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We had previously used an electrophoretic transfer procedure to determine the topographic distribution of low density lipoprotein (LDL) accumulation in the aortic intima of normolipemic swine. In this present study we have employed a similar procedure to assess whether LDL-rich sites consistently demonstrate increased intimal thickening at the iliac bifurcation and common iliac arteries. The topographic distribution of LDL-rich sites was determined in the aortas of six subjects ranging in age from 16 to 36 years, by transferring LDL by electrophoresis from the tissue into an agarose gel containing anti-LDL, and then staining the immunofixed LDL in the gel for lipid. LDL-rich sites were found in all but two of these cases. On the basis of control studies establishing the level of nonspecific staining, we determined that the cutoff between LDL-rich and LDL-poor zones was 37 mg apoB protein/mm2 intimal surface area. Intimal thickening was found to be threefold greater in LDL-rich than in LDL-poor regions. These results confirm and extend earlier immunohistochemical studies suggesting a preferential accumulation of LDL at sites of intimal thickening in human arteries.

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To determine whether in situ tissue lipid characterization is possible, we examined carefully prepared frozen sections from a variety of lipid-rich tissues of the cholesterol-fed rabbit by hot stage polarizing light microscopy and conventional histologic staining. Heating of frozen sections to less than 60 degrees C did not affect tissue architecture or staining characteristics making pathologic and physical chemical correlations possible. The melting temperatures of cholesterol ester inclusions in individual foam cells in rabbit atherosclerotic lesions and adrenal gland could be determined as well as the melting characteristics of crystals and triglyceride in these and other tissues. Differential scanning calorimetry and polarized light transmittance were used to confirm melting temperatures determined by microscopy. Combining data from histologic staining, polarizing light microscopy, and the thermal characteristics of lipid enables the various lipid classes to be identified within individual cells. Differences in melting temperatures between lipids of the same class give indications of the degree of saturation of the lipids. Regional differences of cholesterol ester-melting temperatures in the chow-fed rabbit adrenal cortex were detected which implied differences in chemical composition. Cholesterol feeding raised the melting temperature and tended to abolish the marked regional differences in melting temperature of the cholesterol esters in the adrenal cortex. Rabbit atherosclerotic lesions, induced by balloon deendothelialization and cholesterol feeding, revealed differences in foam cell-melting temperatures within the same lesion. Melting temperatures of cholesterol ester deposited in the liver were more uniform. Each tissue studied revealed distinctly different cholesterol ester-melting characteristics.

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Nuclear magnetic resonance (NMR) images of 93 patients undergoing studies of the abdomen and pelvis were studied for evidence of lesions of the aorta and the iliac and femoral arteries; atherosclerotic lesions were present in 13 of them. The lesions consisted of eccentric and concentric mural thickening with luminal narrowing and discrete plaques protruding into the vessel lumen. This appearance was distinctly different from the morphology of the internal vessel surface and uniformly thin vessel wall in normal patients and volunteers under the age of 30 years. Intraluminal flow signals observed in atherosclerotic and nonatherosclerotic subjects could be distinguished from mural lesions because of their lack of contiguity with the vessel wall and variation in appearance on multiple images obtained with the first and second spin echo. This initial experience suggests a potential role for NMR in the noninvasive imaging of atherosclerotic lesions. The natural contrast between flowing blood and the vessel wall indicates a distinct advantage of NMR for vascular imaging.

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Samples of human Common Iliac Arteries, obtained post-mortem from patients without evidence of atherosclerosis, were analyzed for their content in the various types of collagens using alternatively sonication, pepsin digestion and dithiothreitol reduction followed by sodium dodecyl sulfate polyacrylamide gel electrophoresis of the extracts. A final average proportion of 66% of the total collagen was solubilized. The statistical calculation of the proportions of the different collagens, deduced from the densitometric scannings of the electrophoretic runs, showed that a larger amount of type I was extracted when the extraction was more complete. Type I collagen appeared as the most insoluble collagen in these arteries and its final proportion was probably higher than 60% of the total collagen.

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An endogenous inhibitor of the Na,K pump, postulated to be involved in the etiology of some hypertensive states, has been reported in extracts of mammalian brain. This encouraged us to test its effects on arterial muscles. An acid-acetone extract of guinea pig brain inhibited Na,K-ATPase derived from canine kidney and evoked responses in arterial strips similar to those produced by ouabain. Unlike ouabain, however, it did not prevent muscles in K-free solutions from relaxing when K was re-added. Bioassays on strips of arteries, uterus and portal vein indicated that the extract did not contain sufficient concentrations of norepinephrine, dopamine, serotonin, prostaglandins, angiotensin II, oxytocin or the Na,K-ATPase inhibitor to account for the observed vascular effects. This could not be said of vasopressin. Furthermore, vasopressin and the vasoactive component of the extract were equally sensitive to several peptidases, and conditions which cleave disulfide bridges. A radioimmunoassay verified that the extract contained sufficient vasopressin to cause contractions. Vasopressin did not inhibit the kidney Na,K-ATPase activity. Finally, the Na,K-ATPase inhibitor, but not the vasoactive substance, was present in extracts of vasopressin-deficient Brattleboro rat brains. Therefore, the Na,K-ATPase inhibitor and the vasoactive substance in these extracts were distinctly different.

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A technique using alternatively dithiothreitol reduction, sonication and pepsin digestions permits the extraction of 75% of the insoluble collagen from human common iliac arteries. The pattern of the collagen types obtained by acrylamide gel electrophoresis is far different from that of the aortic wall: type I averages 44 +/- 14% of the total, type III 48.5 +/- 14% and a supplementary fraction, probably belonging to type V, 9 +/- 5% of the total in females and 6 +/- 2% in males.

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Lipoprotein concentrations were compared in serum and in samples of arterial intima obtained during cardiovascular surgery. The interval between blood sampling and surgery did not exceed 48 hours. Apolipoproteins B and C, the major proteins of low density lipoprotein (LDL) and very low density lipoprotein (VLDL) respectivity, were detectable by immuno-assay in all arterial specimens. Highly significant positive correlations existed between the levels of lipoproteins in the intima and in serum. In patients with elevated serum VLDL levels (Type IV hyperlipoproteinaemia) arterial concentrations of apolipoproteins B and C were increased about two-fold; in those with raised LDL levels in serum (Type II), with or without concomitant elevation of VLDL levels, the concentration of apolipoprotein B was increased almost 5-fold in the arterial intima. The arterial wall therefore contains lipoproteins or their immunologically-similar metabolites at concentrations which are determined in part by serum lipoprotein levels.

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We examined the water, sodium, and potassium composition of the thoracic aorta, abdominal aorta (plus iliac arteries), and veins (vena cava and portal vein) from rats with aortic coarctation. The aortas of 10 rats (group A) were coarcted above the renal arteries to produce hypertension. Control groups consisted of 10 rats sham-coarcted above and 10 rats coarcted below the renal arteries. In group A rats heart weights and carotid artery pressures were elevated over controls (P less than 0.01), whereas there were no significant differences in femoral arterial pressures. In group A rats both the hypertensive thoracic aorta and the normotensive abdominal aorta contained about 20% more water per unit of wet weight, and about 35% and 60% more sodium and potassium, respectively, per unit of dry weight than did the corresponding portions of aorta from control rats (P less than 0.01). In group A rats water (P less than 0.01), sodium (P less than 0.02), and potassium (P less than 0.05) contents of veins also were increased. There were no significant correlations between level of carotid arterial pressure and magnitude of changes in arterial and venous composition, nor were there significant differences between the magnitude of changes in the normotensive and hypertensive portions of the aorta. These results indicate that in rats abnormalities in vascular wall salt and water content are not necessarily a direct effect of the elevated pressure in hypertension.

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A comparison of passive mechanics and the effects of smooth muscle (SM) activation has been made in canine carotid and iliac arteries. Measurements of external diameter and axial force were made on isolated cylindrical segments in response to internal pressure variations. Data were obtained during maximal norepinephrine (NE) and potassium (K) activation, and following metabolic inhibition of SM. These data were used to determine wall tangential stress-strain relations. The maximum increase in wall stress after K and after NE was greater for the iliacs and occurred at smaller values of wall strain. Passive stress-strain curves for the iliacs were likewise shifted to smaller strains, suggesting an important role of passive wall elements in setting the length of contractile elements. Diameter responses to K at low values of pressure were the same for iliacs and carotids, but were better maintained at higher pressure levels for the iliacs. Similar findings were found for iliac responses to NE and K. The results suggest that at low values of wall stress, active diameter responses are not strongly dependent on the maximum isometric stress development. However, higher pressure-diameter responses are also determined by the force-generating capacity of SM.

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