食蟹猴肺上皮细胞株MFLE-001的建立及生物学特性研究

该研究建立了食蟹猴肺上皮细胞株并鉴定其生物学特性。以食蟹猴肺部组织进行原代培养,用差速贴壁法纯化细胞株,并通过倒置显微镜、透射电镜、扫描电镜观察,绘制生长曲线,进行核型分析以及肺上皮标志物检测等方法研究其生物学特性。结果显示,体外培养细胞生长均一、稳定,细胞倍增时间为4.8天,免疫组化CK(+)、CK-7(+)、波形蛋白(+),染色体分析表明,具有正常2倍染色体数目(2n=42),符合食蟹猴肺上皮细胞株特性。该研究成功建立第一株食蟹猴肺上皮细胞株,命名为MFLE-001,为肺部疾病研究和药物筛选提供了理想的体外实验模型。     

食蟹猴糖化血红蛋白水平的调查研究

目的调查食蟹猴群体的HbA1c水平并建立其背景数据库。方法采集后肢静脉血以高效液相色谱法测定176只(雌性88只;雄性88只)食蟹猴的HbA1c值。结果雄性食蟹猴的HbA1c平均值高于雌性食蟹猴(4.80%±0.56%vs.4.61%±0.55%),两者间存在显著性差异(P0.05)。不同年龄段食蟹猴的HbA1c统计结果发现,青年组雌雄性食蟹猴的HbA1c平均值均高于老年组,两个年龄段间的雌性和雄性之间均存在显著性差异(P0.05),而中年组雌性与雄性和其他年龄段间的差异不显著。结论不同性别和年龄段间食蟹猴HbA1c水平具有显著差异性,HbA1c水平与性别和年龄具有相关性。     

食蟹猴的基础血糖值调查

目的 调查圈养食蟹猴基础血糖值情况.方法 采用快速血糖仪对153只6～19岁雄性食蟹猴和87只6～24岁雌性食蟹猴的血糖进行测定.结果 不同性别的食蟹猴血糖值存在显著性差异(P＜0.05),其中雌性食蟹猴血糖平均值为4.09 mmol/L±1.03 mmol/L,雄性食蟹猴血糖平均值为3.32 mmol/L±0.59 mmol/L;不同年龄段的食蟹猴血糖值差异显著(P＜0.05),年龄大的食蟹猴血糖值比年龄小的食蟹猴血糖值整体较高;体重指数与基础血糖值之间无显著相关性.结论 食蟹猴基础血糖值与人类基础血糖值相比,水平较低;性别和年龄是影响食蟹猴血糖值的主要因素.食蟹猴基础血糖值调查为糖尿病动物模型的建立及其相关研究提供了有关血糖值的基础数据参考.     

性成熟前食蟹猴生精细胞的发育进程

目的阐明性成熟前食蟹猴生精细胞的发育进程。方法分别采集性成熟前不同年龄（0岁、0.5岁、1岁、1.5岁、2岁、2.5岁、3岁、3.5岁、4岁）食蟹猴睾丸,制作石蜡切片,进行HE染色和PAS/H染色。根据生精细胞的染色特性,分析性成熟前食蟹猴生精细胞的发育进程,并对食蟹猴精原干细胞进行初步鉴定。结果 HE染色结果显示,1岁及以下食蟹猴生精上皮上生精细胞仅有精原干细胞（包括Ad、At及Ap型精原细胞）,1.5岁食蟹猴生精上皮上开始出现B型精原细胞,3岁食蟹猴生精上皮上出现精母细胞,4岁食蟹猴生精上皮上出现从精原干细胞到精子的所有生殖细胞。PAS/H染色结果显示,1～2.5岁食蟹猴Ad型精原细胞胞质呈PAS阳性,At型精原细胞胞质呈PAS弱阳性,Ap型精原细胞胞质呈PAS阴性;其他生精细胞及支持细胞胞质呈阴性;0.5岁及以下,3岁及以上食蟹猴生精细胞的胞质PAS/H染色特性与前者存在差异。结论本文详细阐述了性成熟前食蟹猴生精细胞随年龄增长的渐次性发育模式,并建立了性成熟前食蟹猴精原干细胞原位鉴定的一种新方法,这些研究结果为食蟹猴精原干细胞的其他相关研究奠定了基础。     

恒河猴与食蟹猴微卫星DNA多态性的比较分析

目的分析恒河猴和食蟹猴群体间的遗传多样性,确立一种对恒河猴和食蟹猴种群个体的遗传鉴别方法。方法利用聚合酶链反应（PCR）扩增技术采用15个多态性微卫星DNA位点对50只恒河猴和50只食蟹猴个体进行了DNA多态性的分析,对比两群体间等位基因数目差异。结果筛选的15个具有显著多态性的微卫星DNA位点对恒河猴和食蟹猴种群可以进行DNA多态性分析,其等位基因数目均在7个以上,且两群体间有11个位点的等位基因数存在一定的差异。结论利用这些多态性微卫星DNA位点建立一种有效鉴别恒河猴和食蟹猴种群遗传背景的方法具有一定的可行性。     

妊娠晚期食蟹猴与同龄未孕食蟹猴血液生理和生化指标的比较

目的提供妊娠晚期食蟹猴有关的血液学数据。方法采用Sysmex XE-2100血液分析仪、SysmexCA-7000凝血分析仪,及OL YMPUS AU600型全自动生化分析仪,对15只怀孕晚期食蟹猴及10只正常对照非孕食蟹猴进行血常规、凝血三项及17项血生化指标分析。结果妊娠晚期食蟹猴在WBC、NEU%、LYM%、PT、APTT、Fgb、TP、Alb、BUN、CR、CHOL、Glu、Fe、LDK、HDBH上,与同龄未孕食蟹猴相比差异显著（P〈0.05）,其他指标RBC、HGB、HCT、MCV、MCH、MCHC、PLT、ALT、AST、ALP、T.Bili、TG、P相比,于孕晚期有不同程度改变,但无显著性差异（P〉0.05）。结论初步建立了孕晚期食蟹猴的常规生理学数据,为相关的实验研究提供基础资料。     

自发性与膳食诱发性食蟹猴2型糖尿病其相关基因的表达比较

采用荧光定量PCR技术对自发性和膳食诱发性T2DM食蟹猴外周血白细胞中36个糖尿病相关基因的表达水平进行分析。在36个基因中,糖尿病组的G6PC、CCR2B、CTLA4等19个基因的表达量与对照组相比存在显著差异(P<0.05),且这些基因在诱发组和自发组中的表达模式基本一致。36个基因中,诱发组基因的表达量普遍高于自发组,但大部分基因表达量在两组中差异不显著,表明诱发性和自发性食蟹猴T2DM模型均可作为糖尿病研究较理想的动物模型。因此,通过高能量膳食诱导的食蟹猴糖尿病模型可以替代自发的糖尿病猴,且基因表达量的变化可为疾病的诊断和治疗提供帮助。     

食蟹猴微卫星DNA标记遗传监测及多态性分析

目的研究国内食蟹猴种群的遗传背景特性,建立食蟹猴种群遗传质量监测方法。方法采用微卫星DNA遗传标记技术对50只食蟹猴种群个体进行遗传质量监测及DNA多态性分析。结果从100个微卫星DNA位点中筛选出20个多态性高的位点,其食蟹猴种群个体的等位基因数目为5-10条,个体间均呈现高度的多态性;其观察等位基因数（Na）为5.0～10.0,有效等位基因数（Ne）为4.6118～8.3404,基因多样性（H）为0.7832～0.8801和香隆信息指数（I）为1.5651～2.1592。结论本实验有效地分析了食蟹猴种群的遗传多态性,为今后筛选特异性微卫星位点来建立食蟹猴种群遗传质量监测方法提供了理论依据。     

四种麻醉药物对食蟹猴麻醉效果分析和选择应用

目的分析比对速眠新、氯胺酮、异氟烷和利多卡因4种不同麻醉药对食蟹猴的麻醉效果。方法总结实际工作中分别使用四种不同麻醉药物对食蟹猴作用的麻醉特点。结果速眠新、氯胺酮、异氟烷均能获得较好的麻醉效果,能满足不同手术、采样需要;局麻药利多卡因对食蟹猴麻醉的实际应用不理想。结论食蟹猴的手术及其他侵犯性操作等都应该考虑生物安全和动物福利要求,实行麻醉,但应根据食蟹猴实验内容要求和不同麻醉药特点选择合适的麻醉方法,确保人员和动物安全,实验结果不受影响。     

食蟹猴心血管病风险因素与心血管病相关基因在外周血白细胞表达的相关性

研究心血管病风险因素和心血管病相关基因表达的相关性有助于心血管病风险预警和早期诊断研究。该文采用温和的动脉粥样硬化膳食(0.053mg胆固醇/千焦、40%的能量来源于脂肪)喂养中老年雄性食蟹猴(Macaca fascicularis)(12个月),根据传统心血管病风险因素筛选低、高风险食蟹猴,然后采用荧光定量PCR技术检测113个心血管病相关基因在正常组、低风险和高风险组食蟹猴外周血白细胞内的表达差异。结果在食蟹猴外周血白细胞中共检测到65个心血管病相关基因,其中低、高风险组有16个基因相对于正常组表达上调(P<0.05),19个基因表达下调(P<0.05),另外,还有15个基因表达模式特异(P<0.05)。此外,还检测到42个心血管病相关基因在人和食蟹猴外周血白细胞内均有效表达,其中22个基因在两者之间表达模式一致。上述结果为进一步研究心血管病风险预警和早期诊断指标,缩小了基因遴选范围。     

Detecting ovulation in Macaca nemestrina by correlation of vaginal cytology, body temperature and perineal tumescence with laparoscopy

Vaginal cytology, basal body temperature, and perineal tumescence were correlated with laparoscopic observations during the menstrual cycles of five pigtail monkeys (Macaca nemestrina) of known fertility. Percentages of cells obtained in vaginal smears revealed systematic variation in the presence of cell types in relation to the menstrual cycle. Measuring the percentage of exfoliate vaginal epithelial cells containing pyknotic nuclei proved to be of little value for separating the menstrual cycle into its follicular and luteal phases, nor did body temperature provide an accurate index for the occurrence of ovulation. Perineal tumescence, however, measured from the first day of menses to onset of detumescence, was a reliable indicator of the lengths of the follicular and luteal phases as correlated with laparoscopic confirmation of ovulation. Maximal perineal tumescence usually occurred within 12 hours of ovulation, although on one occasion the two events were separated by 48 hours.     

Ultrastructure of the nonhuman primate vaginal mucosa: epithelial changes during the menstrual cycle and pregnancy

Ultrastructural changes in the vaginal epithelium of the rhesus monkey during the menstrual cycle and pregnancy were studied by scanning and transmission electron microscopy. During the menstrual cycle, the epithelium was keratinized but varied in thickness. Cells of the basal and parabasal layers were polyhedral in shape but as they differentiated they accumulated glycogen and filaments. Cells in the intermediate layers had keratohyaline and membrane-coating granules. Cells in the superficial layers had a thickened cell envelope, abundant keratin filaments, electron-dense intercellular material, and focal tight junctions. The epithelial surface had numerous microridges and numerous adherent bacteria; bacteria were rare on desquamating cells. The epithelium remained keratinized for about the first month of gestation, then underwent "mucification." The cells contained abundant granules and Golgi apparatus. Concomitant with this transformation, bacteria were no longer adherent to the epithelial surface and the surface cells had microvilli instead of microridges. The epithelial changes during pregnancy were roughly associated with the changing pattern of steroid hormone secretion during gestation.     

Age- and hormone-related changes in vaginal smear patterns in the gray-tailed vole, Microtus canicaudus

Female voles, Microtus canicaudus, exhibited age-related changes in vaginal smear patterns when isolated from males after weaning. Between 30 and 50 days of age, nearly all females exhibited persistently leucocytic vaginal smears. By 90-120 days, most females showed vaginal cyclicity with alternating predominance of leucocytes, nucleated epithelial cells or cornified epithelial cells. Most females examined between 150 and 200 days of age exhibited persistent vaginal cornification. The vaginal cyclicity seen in females between 90 and 120 days was not a reflection of cyclic ovulatory changes; plasma progesterone concentrations remained constant, regardless of age or vaginal smear pattern, and corpora lutea were never seen in unmated females. Although progesterone concentrations did not differ among vaginal smear patterns of 120-day-old females, plasma oestrogen values were highest in females exhibiting vaginal cornification.     

Keratinization of rat vaginal epithelium. II. Immunofluorescence study on keratin filaments in cycling and estrogen primed rats

Rat vaginal epithelial layers from animals in different phases of the estrous cycle showed positive immunofluorescence when treated with either monoclonal antibody to intermediate filaments or immunoglobulin G fraction of antiserum raised against epidermal keratin filaments. During estrus, the intensity of fluorescence observed was maximum in the keratinized cellular layers. In estradiol-primed immature and ovariectomized rats the maximum fluorescence intensity was observed in the layers immediately lining the lumen. However, basal layers in ovariectomized rats also showed some fluorescence. Data presented in this communication indicate that the abundance of keratin filaments in vaginal epithelial cells can be modulated by altering the level of estradiol in the system.     

Histomorphology of cervical and uterine epithelia of crab-eating macaque, Macaca fasciularis.

Cytological characteristics and pattern of distribution of different cell types in the epithelia of cervix and uterus of crab-eating macaque (Macaca fascicularis) in follicular and luteal phases of the menstrual cyclic and amenorrhea were studied. The cervix uteri and uterus exhibit remarkable structural differenes in the ciliated, secretory, and ciliated-secretory cells. Since the number of ciliated-sexretory cells in the uterus is higher than in the cervix. It is believed that they form an additional source for the secretion of uterine fluid during the menstrual cylce. Both ciliated and secretory cells undergo degeneration; extensive cytoplasmic vacuolation associated with pycnosis and disorganization of the nuclei encountered.     

Regulation of the bacterial microflora of the vagina in cyclic female rats.

The bacterial microflora was examined in the vagina of cyclic female rats kept under normal laboratory conditions. Large variations occurred during the cycle with high numbers of bacteria (10(5)-10(8) per vagina) during proestrus and estrus and low numbers (10(1)-10(4) per vagina) during the diestrus period. Histological analysis of in situ vaginal tissue and transplanted vaginal tissue revealed an association of high bacterial numbers with the presence of large amounts of cellular debris in the vaginal lumen during the period of epithelial keratinization. Absence of phagocytosis in leucocytes at mestestrus suggested that leucocytes did not play an active role in reduction of bacterial numbers between estrus and metestrus. Accurate measurement of the pH in the vaginal lumen failed to reveal differences which could explain the reduction in bacterial numbers between estrus and metestrus. The cyclic changes in the bacterial population-consisting of species which are normally present in the intestinal flora-- seem to be controlled by cyclic changes in the amounts of cellular debris in the vaginal lumen.     

Localization of estrogen receptor α and progesterone receptor B in bovine cervix and vagina during the follicular and luteal phases of the sexual cycle

Abstract

The localization and distribution of estrogen receptors (ERα) and progesterone receptors (PR-B) in the cervix and vagina of sexually mature bovines during the follicular and luteal phases of the sexual cycle were studied using immunohistocehmistry. The estrous cycle stage of 23 Holstein bovines was assessed by gross and histological appearance of ovaries and blood steroid hormone values. Tissue samples from cervix and vagina were fixed in 10% formaldehyde for routine histological processing. Nuclear staining for ERα and PR-B was observed in the epithelial cells of the surface epithelium, stromal cells and smooth muscle cells. Generally, in the cervix, ERα immunoreactivity was more intense in the epithelial and smooth muscle cells during the follicular phase and in the epithelial cells during the luteal phase (p < 0.05). PR-B immunoreactivity was more intense in the epithelial and smooth muscle cells than in the superficial and deep stromal cells during the follicular and luteal phases (p < 0.05). In the vagina, ERα and PR-B immunoreactivities were more intense in the epithelial cells than in the connective tissue cells and smooth muscle cells during the follicular and luteal phases (p < 0.05). These results indicated that the frequency and intensity of ERα and PR-B immunoreactivity in the cervix and vagina of bovines varied according to the cervical and vaginal cell types and the phases of the sexual cycle.     

Oestrous cycle of captive southern hairy-nosed wombats (Lasiorhinus latifrons) in South Australia, Australia

There is limited information available on the oestrous cycle of female southern hairy-nosed wombats (Lasiorhinus latifrons). This is mainly due to an extremely poor breeding success in captivity and the difficulty in routine recapturing of these cryptic, semi-fossorial animals in the wild. The aim of this study was to characterise the oestrous cycle of this species by monitoring peripheral plasma concentrations of progesterone and oestradiol, assessing changes in vaginal cytology, pouch condition and the urogenital sinus. Eight adult female wombats were monitored during the breeding season (July-December) over 2 years (2002-2003). Samples were collected up to three times a week. Vaginal smears contained several cell types, categorised by morphology, as either superficial epithelial cells or parabasal-intermediate cells. Leucocytes were also counted. Plasma progesterone profiles showed a mean oestrous cycle length of 36.33+/-0.67 days with a peak progesterone concentration of 139.53+/-10.62nmol/L. Levels of oestradiol peaked at a mean level of 467.33+/-44.32pmol/L on average 5 days before a rise in plasma progesterone values. The proportion of epithelial cells in vaginal smears varied throughout the cycle, with a high percentage of superficial epithelial cells observed during the follicular phase. During periods when progesterone concentrations were high, a greater percentage of parabasal-intermediate cells was observed. In conclusion, this study has characterised the oestrous cycle of the southern hairy-nosed wombat and confirmed that changes in vaginal smears together with pouch and urogenital sinus details could be used to determine signs of oestrus in this species.     

小鼠发情周期卵泡发育动态及其对超数排卵的影响

该文探讨了小鼠发情周期中阴门状态、阴道脱落细胞类型变化规律、卵泡发育规律及其相互关系,并比较了发情周期不同阶段的超排效果。结果表明,采用阴门状态观察法和阴道脱落细胞涂片法,能有效判断小鼠发情周期阶段。卵巢组织切片观察结果表明,在发情周期不同阶段,小鼠的卵泡发育和黄体的生成与消退存在明显的规律性变化;小鼠发情周期中,其阴门状态、阴道脱落细胞种类及卵泡发育动态之间存在相关关系;发情周期不同阶段开始超排的小鼠,其配种见栓率和回收胚胎平均数均存在明显差异,发情前期显著优于发情后期与间情期(P<0.05),并高于发情期,但差异不显著(P>0.05),即阴门状态观察法与阴道脱落细胞涂片法均可用于小鼠发情周期阶段的判断,发情前期为最适宜的小鼠超排时期。