Long-term effects of the Diabetes Prevention Program interventions on cardiovascular risk factors: a report from the DPP Outcomes Study.

AIMS: Whether long-term cardiovascular risk is reduced by the Diabetes Prevention Program interventions is unknown. The aim of this study was to determine the long-term differences in cardiovascular disease risk factors and the use of lipid and blood pressure medications by the original Diabetes Prevention Program intervention group. METHODS: This long-term follow-up (median 10 years, interquartile range 9.0-10.5) of the three-arm Diabetes Prevention Program randomized controlled clinical trial (metformin, intensive lifestyle and placebo), performed on 2766 (88%) of the Diabetes Prevention Program participants (who originally had impaired glucose tolerance), comprised a mean of 3.2 years of randomized treatment, approximately 1-year transition (during which all participants were offered intensive lifestyle intervention) and 5 years follow-up (Diabetes Prevention Program Outcomes Study). During the study, participants were followed in their original groups with their clinical care being provided by practitioners outside the research setting. The study determined lipoprotein profiles and blood pressure and medication use annually. RESULTS: After 10 years' follow-up from Diabetes Prevention Program baseline, major reductions were seen for systolic (-2 to -3) and diastolic (-6 to -6.5 mmHg) blood pressure, and for LDL cholesterol (-0.51 to -0.6 mmol/l) and triglycerides (-0.23 to -0.25 mmol/l) in all groups, with no between-group differences. HDL cholesterol also rose significantly (0.14 to 0.15 mmol/l) in all groups. Lipid (P = 0.01) and blood pressure (P = 0.09) medication use, however, were lower for the lifestyle group during the Diabetes Prevention Program Outcomes Study. CONCLUSION: Overall, intensive lifestyle intervention achieved, with less medication, a comparable long-term effect on cardiovascular disease risk factors, to that seen in the metformin and placebo groups.

Bone morphogenetic protein expression in human atherosclerotic lesions.

Artery wall calcification associated with atherosclerosis frequently contains fully formed bone tissue including marrow. The cellular origin is not known. In this study, bone morphogenetic protein-2a, a potent factor for osteoblastic differentiation, was found to be expressed in calcified human atherosclerotic plaque. In addition, cells cultured from the aortic wall formed calcified nodules similar to those found in bone cell cultures and expressed bone morphogenetic protein-2a with prolonged culture. The predominant cells in these nodules had immunocytochemical features characteristic of microvascular pericytes that are capable of osteoblastic differentiation. Pericyte-like cells were also found by immunohistochemistry in the intima of bovine and human aorta. These findings suggest that arterial calcification is a regulated process similar to bone formation, possibly mediated by pericyte-like cells.

TGF-beta 1 and 25-hydroxycholesterol stimulate osteoblast-like vascular cells to calcify.

Previous studies in our laboratory demonstrated messenger RNA for bone morphogenetic protein-2a in human calcified plaque, suggesting that arterial calcification is a regulated process, similar to osteogenesis. To further test this hypothesis, we have isolated and cloned a subpopulation of cells from bovine aortic media that show osteoblastic potential. These novel cells are primarily distinguished from smooth muscle cells by expression of a surface marker preliminarily identified as a modified form of the ganglioside sialyl-lactosylceramide (GM3). Osteoblastic potential was indicated by high levels of alkaline phosphatase and collagen I, expression of osteopontin and osteonectin (SPARC), and production of bone-specific osteocalcin and hydroxyapatite. Cultures of these cells were stimulated to form increased numbers of calcium-mineral-producing nodules by the oxysterol 25-hydroxycholesterol as well as by transforming growth factor-beta 1, both known to be present in atherosclerotic lesions. The stimulation of calcifying vascular cells in the artery wall by these two factors suggests a possible mechanism for the colocalization of calcification with atherosclerosis in vivo.

Mechanism of calcification in atherosclerosis.

Calcification is commonly associated with atherosclerosis, and it has important clinical implications, especially in coronary arteries. The mineral has been identified as the same mineral as in bone, hydroxyapatite, and several features of its development suggest a mechanism similar to osteogenesis and not merely passive precipitation. The artery wall has been shown to contain several bone-related proteins, including those for osteopontin, osteonectin, and osteocalcin, as well as proteoglycan core proteins homologous with bone biglycan. Our laboratory recently demonstrated that a potent osteogenic differentiation factor, bone morphogenetic protein 2a, is expressed in calcified human atherosclerotic lesions, suggesting that arterial calcification may be initiated by an osteogenic differentiation. In addition, a cell capable of calcium mineral formation in vitro has been isolated from bovine and human aorta and identified by immunostaining as having a surface marker characteristic of microvascular pericytes. These findings suggest the possibility that plaque calcification develops when a signal from atherosclerotic plaque or a factor associated with atherosclerosis induces expression of bone morphogenetic protein, leading to osteogenic differentiation of pluripotential, pericytelike cells located in the arterial intima, which then produce bonelike matrix and hydroxyapatite mineral. These findings also raise questions as to whether osteogenic-promoting factors used to prevent osteoporosis may also increase risk of arterial calcification.

Atherosclerotic calcification: relation to developmental osteogenesis.

Calcium deposits of atherosclerotic plaque consist of hydroxyapatite and may appear identical to fully formed lamellar bone, including trabeculae, lacunae, and areas resembling marrow. Possible mechanisms for bone formation in artery walls are developmental retention of pluripotent cells or osteoblastic immigration coupled with loss of molecular regulatory control that unmasks an embryonic osteogenic program. In situ hybridization of calcified human atherosclerotic lesions shows expression of bone morphogenetic protein type 2, a potent osteogenic differentiation factor. Medial cells of bovine aorta cultured (Dulbecco's modified Eagle's medium plus 15% fetal calf serum) for > 2 weeks form nodules similar to those formed by cultured osteoblasts, including the elaboration of hydroxyapatite.

Acellular hemoglobin solution enters compressed lung capillaries more readily than red blood cells.

High lung inflation pressures compress alveolar septal capillaries, impede red cell transit, and interfere with oxygenation. However, recently introduced acellular hemoglobin solutions may enter compressed lung capillaries more easily than red blood cells. To test this hypothesis, we perfused isolated rat lungs with fluorescently labeled diaspirin cross-linked hemoglobin (DCLHb; 10%) and/ or autologous red cells (hematocrit, 20). Septal capillaries were compressed by setting lung inflation pressure above vascular pressures (zone 1). Examination by confocal microscopy showed that DCLHb was distributed throughout alveolar septa. Furthermore, this distribution was not affected by adding red blood cells to the perfusate. We estimated the maximum acellular hemoglobin mass within septa to be equivalent to that of 15 red blood cells. By comparison, we found an average of 2.7 +/- 4.6 red cells per septum in zone 1. These values increased to 30.4 +/- 25.8 and 50.4 +/- 22.1 cells per septum in zones 2 and 3, respectively. We conclude that perfusion in zone 1 with a 10% acellular hemoglobin solution may increase the hemoglobin concentration per septum up to fivefold compared with red cell perfusion.

Transport properties of alveolar epithelium measured by molecular hetastarch absorption in isolated rat lungs.

To evaluate the transport properties of the alveolar epithelium, we instilled hetastarch (Het; 6%, 10 ml, 1 - 1 x 10(4) kDa) into the trachea of isolated rat lungs and then measured the molecular distribution of Het that entered the lung perfusate from the air space over 6 h. Het transport was driven by either diffusion or an oncotic gradient. Perfusate Het had a unique, bimodal molecular weight distribution, consisting of a narrow low-molecular-weight peak at 10-15 kDa (range, 5-46 kDa) and a broad high-molecular-weight band (range 46-2,000 kDa; highest at 288 kDa). We modeled the low-molecular-weight transport as (passive) restricted diffusion or osmotic flow through a small-pore system and the high-molecular-weight transport as passive transport through a large-pore system. The equivalent small-pore radius was 5.0 nm, with a distribution of 150 pores per alveolus. The equivalent large-pore radius was 17.0 nm, with a distribution of one pore per seven alveoli. The small-pore fluid conductivity (2 x 10(-5) ml. h(-1). cm(-2). mmHg(-1)) was 10-fold larger than that of the large-pore conductivity.

Pathophysiology of coronary calcification.

Calcification is a prominent feature of atherosclerosis, frequently associated with myocardial infarction and other adverse cardiovascular outcomes. Currently, calcification is widely viewed as an end-stage, degenerative process which is inevitable in advanced atherosclerosis. Pathologists, however, have long noted that calcification may occur early in atherosclerosis and, at times, may appear histologically identical to organized bone, including areas resembling bone marrow. These observations suggest that rather than being a passive process, atherosclerotic calcification may instead be an active, regulated process similar to that of osteogenesis. Using an in-vitro model of arterial calcification a subpopulation of artery wall cells, capable of producing hydroxyapatite mineral in vitro was discovered. This article discusses some of the cellular and molecular mechanisms of arterial calcification identified utilizing this in-vitro model of vascular calcification.

Employment survey of Montreal dietetic internship graduates, 1975-77.

A mail survey of graduates for the previous three years from Montreal area dietetic internships was carried out in 1977-78. One objective of the survey was to collect data on employment status and job turnover. Mailing lists for graduates were obtained from internship program directors. One hundred and ninety-three questionnaires were mailed. Forty-nine percent of the questionnaires were returned. Sixty-one respondents (64%) reported they had left their first position; 73% of these left within one year. The major reason for leaving first positions was 'temporary position' (43%), and 'family reasons' was cited by 10%. Eighty-one percent of respondents were employed at the time of the survey. The major reasons for unemployment were: 'education' (33%); 'family reasons' (22%), and 'no position' (16%). Family reasons for not working were less common in this survey than in a national survey conducted in 1974. Results presented here indicate that recent dietetic graduates who are married and/or have a family are more likely to work today than in 1974, as is the trend for women in other areas of employment. The implications for new graduates are: higher job turnover, fewer permanent positions, and an increase in the number of post graduate degrees.

The atherosclerosis-calcification link?

Although previously believed to be a passive, degenerative process, there is increasing evidence that arterial calcification is actually an active, regulated process. In this review we address potential mechanisms of arterial calcification and, in particular, ways in which it is similar to bone formation. We also advance the hypothesis that particular factors present in atherosclerosis, such as lipids, may explain the co-localization of calcification and atherosclerosis in vivo.

An evidence-based assessment of the NCEP Adult Treatment Panel II guidelines. National Cholesterol Education Program.

CONTEXT: The Second Report of the National Cholesterol Education Program (NCEP) Expert Panel on Detection, Evaluation, and Treatment of High Blood Cholesterol in Adults (Adult Treatment Panel II) was issued without the benefit of multiple recently published large clinical trials. OBJECTIVE: To analyze the panel's guidelines for treatment of high cholesterol levels in the context of currently available clinical trial results. DATA SOURCES: MEDLINE was searched for all English-language clinical trial data from 1993 through February 1999 relating to the effects of cholesterol treatment on cardiovascular clinical outcomes. STUDY SELECTION: Studies that were selected for detailed review assessed the effects of cholesterol lowering on either coronary events, coronary mortality, stroke, and/or total mortality, preferably by randomized, double-blind, placebo-controlled design. Selection was by consensus of a general internist, a lipid clinic director, and a researcher in atherosclerotic plaque biology. A core of 37 of the 317 initially screened studies were selected and used as the primary means by which to assess the guidelines. DATA EXTRACTION: By consensus of the group, only prespecified end points of trials were included, unless post hoc analysis addressed issues not studied elsewhere. DATA SYNTHESIS: Recent clinical trial data mostly support the Adult Treatment Panel II guidelines for cholesterol management. While existing trials have validated the target low-density lipoprotein cholesterol (LDL-C) goals in the report, studies are lacking that address mortality benefit from reduction below these levels. Few lipid-lowering trials have treated patients with low high-density lipoprotein cholesterol and/or elevated triglyceride levels with LDL-C levels at or below treatment goals. CONCLUSIONS: Lipid-lowering therapy generally should be more aggressively applied to patients with diabetes and/or at the time of coronary heart disease (CHD) diagnosis. The evidence for statin use in secondary CHD prevention in postmenopausal women outweighs current evidence for use of estrogen replacement in this setting. Further studies are needed to address the effects of lipid modification in primary prevention of CHD in populations other than middle-aged men and to study markers of lipid metabolism other than LDL-C.

The formation of hydroxyapatite-calcium polyacrylate composites.

Tetracalcium phosphate (TetCP, Ca4(PO4)2O) reacts rapidly with polyacrylic acid (PAA). Complete reaction results in the formation of hydroxyapatite (HAp) and calcium polyacrylate. Consequently, this combination of reactants can react to form a dental cement. However, reaction occurs so rapidly that it would be difficult to achieve a homogeneous mixture of reactants suitable for use in restorations. In order to explore extending the working time, the effects of prehydrating the TetCP to form surface layers of HAp on the TetCP particles was explored. Prehydration was found to be an effective means of allowing workability. Therefore, the effects of the proportions of TetCP and PAA, with and without HAp filler, on cement properties were investigated. The extents of the reactions were investigated by X-ray diffraction analysis; the extents of PAA neutralization were studied by Fourier transform infra-red spectroscopy (FTIR); pore structures were determined by mercury intrusion porosimetry; microstructures were observed by scanning microscopy, and compressive strengths were determined. After curing for 17 days at room temperature PAA neutralization was almost complete; however, residual TetCP could be detected by X-ray diffraction and PAA by FTIR. As expected, the compressive strengths of the cements showed a dependence on the liquid (water+polymer)-to-solid (TetCP+HAp filler) used. The presence of HAp filler caused a significant decrease in compressive strength and increasing the proportion of HAp filler resulted in a decrease in the compressive strength. The characteristics of the load-deflection curves showed a dependence on the presence of HAp filler. In the absence of filler, two slopes were observed in the curves whereas a linear curve, typical of a ceramic, was observed when HAp filler was present. Mercury intrusion porosimetry (MIP) indicated the majority of the porosity was present in pores larger than 0.1 microm. Porosity increased with increasing liquid-to-solids ratio and with an increasing proportion of HAp filler at a constant liquid-to-solids ratio. Microstructural observations indicated the effect of HAp filler on increasing porosity was the result of porosity present in the filler itself. Thus, poorly consolidated HAp filler contributed to increased porosity and reduced compressive strength.

Intestinal bypass for morbid obesity: a consecutive personal series.

Forty-five patients who underwent end-to-side jejunoileal bypass, with 51 cm in circuit, were followed up from 8 months to 8 years (average 3.4 years). There was no early or late mortality but morbidity was considerable; it included inadequate weight loss or late gain in 22%, malnutrition and liver failure in 11%, severe diarrhea and electrolyte imbalance in 11%. Cholelithiasis and wound complications also occurred. Reanastomosis was necessary in 13% (six patients). The result of the bypass was good in only 20%, satisfactory in 44% and unsatisfactory in 36%. This type of bypass is not adequate treatment for morbid obesity because the proportion of unsatisfactory results (36%) is too high and the number of good results too low and because the outcome is unpredictable. The complication of malnutrition characterized by a decline in body cell mass, an expansion of the extracellular mass and an increase in the ratio of total exchangeable sodium to total exchangeable potassium is quickly and effectively treated by intravenous administration of amino acids or protein hydrolysates. Long-term management of protein malnutrition requires a high protein diet (100 g/d) or reanastomosis.

Serological analysis of the deoxyribonucleic acid polymerase of avian oncornaviruses. I. Preparation and characterization of monospecific antiserum with purified deoxyribonucleic acid polymerase.

Monospecific antiserum was prepared against purified deoxyribonucleic acid (DNA) polymerase from avian myeloblastosis virus (AMV). Immunodiffusion assay with purified DNA polymerase revealed that the anti-DNA polymerase serum formed one precipitation band, whereas no reaction with any of the seven major structural proteins of AMV was observed. The antiserum also demonstrated enzyme-neutralizing antibody activity that was associated with the immunoglobulin G fraction. There was no difference in the neutralization of DNA polymerase activity directed by ribonucleic acid (RNA), DNA, or RNA-DNA hybrid templates.

Fibronectin and collagen I matrixes promote calcification of vascular cells in vitro, whereas collagen IV matrix is inhibitory.

Vascular calcification is a frequent component of atherosclerosis, yet the pathological mechanisms that regulate its formation are poorly understood. Calcification of the vessel wall may represent a process by which cells that normally exhibit a smooth muscle phenotype differentiate into cells that exhibit an osteoblast-like phenotype. One of the determinants of cellular phenotype is extracellular matrix; thus, we undertook the current study to evaluate the influence of extracellular matrix on calcification of vascular cells in vitro. Cell lines derived from bovine aortic media were divided into 1 of 3 groups: those that did not mineralize, those that mineralized slowly, or those that mineralized rapidly. When slowly mineralizing cells were plated onto matrix produced by rapidly mineralizing cells, the time required for mineralization decreased from 33+/-3.0 days to 7.8+/-1.3 days. Matrix produced by rapidly mineralizing cells was found to contain 3 times the amount of collagen I and fibronectin but 70% less collagen IV than nonmineralizing clones. When slowly mineralizing cells were cultured on purified collagen I or fibronectin, mineralized nodule formation, calcium incorporation, von Kossa staining, and alkaline phosphatase activity increased. In contrast, culturing slowly mineralizing cells on purified collagen IV inhibited these mineralization parameters. Furthermore, blocking antibodies to alpha5 integrins significantly inhibited the fibronectin-mediated increases in alkaline phosphatase activity, indicating that integrin-based signaling may be involved. These data suggest that matrix composition can regulate development of arterial calcification and that a subpopulation of vascular cells preferentially produces positively regulating matrix components.

Pulmonary capillary sieving of hetastarch is not altered by LPS-induced sepsis.

BACKGROUND: Gram-negative lipopolysaccharide (LPS) has been demonstrated to increase pulmonary capillary permeability as judged by the increased flow of protein-rich lymph from the lungs of sheep infused with LPS. This finding suggests that LPS-injured pulmonary capillaries might be less restrictive than uninjured capillaries to the filtration of large hetastarch molecules. Hetastarch has a broad molecular mass spectrum (35-1,500 kilodaltons (kDa)), and one way to test the restrictiveness of pulmonary capillaries is to measure the size of the largest hetastarch molecules that cross the microvascular barrier and enter the lymph. To evaluate the effects of LPS, we compared hetastarch molecular distributions in the lung lymph of normal and LPS-injured sheep. METHODS: Adult sheep (38.2 +/- 0.8 kg) were surgically prepared for the collection of lung lymph, with study initiation after a 5- to 7-day recovery period. Hetastarch (6%) was infused (10 mL/kg) 24 hours before study to allow for stabilization of the hetastarch molecular distribution. On the day of study, LPS (Escherichia coli lipopolysaccharide, 2 microg/kg; n = 6) was infused, and plasma and lymph samples were collected for 12 hours. An additional group of animals not infused with LPS (n = 6) served as controls. Hetastarch molecular distributions in plasma and lymph were measured by using high performance size exclusion chromatography. RESULTS: In control sheep, the largest hetastarch molecules in lymph averaged 861 +/- 18 kDa (mean +/- SEM) (plasma, 1,065 +/- 18 kDa). In LPS-treated sheep, the largest hetastarch molecules in lymph averaged 845 +/- 19 kDa (not significant vs. normal) (plasma, 1,025 +/- 14 kDa). Hetastarch concentrations in plasma and lung lymph of normal sheep, respectively, were 0.61 +/- 0.05% and 0.34 +/- 0.07%. In LPS-treated sheep, hetastarch concentrations in plasma and lymph were 0.56 +/- 0.08 (not significant vs. normal) and 0.29 +/- 0.07, respectively (p < or = 0.05). Lymph concentrations were lower after LPS because of increased lymph flows (19.9 +/- 5.4 mL/30 min, compared with 3.6 +/- 0.8 mL/30 min in normal sheep). CONCLUSION: Our results suggest that LPS does not alter the diameter of the largest pores perforating the walls of pulmonary capillaries. Rather, the number of these pores in the capillary wall appears to be increased. This increase would explain why lymph flows rise after LPS with little change in the lymph protein concentration. Our results are also consistent with a filtration model in which capillaries are assumed to be perforated by small pores (protein reflection coefficient = 1) as well as large pores (protein reflection coefficient = 0).

Effect of concentration and hyaluronidase on restriction of hetastarch flux through lung interstitial segments.

The transport properties of lung interstitium were studied by measuring the flow of hetastarch solution (2 and 6%) through 1-cm perivascular interstitial segments of rabbit lungs. Hetastarch (10(4)-10(7) Da) solution has a colloid osmotic pressure similar to that of albumin solution. Driving pressure was 5 cm H(2)O and mean interstitial pressure was 0 cm H(2)O. The flows of 2 and 6% hetastarch solutions were measured before (Q(1)) and after (Q(2)) the addition of 0.02% hyaluronidase. Hetastarch molecular distributions in effluent samples were measured by high-performance size-exclusion chromatography (HPSEC) to determine sieving ratio (C(out)/C(in), downstream-to-upstream concentration ratio). Hyaluronidase significantly (P < 0.0004) increased flow sixfold, but the increase in flow (Q(2)/Q(1)) was reduced through the interstitium around smaller vessels. A similar behavior was observed with the flow of albumin solution without and with hyaluronidase. C(out)/C(in) decreased monotonically with molecular weight, was greater with 6% than with 2% (low colloid osmotic pressure) hetastarch, and increased with hyaluronidase. Modeling the transport through uniform pores, equivalent pore radius was 10 and 15 nm with 2 and 6% hetastarch, respectively, and doubled with hyaluronidase. In conclusion, interstitial pores expand in response to an increase in colloid osmotic pressure both before and after tissue degradation by hyaluronidase.

Ethnic origin and serum levels of 1alpha,25-dihydroxyvitamin D3 are independent predictors of coronary calcium mass measured by electron-beam computed tomography.

BACKGROUND: Blacks have been found to have lower amounts of coronary calcium as well as higher levels of the osteoregulatory steroid 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] than whites. We sought to determine if racial differences in coronary calcium mass could be explained by differences in serum levels of 1,25(OH)2D3. METHODS AND RESULTS: We evaluated standard coronary risk factors, quantified coronary calcium mass with electron-beam computed tomography (EBCT), and measured serum 1,25(OH)2D3 with radioimmunoassay in 283 high-risk subjects (51 [180%] black, 232 [82%] white). Black subjects had lower masses of coronary calcium than whites (14 versus 47 mg; P=.003). Serum 1,25(OH)2D3 levels were slightly higher in blacks (41 versus 38 pg/mL; P=.05). Log 1,25(OH)2D3 levels were inversely proportional to log-transformed calcium mass (r=-.19; P=.001) in both races. Multivariate linear regression demonstrated that both black race (P=.02) and 1,25(OH)2D3 levels (P=.007) contributed inversely and independently to coronary calcium mass. However, an interaction term of racex1,25(OH)2D3 did not significantly contribute to coronary calcium mass, indicating that other undetermined factors in addition to 1,25(OH)2D3 are responsible for ethnic differences in coronary calcium mass. CONCLUSIONS: Both black race and serum levels of 1,25(OH)2D3 are independent negative determinants of coronary calcium mass. Nevertheless, diminished amounts of coronary calcium in blacks are not accounted for by higher 1,25(OH)2D3 levels.

Active serum vitamin D levels are inversely correlated with coronary calcification.

BACKGROUND: Arterial calcification is a common feature of atherosclerosis, occurring in >90% of angiographically significant lesions. Recent evidence from this and other studies suggests that development of atherosclerotic calcification is similar to osteogenesis; thus, we undertook the current investigation on the potential role of osteoregulatory factors in arterial calcification. METHODS AND RESULTS: We studied two human populations (173 subjects) at high and moderate risk for coronary heart disease and assessed them for associations between vascular calcification and serum levels of the osteoregulatory molecules osteocalcin, parathyroid hormone, and 1alpha,25-dihydroxyvitamin D3 (1,25-vitamin D). Our results revealed that 1,25-vitamin D levels are inversely correlated with the extent of vascular calcification in both groups. No correlations were found between extent of calcification and levels of osteocalcin or parathyroid hormone. CONCLUSIONS: These data suggest a possible role for vitamin D in the development of vascular calcification. Vitamin D is also known to be important in bone mineralization; thus, 1,25-vitamin D may be one factor to explain the long observed association between osteoporosis and vascular calcification.