PCSK9突变可减少LDLR总量,进而导致FH.而另一些PCSK9突变影响自身活性,导致低胆固醇血症.本文就PCSK9的基因结构、功能以及该基因不同突变体影响血胆固醇代谢的最新研究进展作一综述.     

Effect of X-ray irradiation on primary mineralization in rat alveolar bone

Summary The effect of X-ray irradiation on the process of primary mineralization in bone was studied by biochemical and ultrastructural methods. A single dose of 1500R was administered to the head region of rats. The animals were examined immediately after irradiation and 1, 2 and 3 weeks later. Fractions of isolated cells and extracellular matrix vesicles were prepared from the maxillary alveolar bone of irradiated and untreated rats by collagenase digestion and differential centrifugation. The protein content and activities of vesicular phosphatases were determined in both fractions. A continuous decrease in the activity of alkaline phosphatase could be observed in both cell and matrix vesicle fractions during a three-week follow up after irradiation. Acid phosphatase activity decreased only in the vesicle fraction.Transmission electron microscopy of irradiated bone tissue revealed that many matrix vesicles were devoid of intact membranes and apatite crystals. Calcifying nodules were abundant in the matrix without their apparent fusion into larger mineralized structures. It is suggested that irradiation interferes with enzymatic processes associated with primary mineralization.     

Single-nucleotide polymorphisms of the proprotein convertase subtilisin/kexin type 5 (PCSK5) gene

The proprotein convertase, subtilisin/kexin type 5, or PCSK5, mediates post-translational endoproteolytic processing for several integrin α subunits. We identified two silent single-nucleotide polymorphisms (SNPs) in PCSK5, which were found to vary in frequency across ethnic groups. The identification of these amplification primers and SNPs provides tools to investigate PCSK5 for association with inflammatory or vascular phenotypes. Received: July 23, 2001 / Accepted: August 28, 2001     

VACTERL/caudal regression/Currarino syndrome-like malformations in mice with mutation in the proprotein convertase Pcsk5

We have identified an ethylnitrosourea (ENU)-induced recessive mouse mutation (Vcc) with a pleiotropic phenotype that includes cardiac, tracheoesophageal, anorectal, anteroposterior patterning defects, exomphalos, hindlimb hypoplasia, a presacral mass, renal and palatal agenesis, and pulmonary hypoplasia. It results from a C470R mutation in the proprotein convertase PCSK5 (PC5/6). Compound mutants (Pcsk5(Vcc/null)) completely recapitulate the Pcsk5(Vcc/Vcc) phenotype, as does an epiblast-specific conditional deletion of Pcsk5. The C470R mutation ablates a disulfide bond in the P domain, and blocks export from the endoplasmic reticulum and proprotein convertase activity. We show that GDF11 is cleaved and activated by PCSK5A, but not by PCSK5A-C470R, and that Gdf11-deficient embryos, in addition to having anteroposterior patterning defects and renal and palatal agenesis, also have a presacral mass, anorectal malformation, and exomphalos. Pcsk5 mutation results in abnormal expression of several paralogous Hox genes (Hoxa, Hoxc, and Hoxd), and of Mnx1 (Hlxb9). These include known Gdf11 targets, and are necessary for caudal embryo development. We identified nonsynonymous mutations in PCSK5 in patients with VACTERL (vertebral, anorectal, cardiac, tracheoesophageal, renal, limb malformation OMIM 192350) and caudal regression syndrome, the phenotypic features of which resemble the mouse mutation. We propose that Pcsk5, at least in part via GDF11, coordinately regulates caudal Hox paralogs, to control anteroposterior patterning, nephrogenesis, skeletal, and anorectal development.     

Case reports of proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibition nonresponse

Proprotein convertase subtilisin/kexin type 9 (PCSK9) inhibitors, a novel class of monoclonal antibodies, reduces low-density lipoprotein cholesterol levels and improves cardiovascular outcomes. Given the short time frame, these agents have been available for use; reports of nonresponse to the PCSK9 inhibitor therapy are scarce in literature. We describe 2 cases with substantially lesser than expected low-density lipoprotein cholesterol lowering on PCSK9 therapy. Nonresponse to PCSK9 inhibition was attributed to autosomal recessive hypercholesterolemia (secondary to low-density lipoprotein receptor adaptor protein 1 mutation) and plasmapheresis after PCSK9 inhibitor drug injections. Additional PCSK9 inhibitor nonresponders are likely to emerge as the use of these agents increases overtime.     

Mutations and polymorphisms in the proprotein convertase subtilisin kexin 9 (PCSK9) gene in cholesterol metabolism and disease

Hypercholesterolemia is one of the major causes of coronary heart disease (CHD). The genes encoding the low‐density lipoprotein receptor and its ligand apolipoprotein B, have been the two genes classically implicated in autosomal dominant hypercholesterolemia (ADH). Our discovery in 2003 of the first mutations of the proprotein convertase subtilisin kexin 9 gene (PCSK9) causing ADH shed light on an unknown actor in cholesterol metabolism that since then has been extensively investigated. Several PCSK9 variants have been identified, some of them are gain‐of‐function mutations causing hypercholesterolemia by a reduction of low‐density lipoprotein (LDL) receptor levels; while others are loss‐of‐function variants associated with a reduction of LDL‐cholesterol (LDL‐C) levels and a decreased risk of CHD. In this review, we focus on reported variants, and their biological, clinical, and functional relevance. We also highlight the spectrum of hypercholesterolemia or hypobetalipoproteinemia phenotypes that are already associated with mutations in PCSK9. Finally, we present future prospects concerning this therapeutic target that might constitute a new approach to reduce cholesterol levels and CHD, and enhance the effectiveness of other lipid‐lowering drugs. Hum Mutat 0, 1–11, 2008. © 2008 Wiley‐Liss, Inc.     

初级纤毛在骨组织力学信号传导中的作用

骨组织不断接受力学刺激并保持骨形成和骨吸收的动态平衡。目前,人们仍不清楚骨组织如何感知力学刺激。越来越多的研究表明初级纤毛可能是骨组织力学刺激感受器,并将细胞外的力学刺激信号转化为细胞内的生化信息,最终骨组织实现其结构的重塑。在此,本文将对初级纤毛研究现状进行综述,并预测初级纤毛未来的研究趋势, 为利用初级纤毛来防治骨质疏松症打下基础。     

The proprotein convertase PC2 is involved in the maturation of prosomatostatin to somatostatin-14 but not in the somatostatin deficit in Alzheimer's disease

A somatostatin deficit occurs in the cerebral cortex of Alzheimer's disease patients without a major loss in somatostatin-containing neurons. This deficit could be related to a reduction in the rate of proteolytic processing of peptide precursors. Since the two proprotein convertases (PC)1 and PC2 are responsible for the processing of neuropeptide precursors directed to the regulated secretory pathway, we examined whether they are involved first in the proteolytic processing of prosomatostatin in mouse and human brain and secondly in somatostatin defect associated with Alzheimer's disease. By size exclusion chromatography, the cleavage of prosomatostatin to somatostatin-14 is almost totally abolished in the cortex of PC2 null mice, while the proportions of prosomatostatin and somatostatin-28 are increased. By immunohistochemistry, PC1 and PC2 were localized in many neuronal elements in human frontal and temporal cortex. The convertases levels were quantified by Western blot, as well as the protein 7B2 which is required for the production of active PC2. No significant change in PC1 levels was observed in Alzheimer's disease. In contrast, a marked decrease in the ratio of the PC2 precursor to the total enzymatic pool was observed in the frontal cortex of Alzheimer patients. This decrease coincides with an increase in the binding protein 7B2. However, the content and enzymatic activity of the PC2 mature form were similar in Alzheimer patients and controls. Therefore, the cortical somatostatin defect is not due to convertase alteration occuring during Alzheimer's disease. Further studies will be needed to assess the mechanisms involved in somatostatin deficiency in Alzheimer's disease.     

The relationship between extracellular matrix vesicles and calcospherites in primary mineralization of neoplastic bone tissue

Summary Primary mineralization in neoplastic tissue was studied in osteosarcoma, correlating observations obtained by SEM to those found with TEM.The process is characterized by extracellular matrix vesicles, distributed in the matrix between the forming neoplastic cells and the calcifying fronts. The occurrence of osmiophilic material and solitary hydroxyapatite crystals within the vesicles is followed by accumulation of apatite crystals, disappearance of the vesicular membrane and formation of calcospherites and calcified fronts. The process described here in neoplastic tissue is essentially similar to primary calcification in normal calcified tissues.     

A recidivant primary cardiac osteosarcoma: the role of bone scans

Primary cardiac tumors are infrequent, less than 15–20% are malignant, and most of them are sarcomas. Primary recidivant cardiac osteosarcomas are extremely rare, only a few cases have been reported, and the prognosis is ominous. We report a case of a primary cardiac osteosarcoma in a 70-year-old woman who was admitted to the hospital for evaluation of congestive heart failure. Despite the wide resection of the tumor, a local and metastatic recurrence was diagnosed. In this report, we illustrate the utility of image techniques for the diagnosis and the monitoring of primary cardiac tumors, especially the role of bone scintigraphy. This technique is not a routine procedure for the cardiologist, but it has been very useful in this case in order to decide the optimal treatment.     

Association of the proprotein convertase subtilisin/kexin-type 2 (PCSK2) gene with type 2 diabetes in an African American population

In a genome-wide scan for type 2 diabetes (T2DM) in African American (AA) families, ordered subsets analysis (OSA) provided evidence for linkage to chromosome 20p in a subset with later age at diagnosis (max LOD 2.57, P=0.008). The proprotein convertase subtilisin/kexin-type 2 (PCSK2) gene is within the LOD-1 interval of this linkage peak. Twenty-nine single nucleotide polymorphisms (SNPs) were genotyped across this gene in 380 unrelated AA individuals with T2DM and end-stage renal disease (T2DM-ESRD), 278 AA controls, 96 European Americans (EA) and 120 Yoruba Nigerian (YRI) controls. In addition, 22 ancestry-informative markers (AIMs) were genotyped in all AA subjects, 120 YRI, and 282 EA controls. ADMIXMAP was used to model the distributions of admixture and generate score tests of allelic and haplotypic association. Association with T2DM was observed among 4 SNPs: rs2021785 (admixture-adjusted Pa=0.00014), rs1609659 (Pa=0.028), rs4814597 (Pa=0.039) and rs2269023 (Pa=0.043). None of the PCSK2 SNPs were associated with age at T2DM diagnosis. A variant in the PCKS2 gene, rs2021785, appears to play a role in susceptibility to T2DM in this AA population.     

Changes in bone remodeling rate influence the degree of mineralization of bone

Mineralization of bone matrix implies two successive steps, a primary mineralization on the calcification front followed by a slow process of secondary mineralization progressively adding about 50-60% of the mineral content on bone matrix. Our model is that antiresorptive agents prolong the lifetime of the basic structural units (BSUs) by causing a marked reduction in the birth rate of basic multicellular units (BMUs), and increase the degree of mineralization of bone (DMB) by allowing a more complete secondary mineralization. Conversely, agents or events provoking an augmentation of the birthrate of BMUs and a decrease of the lifetime of BSUs lead to resorption of new BSUs before they have fully completed their secondary mineralization, leading to the presence of incompletely mineralized BSUs and a low mean DMB value, as measured by quantitative microradiography. Measurements of DMB (distribution and mean value) under circumstances with various remodeling activities favor our model. In postmenopausal osteoporotic women treated during 2 or 3 years with alendronate (10 mg/day), an increase of the mean DMB of approximately 7 to 10% was found, due to a marked reduction in the bone remodeling. In contrast, an activation of bone remodeling as in primary hyperparathyroidism lowered the mean DMB.     

Novel association to the proprotein convertase PCSK7 gene locus revealed by analysing soluble transferrin receptor (sTfR) levels

The level of body iron storage and the erythropoietic need for iron are indicated by the serum levels of ferritin and soluble transferrin receptor (sTfR), respectively. A meta-analysis of five genome-wide association studies on sTfR and ferritin revealed novel association to the PCSK7 and TMPRSS6 loci for sTfR and the HFE locus for both parameters. The PCSK7 association was the most significant (rs236918, P = 1.1 × 10E-27) suggesting that proprotein convertase 7, the gene product of PCSK7, may be involved in sTfR generation and/or iron homeostasis. Conditioning the sTfR analyses on transferrin saturation abolished the HFE signal and substantially diminished the TMPRSS6 signal while the PCSK7 association was unaffected, suggesting that the former may be mediated by transferrin saturation whereas the PCSK7-associated effect on sTfR generation appears to be more direct.     

Temporospatially regulated expression of subtilisin-like proprotein convertase PACE4 (SPC4) during development of the rat submandibular gland.

The temporospatial expression of PACE4, a member of the mammalian subtilisin-like proprotein convertase family involved in the activation of growth/differentiation factors, was investigated by in situ hybridization during the development of the rat submandibular gland (SMG). At the initiation stage (day 15.5 of gestation; E15), PACE4 was intensely expressed in the submandibular epithelium, but weakly expressed in the mesenchymal cells. At E16 when the branching morphogenesis becomes obvious, the expression of PACE4 in the mesenchyme was further decreased, although its level in the submandibular epithelium had not changed remarkably from that at E15. During the next stage of embryonic development (E17-E20), PACE4 was expressed in the cells derived from the submandibular epithelium, which include the proacinar, terminal tubular, and presumptive ductal cells. In the perinatal SMG, PACE4 was still expressed intensely in the terminal portion of the SMG containing the proacinar and terminal tubular cells, whereas its expression in the ductal cells was obviously decreased at the second postnatal day (P2) and at P6. Acinar cells expressing no PACE4 appeared, and their numbers increased following their development (P9-P20). At P30 when the PACE4 expression in the acinar cells was completely suppressed, its expression in the ductal cells became intense again. This temporospatially regulated expression of PACE4 suggests its apparent association with the proliferation, differentiation, and establishment of functional acinar and ductal cells of the SMG.