CHANGES IN THE PROLIFERATION RATE OF MOUSE EPIDERMIS AFTER IRRADIATION: CONTINUOUS LABELLING STUDIES

The proliferative response of mouse skin to damage caused by X-irradiation has been tested by giving repeated injections of tritiated thymidine and scoring the percentage of labelled cells in high resolution autoradiographs. Four, nine and fourteen daily fractions of 300 rads of X-rays were used and labelling commenced 4 days after the last fraction. The epidermis of the upper surface and the sole of the foot were scored separately and were compared with the skin of unirradiated feet. In unirradiated skin the proliferation rate of the basal layer cells is more rapid on the sole than on the upper surface. The cell cycle times deduced from continuous labelling curves were 81 hr and 111 hr respectively and the growth fractions were 97% and 85%. After irradiation with small daily doses the homeostatic response to cell killing was slow. More rapid proliferation occurred after nine fractions in the sole, but was not apparent in the skin of the upper surface until fourteen fractions had been given. After fourteen fractions the cell cycle time was about 24 hr on both surfaces and the growth fraction was about 90%. The initial labelling index after a single thymidine injection was a poor measure of proliferation rate. The delay in the time of onset of faster proliferation is similar, both qualitatively and quantitatively, to that measured previously from the additional dose increments needed if large doses were given at different times after the multifraction treatments (Denekamp, 1973).     

STUDIES ON DNA FROM NORMAL AND SCRAPIE-AFFECTED MOUSE BRAIN

Abstract— DNA has been extracted from normal and scrapie affected mouse brain fractions, 48 h after the injection of [³H]thymidine precursors. The extracted DNA has been subjected to fractionation on hydroxyapatite columns and CsCl gradients. The specific activity of double stranded nuclear DNA is two to three times higher when extracted from scrapie-affected brain than from normal brain, but there is no apparent difference in the number of counts associated with double stranded mitochondrial DNA extracted from similar numbers of normal and scrapie affected brains. DNA from the large granule fraction of scrapie affected brain contains a peak of counts, melting off hydroxyapatite columns before the double stranded peak, consistent with the presence in scrapie brain of trace amounts of a small single stranded DNA.     

CELL POPULATION KINETICS AND DESQUAMATION SKIN REACTIONS IN PLUCKED AND UNPLUCKED MOUSE SKIN

The kinetics of cellular proliferation in plucked and unplucked dorsal skin of mice after local X-irradiation are described, in relation to the time course of the gross desquamation reaction in skin of the dorsum and of the foot.     

FURTHER OBSERVATIONS ON THE LATE LABELLING ASSOCIATED WITH STIMULUS-RESPONSIVE CELLS IN SKIN

The presence of a long-lived DNA precursor pool which may show some specificity for stimulus-responsive cells has been demonstrated. Autoradiography, biochemical analysis, hydroxyurea sensitivity, and the temporal response all suggest that the late incorporation is into DNA in cells in the basal layer of the skin that respond to stimulation. The effect is observed with various doses of tritiated thymidine and both methyl and 6 labelled material. ¹²⁵Iododeoxyuridine also shows late labelling in skin. It is believed that the late labelling is readily distinguishable from reutilization and from possible slow utilization of catabolites into molecules other than DNA. Skin of the ear and dorsum shows similar degrees of late incorporation, while tail and foot skin apparently have smaller long-lived pools. This may indicate smaller stimulus-responsive cell populations. If tritiated thymidine is given to cells after stimulation there is still some slow or delayed uptake. The distribution of labelled cells in the autoradiographs suggests that the late labelling cells may be associated somehow with pre-existing labelled cells (cells in S at the time of thymidine administration).     

CELL POPULATION KINETICS OF PLUCKED AND UNPLUCKED MOUSE SKIN I. UNIRRADIATED SKIN

A detailed study of the cellular proliferation kinetics in interfollicular plucked and unplucked mouse skin has been made in Swiss albino mice, using tritiated thymidine autoradiography. Diurnal variations in mitotic and labelling indices were demonstrated in both systems.
The mean cell cycle times for unplucked and plucked skin were estimated by four different methods and found to be 100 ± 10 and 47 ± 3 hr respectively. Most of the difference was due to the shortening of G₁ phase after plucking. Repeated labelling at intervals shorter than the DNA synthesis times resulted in all the basal layer cells becoming labelled, so that the growth fraction was unity, in unplucked and plucked skin.
A well-defined second wave of labelled mitoses was seen at about 100 hr after labelling the unplucked (i.e. normal) mouse skin.
A double labelling technique using ¹⁴C-TdR and ³H-TdR with a single layer of emulsion gave reasonable values for the duration of the DNA synthesis phase.     

小鼠巨核系祖细胞增殖,分化特性的研究

小鼠骨髓细胞经7d培养后进行细胞形态学观察,可见不同发育阶段的巨核细胞及不同大小的巨核细胞集落。通过计数每个集落中的细胞数,可确定相应祖细胞的有丝分裂能力。结果表明,具有不同有丝分裂能力的祖细胞的体外增殖动力学有所不同。祖细胞的数量与其有丝分裂次数呈负相关(r=-0.986)。进行0、1、2和3次有丝分裂的祖细胞的阿糖胞苷自杀率分别为48.9,58.7,48.0和41.2％;放射敏感性的D_O值(Gy)分别为1.71,1.24,1.03和0.77,D_O值的大小与有丝分裂次数呈负相关(r=-0.958)。经3Gy全身照射后CFU-Meg与CFU-GM的恢复动态过程具有不同特点。     

小家鼠和实验小鼠遗传特性的比较研究

本文用同工酶电泳法、微量细胞毒法和免疫双向扩散法对我国4个动物地理区的6个采集点的156个小家鼠(Mus musculus)进行了遗传特性的调查。结果发现:在全部被测的13个位点中,小家鼠在7个位点上存在着多种实验小鼠中罕见的基因组成;而不同动物地理区和亚区的小家鼠的遗传特性又各不相同。从而指出将小家鼠的特有基因导入实验小鼠,培育新品系的重大意义。     

饲喂泰和乌骨鸡及其黑素对小鼠肝、肠、肾酶组织化学活性的影响

应用酶组织化学方法观察了老年小鼠经饲喂泰和乌骨鸡及其黑素一个半月后,肝、肠、肾SDH、Mg^2 －ATPase、G－6－Pase、5′－Nase和MAO的酶活性变化。结果显示;肝、肠、肾SDH、Mg^2 -ATPase、G－6－Pase、5′-Nase的酶活性增强。MAO的酶活性减弱。结果提示：泰和乌骨鸡及其黑素具有促进机体代谢、维持内环境稳定,延缓衰老的作用。     

STUDIES ON THE LABELLING PATTERNS OF RNA FROM CEREBRAL CORTEX NUCLEI

RNA labelling patterns in nuclei from rat cerebral cortex were investigated subsequent to intracerebral injection of [³H]uridine. Although there was a rapid uptake of label into the ‘heavy’ regions when nuclear RNA was analysed in density gradients, it was not possible to show conclusive evidence for 455 ribosomal precursor RNA. Methyl-ation of 18S and 28S nuclear RNA became evident only after 2 hr and did not appear to involve the intermediacy of RNA species of higher molecular weight.     

用单克隆抗体ELISA研究和分析霍乱弧菌抗原结果

本文用ELISA以12株单克隆抗体(McAb)对55株EI Tor弧菌和用遗传学方法突变的28株霍乱弧菌的菌体抗原进行了研究,同时与四种常规分型方法进行对比试验。用12株McAb可将55株El Tor弧菌分成11种抗原决定簇和18个McAb型,将遗传学方法的突变株分成9种抗原决定簇和10个McAb型。其中一株McAb2B_1H_1F_3能识别所有55株EI Tor弧菌和大部分突变株霍乱弧菌,表明有种的特异性;其余11株McAb与各型霍乱弧菌的抗原反应,均有差异,与这些菌株的反应百分比各不相同(21.0％～100％),表明可能存在着亚型和亚群。文中着重讨论了用McAb研究分析抗原的重要意义和McAb用于诊断的价值。     

正常菌群对常见致病菌拮抗作用的研究

生物拮抗作用是微生物种群关系研究的一个侧面,实验中我们使用从皮肤上分离的常住菌——痤疮丙酸杆菌(A14-1)和表皮葡萄球菌(F65)对常见致病菌如金黄色葡萄球菌(C189)、绿脓杆菌(C8514)和埃希氏大肠杆菌(C1356)的体外拮抗试验表明,它们具有明显的拮抗作用,拮抗作用一般从48小时开始,72小时明显。而皮肤常住菌间无拮抗作用,相反它们还有协同作用,这对于皮肤的自净以及维持皮肤的微生态平衡具有重要的作用。此外,实验中还进行了乳杆菌和双歧杆菌等正常菌群分离株与上述常见致病菌的体外拮抗试验,结果表明也     

STUDIES ON EPITHELIAL CELLS ISOLATED FROM GUINEA PIG SMALL INTESTINE

Sheets of mucosal epithelial cells were released from guinea pig small intestine after incubation with ethylenediaminetetraacetate. Cells in sheets retained their columnar shape for 24 hr at room temperature, and exclusion of nigrosine suggested they had intact plasma membranes. When sheets were disaggregated individual cells had normal morphology for at least 4 hr. During isolation 16% of the total protein and 24% of the total lactic dehydrogenase were lost from the cells, but subsequent enzyme leakage was low. Leakage increased with shaking, incubation at 37°C, or increasing the oxygen tension of the suspending medium, but was minimal when the Na⁺:K⁺ ratio in the medium was 8:1 and the osmolarity was high. Losses of particulate enzyme activities were negligible. Respiration was constant for up to 4 hr and was insensitive to calcium, bicarbonate, oxygen tension, and pH. It was inhibited by cyanide and iodoacetate and varied with the Na⁺:K⁺ ratio of the extracellular fluid and the structural integrity of the cells. All preparations concentrated potassium and excluded sodium, but lost this ability if ouabain was added or cells were broken. Potassium-42 uptake was also sensitive to temperature, ouabain, and structural integrity. The preparations are being used to study cell metabolism in the intestinal epithelium.     

应用氯化锶和放线菌酮对小鼠卵母细胞进行孤雌活化的研究

本试验研究了SrCl_2浓度和作用时间,以及卵龄和蛋白合成抑制剂放线菌酮等对昆明种小鼠卵母细胞活化的影响。研究表明,以含1.6mmol/L SrCl_2的无钙M16液对小鼠卵母细胞活化效果最好(87.0％),显著(P<0.05)优于SrCl_2浓度为1.0、5.0、10.0mmol/L的同种液体。SrCl_2作用时间10分钟显著(P<0.05)好于5、20、30或60分钟。注射hCG后18和20小时卵母细胞的活化率(分别为87.0％和84.6％)显著(P<0.01)高于14或16小时的活化率(分别为4.8％和16.5％)。CHX与SrCl_2联合使用产生显著的协同促进卵母细胞活化作用。     

小鼠精子注入兔卵母细胞受精研究

利用显微操作仪将小鼠精子注入家兔卵母细胞的胞质内和透明带下,对鼠兔异种精卵互作和异种受精胚胎的发育进行了研究,并对注射精子的数量及卵的体外成熟时间等影响鼠兔异种显微受精的因素进行了探讨,结果如下:(1)将小鼠精子分别注入兔卵胞质内和透明带下,均能激活兔卵母细胞,导致精核解聚和原核形成;(2)小鼠精子注入兔卵胞质内和透明带下受精,杂种胚胎体外培养能发育到8-细胞期;(3)鼠兔异种受精4-细胞胚胎染色体标本制备观察结果表明,它们为正常二倍体;(4)鼠兔异种受精4-细胞胚胎的超微结构观察结果表明,它们极近似兔正常4-细胞胚胎的超微结构;(5)将小鼠精子注入兔卵透明带下,注射5—10个精子组卵的受精率(32.4%)和卵裂率(16.2%)均高于注射单个精子组的,但二组间差异不显著(P>0.05);DM 15%NCS液中体外成熟培养11—12h兔卵透明带下注入1—2个小鼠精子后的受精率(42.3%)和卵裂率(30.8%)均高于体外成熟培养24—25h组的,但二组间差异未达到显著水平(P>0.05)。     

鼠肝细胞癌变中DNA甲基化作用的研究

本文用~3H标记的S-腺苷酰甲硫氨酸(~3H-SAM)为甲基供体,以同位素掺入法测定了正常小鼠肝细胞、H_(223)腹水癌细胞及荷癌肝细胞的DNA甲基化酶活力。并用HPLC法测定了上述细胞的DNA甲基化水平。发现:H_(223)腹水癌细胞及荷癌肝细胞的DNA甲基化酶活力和DNA甲基化水平明显低于正常肝细胞。当以抗肝癌药物去甲斑蝥素和斑蝥酸钠处理荷癌小鼠6天后,可使H_(223)腹水癌细胞及荷癌肝细胞的DNA甲基化酶活力回升,但并未检出DNA甲基化水平的回升。     

EXISTENCE OF AN ENDOGENOUS INHIBITOR OF DNA SYNTHESIS IN RABBIT SMALL INTESTINE SPECIFICALLY EFFECTIVE ON CELL PROLIFERATION IN ADULT MOUSE INTESTINE

Aqueous extracts from rabbit organs were prepared by homogenization and centrifugation at 105,000 g . After precipitation with ammonium sulphate, the 0–50 fraction was separated by ultrafiltration through Amicon XM 100 and XM 300 membranes yielding two filtrate fractions (U₁ and U₂) and one retentate fraction (U₃). Only U₁ and U₃ inhibited thymidine incorporation into DNA. After a single injection of U₁ from rabbit small intestine, the uptake of tritiated thymidine was decreased in mouse jejunal and colonic DNA. This effect, totally reversible after 7 hr, was found in neither the kidney nor the testis. The U₁ fractions of colon and non-digestive organs (kidney, testis) were found not to exert a significant inhibition on thymidine incorporation into intestinal DNA in vivo. The U₃ fraction from rabbit small intestine also decreased the uptake of tritiated thymidine in mouse jejunal and colonic DNA in vivo. However, this inhibition was irreversible and not tissue-specific. Slowing of cell migration was also noticed in the jejunum of mice injected with U₁ or U₃, as ascertained radioautographically by determining the position of the leading edge of the labelled cells in U₁- or U₃-injected mice compared with controls. A decrease of mitotic activity in U₁- and U₃-injected mice was recorded 8·5 hr after a single injection of small intestinal fractions. Our results suggest that U₁ and U₃ from rabbit small intestine contain one or more substances which may act on the G₁—S transition of the cell cycle in the mouse intestine. However, only the effect of U₁ is reversible and tissue specific. Our data suggest the existence of a factor, having a low molecular weight, which regulates intestinal cell proliferation.     

PHARMACOLOGICAL STUDIES ON GASTRODIA ELATA BLUME. 2.EFFECTS OF SYNTHETIC GASTRODIN AND ITS GENIN ON THE HEART AND SMALL INTESTINE

Gastrodin is a phenol-glucoside, obtained from Gastrodia elata Blume (Fam.Orchidaceae) by Investigator Zhou Jun and other of The Institute of Botany of Kun-ming, Academia Sinica. The structural formula of gastrodin is p-hydroxymethy phenyl -β-D-glucopyranoside, its chemical formula is C13H18O7, forming a white pin cry-stals, with a melting point of 154-156℃. It is synthetized now.In a previous paper, the authors reported the sedative and anticonvuisant effects of synthetic gastrodin and its genin. The prescnt paper reports the subacute toxicity and the effects of gastrodin and its genin on the heart and small intestine as followg.     

小白鼠发育过程中胃和大肠蛋白酶活性的变化

采用活性电泳方法对小白鼠发育过程中胃和大肠蛋白酶活尾进行了研究。结果发现：（１）出生前、胃和大肠蛋白酶种类少、活性弱,出生后,胃和大肠蛋白酶的种类多,活性增加;（２）胃肠蛋白酶可能与其结构的构建和功能建立有密切关系;（３）食物 刺激可能对胃肠蛋白酶有显著影响。