Lysyl oxidase catalyzes the final enzymatic step required for collagen and elastin cross-linking in extracellular matrix biosynthesis. Pro-lysyl oxidase is processed by procollagen C-proteinase activity, which also removes the C-propeptides of procollagens I-III. The Bmp1 gene encodes two procollagen C-proteinases: bone morphogenetic protein 1 (BMP-1) and mammalian Tolloid (mTLD). Mammalian Tolloid-like (mTLL)-1 and -2 are two genetically distinct BMP-1-related proteinases, and mTLL-1 has been shown to have procollagen C-proteinase activity. The present study is the first to directly compare pro-lysyl oxidase processing by these four related proteinases. In vitro assays with purified recombinant enzymes show that all four proteinases productively cleave pro-lysyl oxidase at the correct physiological site but that BMP-1 is 3-, 15-, and 20-fold more efficient than mTLL-1, mTLL-2, and mTLD, respectively. To more directly assess the roles of BMP-1 and mTLL-1 in lysyl oxidase activation by connective tissue cells, fibroblasts cultured from Bmp1-null, Tll1-null, and Bmp1/Tll1 double null mouse embryos, thus lacking BMP-1/mTLD, mTLL-1, or all three enzymes, respectively, were assayed for lysyl oxidase enzyme activity and for accumulation of pro-lysyl oxidase and mature ϳ30-kDa lysyl oxidase. Wild type cells or cells singly null for Bmp1 or Tll1 all produced both pro-lysyl oxidase and processed lysyl oxidase at similar levels, indicating apparently normal levels of processing, consistent with enzyme activity data. In contrast, double null Bmp1/Tll1 cells produced predominantly unprocessed 50-kDa prolysyl oxidase and had lysyl oxidase enzyme activity diminished by 70% compared with wild type, Bmp1-null, and Tll1-null cells. Thus, the combination of BMP-1/ mTLD and mTLL-1 is shown to be responsible for the majority of processing leading to activation of lysyl oxidase by murine embryonic fibroblasts, whereas in vitro studies identify pro-lysyl oxidase as the first known substrate for mTLL-2.
Differentiation of phenotypically normal osteoblast cultures leads to formation of a bone-like extracellular matrix in vitro. Maximum collagen synthesis occurs early in the life of these cultures, whereas insoluble collagen deposition occurs later and is accompanied by a diminished rate of collagen synthesis. The mechanisms that control collagen deposition seem likely to include regulation of extracellular collagen biosynthetic enzymes, but expression patterns of these enzymes in differentiating osteoblasts has received little attention. The present study determined the regulation of lysyl oxidase as a function of differentiation of phenotypically normal murine MC3T3-E1 cells at the level of RNA and protein expression and enzyme activity. In addition, the regulation of BMP-1/mTLD mRNA levels that encodes procollagen C-proteinases was assayed. The role of lysyl oxidase in controlling insoluble collagen accumulation was further investigated in inhibition studies utilizing beta-aminopropionitrile, a specific inhibitor of lysyl oxidase enzyme activity. Results indicate that lysyl oxidase is regulated as a function of differentiation of MC3T3-E1 cells, and that the maximum increase in lysyl oxidase activity precedes the most efficient phase of insoluble collagen accumulation. By contrast BMP-1/mTLD is more constitutively expressed. Inhibition of lysyl oxidase in these cultures increases the accumulation of abnormal collagen fibrils, as determined by solubility studies and by electron microscopy. Taken together, these data support that regulation of lysyl oxidase activity plays a key role in the control of collagen deposition by osteoblast cultures.
Little is known regarding the profiles of whole body donors in Muslim majority countries where donation is scarce. Therefore, this study aims to profile registered donors in Turkey by means of a survey. The explored data could be used to improve ongoing campaign efforts and ethical practices such as commemoration services. Registered donors of the donation programs at the two faculties of medicine of Istanbul University were compared with the national population and a cluster analysis was performed to reveal any concealed sub‐groups. Data from 188 respondents were analyzed. The majority of registered donors were married (42%), male (65.4%), aged over 50 years (76%), held a tertiary education degree (49.7%), and were irreligious (58.5%). Cluster analysis revealed two groups with significantly different educational levels, marital statuses, and religious choices. Regarding whether their bodies could be used for education or research, the majority (64.5%) of the respondents left the decision to the anatomy department. Similarly, 73.8% approved indefinite use of their organs, body parts and/or skeletons. The respondents were also willing to share their medical history (94.2%) and personal information (81.6%) if needed. Motivational themes for body donation including usefulness, impermanence, religion, awareness, and kinship were devised after a thematic analysis. Among the respondents, 56.5% were registered organ donors and 63.3% were frequent blood donors. The results of this study provide data that may help revising informed consent forms, developing and implementing thanksgiving ceremonies, and selecting additional targets for supporting body donation campaign activities such as organ and blood donation units.
The LFCN may emerge from the lateral border of the psoas major above or below the IC. The AC/AB ratio can help surgeons to find the LFCN in patients with different body types.
Maximum collagen synthesis and maximum accumulation of insoluble collagen occur at different phenotypic stages in developing osteoblastic cell cultures. Insoluble collagen accumulation depends in part on the activity of extracellular enzymes including procollagen N-proteinases, procollagen C-proteinase (derived from the BMP1 gene), and lysyl oxidase. In addition to its action on procollagen, procollagen C-proteinase processes prolysyl oxidase to mature 32-kDa lysyl oxidase. The regulation of extracellular activities that control insoluble collagen accumulation has not been studied extensively. The present study compares molecular events that control production of a collagenous mineralized extracellular matrix in vitro among five different murine osteosarcoma cell clones derived from the same tumor, but which differ in their ability to produce an insoluble mineralized matrix. Levels of insoluble type I collagen, insoluble calcium, bone morphogenetic protein 1 (BMP-1), and lysyl oxidase expression, lysyl oxidase biosynthesis, lysyl oxidase activity, and prolysyl oxidase processing activity were determined. Results surprisingly indicate that lysyl oxidase activity is not related closely to lysyl oxidase messenger RNA (mRNA) levels among the different cell clones. However, it appears that BMP-1-dependent prolysyl oxidase processing could contribute to the observed lysyl oxidase activity. Highest collagen and BMP-1 mRNA levels, prolysyl oxidase processing activity, and lysyl oxidase activity occurred in a cell clone (K8) that showed the highest levels of insoluble collagen accumulation. Culture media from a cell clone (K37) that accumulates little insoluble collagen or calcium but expresses high levels of lysyl oxidase mRNA contained low molecular weight fragments of lysyl oxidase protein and showed low lysyl oxidase activity. By contrast the K14 cell line exhibits relatively high lysyl oxidase activity and collagen accumulation, but low levels of mature lysyl oxidase protein.
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