The identification of cholesterol as a constituent of atheromatous plaques has aroused recurrent waves of suspicion that lipid metabolism is in some way responsible for the development of atherosclerosis and coronary artery disease. Lipids are transported in serum as constituents of three major classes of compounds: cholesterol and its esters, phospholipids, and triglycerides or neutral fat. Most investigations in recent years have centered about the notion that serum cholesterol plays a causative role in the development of coronary artery disease in spite of numerous studies focusing attention on triglycerides. Included in these studies are reports of increased turbidity, increased and prolonged chylomicronemia, and increased ratio of \g=b\to \g=a\-lipoproteins. Although triglycerides rather than cholesterol are chiefly responsible for turbidity of serum and constitute an important fraction of \g=b\-lipoproteins, the quantitative estimation of triglycerides has been largely neglected in studies of serum lipids in coronary artery disease.The present research was undertaken to observe the interrelations of the three major serum lipid fractions 14 ; their interrelations had been studied in certain metabolic dis¬ orders in 1943.5,e The neglected fraction, triglycérides, appeared to be most frequently elevated in coronary artery disease.7 Materials and MethodsThe subjects were 100 patients with a history of a documented myocardial infarction 1 day to 12 years previously, confirmed by electrocardiographic studies and laboratory and clinical findings. The patients were obtained during the past five years from the clinics and inpatient services of the Grace-New Haven Hospital, and we include all patients meeting the above requirements whose physicians requested lipid determinations. The 100 patients, 82 men and 18 women, ranged from 21 to 78 years of age, the greatest number being in the 50's. Eleven were diabetics.Control subjects consisted of a heterogeneous group of healthy medical school students and per¬ sonnel, healthy industrial workers, and hospital patients without evidence of focal vascular disease and without known metabolic disorders. Ages ranged from 20 to 78.Blood was drawn before breakfast, and the serum was analyzed for total fatty acids after hydrolysis, for total cholesterol, and for lipid phosphorus.2 Triglycéride fatty acids were esti¬ mated by subtracting the fatty acids attributed to cholesterol esters and phospholipids from the total fatty acids. Cholesterol-phospholipid ratios were not remarkable. Normal ranges for cholesterol and triglycérides were established as two standard deviations above CHOLESTEROL MG % 40 AGE Fig. 1.-Concentration of cholesterol by age in the sera of 82 men, including 9 diabetics, with a history of myocardial infarction (upper graph) and of 134 normal men (lower graph). In each a horizontal line indicates the upper limit of nor¬ mal, 269 rng. %.

Chromium deficiency may cause insulin resistance, hyperinsulinemia, impaired glucose tolerance, and hyperlipidemia, recovered by chromium supplementation. The effect of chromium supplementation on serum lipids and glucose tolerance was tested in a double-blind 12-wk study of 23 healthy adult men aged 31 to 60 yr. Either 200 micrograms trivalent chromium in 5 ml water (Cr) or 5 ml plain water (W) was ingested daily 5 days each week. Half the subjects volunteered for glucose tolerance tests with insulin levels. At 12 wk high-density lipoprotein cholesterol increased in the Cr group from 35 to 39 mg/dl (p less than 0.05) but did not change in the water group (34 mg/dl). The largest increase in high-density lipoprotein cholesterol and decreases in insulin and glucose were found in those subjects having normal glucose levels together with elevated insulin levels at base-line. The data are thus consistent with the hypothesis that Cr supplementation raises high-density lipoprotein cholesterol and improves insulin sensitivity in those with evidence of insulin resistance but normal glucose tolerance.

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To assess the effect of dietary fiber on glucose tolerance four different meals of varying fiber content but identical protein fat and carbohydrate content were fed to eight healthy men aged 22 to 45. Each meal provided 75 g of carbohydrate as liquid glucose formula, as brown rice, pinto beans, or All Bran. The mean plasma glucose and insulin responses were highest following the formula, and least for All Bran and pinto beans. Rice produced nearly as great a rise in insulin and glucose as did the formula. The rank of each meal by content of neutral detergent fiber was nearly the inverse of the rank by magnitude of the insulin response evoked, fiber content being greatest in All Bran (18 g) and pinto beans (16.2 g), low in rice (2.8 g) and absent from the formula. It was concluded that dietary fiber dampened the insulin response to a high carbohydrate meal.

The interrelationships among fatness measures, plasma triglycerides and high density lipoproteins (HDL) were examined in 131 normal adult subjects: 38 men aged 27-46, 40 men aged 47-66, 29 women aged 27-46 and 24 women aged 47-66. None of the women were taking estrogens or oral contraceptive medication. The HDL concentration was subdivided into HDL2b, HDL2a and HDL3 by a computerized fitting of the total schlieren pattern to reference schlieren patterns. Anthropometric measures employed included skinfolds at 3 sites. 2 weight/height indices and 2 girth measurements. A high correlation was found among the various fatness measures. These measures were negatively correlated with total HDL, reflecting the negative correlation between fatness measures and HDL2 (as the sum of HDL2a and 2b). Fatness measures showed no relationship to HDL3. There was also an inverse correlation between triglyceride concentration and HDL2. No particular fatness measure was better than any other for demonstrating the inverse correlation with HDL but multiple correlations using all of the measures of obesity improved the correlations. Partial correlations controlling for fatness did not reduce any of the significant correlations between triglycerides and HDL2 to insignificance. The weak correlation between fatness and triglycerides was reduced to insignificance when controlled for HDL2.

The present report is an analysis of the course of peripheral vascular disease (PVD) in 619 patients with non-insulin-dependent diabetes (NIDDM) recruited within 1 yr of diagnosis and followed quarterly for up to 14 yr (X = 10.5 yr). At 13 yr duration, the actuarially determined cumulative risks for intermittent claudication (IC), nonpalpable dorsalis pedis pulse (NPUL), and arterial calcification (CALC) were, respectively, 37.9%, 34.5%, and 60.9% for men and 24.3%, 37.6%, and 32.2% for women. Major amputations (AMP) occurred in only 1.3% of the patients, equivalent to approximately one case per 1000 patients per year. The corresponding incidences of IC, NPUL, and CALC were, respectively, 29, 27, and 47 per 1000 men and 19, 27, and 25 per 1000 women per year. CALC and NPUL were strongly related to mortality. Baseline risk factors with probability levels that suggest a relationship to PVD were, in women, age versus CALC (P less than 0.01), age versus NPUL (P less than 0.05), weight versus NPUL (P less than 0.05), systolic BP versus CALC (P less than 0.01), summed glucose tolerance test versus CALC (P less than 0.01), and triglyceride level versus CALC (P less than 0.05). In men, the only significant risk factors were diminished vibration perception, which was related to NPUL (P less than 0.05), and the serum triglyceride level, which was related to IC (P less than 0.05). In patients who are carefully followed prospectively, IC is far more common, but AMP is far less common than has been generally appreciated. Further studies are needed to clarify the roles of the diverse risk factors that are possibly related.

In the course of studies on lactescence of serum, it was found that the insoluble lipids displaced serum water to a degree which was occasionally great enough to be of considerable practical importance. The immediate consequence of this displacement was the finding of spuriously low concentrations of water-soluble components of serum, although their concentrations were normal when repeated on sera from which the insoluble lipids had been removed by ultracentrifugation.These observations clearly showed the need for a simple, rapid method for determination of serum water that would be applicable to sera with a high concentration of lipids. The method using Karl Fischer reagent (1) (3).previously described (7). The original and subnatant fluids were subjected to analysis.C. Gravimetric estimation of serum water:The water was evaporated and the weight loss used to calculate the serum water in grams per 100 ml. of serum (8). D. Calculation of serum water from serum protein:Serum water was calculated from the following formula, the derivation of which is explained elsewhere (8): Ws = 98.5 -0.745 P. in which W. = serum water in gm. per 100 ml. of serum, and P. = serum protein in gm. per 100 ml. E. Osmometric method for estimation of serum water:Theory: The osmotic pressure of serum is determined before and after the addition of a known amount of dry sodium chloride designed approximately to double the osmolarity of serum. The only change taking place upon this addition is an increase in the molal concentration of sodium chloride. Since the osmotic pressure of a solution is determined by the molal concentration of solute particles, the exact molal concentration of the added sodium chloride can be calculated from the increase it causes in the osmotic pressure of serum. Knowing both the absolute amount and the molal concentration of the added salt, the volume of water in which it was dissolved can then be calculated.A freezing point osmometer was used for the determination of osmotic pressure. Although osmotic pressure is not actually measured, advantage is taken of the proportionality between osmotic pressure and freezing point depression in such a way that the results are read directly as milliosmols per liter.Reagents: 1. 0.161 molar sodium chloride solution. Dilute 9.404 gm. dry sodium chloride (reagent grade) to one liter. Double distilled water.Special equipment: A Fiske Associates freezing-point osmometer was used for the determinations of osmolarity. Procedure: To prepare the dry sodium chloride exactly 2 ml. of the sodium chloride solution were pipetted into test tubes to be used later in the osmometer. These were brought to dryness in an oven at 980 C. and subsequently stored in a desiccator until ready for use. The addition of exactly 2 ml. of double-distilled water to the dry salt results in a 0.161 molal solution of sodium chlo-1483

It has long been recognized that the cloudy or lactescent serum occurring in certain diseases contains an abnormally large amount of neutral fat (1), though a quantitative relationship between neutral fat and lactescence has not been established. In the present investigation the ultracentrifuge has been utilized to study the suspended lipid particles which are responsible for such lactescence. By analyzing a variety of clear and lactescent sera having high concentrations of one or more lipid components the relationship of neutral fat to lactescence has been verified and quantitatively defined. METHODSSerum lipids were determined before and after the removal of the suspended lipid particles by flotation in the ultracentrifuge. The technique of Van Eck, Peters, and Man (2) was modified as described below to permit the more complete separation of the lipids present as visible particles from the lipids present in clear solution. The particulate lipids which caused turbidity and were removed by centrifugation shall be referred to as "insoluble" lipid, while the lipids present in the clear subnatant after centrifugation shall be designated "soluble" lipid. The visual appearance of the serum was thus the criterion used in determining the state of solubility. No effort was made to classify the differential flotation characteristics of the lipids, the ultracentrifuge being used simply to remove the particles causing turbidity. Although no measurement of particle size was made, it is assumed that the insoluble lipids ranged from 0.1 IA in diameter, the size just sufficient to scatter light (3), to 1 1A, the size of the larger chylomicrons.Duplicate 5 ml. portions of undiluted serum, in no instance more than 24 hours old, were measured accurately into two 13.5 ml. lustroid tubes. The serum was spun for one hour at 18,000 rpm (about 20,000 g.) at room temperature in a Spinco Model L Preparative Ultracentrifuge. After centrifugation the insoluble matter was visible on top as a buttery yellow layer in very lactescent serum, and as a faint white film in,clear or An exact quantity of the subnatant fluid was withdrawn into the pipette, leaving the "cream" layer floating undisturbed on top of the remaining serum. As the subnatant fluid was removed, exquisite care was taken to keep the tip of the outer needle just below the "cream" layer. In this way the fatty material which adhered to the outer needle was excluded from the clear subnatant fluid.When the serum was moderately or grossly lactescent, the original serum and the clear subnatant fluid obtained after centrifugation were analyzed for lipids. The insoluble lipids were calculated by subtracting the lipids of the clear subnatant fluid from those of the original serum.When the serum was clear or slightly lactescent and the anticipated difference between the original serum and the subnatant fluid was small, exactly 3 ml. of the subnatant fluid were removed from the bottom of the tube, and the "cream" layer was resuspended in the remaining 2 ml. of serum by gentle agitation of ...