Differential modulation of carbachol and trans-ACPD-stimulated phosphoinositide turnover following traumatic brain injury

In the fluid percussion model of traumatic brain injury (TBI), we examined muscarinic and metabotropic glutamate receptor-stimulated polyphosphoinositide (PPI) turnover in rat hippocampus. Moderate injury was obtained by displacement and deformation of the brain within the closed cranial cavity using a fluid percussion device. Carbachol and (±)-1-Aminocyclopentane-trans-1,3.-dicarboxylic acid (trans-ACPD)-stimulated PPI hydrolysis was assayed in hippocampus from injured and sham-injured controls at both 1 hour and 15 days following injury. At 1 hour after TBI, the response to carbachol was enhanced in injured rats by up to 200% but the response to trans-ACPD was diminished by as much as 28%. By contrast, at 15 days after TBI, the response to carbachol was enhanced by 25% and the response to trans-ACPD was enhanced by 73%. The ionotropic glutamate agonists N-methyl-D-aspartate (NMDA), and -amino-3 hydroxy-5-methyl-4-isoxazolepropionate (AMPA), did not increase PPI hydrolysis in either sham or injured rats and injury did not alter basal hydrolysis. Thus, hippocampal muscarinic and metabotropic receptors linked to phospholipase C are differentially altered by TBI.Abbreviations used TBI traumatic brain injury - EAA excitatory amino acids - PPI polyphosphoinositides - IP inositol phosphates - NMDA N-methyl-D-aspartate - AMPA -amino-3-hydroxy-5-methylisoxazole-4-propionate - trans-ACPD (±)-1-Aminocyclopentanetrans-1,3-dicarboxylic acid - LTP long term potentiation     

Strong adjuvant action of Klebsiella O3 lipopolysaccharide and its inhibition of systemic anaphylaxis

Abstract Immunization with lipopolysaccharide from Klebsiella O3 as an immunological adjuvant did not cause the death of mice in systemic anaphylaxis to bovine serum albumin. On the other hand, most mice immunized with lipopolysaccharide from Escherichia coli O111, Klebsiella O4 and Salmonella minnesota did die. Klebsiella O3 lipopolysaccharide enhanced IgM and IgG antibody response to BSA more markedly than Escherichia coli O111 lipopolysaccharide, while it affected the production of IgE antibody only slightly. Therefore, it is suggested that the inhibition of systemic anaphylaxis by Klebsiella O3 lipopolysaccharide adjuvant might be related to its strong adjuvant action on IgM and IgG class antibody production, and that high levels of circulating IgM and IgG antibodies might act as blocking antibodies in the development of IgE-mediated systemic anaphylaxis.     

浙北平原水杉人工林生物量的研究

 本文对浙江省北部平原水杉人工林的生物量和生产力进行了测定和研究。按平均标准木法分别调查了干、枝、叶、根的生物量。据调查材料,用相对生长规律建立了估测单株林木各器官干重的回归方程,方程的相关系数和估测精度符合要求,具实用价值。研究结果表明：水杉人工林生物量随年龄的增长而增加,林带18年生后增加速度变缓,片林生物量普遍大于林带生物量,生物量组成比例因年龄而异。随着年龄的增加,年平均净生产量、叶面积指数、光能利用率均逐步加大,至18年生时,上述指标分别为17.51t／ha·a、9.1和0.77%。叶净光合生产率以速生阶段为最大,衰退阶段为最低,当叶净光合生产率急剧下降时,可实施主伐更新。     

Purification and structural characterization of progastrin-derived peptides from a human gastrinoma

Several peptides derived from the gastrin-predicted preprohormone sequence were isolated from a human gastrinoma by gel permeation, anion exchange, and reverse phase chromatography. The peptides were identified and characterized structurally by a combination of radioimmunoassays, mass spectral analysis, and microsequence analysis. The largest peptide, progastrin-(1-35) (cryptagastrin), extends from the putative processing site for the signal peptidase to the double basic residues adjacent to the amino terminus of gastrin 34. A shorter form of this peptide, progastrin-(6-35) (cryptagastrin-(6-35), was also isolated in smaller amounts. In addition, sulfated and nonsulfated gastrin 17 amides (progastrin-(55-71)) and the glycine-extended nonsulfated gastrin 17 (progastrin-(55-72)) were identified by radioimmunoassay, and their structures were confirmed by mass spectral analysis. Isolation of cryptagastrin indicates that the signal peptide of human preprogastrin contains 21 amino acid residues, and progastrin, therefore, contains 80 amino acids. There is minimal processing of the cryptic peptide preceding the sequence of gastrin 34. An amidated gastrin form larger than gastrin 34 could contain 71 amino acids. No evidence was obtained for processing that would produce gastrins containing more than 34 but less than 71 amino acid residues.     

Covalently cyclized agonist and antagonist analogues of bombesin and related peptides.

During a search for possible cyclization points in shortened, potent bombesin agonists and antagonists, it was found that the joining of amino acid residues in positions 6 and 14 by various means resulted in retention of significant binding affinity for rat pancreatic acini and murine Swiss 3T3 cells. In one series of analogues, Cys residues in these positions were used for bridging via a disulfide bond. (D)-C-Q-W-A-V-G-H-L-C-NH2 retained significant binding affinity for rat pancreatic acini cells and was a full amylase releasing agonist (EC50 187 nM). Potency was markedly increased by substituting D-Ala for Gly (EC50 67 nM compared to 10 nM for its linear counterpart) and was decreased by substituting L-Cys for D-Cys in this analogue (EC50 214 nM), thus strongly suggesting stabilization of peptide folding by the D residues. Elimination of the COOH-terminal amino acid produces competitive antagonists in the linear analogues; however, (D)-C-Q-W-A-V-G-H-C-NH2 was devoid of activity. Likewise, cyclization to position 13 with the 14 amino acids intact to give (D)-C-Q-W-A-V-G-H-C-L-NH2 resulted in an almost inactive peptide. On the other hand, as in the linear series, the reduced peptide bond analogue, (D)-C-Q-W-A-V-(D)-A-H-L-psi (CH2NH)-C-NH2, was a receptor antagonist (IC50 5.7 mM), albeit much weaker than the corresponding linear analogues, but with no residual agonist activity. Direct head-to-tail cyclization was also tried. Both cyclo[(D)-F-Q-W-A-V-G-H-L-L] (EC50 346 nM) and the shorter cyclo [Q-W-A-V-G-H-L-L] (EC50 1236 nM) were full agonists. Elimination of the COOH-terminal residue in cyclo[(D)-p-Cl-F-Q-W-A-V-(D)-A-H-L] produced an agonist (EC50 716 nM) rather than an antagonist. These results provide support for the proposal that both bombesin agonists and antagonists adopt a folded conformation at their receptor(s). Furthermore, the retention of appreciable potencies using several cyclization strategies and chain lengths suggests that further optimization of these structures both in terms of potency and ring size is possible. Since these peptides have increased conformational restriction, they should begin to serve as useful substrates for NMR and molecular modeling studies aimed at comparing the obviously subtle differences between agonist and antagonist structures.     

Binding and complement activation by C-reactive protein via the collagen-like region of C1q and inhibition of these reactions by monoclonal antibodies to C-reactive protein and C1q

Ligand-complexed C-reactive protein (CRP), like aggregated or complexed IgG, can react with C1q and activate the classical C pathway. Whereas IgG is known to bind to the globular region and not to the collagen-like region (CLR) of C1q, the site of interaction of C1q with CRP has not been defined. CRP-trimers were prepared by cross-linking and found to bind to C1q and to activate the C system. Heat-aggregated IgG (Agg-IgG) did not block the binding of CRP-trimers to C1q, nor did CRP-trimers block binding of Agg-IgG to C1q, suggesting that CRP and IgG bind at different sites. ELISA and Western blot analysis showed that CRP-trimers bound to the CLR, whereas Agg-IgG bound only to the globular region; similarly, anti-CLR mAb inhibited binding of CRP-trimers to C1q whereas anti-globular region mAb did not. Reactivity with CRP-trimers as well as with Agg-IgG was retained after reduction/alkylation and SDS treatment of C1q. A group of 22 anti-CRP mAb directed against at least six distinct native-CRP epitopes and eight distinct neo-CRP epitopes was tested for ability to inhibit the CRP-CLR interaction; one mAb, anti-native CRP mAb 8D8, with strong inhibitory activity was identified. Fab' of 8D8 blocked binding of CRP-trimers to intact C1q as well as CLR, and also inhibited CRP (CRP-trimers and CRP-protamine complexes) induced C activation, but had no effect on C1q binding or C activation by Agg-IgG. These results indicate that a conformation-determined region on CRP binds to a sequence-determined region on the CLR of C1q in an interaction which leads to C activation. Anti-CRP and anti-C1q mAb that specifically inhibit this interaction are described.     

草鱼免疫应答的初步研究

研究了草鱼在不同水温条件下受抗原刺激后其中和抗体的变化。15℃培养条件下中和抗体上升缓慢,9周内滴度低于1:8;20℃时,3周后抗体可上升到1:256,最高达1:5270,而在25℃时,1周中和抗体即达到1:570,最高可达1:20000以上。并探索了从草鱼血清中提纯抗体的条件,研究其抗体的特性。草鱼血清中的抗体为大分子蛋白,容易解离为抗原性相同,分子量近似于人IgG的较小分子,含有较多的二硫键,具有类似IgM的某些特性。     

Families of metalloendopeptidases and their relationships.

Crystal structures available for four metalloendopeptidases have revealed zinc ligands for these enzymes. New sequence information has made it possible to compare the primary structures of the zinc-binding site in metalloendopeptidases. A scheme based on the zinc-binding site is proposed to classify metalloendopeptidases into five distinct families: thermolysin, astacin, serratia, matrixin, and snake venom metalloproteinases. Two histidines and one glutamate are zinc-ligands in the thermolysin family. Three histidines and one tyrosine are zinc ligands in the other four families, which are further distinguished by the identity of the residue following the third histidine and by the environment surrounding the tyrosine.     

Nuclear totipotency of cultured rabbit morulae to support full-term development following nuclear transfer.

The rabbit was used as a model for nuclear transfer. A critical step in nuclear transfer is oocyte activation, which was evaluated in this research. Optimal field strength of an electric stimulus for activation was examined. A significantly higher activation rate in all criteria tested was achieved when oocytes were activated electrically with a field strength of 2.4 kV/cm versus 1.2 or 1.8 kV/cm. Also, electrical stimulation with combined alternating current (AC) and direct current (DC) was superior to DC stimulation alone for activation. In another study involving 586 oocytes, exposure of oocytes to cytochalasin B for 1 h followed by activation with electrical stimulation significantly improved development of the oocytes to blastocyst stage compared to oocytes without cytochalasin B pre-exposure (38% vs 26%, p less than 0.05). Cytochalasin B exposure alone (control), however, had no effect on activation. Exposing oocytes to activation medium without electrical stimulation also activated some oocytes. In the nuclear transfer experiment, blastomeres from 8-cell embryos cultured for 20-24 h to the 32-64-cell stage were used as nuclear donor cells. Of 491 oocytes used, 459 (93%) survived the enucleation and fusion procedure, 370 (81%) fused, and 284 (77%) developed into 2-4-cell embryos. A total of 243 of these 2-4-cell embryos were transferred to 15 pseudopregnant recipients and produced 8 young (3%). Although the efficiency is low, this study demonstrated that rabbit morulae cultured for 20-24 h to the 32-64-cell stage as nuclear donors for transfer remain totipotent.