Acquired α2 macroglobulin on the surface of human lymphoblastoid cells

The molecular nature of the membrane antigen that is acquired from FCS-containing media by human lymphoblastoid cells has been investigated. The presence of bovine α₂, macroglobulin on the surface of Namalva cells was demonstrated by radioimmunoassay using specific antisera. Alternatively, cell-bound bovine α₂,M could be detected by the more sensitive heterophile rosette assay described previously. Namalva cells grown in NHS-containing media acquired bovine α₂ M upon subsequent incubation with the purified protein in a dose- and time-dependent way. Acquisition of α₂ M was demonstrated using both viable and formaldehyde-fixed cells. Purified fetuin, which carries a heterophile epitope shared with bovine α₂ M as well as with other glycoproteins, failed to bind with the membrane of Namalva cells. The possible role of acquired α₂ macroglobulin on cells has been discussed.     

Novel L2HGDH mutations in 21 patients with L-2-hydroxyglutaric aciduria of Portuguese origin

We studied 21 patients, from 18 families, with L-2-hydroxyglutaric aciduria (L-2-HGA), a rare neurometabolic disorder with a homogeneous presentation: progressive neurodegeneration with extrapyramidal and cerebellar signs, seizures, and subcortical leukoencephalopathy. Increased levels of L-2-hydroxyglutaric acid in body fluids proved the diagnosis of L-2-HGA in all 21 patients. We analyzed the L-2-HGA gene (L2HGDH), recently found to be mutated in consanguineous families with L-2-HGA, and identified seven novel mutations in 15 families. Three mutations appeared to be particularly prevalent in this Portuguese panel: a frameshift mutation (c.529delC) was detected in 12 out of 30 mutant alleles (40%), a nonsense mutation (c.208C>T; p.Arg70X) in 7/30 alleles (23%), and a missense mutation (c.293A>G; p.His98Arg) in four out of 30 mutant alleles (13%), suggesting that common origin may exist. Furthermore, two novel missense (c.169G>A; p.Gly57Arg, c.1301A>C; p.His434Pro) and two splice error (c.257-2A>G, c.907-2A>G) mutations were found. All the mutations presumably lead to loss-of-function with no relationship between clinical signs, progression of the disease, levels of L-2-HGA and site of the mutation. In the three remaining families, no pathogenic mutations in the L-2-HGA were found, which suggests either alterations in regulatory regions of the gene or of its intervening sequences, compound heterozygosity for large genomic deletion and, or further genetic heterogeneity.     

Codon optimization of the human papillomavirus 11 (HPV 11) L1 gene leads to increased gene expression and formation of virus-like particles in mammalian epithelial cells

The 505 amino acid L1 protein of the human papillomavirus type 11 (HPV 11) is the major capsid polypeptide that has been shown to self-assemble into virus-like particles (VLPs) in vivo and in vitro. While L1 is essential for viral infection, expression studies in mammalian cells have been hampered by different codon preference between the virus and its host. To optimize L1 gene expression in mammalian cells, we converted wild-type HPV 11 L1 (11 L1wt) codons to those more common in human genes. The modified HPV 11 L1 gene (11 L1h) generated protein levels that were at least 100-fold higher than those of wild-type HPV 11 L1, while no obvious differences were seen in the level of mRNA. HPV 11 L1 protein was detected in mammalian epithelial and fibroblast cells, by immunoblotting and indirect immunofluorescence (IIF) techniques. Unlike the situation in situ, IIF revealed the presence of L1 mainly at perinuclear sites. Virus-like particles assembled intranuclearly only to a low extent, as indicated by transmission electron microscopy. DNA vaccination using the HPV 11 L1h gene yielded a drastic increase in L1-specific antibody production in mice as compared to immunization with the wild-type gene.     

Immunocytochemical localization of cell adhesion molecule L1 in developing rat pyramidal tract

L1 is a representative of a family of carbohydrate neural cell adhesion molecules. The expression of L1 was studied during postnatal development of the rat pyramidal tract by immunohistology using polyclonal antibodies to L1 in spinal cord cervical intumescences. On postnatal day 1 (P1), L1 immunoreactivity was present in the entire dorsal funiculus, consisting of the ascending fasciculus gracilis and fasciculus cuneatus and the descending pyramidal tract. At that time the cervical pyramidal tract contains the first outgrowing corticospinal axons. At P4 both the fasciculus gracilis and the pyramidal tract are immunoreactive whereas the fasciculus cuneatus is negative. At P10 the pyramidal tract is intensely labelled whereas both ascending bundles are negatively stained. In the period between P4 and P10 the pyramidal tract is characterized by a massive outgrowth of corticospinal axons. During pyramidal tract myelination, between P10 and the end of the third postnatal week (P21), L1 immunoreactivity is progressively reduced. These observations suggest that L1 may play a prominent role in outgrowth, fasciculation and the onset of myelination of rat pyramidal tract axons. The differential L1 immunoreactivity of the pyramidal tract and the earlier developing ascending systems in rat dorsal funiculus indicate that this polyclonal antiserum is a useful differentiating marker for outgrowing fibre tracts.     

表面磁性膜医用316L不锈钢支架的生物相容性

目的 对表面磁性膜血管内支架进行生物相容性研究,为该支架的临床应用提供实验依据.方法 通过溶血实验、动态凝血时间实验、急性全身毒性实验、皮内刺激实验、细胞毒性实验、热源实验、过敏实验、体内植入实验综合评价表面磁性膜血管内支架的生物相容性.结果 表面磁性膜血管内支架无溶血反应及凝血功能的改变,无急性全身毒性反应,无热源反应,支架材料中不存在致敏性物质;支架材料动物体内植入在初期有轻度的炎性反应,12周后炎性反应基本消失,未见炎性细胞浸润积聚现象.结论 表面磁性膜血管内支架具有良好的生物相容性,其应用于临床具有可行性和安全性.     

Papain-induced lung injury: alterations in connective tissue metabolism without emphysema

The acute effects of the deposition of papain in the lungs of 4-month-old rabbits have been investigated with an amount not sufficient to cause emphysematous changes. During the first 2 days of in vivo deposition when hemorrhaging and inflammation occur, the total amounts of collagen and elastin in the lung, into which the papain had been instilled, did not change but the concentrations of collagen and elastin decreased 20 and 25% respectively. The rates of synthesis of collagen and other proteins doubled. By 4 weeks, collagen and elastin concentrations returned to, or were slightly above, control levels. The histological appearance and mean linear intercepts of lung sections and the compliance curves of excised lungs of the treated animals were similar to controls by 2 weeks after papain administration. The rabbit lung thus appears able to repair mild injury due to proteolytic enzymes.     

A comparison of the bronchodilator activity of Sch 1000 and salbutamol.

The effects of the beta 2-adrenergic agonist, salbutamol, 200 mug, and the cholinergic antagonist, Sch 1000, 40 mug, have been compared in 25 asthmatic patients using a single dose, double-blind, crossover trial design. Salbutamol aerosol produces a greater degree of bronchodilatation than Sch 1000 aerosol during the initial three hours following drug administration. There is no significant difference in the bronchodilator effects of the two drugs in the interval four to eight hours after drug administration. Nonatopic patients showed less difference in bronchodilator response to each of the two drugs than atopic patients. Neither drug showed any significant adverse effect on blood pressure, pulse rate, or electrocardiogram. In six asthmatic patients the effect of the combination of salbutamol, 200 mug, and Sch 1000, 40 mug, was evaluated. The combination produced a longer duration of bronchodilatation than either drug alone when compared to placebo.     

Crystal Growth in Poly(L‐lactide) Thin Film Revealed by in situ Atomic Force Microscopy

Direct visualization of crystal growth in poly(L ‐lactide) thin films was carried out by using a temperature‐controlled atomic force microscopy (AFM). At the initial stage of crystallization, edge‐on lamellar crystals have nucleated and elongated. Subsequently, the edge‐on lamellar crystals showed S‐shaped morphology and changed their orientation from edge‐on manner to flat‐on one. The curvature of edge‐on lamellar crystal has been discussed in terms of inclination and distortion of polymer chains in the crystal. In addition, mechanism on the formation of flat‐on crystal from edge‐on lamellae was proposed as derivative growth on the basis of in situ AFM observation of crystal growth and enzymatic degradation.

     

Analysis of CD154 and CD40 expression in native coronary atherosclerosis and transplant associated coronary artery disease

T cells have roles in the pathogenesis of native coronary atherosclerosis (CA) and transplant-associated coronary artery disease (TCAD). The mechanisms by which T cells interact with other cells in these lesions are not fully known. CD154 is an activation-induced CD4⁺ T cell surface molecule that interacts with CD40⁺ target cells, including macrophages and endothelial cells, and induces the production of pro-inflammatory molecules, including CD54 (ICAM-1) and CD106 (VCAM-1). To investigate whether CD154-CD40 interactions might be involved in the pathogenesis of CA or TCAD we performed immunohistochemical studies of CD154 and CD40 expression on frozen sections of coronary arteries obtained from cardiac allograft recipients with CA (n=10) or TCAD (n=9). Utilizing four different anti-CD154 mAb we found that CD154 expression was restricted to infiltrating lymphocytes in CA and TCAD. CD40 expression was markedly up-regulated on intimal endothelial cells, foam cells, macrophages and smooth muscle cells in both diseases. Dual immunolabeling demonstrated many CD40⁺ cells co-expressed CD54 and CD106. The extent of CD40, CD54 and CD106 expression showed statistical significant correlation with the severity of disease and the amount of intimal lymphocytes. Together these studies demonstrate the presence of activated CD154⁺ and CD40⁺ cells in both CA and TCAD lesions and suggest that CD154-mediated interactions with CD40⁺ macrophages, foam cells, smooth muscle cells and/or endothelial cells may contribute to the pathogenesis of these diseases. Received: 17 December 1999 / Accepted: 20 January 2000     

Purification and enzymatic activity of an NADH-fumarate reductase and other mitochondrial activities of Leishmania parasites

A 65 kD membrane-associated NADH-fumarate reductase subunit, which has a molecular weight similar to that of one of the enzyme subunits from bacteria, was purified from Leishmania donovani promastigotes. NADH-fumarate reductase and other mitochondrial enzymatic activities of L. major and L. donovani promastigotes and amastigotes were investigated. The presence of NADH-fumarate reductase was demonstrated in digitonin-permeabilized L. major promastigotes and mitochondria of L. major and L. donovani promastigotes and amastigotes. The activity of solubilized NADH-fumarate reductase was measured in L. major and L. donovani promastigotes. Succinate exhibited a clear concentration-dependent inhibitory effect on fumarate reductase, whereas fumarate also exhibited a clear concentration-dependent inhibitory effect on succinate dehydrogenase. The data indicate that fumarate reductase is an obligatory component of the respiratory chain of the parasite. Since the enzyme is an important component in the intermediate metabolism in the Leishmania parasite and is absent in mammalian cells, it could be a potential target for antileishmanial drugs.