Atherothrombosis is a multifactorial disease and risk factors for atherothrombosis are now estimated to be in the hundreds. In addition to classical risk factors like serum lipids (total cholesterol and triglycerides), lipoproteins, and apolipoproteins, a variety of inflammatory molecules such as C-reactive protein and serum amyloid A, hemostatic factors and various specific cell types have been also considered for risk and/or require management. Studying the cardiovascular lipid profile of patients with rheumatoid arthritis (RA), we observed diagnostically important laboratory interferences and studied their nature and source. Initially, we attempted to quantitate lipoprotein-cholesterol fractions in previously frozen heparinized plasma of these patients using an electrophoretic enzymatic cholesterol method (REP Vis Cholesterol, Helena). However, our attempt has failed because of incomplete separation of lipoproteins in all 54 specimens tested, apparently due to the phenomenon of cryogelation. Cryogel is a co-precipitate of extra domain A-containing (cellular) fibronectin, plasma fibronectin, fibrinogen and heparin. Cryogelation has been described to occur in the presence of heparin at low temperatures and has been found to be common in patients with RA. Our failure with the electrophoretic lipoprotein-cholesterol test prompted studying possible interference of cryogelation with other laboratory tests. Serum and heparinized plasma were prepared from 6 RA patients using different blood collection and blood processing/storage protocols. These protocols included preparation of serum from separator gel tube and preparation of heparinized and EDTA plasma at room temperature and at 37 oC and storage of the specimens in refrigerator or frozen at -70 oC prior to laboratory testing. A panel of 20 routine chemistry tests and tests for total cholesterol (TC), triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) were performed using the Synchron LX-20 automated chemistry analyzer (Beckman Coulter). Select sera and plasmas were also analyzed for troponin I, thyroid-stimulating hormone (TSH), adrenocorticotropic hormone or ACTH and cortisol (Immulite 2500, DPC/Siemens), troponin T, pro-brain natriuretic peptide (BNP), creatine kine-MB or CK-MB isoenzyme, and beta-human chorionic gonadotropin or beta-HCG (Elecsys 2010, Beckman Coulter), brain natriuretic peptide or BNP (Axsym, Abbott), high-sensitivity C-reactive protein or hsCRP and rheumotaid factor or RF (Immage, Beckman Coulter), serum amyloid A or SAA (BN-II, Dade-Behring), and electrophoretic protein fractions (Hydragel, Sebia). The presence of visible cryogel in heparinized plasma specimens from the heparinized plasma specimens necessitated high-speed centrifugation prior to testing. Apart from expected differences between serum and plasma for potassium and total protein, the results for most other routine chemistry tests, TC, TG, various immunoassays and protein electrophoresis were insignificantly different, either statistically or clinically, among various protocols. Compared to the reference serum specimens (serum processed and stored at 4 oC), all refrigerated or frozen heparinized plasma specimens gave higher (8%) HDL-C results and tended to give lower (4%) LDL-C results; with more pronounced effects for heparinized plasma specimens stored and processed at 37oC. It appears that, apart from lipoprotein fraction testing, even refrigerated or previously frozen heparinized plasma samples are suitable for most routine and specialized chemistry tests after removal of the cryogel that may cause sampling problems. ? In a collaborative study, we found that cardiac rehabilitation in patients receiving stable statin therapy and with low-density lipoprotein cholesterol at goal increases endothelial progenitor cell number, endothelial progenitor cell survival, and endothelial differentiation potential, associated with increased nitric oxide in the blood. Although this response was observed in most patients, a significant minority showed neither endothelial progenitor cell mobilization nor increased nitric oxide in the blood. In another collaborative study, we found that both serum SAA and CRP are significantly elevated in unmedicated, remitted women with major depressive disorder (MDD) versus controls. These findings suggested a sustained low-grade pro-inflammatory state in women with MDD that may be related to the increased coronary artery disease (CAD) risk observed previously in this population. In still another collaborative study, we studied the ability of the rodent (mouse) and human scavenger receptors SR-BI (called CLA-1 in humans) and its splicing variant SRB-II (called CLA-2 in humans) to mediate bacterial adhesion and cytosolic bacterial invasion in mammalian cells. Our results indicate that these receptors, in addition to their already established function as high-density lipoprotein receptors, also are involved in the defense mechanism of mammalian host cells and, by facilitating bacterial adhesion and cytosolic invasion, they may play an important role in infection and sepsis.
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