影响反转录过程多种因素的探讨

本文探讨了影响反转录反应的各种因素。温度的适当提高和模板的严格处理可以破坏RNA的二级结构,并提高引物的特异性结合;模板的纯度可以避免杂质对酶的特异性的影响;巢式引物可以在PCR过程中很有效地避免反转录过程中产生的非特异性产物的扩增,而 提高最终产物的特异性。     

FURTHER EVALUATION OF POLYPEPTIDE SYNTHESIS IN CEREBRAL ANOXIA, HYPOXIA AND ISCHEMIA

The alteration of polypeptide synthesis was evaluated with microsomes isolated from anoxic rabbit, hypoxic rat and ischemic gerbil brains to estimate the extent of functional or structural changes in polyribosomes in situ and the extent of artifact during tissue preparation. By using two-stage experimentation with combination of control and pathological microsomes and supernatant, it was found that the previously observed effects on microsomal or polyribosomal polypeptide synthesis in the above pathophysiological conditions were mainly the reflection of the alteration of polyribosomes in situ rather than the artifact during tissue preparation by degradative processes. In support of this finding. the use of inhibitors of degradative enzymes did not significantly protect microsomes either in normal or in pathological conditions. It was noted that the decline of tissue pH, to a certain extent, could be correlated with dysfunction of polyribosomes both in situ and during tissue preparation in cerebral hypoxia and anoxia. Since there is little change in ATP level, it was postulated that the alteration of pH in situ is responsible for the observed suppression of polypeptide synthesis in vitro at least in cerebral hypoxia. This hypothesis was supported by the subsequent experiments with incubation of brain slices and homogenization of brain tissue under various pH. It was emphasized that the environmental biochemical elements surrounding polyribosomes in cytoplasm should be evaluated as possible contributing factors for polyribosomal dysfunction in such pathological conditions as cerebral anoxia, hypoxia or ischemia if the alteration of energy state does not explain the phenomenon entirely.     

5'UTR中AUG的分布及其对翻译起始的影响

以细菌和古菌基因组5’UTR序列作为研究对象,分析在5’UTR的3个不同阅读框架中三联体AUG的分布,发现无论是细菌还是古菌基因组都在阅读框1中有非常明显的AUG缺失（depletion）。AUG的缺失表明在起始密码子上游的AUG很可能会对基因的翻译起始产生影响。分析得知：绝大部分的AUG都是以uORF（upstream open readingframe）的形式出现的,uAUG（upstreamAUG）的数量很少,特别是在阅读框1中,而且在细菌基因组的阅读框1中uAUG较多地出现在了含有SD序列的基因上游。比较发现,uAUG引导的序列在同义密码子使用上的偏好性较真正的编码序列差,这可能表明细菌和古菌在同义密码子使用上的偏好性也是决定基因准确地翻译起始的重要因素之一。     

THE EFFECT OF INSULIN, ALLOXAN DIABETES, AND ANOXIA ON THE ULTRASTRUCTURE OF THE RAT HEART

Hearts from normal and alloxan diabetic rats were perfused in vitro with a bicarbonate-buffered medium containing glucose. Transport of glucose through the cell membrane was stimulated with insulin or by induction of anaerobiosis. The organs were rapidly fixed and examined by electron microscopy. Transport stimulation was not associated with any increase in the number of sarcolemmal invaginations or subsarcolemmal cytoplasmic vesicles. It was concluded that glucose transport and the effects of insulin or anoxia do not involve pinocytosis. The relationship of pinocytosis to glucose transport is discussed. The appearance of numerous lipid inclusions at the Z line level of the sarcomeres in the diabetic and anoxic myocardia is described.     

蛛网膜下腔出血对大鼠脑血流量和体感诱发电位的影响

目的：探讨蛛网膜下腔出血（ＳＡＨ）后脑血流量、体感诱发电位（ＳＥＰ）潜伏期的改变及其与一氧化氮（ＮＯ）的关系。方法：对假手术对照组和ＳＡＨ模型组大鼠检测２４ｈ局部脑血流量（ｒＣＢＦ）、ＳＥＰ潜伏期和血清及脑组织ＮＯ含量动态变化。结果：非开颅刺破Ｗｉｌｌｉｓ环的方法可成功地诱发ＳＡＨ。ＳＡＨ后ｒＣＢＦ立即降低,在２４ｈ内无恢复趋势。ＳＥＰ潜伏期于ＳＡＨ后１ｈ开始至２４ｈ明显延长。血清和脑组织ＮＯ含量     

L-Glu促进纯化培养的海马和皮质星形胶质细胞增殖的研究

目的研究L-Glu对纯化培养的皮质和海马星形胶质细胞促增殖作用的机制.方法将纯化培养的星形胶质细胞分为7组:①无血清培养基组,②含L-Glu(1mmol/L)的无血清培养基组,③含MCCG(100μmol/L)的无血清培养基组,④含MPEP(100μmol/L)的无血清培养基组,⑤含L-Glu MCCG(分别为1mmol/L,100μmol/L)的无血清培养基组,⑥含L-Glu MPEP(分别为1mmol/L,100μmol/L)的无血清培养基组,⑦含L-Glu MCCG MPEP(分别为1mmol/L,100μmol/L及100μmol/L)的无血清培养基组.流式细胞仪检测细胞周期的变化.结果 L-Glu促进纯化培养的星形胶质细胞的增殖,单独加入mGluRs的竞争性拮抗剂MCCG或MPEP则L-Glu的促增殖作用减弱,同时加入两种拮抗剂则L-Glu的促增殖作用消失.结论 L-Glu通过作用于mGluR3和mGluR5促进星形胶质细胞的增殖,两者可能在星形胶质细胞增殖方面具有协同作用.     

缺氧对离体鸡胚前脑神经细胞生长的影响

在缺氧(95％N_2和5％CO_2)条件下进行8d鸡胚前脑的纯神经细胞培养。培养24,48和72h后,MTT微量比色法检测显示,甲(目替)明显减少,表明神经元已遭严重损害,同时,培养液中葡萄糖被急剧地消耗。但是,即使把糖浓度提高到800—1200mg/100ml,缺氧仍造成神经细胞死亡。结果提示,胚胎脑神经细胞对缺氧敏感,胶质细胞对缺氧神经元的保护可能不在于提供糖原。     

ANOXIA AND BRIGHTNESS DISCRIMINATION

1. Brightness discrimination has been studied with individuals breathing oxygen concentrations corresponding to 7 altitudes between sea level and 17,000 feet. The brightnesses were 0.1, 0.01, and 0.001 millilambert involving only daylight (cone) vision. 2. At these light intensities, brightness discrimination begins to deteriorate at fairly low altitudes. The deterioration is obvious at 8,000 feet, and becomes marked at 15,000 feet, where at low brightness, the contrast must be increased 100 per cent over the sea level value before it can be recognized. 3. The impairment of brightness discrimination with increase in altitude is greater at higher altitudes than at lower. The impairment starts slowly and becomes increasingly rapid the higher the altitude. 4. Impairment of brightness discrimination varies inversely with the light intensity. It is most evident under the lowest light intensities studied, but shows in all of them. However, it decreases in such a way that the deterioration is negligible in full daylight and sunlight. 5. The thresholds of night (rod) vision and day (cone) vision are equally affected by anoxia. 6. The quantitative form of the relation between brightness discrimination ΔI/I and the prevailing brightness I remains the same at all oxygen concentrations. The curve merely shifts along the log I axis, and the extent of the shift indicates the visual deterioration. 7. The data are described in terms of retinal chemistry. Since anoxia causes only a shift in log I it is shown that the photochemical receptor system cannot be affected. Instead the conversion of photochemical change into visual function is impaired in such a way that the conversion factor varies as the fourth power of the arterial oxygen saturation.     

EFFECTS OF SHOCK AND ANOXIA ON NUCLEOTIDES AND CREATINE PHOSPHATE IN THE ISOLATED BRAIN OF THE DOG1

Surgically isolated canine brains were maintained with compatible donor blood from an extracorporeal perfusion system. Small samples of frozen cerebral cortex were removed with a newly-developed Freon cryoprobe and were analysed for acid-soluble nucleotides, creatine phosphate and inorganic phosphate. Most animals were in the early stages of shock unless they had received preoperative α-adrenergic blockade with phenoxybenzamine hydrochloride (Dibenzyline). Values for high-energy phosphates were in the normal range only when the animal had been premedicated with phenoxybenzamine hydrochloride. During a 4-min period of anoxia (induced by blood which had been equilibrated with 95% N₂ and 5% CO₂), the cerebral cortex rapidly became iso-electric, and the levels of creatine phosphate and ATP decreased concomitantly with increases in levels of ADP and P_i. These electrical and chemical changes were rapidly and completely reversed by reoxygenation. The levels of high-energy phosphates provide a sensitive criterion of functional adequacy that may be more readily quantitated than cerebral electrical activity (EEG). EEG recovery did not correlate closely with rephosphorylation.     

甘珀酸干预对大鼠脑缺血再灌注损伤的影响

目的观察缝隙连接阻断剂甘珀酸对局灶性脑缺血/再灌注损伤的影响。方法采用大鼠大脑中动脉阻塞再灌流模型（MCAO）,将动物随机分为脑缺血60min再灌注（MCAO）组,脑缺血再灌注加甘珀酸干预（MCAO＋CBX）组和假手术组（sham）。采用尼氏染色显示脑梗死灶并计算梗死灶体积;应用免疫荧光与TUNEL染色法分别观察脑缺血后3d与7d不同时间点缺血边缘区胶质纤维酸性蛋白（GFAP）的表达和细胞凋亡情况。结果（1）缺血后3d、7d MCAO＋CBX组大鼠梗死体积小于MCAO组,3d、7d MCAO＋CBX组大鼠梗死体积较MCAO组分别缩小5%和4.6%;（2）缺血后3d、7d于缺血边缘区可见大量TUNEL阳性染色细胞,且MCAO组大鼠缺血边缘区细胞凋亡数目明显多于MCAO＋CBX大鼠（P〈0.001）;（3）缺血后3d和7d组缺血边缘区GFAP表达明显增强,3d的MCAO组与MCAO＋CBX组大鼠缺血边缘区GFAP的表达均较假手术组强（P〈0.05）,7d的MCAO＋CBX组大鼠缺血边缘区GFAP的表达较假手术组强（P〈0.001）,但明显弱于MCAO组大鼠（P〈0.01）;结论缝隙连接阻断剂甘珀酸可减少大鼠大脑中动脉阻塞后脑梗死体积,其机制可能与阻断缝隙连接后缺血边缘区神经元凋亡降低有关,星型胶质细胞的反应性变化参与了该过程。     

7SRNA在癌细胞中含量改变及其对染色质转录活性的调节

7s RNA与哺乳动物基因组中有300,000重复拷贝的Alu DNA顺序高度互补。Alu DNA的功能是与结构基因的转座和表达调控密切有关。因此7s RNA对基因表达可能有调节作用。本文提供实验:1.肝癌细胞核内7s RNA含量比正常核少,意味着减弱了对癌细胞基因表达的控制。2.7s RNA比其他的核小分子量RNA更紧密与染色质结合。3.7sRNA对离体染色质的转录活性有促进和抑制的两相作用,这些结果直接和间接表明7s RNA对基因表达起一定的调节作用。由于Alu DNA在基因组中分布是广泛性的,因此认为7s RNA的调控作用也应是一般控制性质的。     

血管紧张素Ⅱ对大鼠下丘脑内血管升压素基因转录的影响

实验在雄性ＳＤ大鼠中进行,用核酸斑点杂交技术观察下丘脑组织中血管升压素（ＡＶＰ）基因转录水平变化,用异羟基洋地黄毒甙（ＧＩＧ）标记的２６个碱基长寡聚核苷酸作为检测探针。实验中观察到,用渗透压微泵向大鼠侧脑人连续注射微量血管紧张素Ⅰ（０．２ｎｍｏｌ／ｈ）２ｄ后,可引起动物饮水量显著增加,下丘脑组织中ＡＶＰ基因转录水平高,但无统计学显著意义。将实验动物日饮水量限制在与对照动物相同的每日饮水量之后,侧脑     

牛磺酸调节缺氧性肺、脑血管反应的机理研究

本实验从磷脂酶Ａ＿２（ＰＬＡ＿２）、前列腺素（ＰＧｓ）、白三烯（ＬＴｓ）和过氧化脂质（ＬＰＯ）方面探讨了牛磺酸调节肺、脑血管对急、慢性缺氧反应的机制。急性缺氧时狗出肺与出脑血中ＬＰＯ增加,ＰＬＡ,活性有升高趋势,但出脑与出肺（入脑）血相比无显著性差异。出肺与出脑血中ＬＴＣ＿４、ＴＸＢ＿２、６－Ｋｅｔｏ－ＰＧＦ＿（１ａ）及ＴＸＢ＿２／６－Ｋｅｔｏ－ＰＧＦ＿（１ａ）比值均升高。慢性缺氧大鼠肺、脑组织中ＰＬＡ＿２活性均升高。牛磺酸增加缺氧时６－Ｋｅｔｏ－ＰＧＦ＿（１ａ）,减弱其它变化。提示牛磺酸对缺氧性肺缩血管反应的调节作用可能与降低缺氧时ＰＬＡ＿２活性,抑制脂质过氧化和ＬＴＣ＿４、ＴＸＡ＿２生成,降低ＴＸＡ＿２／ＰＧＩ＿２比值有关;而牛磺酸减弱缺氧性脑舒血管反应不是直接通过上述变化起作用的。     

THE EFFECT OF CARBON DIOXIDE TENSION ON THE METABOLISM OF CEREBRAL CORTEX AND MEDULLA OBLONGATA

Manometric measurements were made of oxygen uptake (Q _OO2) and aerobic lactic acid output (Q_G) by slices of cerebral cortex and medulla oblongata of the cat in the presence of mixtures of 1, 5, and 20 volumes per cent of carbon dioxide in oxygen. The concentrations of NaHCO₃ and NaCl in the medium were varied to maintain constant pH and sodium ion concentrations. The calcium ion concentration was 0.0002 M. At pH 7.5 under these conditions, an increase in carbon dioxide from 1 per cent to 5 per cent doubled the Q_G of both tissues but did not alter Q _OO2; an increase from 5 per cent to 20 per cent carbon dioxide had no further effect on Q_G in either tissue or Q _OO2 of cortex, but did depress the Q _OO2 of medulla. At pH 8.1, an increase in carbon dioxide from 1 per cent to 5 per cent raised the Q _OO2 and Q_G of cortex by about 60 per cent. Measurements at low oxygen tension carried out previously in phosphate medium were repeated in bicarbonate medium to obtain data for the combined output of lactic acid and carbon dioxide (Q_A). When the oxygen in the gas phase was decreased from 95 to 3 volumes per cent, the lactic acid output as measured colorimetrically increased by 114 mg./gm. in cortex and by 8 mg./gm. in medulla; Q_A increased from 12.3 to 13.5 in cortex and decreased from 5.1 to 3.8 in medulla.     

ANOXIA AND THE CONVULSIVE STATE

The author believes he has clinical and experimental evidence to suggest that one of the underlying causes of idiopathic epilepsy lies in lack of sufficient oxygenation of the cerebrum before or during birth.     

曲霉淀粉酶基因与转录调控研究进展*

本文总结了到目前为止已克隆到的曲霉淀粉酶基因,并阐述了已发现的与调控相关的元件和因子如：顺式作用正调控元件和反式作用正调控因子等,在此基础上提出了曲霉淀粉酶基因的转录调控方式,并简述了曲霉淀粉酶基因的应用前景。 淀粉酶是水解淀粉和糖原酶类的通称,广泛存在于动植物和微生物中（张树政 1984）。由于淀粉酶在工农业上的重要价值,因此很早就有人对淀粉酶进行研究。我国50年代初,先后研究和生产米曲霉的α-淀粉酶用作消化剂和葡萄糖生产,将细菌产生的α-淀粉酶用于纺织品退浆、饴糖制造等(张树政 1984);随着分子生物学技…     

低氧条件下大脑皮层微血管平滑肌细胞内皮细胞中[Ca~(2 )]_i变化的研究

目的和方法：本研究采用离子探针Ｆｕｒａ２／ＡＭ 结合计算机图象分析技术,并通过施加ＮＯ合酶抑制剂ＬＮＮＡ和ＮＯ的作用靶———鸟苷酸环化酶（ＧＣ）的抑制剂美兰（Ｍｅｔｈｙｌｅｎｅ Ｂｌｕｅ;ＭＢ）,观察经培养的大鼠大脑皮层微血管内皮细胞和平滑肌细胞中的［Ｃａ２＋］ｉ 在低氧作用后的变化以及与有关血管舒张因子ＮＯ和ｃＧＭＰ之间的关系。结果：低氧时大脑微血管内皮细胞和平滑肌细胞内的Ｃａ２＋ 浓度有所下降,变化幅度的大小与低氧的程度及低氧作用的时间有关,且可以被ＬＮＮＡ和ＭＢ所抑制。结论：低氧时大脑微血管的舒张反应与ＮＯ的产生有关,ＮＯ通过细胞内的多种机制,最终使得胞内Ｃａ２＋ 下降而导致血管舒张