THE INFLUENCE OF ESTERIFICATION OF CARBOXYL GROUPS OF RIBONUCLEASE-A ON ITS STRUCTURE AND IMMUNOLOGICAL ACTIVITY

The effect of modification of carboxyl groups of Ribonuclease-A^a on the enzymatic activity and the antigenic structure of the protein has been studied. Modification of four of the eleven free carboxyl groups of the protein by esterification in anhydrous methanol/0.1 M hydrochloric acid resulted in nearly 80% loss in enzymatic activity but had very little influence on the antigenic structure of the protein. Further increases in the modification of the carboxyl groups caused a progressive loss in immunological activity, and the fully methylated RNase-A exhibited nearly 30% immunological activity. Concomitant with this change in the antigenic structure of the protein, the ability of the molecule to complement with RNase-S-protein increased, clearly indicating the unfolding of the peptide “tail” from the remainder of the molecule. The susceptibility to proteolysis, accessibility of methionine residues for orthobenzoquinone reaction and the loss in immunological activity of the more extensively esterified derivatives of RNase-A are suggestive of the more flexible conformation of these derivatives as compared with the compact native conformation. The fact that even the fully methylated RNase-A retains nearly 30% of its immunological activity suggested that the modified protein contained antibody recognizable residual native structure, which presumably accommodates some antigenic determinants.     

DEAMIDATED ACTIVE INTERMEDIATES IN THE IRREVERSIBLE ACID DENATURATION OF RIBONUCLEASE-A

A study has been made on the changes in the enzymatic activity of Ribonuclease-A**-(RNase-A) exposed to highly acidic (pH < 1) aqueous environment. Irreversible alterations of activity were observed when the protein was exposed to an acidic medium for a long period (20 to 60 h). Even prior to these changes in activity RNase-A was found to form intermediates which had very nearly the same activity as the native protein. The primary process in the acid denaturation of RNase-A was observed to be deamidation of the protein leading to the formation of active chromatographically distinct derivatives. The initial product of deamidation, a monodeamidated derivative, has been isolated by chromatography on Amberlite XE-64. This initial deamidation reaction proceeded with very high specificity. The subsequent deamidation reaction is comparatively slower, so that nearly 50% of the native protein could be converted to this derivative before any subsequent deamidation took place. This monodeamidated derivative has been designated RNase-Aa₁. The conversion of RNase-A to RNase-Aa₁ was not accompanied by any changes in the primary structure other than the observed deamidation. Apart from the differences in chromatographic and electrophoretic mobilities, RNase-Aa₁ was found to have very nearly the same activity and physico-chemical properties as the native enzyme. Significance of this specific and faster deamidation of RNase-A in this denaturing medium as well as the biological significance of such deamidation reactions of proteins are discussed.     

ASSESSING VASCULAR REACTIVITY OF ARTERIES IN THE SMALL VESSEL MYOGRAPH

1. Using a small vessel myograph, experiments were performed on rat small mesenteric arteries (and in some cases dog small coronary arteries) to assess the dependence of vasoconstrictor potency (EC50) and maximum response (Emax) on the initial passive conditions and on the mode of recording (i.e. isometric, isobaric or isotonic). 2. Maximum active isometric tension development to methoxamine occurred at different points on the passive diameter--tension curve depending on the origin of the vessel. The point of maximum sensitivity of methoxamine did not coincide with the point of maximum tension development on the passive diameter-tension curve. 3. Vascular reactivity to methoxamine was assessed under isobaric, isotonic and isometric conditions using a new computerized myograph. Methoxamine was significantly more potent, but only by a factor of twofold, when assessed under isometric conditions. In addition, the maximum response to methoxamine, in terms of diameter change, was always greater under isobaric than under isotonic conditions. 4. The results show that, in studies comparing vascular reactivity of vasoactive drugs, the results depend, to some extent, on the initial passive conditions selected. In terms of assessing the pharmacological activity of drugs on isolated blood vessels, the use of common isometric recording procedures are adequate. However, the use of isobaric, isotonic and isometric recording procedures have shown the complexities of vascular reactivity which depend on the passive and active properties of the blood vessel. These factors may need to be taken into account when comparing the reactivity of isolated blood vessels.     

单环非典型内酰胺的抗菌活性与化学活性关系研究

对已合成的六个单环非典型内酰胺使用量子化学的ＭＮＤＯ方法进行计算，所得结果支持作者关于化学活性是决定单环非典型内酰胺抗菌活性主要因素的设想。     

头孢菌素和青霉素交叉过敏反应的结构基础探讨

利用亲和层析、ELISA、反向间接血凝试验和PCA试验等免疫学试验方法,证明头孢菌素和青霉素间有弱的交叉过敏反应。两类抗生素形成抗原或多价半抗原时,抗原决定簇中由β—内酰胺环形成的新抗体结合位点是交叉过敏反应的重要抗体结合位点;β—内酰胺环结构是交叉过敏反应的结构基础。 交叉过敏反应的强弱除了由抗原间的相似性所决定,还和机体中特异性抗体的抗原结合部位特异性有关。通过对18只兔抗BPO—BSA抗血清和特异性已知的青霉素单克隆抗体的比较分析,认为对青霉素和头孢菌素间的交叉过敏反应现象应有足够的重视。     

活血化瘀治则的药理学研究——(一)川芎、人参、丹参配伍对犬急性实验性心肌缺血的影响

本实验选用川芎(行气活血),川芎人参(益气活血),川芎丹参(养气活血),作为活血三治则的代表药,进行了犬急性心肌梗塞的实验研究。显示三组中药均可减轻急性心肌缺血时的∑—ST和N·ST的变化,可以不同程度地缩小缺血范围和梗塞范围,而它们的作用途径并不完全相同。经比较研究证明,在抗心肌缺血方面,益气活血及养气活血较单纯行气活血为好,与中医标本兼治理论一致。     

EFFECT OF FISH OIL FEEDING ON BLOOD PRESSURE AND VASCULAR REACTIVITY IN SPONTANEOUSLY HYPERTENSIVE RATS

1. To examine possible antihypertensive mechanisms of fish oil feeding, we have studied vascular reactivity of aortic rings and blood pressure of spontaneously hypertensive rats (SHR). 2. SHR were fed a synthetic diet supplemented with either (10% by weight) 'Max EPA' fish oil or hydrogenated coconut oil (saturated fat) for 4 weeks. 3. Mean systolic blood pressure of fish oil fed rats was 9 mmHg lower than saturated fat fed controls. 4. Aortic rings of control SHR had a biphasic response to acetylcholine (ACh), relaxing at lower concentrations but contracting at concentrations higher than 3 x 10(-7) mol/L. No such contractions were seen in tissues of fish oil fed rats. The contractions were abolished by indomethacin, suggesting that they were caused by a cyclo-oxygenase product. 5. Tissue analysis showed that both aortic and serum generation of thromboxane B2(TxB2) was approximately three times less in fish oil fed rats than in control tissues. 6. These results indicate that the lowering of blood pressure in fish oil fed SHR could in part be due to decrease in production of thromboxane (TxA2), a potent vasoconstrictor, hence influencing vascular tone and compliance of the aorta.     

INCREASE IN REACTIVITY OF HUMAN PLATELET GUANYLATE CYCLASE DURING AGGREGATION POTENTIATES THE DISAGGREGATING CAPACITY OF SODIUM NITROPRUSSIDE

1. Basal and stimulated guanylate cyclase activity during ADP-induced human platelet aggregation in comparison with the actions of sodium nitroprusside (SNP) on platelets was investigated. 2. Sodium nitroprusside exhibited both ex vivo and in vitro antiplatelet effects, as assessed by inhibition of subsequent ADP-induced aggregation in platelet-rich plasma. A strong correlation between decrease in aggregation and increase in platelet guanylate cyclase activity in the presence of SNP was obtained. 3. When SNP was administered after the induction of aggregation, it caused acceleration of disaggregation (in reversible aggregation) and produced disaggregation (under conditions of otherwise irreversible aggregation) which was time-dependent. 4. Platelet aggregation was accompanied by a transient increase in platelet cyclic GMP content and guanylate cyclase activation by the nitric oxide (NO) donor SNP. Changes in guanylate cyclase activity were haem-associated and probably reflected saturation of enzyme by haem. 5. Maximal SNP disaggregating effect coincided with peak guanylate cyclase responsiveness to SNP. 6. The present investigation provides evidence that increased responsiveness of platelet guanylate cyclase to NO during aggregation facilitates disaggregation in the presence of SNP. Thus, availability of NO (endogenous or exogenous) at sites of incipient platelet aggregation in vivo may play a pivotal role regarding limitation of this process.     

USE OF THIOCYANIC ACID TO FORM 2-THIOHYDANTOINS AT THE CARBOXYL TERMINUS OF PROTEINS

The chemistry of the formation of 2-thiohydantoins on the carboxyl terminal of peptides or proteins was investigated. It was found that thiocyanic acid was much more reactive for the formation of 2-thiohydantoins than were the thiocyanate salts. The physical reasons for this observation are explained. The kinetics of the reaction of a number of proteins, and some of their fragments, with thiocyanic acid were also determined. Simple and safe procedures for the preparation of anhydrous thiocyanic acid solutions were devised. The prospective application of these procedures to sequencing from the carboxyl terminal of a polypeptide is discussed.     

EFFECT OF PURE EICOSAPENTAENOIC ACID FEEDING ON BLOOD PRESSURE AND VASCULAR REACTIVITY IN SPONTANEOUSLY HYPERTENSIVE RATS

1. This study examined the effects of eicosapentaenoic acid (EPA) treatment on vascular reactivity and blood pressure in spontaneously hypertensive rats (SHR). 2. Twenty SHR were given pure EPA as the methyl ester (280 mg/kg) by gavage for 10 days. An equal number of control rats received vehicle alone. EPA treatment had no effect on blood pressure compared with control rats. 3. Aortic rings from EPA-treated rats, precontracted with PGF2 alpha showed increased endothelium-dependent relaxations to acetylcholine. Endothelium-independent relaxations to sodium nitroprusside were not altered. Rings from rats fed pure EPA did not show any differences in vasoconstrictor responses to noradrenaline or serotonin. 4. Serum thromboxane B2 (TXB2) levels fell 17% in animals given pure EPA, but prostacyclin production was not affected. These responses are less than those seen following Max EPA fish oil. 5. Thus, pure EPA treatment did not lower blood pressure, but may have a direct effect on aortic endothelia and cause increased endothelium-dependent relaxations in response to acetylcholine in SHR.     

A PREPARATIVE METHOD FOR THE ISOLATION OF CARBOXYL TERMINAL TRYPTIC PEPTIDES FROM PROTEINS

A method has been developed for isolating glycinamide-blocked carboxyl-terminal tryptic peptides from proteins. It has been successfully applied to four proteins of known carboxyl-terminal sequence, yielding the carboxyl-terminal peptide in up to 68% yield. The separation method is designed to be selective for the desired peptide and to handle any quantity of protein.     

EFFECTS OF HOE 140 ON SYSTEMIC DEPRESSOR RESPONSES TO BRADYKININ AND MESENTERIC VASCULAR REACTIVITY IN VIVO IN PREGNANT WISTAR-KYOTO RATS

1. To examine possible changes in vascular reactivity to exogenous bradykinin (BK) and the possible role of endogenous BK in reduced mesenteric vascular reactivity in pregnant rats. The authors studied the effects of Hoe 140 on systemic depressor responses to BK and on mesenteric vascular reactivity in in situ blood-perfused mesenteric resistance vessels of 18–20 day pregnant and age-matched non-pregnant Wistar-Kyoto rats (WKY). 2. Mean intra-arterial blood pressure (MBP) of pregnant rats was lower than non-pregnant controls. Basal mesenteric perfusion pressure (BPP) was slightly, but not significantly, reduced in the pregnant group. Neither MBP nor BPP was significantly influenced by Hoe 140 (1 mg/kg. s.c.) 3. Systemic depressor responses to BK (1–30 μg/kg, i.v.) were significantly increased in pregnant rats at 1 and 3 μg/kg. Hoe 140 completely abolished systemic depressor responses to BK in either pregnant or non-pregnant animals. 4. Mesenteric vascular responses to regional administration of noradrenaline, electrical stimulation of sympathetic nerve and angiotensin II were overall decreased in pregnant compared with non-pregnant groups, but those responses were not significantly affected by Hoe 140 in either groups. 5. The results suggested that although systemic depressor responses to exogenous BK were increased, endogenous BK does not contribute to decreased mesenteric vascular reactivity in vivo in pregnant WKY.     

EFFECT OF Na+ PUMP SUPPRESSION ON REACTIVITY OF RAT TRACHEALIS TO COOLING

1. This study tests the hypothesis that suppression of Na+ pump would increase the rate of tension development and the magnitude of contraction induced by cooling in airway smooth muscle. 2. Rat tracheal preparations were incubated in ouabain for 8 h and tested hourly for their response. In a representative specimen the rate of tension development increased from the control value of 0.7 mg/s to 5.5 mg/s after 7 h of incubation in ouabain concentration of 4 X 10(-4) mol/L; likewise, the magnitude of contraction increased from the control value of 80 mg to 550 mg. 3. Using ouabain concentrations between 9 X 10(-5) mol/L and 6 X 10(-3) mol/L, the rate of tension development and the magnitude of contraction first increased in a dose-dependent manner up to 8 X 10(-4) mol/L, then declined with higher doses but the responses were still greater than the control values at all concentrations. 4. After 3 h incubation in ouabain at 8 X 10(-4) mol/L, the mean rate of tension development and the mean magnitude of contraction increased to 647% and 578% of the control value, respectively. 5. These results indicate that depression of Na+ pump results in hypersensitivity and hyper-reactivity of the airway smooth muscle to cooling.     

西咪替丁对大鼠体内呋喃氟脲嘧啶药代动力学的影响

本文用交叉实验设计法研究了西咪替丁(Cim)对9只大鼠口服呋喃氟脲嘧啶(FT—207)药代动力学的影响。反相HPLC法测定血清FT-207浓度,其药时曲线符合一室开放模型。单次口服Cim80mg/kg或连用Cim80 mg/kg·d~(-1)×5d,均增加血清FT—207浓度和AUC。连用时,FT-207的C_(max)和AUC分别增加21.0%(P<0.05)和34.1%(P<0.025),Te_(1/2)缩短(P<0.005);单次服Cim时,FT-207的药代学改变与连用时相似,但差异无显著性。文中讨论了二药相互作用的机制及临床意义。     

中国木贼科植物的数量分类学研究

选择了中国木贼科两属11种植物共11个分类运算单位;选取了形态学、组织学、化学成分等方面的35个性状;采用单联法、全联法、WPGMA 法、UPGMA 法、离差平方和法、可变类平均法和可变法等七种方法,进行了分类运算。最后,绘出了 UPGMA 法的树系图和结合线图。研究结果支持将木贼科分为两属:木贼属和问荆属。     

EFFECT OF PERINDOPRIL ON THE DEVELOPMENT OF ATHEROSCLEROSIS IN THE CHOLESTEROL-FED RABBIT

1. The aim of the study was to examine the effect of the angiotensin-converting enzyme inhibitor perindopril on the development of atheroma in the cholesterol-fed rabbit.
2. The normal human carotid artery, like most large human arteries, has a preformed diffuse intimal thickening. To model this thickening, the right carotid artery of the 12-week-old rabbit had an expanded balloon catheter passed down it to remove the endothelium and partially damage the media.
3. Fourteen weeks after this operation, a myointimal thickening similar in almost all respects to the human intimal thickening had developed. The rabbits were then divided into six groups of six rabbits fed on: (i) a 1% cholesterol diet; (ii) a 1% cholesterol diet plus a hypotensive dose of perindopril (0.3 mg/kg per day); (iii) a 1% cholesterol diet plus a non-hypotensive dose of perindopril (0.01 mg/kg per day); (iv) a normal diet; (v) a normal diet plus a hypotensive dose of perindopril (0.3 mg/kg per day); and (vi) a normal diet plus a non-hypotensive dose of perindopril (0.01 mg/kg per day).
4. After 6 weeks of treatment the animals were sacrificed. There were ameliorating effects of both hypotensive and non-hypotensive doses of perindopril on the development of plaques, as determined by the area of intima covered by Oil Red-O-staining plaque and light microscopy.
5. Cell culture studies indicated that perindopril has no effect on smooth muscle proliferation, but increases collagen and non-collagen synthesis by smooth muscle cells and decreases their binding of the atherogenic lipoprotein beta-very low density lipoprotein (β-VLDL).
6. The results suggest that perindopril has a beneficial effect in decreasing the severity of atherosclerotic lesions in the cholesterol-fed rabbit, possibly by affecting lipoprotein metabolism.