Beta Decay of Molecular Tritium.

The beta decay of tritium in the form of molecular T_{2} is the basis of sensitive experiments to measure neutrino mass. The final-state electronic, vibrational, and rotational excitations modify the beta spectrum significantly and are obtained from theory. We report measurements of the branching ratios to specific ionization states for the isotopolog HT. Two earlier, concordant measurements gave branching ratios of HT to the bound HHe^{+} ion of 89.5% and 93.2%, in sharp disagreement with the theoretical prediction of 55%-57%, raising concerns about the theory's reliability in neutrino mass experiments. Our result, 56.5(6)%, is compatible with the theoretical expectation and disagrees strongly with the previous measurements.

Viscosity of argon at high temperatures.

New data for the viscosity of argon at high temperatures indicate that the accepted data are substantially too low at temperatures above 600 degrees K.

Animals at very high pressures of helium and neon.

At pressures up to 125 atmospheres, helium failed to anesthetize mice; at slightly higher pressures (135 to 145 atmospheres) it proved lethal. With Italian newts (Triturus italicus), whose sensitivity to anesthesia by nitrogen is similar to that of mice, responsiveness was lost at pressures between 165 and 245 atmospheres, whether the pressure was achieved with helium or neon, or hydrostatically. It was concluded that the anesthetic pressures of helium and neon, for mice and newts, are higher than the tolerable mechanical pressures.

Basal cell carcinoma does metastasize.

Basal cell carcinoma (BCC) rarely metastasizes. However, this unfortunate outcome can occur, usually in neglected tumors. We report a 52-year-old man with a BCC on the left chest that enlarged and then ulcerated over a 6-year period. Metastasis of the tumor to lymph nodes in the left axilla resulted, but the patient remains free of disease 24 months after wide excision, lymph node dissection, and local radiation therapy to the axilla.

Low-density lipoprotein concentration in interstitial fluid from human atherosclerotic lesions. Relation to theories of endothelial damage and lipoprotein binding.

Increased endothelial permeability to low-density lipoprotein (LDL) is believed to be an initiating factor for atherosclerotic lesions. Concentrations of LDL, alpha 2-macroglobulin and albumin were measured by immunoassay in interstitial fluid collected from normal intima and atherosclerotic lesions of human aortas. The concentration of LDL in interstitial fluid from normal intima was twice the concentration in the patient's serum. In early proliferative (gelatinous) lesions the amount of interstitial fluid was consistently increased but its LDL concentration varied between 80 and 200% of adjacent normal intima. Highest concentrations of LDL were found in interstitial fluid from more advanced proliferative lesions, but the amount was reduced, suggesting a shift in tissue water. LDL was consistently low in interstitial fluid from fatty streaks comprised of lipid-filled cells, and in four of 12 lesions it was absent although alpha 2-macroglobulin and albumin concentrations were normal. Electrophoretic mobility of LDL, reflecting surface charge, was unchanged or increased in interstitial fluid from normal intima and fatty streaks, but decreased in gelatinous lesions. The ratio of LDL to alpha 2-macroglobulin and albumin in interstitial fluid was higher than in adjacent intact tissue. The results do not support the idea that increased endothelial permeability to LDL initiates atherogenesis.

Fibronectin distribution in human aortic intima and atherosclerotic lesions: concentration of soluble and collagenase-releasable fractions.

Fibronectin is associated with cell attachment and migration and interacts with fibrin, collagen and glycosaminoglycans; thus, it may be a factor in the focal proliferation of smooth muscle cells and collagen in atherosclerosis. We have measured, by rocket immunoelectrophoresis, the concentrations of soluble and collagenase-releasable fibronectin in normal human aortic intima and different types of atherosclerotic lesions. Soluble fibronectin concentration showed no significant difference between normal intima and lesions, but was 6-8-times higher than expected on the basis of plasma concentration and molecular mass. The concentration free in the interstitial fluid was about 3-times the expected level, suggesting that it originates from local synthesis as well as plasma insudation. In tissue, about half the fibronectin appeared to be reversibly associated with tissue components. Incubation with collagenase released fibronectin equal to twice the soluble fraction from normal intima and early proliferative lesions. In more advanced plaques that were accumulating lipid, the amount released was significantly higher (P less than 0.05) and more than 3-times the soluble fraction, suggesting that it might be involved in lipid accumulation. However, there was no correlation between release of fibronectin and bound low-density lipoprotein.

Mononuclear leukocyte beta 2-adrenergic receptors and adenylate cyclase sensitivity in insulin-dependent diabetes mellitus.

Patients with insulin-dependent diabetes mellitus (IDDM) have been found to have a heightened hyperglycemic response to epinephrine. To determine if patients with IDDM have increased sensitivity of cellular beta 2-adrenergic receptor-effector systems, we assessed beta 2-adrenergic receptors and adenylate cyclase sensitivities to isoproterenol in partially purified mononuclear leukocyte (MNL) plasma membranes from 10 patients with IDDM (without adrenergic neuropathy) and 10 matched nondiabetic controls. MNL beta 2-adrenergic receptor densities (Bmax = 48 +/- 8 fmol [3H] DHA/mg protein in IDDM, 44 +/- 3 fmol [3H] DHA/mg protein in controls) and binding affinities (apparent KD = 0.3 +/- 0.07 nM in IDDM, 0.3 +/- 0.04 nM in controls) did not differ. Further, MNL adenylate cyclase activities were not significantly different either at baseline (325 +/- 86 pmol/mg protein/15 min in IDDM, 275 +/- 49 pmol/mg protein/15 min in controls) or in response to isoproterenol (842 +/- 229 pmol/mg protein/15 min in IDDM, 608 +/- 86 pmol/mg protein/15 min in controls). Thus, the data do not support the presence of a generalized alteration of beta-adrenergic receptors or adenylate cyclase sensitivity in IDDM. To the extent that MNL beta 2-adrenergic receptors and adenylate cyclase activities reflect those of extravascular catecholamine target cells, these findings suggest that the heightened hyperglycemic response to epinephrine exhibited by patients with IDDM is not due to increased sensitivity of cellular beta 2-adrenergic receptor-effector systems and is best attributed to the altered hormonal milieu of the insulin-deficient state.

Separation of newborn rat epidermal cells on discontinuous isokinetic gradients of PERCOLL.

A one-step procedure has been developed for the separation of epidermal cells using PERCOLL (a new colloidal silica medium of low viscosity, osmolarity, and toxicity) for density gradient centrifugation. Newborn rat epidermal cells were dispersed with trypsin-EDTA and separated into 4 fractions in discontinuous isokinetic gradients. The cell fractions were characterized by their appearance in photomicrographs and their distribution by number and size. Preferential incorporation of 3H-thymidine and 14C-glycine, by basal and granular cells respectively, confirmed the identification of cell types. The basal cells, which were collected in the densest fraction (1.090), were the most homogeneous population with a mean diameter between 7-8 mum and showed 98% viability. The granular cells predominated in the least dense fraction (1.023). The intermediate fractions contained spinous cells admixed with the other cell types.

Evidence for histamine in the urticating hairs of Hylesia moths.

An urticarial dermatosis after contact with the urticating hairs of the adult female Hylesia moth may occur by several mechanisms including the intradermal injection of inflammatory mediators through the urticating hairs. Extracts were prepared from whole moths, urticating hairs, and other moth parts. Each of these extracts was subjected to a radioenzyme assay for histamine. Histamine was present in extracts made from whole moths and from urticating hairs. Extracts made from other moth parts contained no histamine. Cutaneous wheals occurred after intradermal injections of histamine and various concentrations of Hylesia extract (HE) into the backs of cynomolgus monkeys. This whealing response was suppressed by pretreatment of the animals with diphenhydramine hydrochloride, but not by pretreatment with indomethacin. Histologic examinations showed a perivascular lymphocytic infiltrate around dilated capillaries without evidence of mast cell degranulation in HE-injected sites but not in controls. These findings provide evidence that histamine may be the mediator responsible for the urticarial lesions seen after contact with Hylesia moths.

beta 2- and alpha 2-adrenergic receptors and receptor coupling to adenylate cyclase in human mononuclear leukocytes and platelets in relation to physiological variations of sex steroids.

In view of evidence, largely in animals, indicating effects of sex steroids on adrenergic receptors, we measured mononuclear leukocyte (MNL) beta 2-adrenergic receptors and adenylate cyclase sensitivity to stimulation by isoproterenol as well as platelet alpha 2-adrenergic receptors and sensitivity of sodium fluoride-stimulated adenylate cyclase to inhibition by epinephrine in 3 groups of normal humans with physiologically disparate levels of testosterone, estradiol, and progesterone (10 normal men and 10 normal women, the latter sampled in both the follicular and luteal phases of their menstrual cycles). Differences in testosterone, estradiol, and progesterone were as expected; testosterone levels were 10-fold higher in men, and progesterone levels were 20-fold higher in luteal phase women. T4, cortisol , and norepinephrine levels did not differ. Basal plasma epinephrine concentrations were slightly but significantly higher in luteal phase women [34 +/- 5 (+/-SE) pg/ml] than in follicular phase women (16 +/- 3 pg/ml; P less than 0.01) or men (20 +/- 3 pg/ml; P less than 0.05). There were no significant differences among these 3 groups in the densities or affinities of MNL beta 2-adrenergic or platelet alpha 2-adrenergic receptors or in the corresponding MNL and platelet adenylate cyclase sensitivities. Thus, there is not a generalized effect of physiological variations of testosterone, estradiol, and progesterone on adrenergic receptors or adenylate cyclase. To the extent that the adrenergic receptors and adenylate cyclase activities of circulating cells reflect those of extravascular catecholamine target cells, these data provide no support for a role of physiological variations of testosterone, estradiol, or progesterone in the regulation of catecholamine action in humans.

Haemostatic factors and atherogenesis.

It is increasingly realised that fibrin deposition and fibrin lysis are major factors in vascular pathology. In addition to thrombotic occlusion fibrin is a component of atherosclerotic lesions, but the increased interest in components of the haemostatic system was mainly triggered by clinical use of fribrinolytic agents, and the problems of re-stenosis following angioplasty. This review focuses on the main components of the fibrinolytic system--tissue plasminogen activator (tPA), urokinase (uPA) and plasminogen activator inhibitor (PAI-1)--and on thrombin. These factors are not only involved in fluid phase clotting and clot lysis; they react specifically with cells and matrix components. During the last 5 years, the main period under review, there have been numerous studies on their interactions with endothelial and smooth muscle cells in culture, in whole tissues and in vivo, and with arterial extracellular matrix of which a major component is fibrin. Plasminogen activators bind to cell surface receptors, influence cell migration and release active thrombin from fibrin. Thrombin emerges as a pluripotent factor which modulates many aspects of endothelial and smooth muscle cell behaviour, including release and synthesis of fibrinolytic components, and stimulation of cell proliferation.

Does lipoprotein(a) (Lp(a)) complete with plasminogen in human atherosclerotic lesions and thrombi?

Thrombotic occlusion is the major cause of myocardial infarction (MI), and fibrin accumulation appears to play a significant role in development of atherosclerotic lesions. Any factor that reduces the lysis of fibrin may thus increase the risk of MI, and it has been suggested that this accounts for the atherogenicity of the lipoprotein variant Lp(a). The characteristic feature of Lp(a) is an apoprotein which is homologous with part of the plasminogen molecule, and experiments in vitro suggest that it interferes with uptake and activation of plasminogen on cell surfaces and fibrin. The presence of Lp(a) also seemed to offer an explanation for the apparent absence of plasminogen from 70-80% of intimal samples. We have compared the levels of Lp(a) and plasminogen in normal intima and atherosclerotic lesions. In aortic intima there was no relation between Lp(a) and plasminogen, which was absent in some samples with no Lp(a), and present in others with high levels. In intravascular thrombi plasminogen was present at a rather constant concentration (16.3 +/- 4.6 micrograms/100 mg wet tissue), whereas Lp(a) varied over a 100 fold range (0-104 micrograms/100 mg). Plasminogen binds to fibrin and is activated on the fibrin clot, so levels in extracts may not fully represent Lp(a)/plasminogen interactions. After extraction the residual tissues and thrombi were treated with 1 M epsilon-aminocaproic acid (epsilon-aca) to elute lysine-bound components. Lp(a) was eluted from all but one intimal sample, confirming previous findings on its binding to fibrin in lesions, but there was no relation between the amounts of Lp(a) and plasminogen in the tissue eluates. Paradoxically, in the thrombi there was a weak positive correlation between Lp(a) and plasminogen in epsilon-aca eluates (r = 0.504, P = 0.05). These results do not support the hypothesis that Lp(a) displaces plasminogen in vivo, but the large amount of Lp(a) eluted by epsilon-aca suggests that its atherogenicity resides in preferential binding to fibrin, leading to increased lipid accumulation in lesions.

Distribution of plasma proteins across the human aortic wall--barrier functions of endothelium and internal elastic lamina.

The concentrations of plasma proteins of different molecular weights were measured in layers across the human aortic wall. On a volumetric basis the concentration of low density lipoprotein (LDL) in inner intima, adjacent to the endothelium, was almost twice the concentration in the patient's plasma. With the exception of transferrin, which behaved anomalously, the concentration of each protein was a linear function of is plasma concentration and molecular weight, so that the relative retention of albumin was only 15% of LDL retention and its concentration in inner intima less than one-quarter of the plasma concentration. Between the inner (luminal) and outer layers of intima the concentration of all proteins decreased by about 40%. In aortas in which the internal elastic lamina (IEL) appeared to be intact it provided an almost total barrier to LDL, but for smaller proteins the concentrations in the layer immediately outside it were inversely related to molecular weight; the concentration of LDL was only 0.3% of the intimal concentration whereas albumin was 26% of the intimal concentration. However, in aortas in which the IEL appeared morphologically frayed, fragmented or discontinuous there was an 80% increase in albumin, and a 25-fold increase in LDL in this layer. The differential barrier functions of endothelium and IEL produce bizarrely different macromolecular environments for smooth muscle cells in intima and media.

Variability of the electrophoretic mobility of low density lipoprotein. Comparison of interstitial fluid from human aortic intima and serum.

Interstitial fluid was collected from human aortic intima and the low density lipoprotein (LDL) was compared with serum LDL by two-dimensional immunoelectrophoresis. In half the samples derived from normal intima the migration rate was within +/- 10% of migration of LDL in serum, but in the remaining samples it was higher, indicating an increase in net negative charge on the molecule. By contrast, in most samples of interstitial fluid from gelatinous thickenings the migration rate of LDL was lower than in serum, indicating reduction in net negative charge. To test for the effect of different electrolyte concentrations in interstitial fluid and serum, migration of alpha 2-macroglobulin (alpha 2-M) was measured simultaneously in both. This showed no significant difference between interstitial fluid and serum, and varied by less than 2% between different serum samples. However, the mobility of LDL relative to alpha 2-M showed a remarkable variation, ranging from 80.9 to 127.1% of alpha 2-M mobility in interstitial fluid and from 58.6 to 115.9% in 23 serum samples. In serum, LDL relative mobility showed no correlation with general acid-base status, as indicated by serum bicarbonate levels, with fatty meals or diabetic keto-acidosis, or with extent of glycosylation of haemoglobin. This suggested that the variation in surface charge is intrinsic to the molecule and not a transient reflection of the plasma environment. Experimental alterations in the surface charge of LDL change its interaction with smooth muscle cells and macrophages in vitro, and its mitogenic properties. This raises the possibility that physiological variation in the surface charge of LDL in different subjects could alter its atherogenic potential.

Destruction of endogenous low density lipoprotein in incubated intima.

The effect of incubation on the content of endogenous intact plasma lipoprotein (LP) has been examined in minced samples of normal intima and lesions from 38 patients. Both the electrophoretically mobile and the immobilized LP fractions decreased on incubation, and the rate of destruction was proportional to LP concentration (r=0.832, p less than 0.001). Mincing the intima with EDTA before incubation increased the rate of destruction about 4-fold in fibrous lesions but not in lesions containing numerous fat-filled cells. The destruction of LP was highly dependent on pH; the rate was highest below pH 5.5 and destruction was almost completely inhibited above pH 6.4. In standard cathepsin assays haemoglobin substrate was hydrolysed at a rate comparable to the rate of destruction of LP. The results suggest that LP may be degraded by a lysosomal cathepsin in intima.

Factors influencing the accumulation in fibrous plaques of lipid derived from low density lipoprotein. II. Preferential immobilization of lipoprotein (a) (Lp(a)).

The extracellular lipid that accumulates in fibrous atherosclerotic lesions appears to be derived directly from plasma low density lipoprotein (LDL). One factor that may influence the lipid deposition is immobilization of part of the LDL in lesions, and an immobilized fraction can be released by incubation with the fibrinolytic enzyme, plasmin, suggesting that it is associated with fibrin. The lipoprotein variant Lp(a) is associated with increased risk of arterial disease, and its characteristic apoprotein, apo(a), is structurally related to plasminogen, suggesting that it might bind to the plasminogen binding sites on fibrin. In this study we have compared blood Lp(a) and the soluble and plasmin-releasable Lp(a) in 45 samples of normal intima and different types of lesion. Levels of soluble and plasmin-releasable Lp(a) were dependent on both blood level and type of tissue sample. Although the amount of soluble LDL was 5-20 times higher than Lp(a) in intima, the amounts released by plasmin were similar, and Lp(a) appears to account for most of the apo B-containing lipoprotein that is immobilized in lesions.

Fibrinolysis and plasminogen concentration in aortic intima in relation to death following myocardial infarction.

In samples of human aortic intima fibrin/fibrinogen degradation products (FDP) were assayed by isoelectric focussing/immunoelectrophoresis as a possible measure of endogenous fibrinolysis, and plasminogen concentration was assayed by rocket immunoelectrophoresis as a possible marker for fibrinolytic potential. No consistent differences were found between normal intima and different types of atherosclerotic lesion, but there was marked variation between patients, and multiple samples from the same aorta showed similar levels. There was no significant correlation with age, sex, or time after death. Low concentrations of FDP and failure to recover measureable amounts of plasminogen from intima were highly associated with death in patients who had suffered a recent myocardial infarction. In aortas from which 3 or more samples of intima and lesions were obtained (n = 16), no FDP were found in 3 (total of 12 samples); all of these were from patients who died following myocardial infarction. Low levels were present in the 4th patient with myocardial infarction. No plasminogen was found in 10 of 11 aortas from patients dying after myocardial infarction (total of 46 samples with no plasminogen), but it was present in 10 of 17 aortas from patients dying of other causes (X2 = 7.6, P less than 0.01). Where both were assayed, FDP were not found in any samples which did not contain plasminogen. Low levels of FDP and absence of plasminogen were associated with increased involvement with atherosclerosis. There was no relation between intimal and serum plasminogen levels, and prothrombin and low density lipoprotein were present in all samples from which no plasminogen was recovered. The results indicate that in some patients, particularly those dying after myocardial infarction, there is decreased fibrinolysis and fibrinolytic potential in the arterial intima, and this may result in increased intimal accumulation of fibrin.

Factors influencing the accumulation in fibrous plaques of lipid derived from low density lipoprotein. I. Relation between fibrin and immobilization of apo B-containing lipoprotein.

The lipid that accumulates in some fibrous atherosclerotic lesions appears to be derived from plasma low density lipoprotein (LDL). An early stage in lipid accumulation may be immobilization of a fraction of the LDL, and this is released by incubation with proteolytic enzymes, of which the most effective is the fibrinolytic enzyme, plasmin. We have examined the relationship between release of fibrin degradation products (FDP) and LDL in controlled plasmin incubations of 42 samples of normal intima and atherosclerotic lesions from aortas of 10 patients. In three patients (group 1) no LDL was released from any of the 11 tissue samples although they comprised lesions as well as normal intima. In 2 more patients (group 2) LDL was consistently low. However, in 5 patients (group 3) substantial amounts of LDL were released from all 21 tissue samples, and there was a significant correlation between the amounts of FDP and LDL (P less than 0.001). In spite of this correlation there were marked differences in the ratio FDP/LDL, but analysis by SDS-polyacrylamide gel electrophoresis and immuno blotting of the FDP released showed no consistent pattern related to LDL binding. Although the ratio FDP/LDL showed a 4-fold range, in 6 lesions subjected to successive 2-h incubations with plasmin the ratio within each lesion remained constant, supporting the concept that fibrin and LDL are linked.

Presence of growth-stimulating fibrin degradation products containing fragment E in human atherosclerotic plaques.

The key event in the formation of stenosing atherosclerotic lesions is widely thought to be smooth muscle cell proliferation, but the factors primarily responsible for initiating this remain uncertain. Previously we have shown that aqueous extracts of proliferative types of human atherosclerotic plaque stimulate cell proliferation in the chick chorioallantoic membrane (CAM). This has been attributed largely to the fibrin degradation products in the extracts, components removeable by affinity chromatography. We now demonstrate that the fibrinogen content of the extract, removeable by clotting out with thrombin, also makes a contribution to the activity by forming fibrin on the surface of the CAM. Affinity chromatography experiments using anti fragment D and E antisera indicate that activity resides in the E-containing fibrin fragments, consistent with previous work with FDP prepared in vitro.