Characterization of human alpha fetoprotein charge microheterogeneity during fetal development.

Aliquotes of human amniotic fluid (AF), fetal serum (FS), and cord blood (CB) were obtained as by-products of routine clinical diagnostic procedures at term or in the second trimester of pregnancy. When samples of CB were applied to a pH 5.5-4 chromatofocusing gradient, three isoforms of AFP could be resolved; a pl 4.57 form (isoform IA, 52% AFP), a pl 4.27 form (isoform IB, 43% AFP), and one species that was bound to the column but could be eluted with 1.0 M NaCl (isoform II, pl less than 4.00, 5% AFP). Term AF displayed a profile similar to that observed in term CB. When samples of 15-20-week gestation AF were chromatofocused, the immunoreactive AFP recovered was distributed between isoform IA and IB (60%) and isoform II (40%). FS and AF obtained from same pregnancy (23-26 weeks) displayed an identical chromatofocusing profile. Aliquotes of AF subjected to conA revealed 83% reactive variants compared with greater than 95% reactive variants for CB. FS displayed a conA profile identical to CB. When individual CB charge isoforms were isolated and subjected to conA analysis, greater than 97% of the AFP bound to conA. In contrast, when AFP isoform IA and IB were isolated from midgestation AF, approximately 22% of the AFP did not bind to the lectin while 100% of isolated AFP isoform II eluted as the reactive variant. These data suggest that human AFP exists as at least three charge and two lectin variants and that the charge profile may change during fetal development.     

Ethanol Inhibits Basic Fibroblast Growth Factor-Mediated Proliferation of C6 Astrocytoma Cells

Abstract: Early ethanol exposure alters the proliferative activity of glial and neuronal precursors in the developing CNS. The present study tests the hypothesis that ethanol-induced alterations in cell proliferation result from interference with growth factors. An in vitro model of astroglia (C6 astrocytoma cells) was used to study the effects of ethanol on proliferation mediated by basic fibroblast growth factor (bFGF). bFGF stimulated the proliferation of C6 cells. This bFGF-enhanced proliferation was evident by increases in total cell number, DNA synthesis (as measured by [³H]thymidine incorporation), and the number of cells that took up bromodeoxyuridine. A synthetic peptide that specifically blocked the binding of bFGF to its high-affinity receptor completely abolished the proliferation-promoting effect of bFGF. The action of another mitogen for C6 cells, insulin-like growth factor-1, was not affected by this peptide. Therefore, the bFGF-stimulated proliferation was mediated through a specific bFGF receptor. Ethanol inhibited bFGF-mediated proliferation in a concentration-dependent manner. Ethanol concentrations of 100 and 200 mg/dl partially inhibited bFGF-mediated proliferation (by 58 and 74%, respectively), whereas concentrations of ≥400 mg/dl completely abolished the growth-stimulating effect of bFGF. Our data show that ethanol alters proliferative activity of C6 cells by disrupting the action of bFGF. The target of ethanol neurotoxicity is a receptor-mediated activity. bFGF can affect cell proliferation by a non-receptor-mediated intracellular pathway, but ethanol does not have an impact on this pathway.     

猴头菇对小鼠抗疲劳作用的实验研究

分别以猴头菇干粉（猴头菇Ⅰ组）和猴头菇浸出液（猴头菇Ⅱ组）饲喂小鼠,观察猴头菇对小鼠血清乳酸脱氢酶（ＬＤＨ）活力、血乳酸、血清尿素氮（ＢＵＮ）、肝糖原、肌糖原含量及运动耐力的影响。结果表明：实验６０ｄ后,猴头菇Ⅰ、Ⅱ组ＬＤＨ活力、肝糖原及肌糖原含量明显高于对照组（Ｐ＜０．０５或Ｐ＜０．０１）;运动后血乳酸的水平和ＢＵＮ的增量明显低于对照组（Ｐ＜０．０５或Ｐ＜０．０１）;运动后血乳酸消除速率显著高于对照组（Ｐ＜０．０５）;在运动耐力测定时在水中淹死的时间比对照组长得多（Ｐ＜０．０５）。提示：猴头菇具有明显的增强运动能力和解除疲劳的作用。     

Membrane receptors for estrogen,progesterone, and testosterone in the rat brain: Fantasy or reality

Summary 1. There are numerous circumstantial evidence supporting the concept that steroid hormones control cellular function by means other than the nuclear receptor steroid binding mechanism. It is the intent of this report to present evidence indicating that steroids bind to specific sites in neuronal membranes.2. Some of the criteria to define steroid membrane receptors using steroid-BSA conjugates that can be radioiodinated to desired specific activity have been fulfilled for each of the three sex steroids using crude synaptosomal membrane preparations (P2 fractions) from the CNS of female and male rats. Ligand binding for each of the three steroids indicate high-affinity and high-capacity sites with distinct brain selectivity and stereospecificity. For example, 17-E-6-[¹²⁵I]BSA binds hypothalamic P2 fractions (HYP-P2) with an estimatedK _d of about 3±0.7 nM (X ± SE;n=3), whereas the cerebellum P2 (CB-P2) fractions bind the ligand with aK _d of 34±7 nM and, aB _max of 3 and 42 pmol/mg protein, respectively. Estrogen and testosterone binding fit best a one-single site, while progesterone binding sites can be best represented by a two-binding site, one high-affinity (K _d=1–2 nM) and one low affinity (K _d=62 nM), in CB-P2 fractions from intact adult female rat brain. Kinetics studies for T-3-[¹²⁵I]BSA indicate that the estimatedK _d of 30±2 nM for the olfactory bulb P2 fractions (OB-P2) from male rats is in good agreement withK _d values computed from Scatchard-derived data using the LIGAND algorithm.3. 17-E-6-[¹²⁵I]BSA binding sites are stereospecific and appears to be present as early as 5 days of age in both the OB- and the CB-P2 fractions without changes during development. In contrast, P-6-[¹²⁵I]BSA binding sites are practically absent during days 5 and 12 and appear by day 22.4. Finally, membrane receptor molecules for estrogen and progesterone have been isolated and purified by affinity chromatography and characterized by PAGE and Western blot. Microsequencing of one of the membrane estrogen binding proteins indicates that the high-affinity site corresponds to the OSCP subunit of the proton ATP synthase.5. It remains to be determined if P and T also bind to this complex enzyme or if they bind to other subunits of the family of proton ATPases. Overall the data indicate that steroid hormones conjugated to BSA are important tools to study the reality of membrane steroid receptors.     

Study of the role of iron in the anticryptococcal activity of human serum and fluconazole

Anticryptococcal activity of human serum and apotransferrin in RPMI 1640 was studied in vitro. The effects of varying concentrations of FeCl₃ on this activity was investigated. Possible synergy of serum and apotransferrin with fluconazole was also measured. The fungistatic activity of human serum, whether lyophilized, stored at 4 °C, fresh frozen or purchased from commercial sources vs. Cryptococcus neoformans was comparable. There was no significant loss of fungistatic activity after freezing and thawing the serum up to 10 times. The fungistatic activity of human serum was similar when tested in different tissue culture media with the exception of Medium 199. The addition of apotransferrin (2.0 or 0.2 mg/ml) to RPMI 1640 had an inhibitory effect on cryptococcal growth. This effect was reversed by 20 M of FeCl₃ at both apotransferrin concentrations. By contrast, addition of FeCl₃ to human serum and RPMI 1640 did not reverse inhibition of growth. Fluconazole synergized with the human serum preparations described, but not with pooled commercial serum, for fungicidal activity. Synergistic activity of fluconazole and human serum was not affected by the addition of FeCl₃. Apotransferrin did not show any synergistic fungicidal activity with fluconazole.     

一种“无血清”培养基对肿瘤细胞生长的支持

本文报告一种为适于杂交瘤细胞生长“无血清”培养基－适量的成牛血清低分子成分超滤液、１０ｍｇ／Ｌ转铁蛋白（Ｔ）、１０ｍｇ／Ｌ胰岛素（Ｉ）、２０μｍｏｌ／Ｌ乙醇胺（Ｅ）、４０ｎｍｏｌ／Ｌ硒酸钠（Ｓ）等诸补充成分替代胎牛血清加到基础液中－也完全适用于培养肿瘤细胞,且观察到与之相似的规律性结果,癌细胞在本“无血清”培养若的生长水平达到在含胎牛血清培养基中生长的水平。对于癌细胞的生长,ＬＭＷ－ＣＳ与Ｔ、Ｉ、     

Extrahepatic tissue copper concentrations in white perch with hepatic copper storage

Hepatic copper storage in man (Wilson's disease), Bedtington and West Highland white terriers, and white perch ( Morone americana ) is characterized by the progressive accumulation of copper in hepatic lysosomes bound to cytoprotective metallothionein. In man, saturation of the liver storage capacity results in the distribution of copper to extrahepatic tissues with multiple organ system dysfunction. To determine if extrahepatic tissue copper concentrations also increase in white perch, copper and zinc levels in liver, brain, heart, gills, serum, and bile were determined by atomic absorption spectrophotometry and compared to striped bass ( Morone saxatilis ). Results showed that brain copper concentrations in. white perch were elevated and significantly correlated with liver copper. Bile and serum copper also increased significantly with liver copper. Copper levels in heart and gill tissues were low. Liver zinc was increased in white perch but not to the same magnitude as copper, and was correlated significantly with liver copper; possibly a non-specific secondary increase related to an overall increase in hepatic metallothionein. Histochemical staining of liver with rubeimc acid for copper was proportional to copper concentrations, and clusters of positive mononuclear cells were also seen in brain and spleen. Foci of macrophages in spleen were also intensely positive with Perl's iron stain which may have been indicative of haemolysis. The patterns of copper distribution seen in white perch present a useful comparative model to study alterations in copper metabolism.     

Efficiency of serum copper/zinc ratio for differential diagnosis of patients with and without lung cancer

We examined serum copper (Cu), serum zinc (Zn), and the serum copper/zinc ratio (Cu/Zn) in 162 patients. All of them were seen to have an abnormal shadow in the chest X-ray films, that is, 109 patients with lung cancer (LC) and 53 patients with no lung cancer (NLC). The mean Cu and Cu/Zn in LC patients were significantly higher than those in NLC patients (p<0.05). In LC patients, Cu and Cu/Zn were higher and Zn was lower in advanced tumors than early ones. There was a significantly clear relation between Cu or Cu/Zn and the tumor (T) stages. When the relative risk (RR) of LC was estimated, it was seen that the higher Cu and Cu/Zn became, the higher RR became. Furthermore, we showed the sensitivity of the receiver operator characteristic of the test (ROC) curve for Cu, Cu/Zn, and carcinoembryonic antigen (CEA) to diagnose LC, as explained in a paragraph of methods.The determinations of Cu, Zn, and Cu/Zn are simple and inexpensive. They also appear to have a great diagnostic value in determining the local invasion of LC and as a screening test in the high-risk patients for LC.     

Ectopic expression of β-lactoglobulin/human serum albumin fusion genes in transgenic mice: hormonal regulation andin situ localization

We produced transgenic mice carrying the native sheep -lactoglobulin (BLG) or fusion genes composed of the BLG promoter and human serum albumin (HSA) minigenes. BLG was expressed exclusively in the mammary glands of the virgin and lactating transgenic mice evaluated. In contrast, transgenic females carrying the BLG/HSA fusion constructs also expressed the HSA RNA ectopically in skeletal muscle, kidney, brain, spleen, salivary gland and skin. Ectopic expression of HSA RNA was detected only in strains that express the transgene in the mammary gland. There was no obvious correlation between the level of the HSA RNA expressed in the mammary gland and that found ectopically. In three transgenic strains analysed, the expression of HSA RNA in kidney and skeletal muscle increased during pregnancy and lactation, whereas in the brain HSA expression decreased during lactation in one of the strains. HSA protein was synthesized in skeletal muscle and skin of strain #23 and its level was higher in lactating mice compared with virgin mice. Expression of HSA was also analysed in males and was found to be more stringently controlled than in females of the same strains.In situ hybridization analyses localized the expressed transgene in the skin, kidney, brain and salivary glands of various transgenic strains. Distinct strain-specific and cell-type specific HSA expression patterns were observed in the skin. This is in contrast to the exclusive expression of the HSA transgene in epithelial cells surrounding the alveoli of the mammary gland. Taken together, these results suggest that the absence of sufficient mammary-specific regulatory elements in the BLG promoter sequences and/or the juxtaposition of the BLG promoter with the HSA coding sequences leads to novel tissue- and cell-specific expression in ectopic tissues of transgenic mice.