Lp(a) lipoprotein enters cultured fibroblasts independently of the plasma membrane low density lipoprotein receptor

Lp(a) lipoprotein shares the apoB antigen with low density lipoprotein (LDL). The Lp(a) antigen is unique for Lp(a) lipoprotein. Fibroblast association (i.e. plasma membrane binding plus intracellular accumulation), plasma membrane binding, intracellular accumulation and degradation of 125I-Lp(a) lipoprotein were studied in strains from subjects with or without autosomal dominant hypercholesterolemia (HC). Subjects without HC (non-HCs) have cell surface receptors for low density lipoprotein (LDL receptors). On the average, HC heterozygotes have half-normal LDL receptor activity and "receptor-negative" HC homozygous cell strains lack functional receptors. Fibroblast processing of 125I-Lp(a) lipoprotein was compared to fibroblast processing of 125I-LDL. LDL receptor-dependent processing of 125I-LDL was saturated at about 50 microgram apo 125I-LDL.ml-1 in non-HC fibroblasts. 125I-Lp(a) lipoprotein was, however, largely processed independently of receptor mechanisms by non-HC cells (highest concentration examined 150 microgram apo 125I-Lp(a) lipoprotein . ml-1). Lp(a) lipoprotein did not interfere with 125I-LDL for fibroblast association, but inhibited 125I-LDL degradation. The interference with 125I-LDL degradation was time dependent. Only slightly higher 125I-Lp(a) lipoprotein processing values were found in non-HC and HC heterozygous strains than in "receptor-negative" HC homozygous strains. However, non-HC cells had more than tenfold higher 125I-LDL processing values than "receptor-negative" HC homozygous cells.     

In vitro studies of the interaction of calcium ions and other divalent cations with the Lp(a) lipoprotein and other isolated serum lipoproteins

The interaction of isolated Lp(a) lipoprotein with different divalent cations was studied and compared to that of other isolated lipoprotein classes.
Purified Lp(a) lipoprotein was found to be most sensitive to the metal ions tested, and the Lp(a) lipoprotein was the only lipoprotein which was precipitated by calcium ions alone. The precipitation apparently depends on the ionic radii of the cations used as well as on the lipoprotein class tested. The precipitation reaction between calcium ions and the Lp(a) lipoprotein, and the interaction between calcium ions and LDL (without precipitation) seem to follow the known rules for small ion - macromolecule interaction reasonably well. The calcium ion - Lp(a) lipoprotein interaction results in a small aggregate. The binding is of ionic type and the precipitation reaction is initially reversible. It was estimated that LDL particles have a mean of 290 equivalent and non-interacting binding sites for calcium ions.
The above observations concerning the Lp(a) lipoprotein may be of interest in view of the significantly higher frequency of early coronary heart disease in Lp(a+) than in Lp(a-) individuals, and in view of the previously reported biochemical differences between individuals of different Lp phenotype.     

Ultrastructural studies on the evolution of amyloidosis in the cyclic hematopoietic (CH) dog

Summary Electron microscopy studies were made on tissues of cyclic hematopoietic (CH) dogs of various ages presenting a high incidence of spontaneous amyloidosis. The distribution and morphologic characteristics of amyloidosis in this animal model closely correspond to the secondary and familial forms of the disease in humans. Plasma cells and, particularly, macrophages presented marked changes during the evolution of amyloid deposition. Residual bodies in the macrophages contained abundant cell debris, a result of both endocytic and autophagocytic activities. Intracellular amyloid fibrils were not observed by conventional electron microscopy. A few reticular cells contained intracytoplasmic fibrils which were morphologically different from amyloid. There was no correlation between the amount of intracellular fibrils and the size of the extracellular amyloid deposits. On the contrary, a temporal association between the magnitude of the amyloid deposits and cytoplasmic changes in the macrophages at sequential stages of the evolution of the disease was evident. It is suggested that the hematopoietic defect in the CH dog could play an important role in the production of amyloidosis, making this animal an excellent experimental model for studies of that disease.Supported by Grants NIH RR00874, NIH HL 15647, and NIH FR5541 from the National Institutes of Health, DHEW     

脂蛋白（a）及低密度脂蛋白与成纤维细胞表面结合特性的比较

脂蛋白（a)[Lipoprotein(a),Lp(a)]的代谢途径尚有争议。本文比较研究了Lp(a)和低密度脂蛋白（Low Density Lipoprotein,LDL)与成纤维细胞表面的结合特性,结果显示,Lp(a)与成纤维细胞呈特异性,高亲和力,可饱和性结合,但是Lp(a)与细胞的亲和力,最大结合容量均比LDL低,竞争性结合试验显示LDL只能部分抑制Lp(a)与成纤维细胞表面特异性的结合;细胞表面LDL受体活性的下调,可使LDL与细胞的结合量下降58％,而Lp(a)的结合量仅下降12．3％,这些结果表明LDL受体只能参与部分Lp(a)与细胞表面的结合及代谢。     

高脂血症兔与正常兔动脉平滑肌细胞体外生长及形态学观察

本文通过体外培养比较高脂血症兔动脉壁未发生斑块区的平滑肌细胞与正常动脉壁平滑肌细胞的生长代谢特性。结果表明,高脂血症兔SMC生长率明显高于正常兔SMC。并有高尔基氏体发达,内质网及线粒体增多且以不同程度扩张呈空泡状等形态改变。免疫胶体金标记观察提示高脂血症兔SMC在体外仍有较强的分解、代谢低密度脂蛋白及其载脂蛋白的能力。     

Ultrastructural studies on the formation of myofilaments and myofibrils in the human embryonic and adult hypertrophied heart

Summary The myofibrillogenesis in the human embryonic heart is described. The synthesis of thin filaments, which are the first to appear, takes place in close proximity to smooth surfaced SR tubules. Z-band material is closely related to the thin filaments and appears first as irregularly distributed patches in the filamenteous mass. Further cellular differentiation includes an organization of the thin filaments/Z-band material. The synthesis of thick filaments, which follows that of the thin filaments, takes place in ribosome rich areas of the cell. They are rapidly incorporated into the strings of organized thin filaments/Z-band material. The periodic binding sites on both kinds of filaments are believed to play an important role in the precise ordering of the filaments.The formation of myofilaments in the adult hypertrophied human heart is also described. The similarities between this process and that observed in the embryonic heart are striking, and we believe it to be the same process.     

Lp(a) lipoprotein is a major risk factor for cardiovascular disease: pathogenic mechanisms and clinical significance

The results of two previous and two recent studies of middle-aged males and females are presented to exemplify the clinical importance of lipoprotein (a) (Lp(a)) as a risk factor for atherosclerosis and coronary heart disease. In these studies various conventional and recently suggested risk factors were included and different methods for Lp(a) quantification were used. Lp(a) was a significant risk factor in all four studies. In the recent prospective case-control study, Lp(a) and cholesterol were found to act synergistically and predict primary acute myocardial infarction in Swedish males. A cholesterol level above 6.5 mmol/1 increased the risk of acute myocardial infarction if the Lp(a) level was above 200 mg/1. The plasma apo A-I level was a protective factor. In the other recent case-control study, an Lp(a) level above 500 mg/1 was a highly significant risk factor in Black and White US women with myocardial infarction or advanced coronary artery disease in addition to low density lipoprotein cholesterol levels above 130 mg/dl. A high apo A-I level was a protective factor. In these studies no other factors tested reached significance in multivariate logistic regression analysis. A hypothetical association between high Lp(a) levels and intracellular infection with Chlamydia pneumoniae is discussed. The results suggest that the Lp(a) level is useful in identifying high-risk individuals. Lowering low density lipoprotein cholesterol below 100 mg/dl (7lt;2.6 mmol/1) seems to be most important in both males and females with high-risk Lp(a) levels.     

In vitro studies of the interaction of isolated Lp(a) lipoprotein and other serum lipoproteins with glycosaminoglycans

The interaction of isolated Lp(a) lipoprotein or other lipoprotein classes with different glycosaminoglycans (GAG) bound to activated Sepharose was studied. In contrast to LDL, the Lp(a) lipoprotein did not bind to the GAG tested if sodium was used as a buffer cation. In the presence of Ca++, however, even the Lp(a) lipoprotein was bound to GAG. This type of binding, probably mediated by divalent cation bridges, is apparently not a simple function of the GAG used. Addition of GAG in solution revealed that this binding may be the only one existing under physiological conditions, and it appears possible that the Lp(a) lipoprotein is bound more firmly to GAG than is LDL under such conditions.     

Observations on the ultrastructure and function of the so-called “microfold” or “membraneous” cells (M cells) by means of peroxidase as a tracer

Summary 79 NMRI mice and Wistar rats were used for ultrastructural investigations of the sequential uptake of horseradish peroxidase (HRP) by M cells. In addition the ultrastructure of the so-called tuft-cells was reported.HRP, a foreign protein antigen, was applied either by injection (Owen 1977), or by stomach tube. After variable exposure times (5 min to 3 h) segments of the distal small intestine, containing Peyer's patches, mesenteric lymph nodes and liver tissue were removed. After fixation, they were reacted with H₂O₂-3,3-diaminobenzidine tetrachloride and were examined by light and electron microscopy for HRP reaction products. The uptake of HRP mainly occurs through the M cells in the dome epithelium of Peyer's patches with a continual transport of the antigenic material into lymphoid cells, macrophages, and dendritic reticulum cells. In the 3 h specimens a few single HRP-positive lymphoid cells can be observed within the efferent lymphatics of Peyer's patches. In addition, a continual uptake of HRP by necrobiotic enterocytes was observed. It has also been shown that after 3 h HRP is located inside the Kupffer cells of the liver. These findings also support the presumption that antigenic material can be transmitted via the portal circulation. However, definite, quantitatively and permanently recorded uptake of HRP by brush border cells was not be observed.To exclude a toxic effect of the applied HRP on the enterocytic epithelium additional resorptive-physiological investigations were performed using the in vivo-perfusion-recirculation method and in vitro-accumulation of L-phenylalanine.This study was supported by a grant from Deutsche Forschungsgemeinschaft (Ot 53/4-6)     

Studies on the structure and function of the apolipoprotein(a) gene

Lp(a) is an LDL-like lipoprotein that is a major inherited risk factor for atherosclerosis. It is distinguished from Lp(a) by the addition of apolipoprotein(a). The gene structure of apolipoprotein(a) is homologous to plasminogen, and competition with plasminogen activity may account for some of the pathophysiology associated with Lp(a). Six highly related genes have now been identified, and at least four are found in close proximity in overlapping genomic clones. Studies have begun on the regulation of apolipoprotein (a) gene expression, and the human apolipoprotein(a) gene has been inserted into transgenic mice, where it leads to the development of arterial lesions.     

The effect of percutaneous oestrogen replacement therapy on Lp(a) and other lipoproteins

Objectives: To determine the effect of percutaneous oestrogen replacement therapy on lipoprotein (a) and other plasma lipoproteins. Methods: Open longitudinal prospective study conducted at the hormone replacement clinic of the Prince of Wales Hospital, New Territories, Hong Kong. Thirty women who had undergone a total abdominal hysterectomy and bilateral salpingo-oophorectomy for benign gynaecological conditions were treated with 1.5 mg of percutaneous 17β-oestradiol gel applied daily for a period of 12 consecutive months. Measurements of plasma lipoproteins were made before the commencement of treatment and repeated at 6- and 12-month intervals. Results: There was a significant reduction in the concentrations of Lp(a) during the first 6 months of treatment, with median values falling from 7.87 mg dL⁻¹ to 6.16 mg dL⁻¹ (P = 0.004, 0–6 months). During the second 6 months, the median concentration increased to 9.38 mg dL⁻¹, (P = 0.072, 66-12 months), which did not significantly differ from the baseline level (P = 0.545, 0–12 months). Significant reductions in the concentrations of apoprotein A-I (apo A-I), apoprotein B (apo B), high density lipoprotein cholesterol (HDL-C), and HDL₃-C were also present after 6 months (P = 0.043, 0.049, 0.028, 0.013, respectively), but there were no differences between the baseline values of these lipoproteins and those at the completion of the study (P = 0.948, 0.244, 0.839, 0.117 respectively). Drug compliance was maintained throughout the study, with similar mean oestradiol concentrations at 6 and 12 months. Conclusions: The percutaneous administration of 17β-oestradiol has variable short term effects on plasma lipoproteins which are not maintained over a longer duration of treatment. By avoiding the ‘first pass’ effect on the liver, this method of delivery does not appear to produce the sustained changes in lipoproteins seen with oral treatment.     

Confirmation of an influence of the inherited Lp(a) variation on serum insulin and glucose levels

Previously reported analyses on three different series of people suggested that fasting serum insulin levels are lower in males with (high levels of) serum Lp(a) lipoprotein (Lp(a+)) than in males without detectable Lp(a) lipoprotein (Lp(a-)). The same was observed during an oral glucose tolerance test. Also, blood glucose concentrations tended to be lower in males with high levels of Lp(a) lipoprotein than in those in whose serum no Lp(a) lipoprotein could be detected. In this paper, we present data which appear to confirm the previously reported results. A significant correlation was found between the fasting triglyceride level and the sum of insulin values determined during the oral glucose tolerance test in healthy Lp(a-) but not in Lp(a+) individuals. The present data, together with those previously reported on an effect of the Lp(a) locus on serum lipid levels and on propensity to contract coronary heart disease, indicate that the genetically determined Lp(a) lipoprotein may be of considerable clinical importance.     

The inheritance of sinking-pre-beta lipo-protein and its relation to the Lp(a) antigen*

We have investigated the genetics of plasma sinking-pre-beta lipoprotein (spβ) as determined by the method of Breckenridge and Maguire, using several approaches: (i) a population study, (ii) a twin study and (iii) the use of family data. In addition, by the use of split samples, the spβ level as determined by us was correlated with Lp(a) typing carried out in Oslo by Dr. Kåre Berg. Although the spβ level is a continuous character, the results clearly showed it to be to a considerable extent under the control of the major autosomal gene pair constituted by the alleles Lp^a and Lp, which mainly control the production of the Lp(a) antigen, Lp^a being dominant. In our data the boundary between the LpLp and Lp^a Lp genotypes appeared to fall between the spβ 2 and 3 mg% levels, while that between Lp^a Lp and Lp^a Lp^a was in the 15 mg% region. These boundaries, which were inferred from both typing and population statistics, received good confirmation from the family data. It appears that some 88% of the variation in spp level is directly ascribed to segregation of Lp^a and Lp. On the basis of the twin study and other data, we conclude that the residual observed variation in spβ is almost entirely ascribed to analytical error of determination and polygenic effects, the influence of environment being negligible. The heritability is close to 100%.     

Detection of the Pro664-Leu mutation in the low-density lipoprotein receptor and its relation to lipoprotein(a) levels in patients with familial hypercholesterolemia of Dutch ancestry from The Netherlands and Canada

From a large cohort of hyperlipidemic patients, who attended the Lipid Research Clinic in Amsterdam, The Netherlands and in Vancouver, Canada, 915 consecutive patients with familial hypercholesterolemia (FH) of Dutch descent, were selected. This group of FH patients was screened for the presence of a cytosine to thymidine nucleotide substitution in exon 14 of the LDL-receptor gene, in order to determine the frequency of this mutation in patients of Dutch descent and to investigate the relationship between the mutation and the level of lipoprotein(a). The mutation was detected in seven individuals. All patients with this mutation shared the same haplotype, which is suggestive of an ancient mutation. The index patients and a large kindred with this mutation were further analyzed at the biochemical and clinical level. Except for total and LDL-cholesterol, there was no statistically significant difference in biochemical parameters between family members with and without FH. In contrast to previous reports, there was no difference in plasma levels of lipoprotein(a) between patients with the mutation in exon 14 and unaffected individuals.     

Detection of the Pro664-Leu mutation in the low-density lipoprotein receptor and its relation to lipoprotein(a) levels in patients with familial hypercholesterolemia of Dutch ancestry from The Netherlands and Canada

From a large cohort of hyperlipidemic patients, who attended the Lipid Research Clinic in Amsterdam, The Netherlands and in Vancouver, Canada, 915 consecutive patients with familial hypercholesterolemia (FH) of Dutch descent, were selected. This group of FH patients was screened for the presence of a cytosine to thymidine nucleotide substitution in exon 14 of the LDL-receptor gene, in order to determine the frequency of this mutation in patients of Dutch descent and to investigate the relationship between the mutation and the level of lipoprotein(a). The mutation was detected in seven individuals. All patients with this mutation shared the same haplotype, which is suggestive of an ancient mutation. The index patients and a large kindred with this mutation were further analyzed at the biochemical and clinical level. Except for total and LDL-cholesterol, there was no statistically significant difference in biochemical parameters between family members with and without FH. In contrast to previous reports, there was no difference in plasma levels of lipoprotein(a) between patients with the mutation in exon 14 and unaffected individuals.     

Ontogeny of the endocrine cells of the intestine and rectum of sea bass (Dicentrarchus labrax L.): an ultrastructural study

Several endocrine cell types were ultrastructurally characterized during the differentiation of the intestine and rectum of sea bass (Dicentrarchus labrax L.) larvae. Only one cell type (type I) was found in the posterior region of the undifferentiated gut of 5-day-old larvae (phase I). Types V and VI were found in both the intestine and rectum, types II, III and IV in the intestine, and types VII and VIII in the rectum of 9- and 12-day-old larvae (phase II), the rectum alone showing signs of functional differentiation. In phase III larvae, in which both the intestine and rectum were differentiated, types IX, X, XI, XII, XIII, XIV and XV were found in the intestine, only types X, XI and XII being seen in the rectum. Besides these, a new cell type, XVI, was observed in the intestine of 55- and 60-day-old larvae (phase IV), in which the digestive tract was completely differentiated. The endocrine cells appearing in phases I and II showed very scarce secretory granules and the ultrastructural features of undifferentiated cells. Some endocrine cell types in the earliest developmental stages were related to some of those found later. A maturational process of the endocrine cell types paralleled the differentiation of the intestine and rectum, with an apparent increase in the number of secretory granules accompanying organelle development.     

Apolipoprotein(a) is a human vascular endothelial cell agonist: studies on the induction in endothelial cells of monocyte chemotactic factor activity

Elevated levels of lipoprotein(a), Lp(a), are associated with premature atherosclerosis; however, the mechanisms of its atherogenicity are not known. Recruitment of monocytes to the blood vessel wall is an early event in atherogenesis. Since Lp(a) is associated with macrophages in the plaque, we have examined the effect of Lp(a) on inducing monocyte chemotactic activity (MCA) in vascular endothelial cells. We report that Lp(a) and apo(a) induced human umbilical vein (HUVEC) and coronary artery endothelial cells to secrete monocyte chemotactic activity as early as 30 min of incubation. In the absence of cells, Lp(a) had no direct monocyte chemotactic activity. Actinomycin D and cycloheximide inhibited the HUVEC response, indicating that protein and RNA synthesis were required. Endotoxin was shown not to be responsible for the induction of monocyte chemotactic activity. Granulocyte monocyte-colony stimulating factor antigen was not detected in the Lp(a)-conditioned medium, nor was monocyte chemoattractant protein-1 mRNA induced by Lp(a). These results suggest that Lp(a) may be involved in the recruitment of monocytes to the vessel wall, thus providing a novel mechanism for the participation of Lp(a) in the atherogenic process.     

Immunological and virological investigation in patients with lymphoadenopathy syndrome and in a population at risk for acquired immunodeficiency syndrome (AIDS), with particular focus on the detection of antibodies to human T-lymphotropic retroviruses (HTLV III)

Thirteen patients affected with unexplained lymphoadenopathy, fever, weight loss, and diarrhea (lymphoadenopathy syndrome; LAS) were evaluated for the possible appearance of acquired immunodeficiency syndrome (AIDS) and for immunological and virological characterization. The patients belonged to categories of individuals at risk for AIDS and were homosexual and/or drug abusers or hemophiliacs. Lymph node biopsy showed the histological picture of a follicular hyperplasia. The study of cell-mediated immunity (CMI), humoral immune response, and natural killer (NK) activity demonstrated a significant decrease in T cells with the helper/inducer phenotype (OKT ₄ ⁺ cells) and a relatively increased number of lymphocytes with the suppressor/cytotoxic phenotype (OKT ₈ ⁺ cells). NK activity was significantly lower than in normal controls. Thein vitro response to policlonal activators (phytohemagglutinin; PHA) and the cutaneous responsiveness to recall skin tests were impaired, whereas immunoglobulin production was increased, mainly in the IgG fraction. Virological studies showed high serum antibody titers to cytomegalovirus (CMV) but a lack of specific CMI as assayed by the leukocyte migration inhibition test (LMIT). CMV was also isolated from the urine specimen of one patient. The antibody pattern to Epstein-Barr virus (EBV) showed the uncommon contemporary presence of both Epstein-Barr nuclear antigen (EBNA) and early antigen (EA) antibodies. Antibodies to human T-lymphotropic retroviruses (HTLV III) were positive in 10 patients and the virus was isolated in 3 of them. In some patients the presence of serum antibodies to HTLV III was not associated with an impairment of the immune function. A group of individuals at risk for AIDS without LAS was also evaluated for the presence of HTLV III antibodies; the percentage of positive sera was 11.4. Nevertheless, individuals without specific antibodies had immunological abnormalities resembling those of LAS HTLV III-positive patients. The possible implications of these findings are discussed.