Ex vivo localization of the mouse saposin C activation region for acid beta-glucosidase

Saposin C is a biological activator of acid beta-glucosidase (GCase), the lysosomal hydrolase with activity towards glucosylceramide (GC). In addition, saposin C possesses a functional domain that determines the in vitro and ex vivo neuritogenic effects of prosaposin, the precursor of saposins A, B, C, and D. The domains for enzymatic activation and neuritogenic function segregate in vitro, respectively, to the carboxyl- and amino-terminal halves of human and mouse saposin C. A chimeric mouse saposin C(1-8)B(8-28)C(30-80) was created to obliterate the neuritogenic region by substituting amino acids 9-29 of saposin C with amino acids 8-28 of saposin B. This saposin showed normal in vitro enzymatic activation effects toward GCase, but no neuritogenic activity. An altered prosaposin was made to contain the chimeric saposin C region. Expression of this altered or wild-type prosaposin was driven by the PGK-1 promoter as a transgene in prosaposin knock-out mice. In cultured fibroblasts from such mice, expressed saposins localized to the lysosomal compartments. Metabolic lipid labeling using L-[3-(14)C]serine showed retention or clearance of GC in prosaposin deficient or transgene reconstituted cells, respectively. In addition, sulfatide catabolism, that requires saposin B and arylsulfatase, was also normalized in prosaposin KO cells reconstituted with the transgenes. These data show that the transgenic prosaposins were expressed and processed to functional saposins in fibroblasts. These results also show that the enzymatic activation domain is located at carboxyl-terminal half of saposin C and functions only in the context of the general saposin structure.     

Association of haplotypes in the beta-chemokine locus with multiple sclerosis

Linkage studies in multiple sclerosis (MS) identified several susceptibility loci. One of these regions includes chromosome 17q11 where a meta-analysis of data from three genome scans suggested linkage. This region encodes a cluster of genes for beta-chemokines or CC chemokine ligands (CCLs), which may be involved in the development of MS lesions. Here we aimed to test if CCL alleles and haplotypes are associated with MS. Using methods of linkage and association, we observed deviations from the expected 50% transmission of haplotypes from unaffected parents to their affected children at CCL2, CCL11-CCL8-CCL13 and CCL3 within the investigated 1.85 MB chromosomal segment. Analyses of the linkage disequilibrium map support that variants with possible relevance to MS can be located within these subregions. Identification of MS associated CCL variants may have direct clinical significance, as it can lead to the design of small competitive antagonists of these molecules with beneficial effects in the treatment of patients with early and active disease.     

内皮祖细胞在血管组织工程的应用

内皮祖细胞是一群具有游走特性,能进一步增殖分化成为成熟内皮细胞的幼稚内皮细胞.内皮祖细胞参与了出生后缺血组织的血管发生和血管损伤后的修复.内皮祖细胞的发现为血管组织工程种子细胞增添了一个新来源.本文重点介绍成体内皮祖细胞的来源、鉴定、生物学特性、功能以及在血管组织工程中的应用.     

一步温育EIA法检测HBsAg

本文报道用尼龙网为载体的抗ＨＢｓＡｇ抗体膜作免疫吸附剂用于酶联免疫测定，对检测ＨＢｓＡｇ的免疫反应程序和反应如酶标／抗原溶液配比量和温育反时间等进行了研究。结果得到一步温充ＥＩＡ法的灵敏度与二步温育ＥＩＡ法相比没有下降，检测时间由原来的约３ｈ缩短至约５０ｍｉｎ。     

Quantitative distribution of rat brain monoamine oxidase A by [14C]clorgyline autoradiography

The distribution of functionally active monoamine oxidase type A (MAO-A) was investigated by in vivo quantitative autoradiography using [¹⁴C]clorgyline in normal, conscious rat brain. [¹⁴C]clorgyline was synthesized by the methylation reaction of N-desmethylclorgyline using [¹⁴C]methyliodide. Sixty minutes after [¹⁴C]clorgyline administration (1.58 MBq/animal i.v.), the brains were removed and prepared for autoradiography by washing the brain sections with 5% trichloroacetic acid solution to remove the nonbinding free tracer. The amount of MAO-A was calculated from the regional acid-insoluble tissue radioactivity and the specific activity of the tracer. The highest amount of MAO-A (5.84 nmol/g tissue) was found in the locus coeruleus. The interpeduncular nucleus, habenular nucleus, fasciculus retroflexus, and solitary tract nucleus possessed over 1.6 nmol/g tissue of MAO-A. Among 23 regions of interest, the lowest amount of MAO-A (0.37 nmol/g tissue) was found in the globus pallidus. The findings of this study suggest that the pattern of MAO-A parallels both in neuroanatomical distribution and in density that of norepinephrine and serotonin innervation. The MAO-A concentration was, however, relatively low in the dopamine-related areas. This corresponded to the previous results obtained by histochemical analysis. In addition, among the white matter structures, a high amount of MAO-A was found specifically in the fasciculus retroflexus.     

低频超声透皮给药的研究进展与应用

低频超声可以增强包括大分子药物在内的许多药物的透皮传输,其主要机制是超声的空化作用,大多数人认为是通过改变角质层角化细胞排列结构来提高皮肤渗透能力的.低频超声透皮给药已被人们用于离体实验和动物活体实验,到目前为止,无论是小分子透皮传输还是大分子透皮传输都有很多成功的例子.但是真正通过低频超声透皮导入药物进行治疗的临床应用报道很少,需要更进一步大量的临床试验以确定其安全性与实用价值.一旦其安全性得以证实,合适的低频超声透皮仪研制成功,低频超声快速透皮必将成为一种安全、有效、可控、经济的新型给药方式.     

Self-assembled biomimetic monolayers using phospholipid-containing disulfides

Several phospholipid-based disulfide molecules were synthesized and attached onto the gold-coated silicon wafer using the self-assembling method. The syntheses of these surface-modifying agents were conducted by introducing bromoethylphosphorate (PBr), phosphorylcholine (PC) or phosphorylethanolamine (PE) groups on the terminals of a dialkyl disulfide. After disulfides adsorption onto gold substrate surfaces, the composition, the film thickness, and the conformational order of self-assembled monolayer surfaces were explored and discussed in detail based on reflection-absorption infrared spectroscopy, contact angle measurement, Auger electron spectroscopy, X-ray photoelectron spectroscopy, and so on. The monolayer having the PBr end group could also be converted to a PC surface by treating with trimethylamine. The model functional surfaces of Au-SC11-PC, -PE, -PBr, -OH or corresponding mixed layers were used to mimic biomembrane surfaces. The monolayer having PC groups was found to reduce fibrinogen adsorption as evaluated from protein adsorption experiments using quartz crystal microbalance. It also showed relatively low platelet adherence compare to the glass, PBr and PE surfaces. The cell viability test also revealed that the PC surface displayed lower cytotoxicity than other surfaces.     

过继转输胚胎抗原耐受T细胞诱导自然流产孕鼠母胎免疫耐受

探讨过继转输胚胎抗原耐受T细胞对小鼠自然流产模型妊娠预后及宿主孕鼠免疫细胞对父系抗原免疫耐受状态的影响。以CBA/J×BALB/c为正常妊娠模型 ,CBA/J×DBA/ 2为自然流产模型 ,将自然流产模型CBA/J孕鼠于孕 4d (着床期 )分别腹腔注射大鼠抗小鼠CD80和CD86mAb或大鼠同型IgG。于孕 9d ,应用免疫磁珠阴性分选三组孕鼠的脾脏T细胞 ,并将三组T细胞分别转输至孕 4d的CBA/J×DBA/ 2孕鼠。于宿主孕鼠孕第 9天 ,采用单向混合淋巴细胞反应分析宿主孕鼠脾脏免疫细胞对父系抗原的增殖能力 ,并用流式细胞术分析经父系抗原刺激的宿主孕鼠脾脏T细胞内IL 2表达水平。于孕 1 4d分别观察宿主孕鼠的胚胎吸收率。结果显示 ,过继转输胚胎抗原耐受T细胞和转输正常妊娠模型孕鼠的T细胞均可诱导宿主孕鼠脾脏免疫细胞对父系抗原的增殖能力及IL 2的表达显著下降 (P <0 0 5 ) ,孕 1 4d胚胎吸收率也显著下降 (P <0 0 5 )。这些结果表明 ,于孕早期过继转输胚胎抗原耐受T细胞和转输正常妊娠模型孕鼠的T细胞能诱导宿主孕鼠母 胎免疫耐受 ,防止母体对胚胎的免疫排斥 ,从而使自然流产模型的妊娠预后达到正常妊娠水平。