表皮生长因子对无透明带小鼠胚胎体外发育的影响

为了观察不同浓度的表皮生长因子对无透明带小鼠胚胎体外发育的影响,试验将小鼠原核期胚胎用链霉蛋白酶去除透明带后,分别置于含有不同浓度的表皮生长因子无血清胚胎体外培养液中培养,观察各期胚胎的发育情况.结果表明:除0.01 ng/mL表皮生长因子添加组胚胎的2细胞发育率和囊胚回收率与添加0.1 ng/mL表皮生长因子处理组差异不显著外(P>0.05),其体外胚胎各期的发育率、囊胚回收率和囊胚细胞数与其他处理组比较均差异显著(P<0.05).说明一定浓度的表皮生长因子可以提高无透明带小鼠胚胎体外培养各期的发育率,但是高浓度的表皮生长因子对无透明带小鼠胚胎的体外发育有一定的抑制作用.     

金丝桃素可溶性粉体外抗犊牛病毒性腹泻-黏膜病病毒的实验研究

为了探讨金丝桃素可溶性粉抗犊牛病毒性腹泻-黏膜病病毒(BVD-MDV)的作用机制,试验过程中采用先给药后接毒、先接毒后给药和同时给药接毒3种不同的途径,分别在72、96、120 h观察病毒致牛肾原代细胞(MDBK)的病变(CPE)情况.结果表明,病毒的TCID50为10-6/0.1 mL,金丝桃素可溶性粉的TC0为2.5 g/L;药物质量浓度在0.3～2.5 g/L范围内同时给药和接毒组细胞生长良好,细胞单层完整,视野中未见大量圆细胞或死细胞,而其余2种给药途径细胞病变明显.结果提示,金丝桃素可溶性粉对BVD-MDV有显著的直接杀灭作用.     

不同培养液对小鼠受精卵体外发育的影响

小鼠卵母细胞体外受精后通过比较4种培养液(M16、mCZB、HTF、mCZB*)对小鼠受精后胚胎发育的影响,以期建立最优的体外受精培养液系统。结果表明:受精卵在mCZB*、M16、mCZB、HTF培养液中2-4细胞发育率分别为60.26%、50.00%、48.72%和52.78%。mCZB*比其他3种培养液效果显著(P<0.05),其他3个培养液之间差异不显著(P>0.05)。在mCZB*、M16、mCZB、HTF培养液中囊胚发育率分别为44.37%、27.03%、26.28%和22.22%,各培养液处理组之间差异显著(P<0.05)。本试验提示mCZB*为小鼠体外受精胚胎发育的最佳培养液。     

高能微波对体外培养乳鼠心肌细胞生长活力的影响

体外培养的心肌细胞可保持心肌结构及功能上的诸多特点 ,有自发性节律性搏动 ,且不受神经、体液等体内因素的影响 ,因而在实验研究上得到了广泛应用。目前 ,乳鼠心肌细胞的培养方法较多 [1 ] ,但由于心脏细胞成分复杂 ,心肌细胞培养过程较为繁琐 ,应根据实验目的摸索简便快捷的培养方法。随着微波技术的应用 ,微波污染也日益增多 ,并成为日常影响人畜身体健康的主要物理因素之一 [2 ]。本试验对乳鼠心肌细胞的培养方法作了改进 ,并观察了高能微波对乳鼠心肌细胞生长活力的影响。1　材料与方法1 .1　动物　生后 1～ 3d的 Wistar大鼠 (二级 )…     

影响小鼠卵母细胞体外成熟和体外受精效果的因素分析

研究卵母细胞发育程度和不同营养因子对小鼠卵母细胞体外成熟及体外受精率的影响。小鼠卵母细胞分别在含有FSH、BSA和胰岛素的培养液中体外成熟 ,在Whitten氏液中体外受精 ,比较体外成熟率、体外受精率。卵丘卵母细胞 (COC)较裸卵 (DO)的体外成熟率、体外受精率都低。FSH和胰岛素均能提高小鼠卵母细胞的体外成熟率、体外受精率。BSA可以降低小鼠卵母细胞体外受精率。GV期卵母细胞的体外受精率显著低于体外培养的排出第一极体的卵母细胞。 (P2 -cell <0 0 5 ,P受精 <0 0 5)。     

Effect of zearalenone on the growth of mouse embryos from blastocysts to the egg cylinder stage in vitro

Embryos were harvested at the blastocyst stage from nontreated outbred mice and were grown in vitro for 4 days. Embryos cultured in control medium hatched and grew to the egg cylinder stage. Purified zearalenone (ZEN) added to the culture medium at concentrations of 8.5 to 68 micrograms/ml decreased the number of embryos growing, with a 50% decrease in the number growing in 32 micrograms of ZEN/ml of medium. Embryos growing in ZEN had decreased numbers of cells derived from the inner cell mass, normal growth of the trophoblast, less cellular differentiation than was seen in control embryos, and increased numbers of phagosomes. Undifferentiated cells of the inner cell mass of control and treated embryos were of the same size, as determined by morphometric analysis. Addition of 25 micrograms of estradiol/ml of culture medium caused no decrease in number of embryos growing or in embryonic size. Saturation of culture medium with ZEN (68 micrograms/ml) did not inhibit the growth of a tissue culture line of goat synovial cells. Seemingly, ZEN at concentrations near saturation inhibited the growth of mouse embryos in vitro. This effect was not duplicated with similar concentrations of estradiol and was not manifested in culture-adapted cells.     

Effect of nutrient intake during pregnancy on fetal and placental growth and vascular development

Remarkable diversity of size and health of offspring exists after normal pregnancies. When pregnancies are complicated by an extrinsic variable such as inappropriate maternal nutrition, birth weight and health of the neonate are substantially affected. The placenta is the organ through which respiratory gases, nutrients, and wastes are exchanged between the maternal and fetal systems. Thus, transplacental exchange provides for all the metabolic demands of fetal growth. Transplacental exchange is dependent upon uterine and umbilical blood flow, and blood flow rates are in turn dependent in large part upon vascularization of the placenta. Therefore, factors that influence placental vascular development will have a dramatic impact on fetal growth and development, and thereby on neonatal mortality and morbidity. Recent work from our laboratories has focused on the effects of nutrient intake during pregnancy on placental growth and vascular development. Both nutrient restriction of the adult dam and overnourishment of the adolescent dam during pregnancy suppress placental cell proliferation and vascularity. Furthermore, placental expression of angiogenic factors and their receptors, factors that are known to affect vascular growth, are perturbed by level of nutrition. Studies in this area will lead to improved methods to manage nutritionally-compromised pregnancies.     

Long-term effects of in vitro growth of mouse oocytes on their maturation and development

Since very few oocytes grow completely in vivo, in vitro growth (IVG) of ovarian oocytes may provide a new source of functional oocytes. The long-term effects of in vitro maturation (IVM) of oocytes and in vitro culture of fertilized eggs have been reported; however, the effects of IVG of oocytes are unknown. Here in, we report the long-term effects of IVG of oocytes. Ovaries from 1-day-old mice containing non-growing oocytes were cultured for 10 days; the isolated follicles were then cultured for 11 days. Secondary follicles from 10-day-old mice were also cultured for 11 days. The nuclei of oocytes collected from the IVG and Graafiais follicles of adult mice were transferred to enucleated oocytes grown in vivo, respectively. Developmental competence was examined following IVM of the reconstituted oocytes. Chronologically, oocytes of 1-day-old, 10-day-old and adult mice were cultured for 22, 12 and 1 day(s). The result showed that the reconstituted eggs developed into pups at high rates after nuclear transfer and in vitro fertilization (IVF) in all the experimental groups (29-45%). However, the pups from reconstituted eggs containing the nuclei of 22-day cultured oocytes were heavier than the control pups (P<0.05). We concluded that long-term culture of oocytes did not affect their nuclear ability to develop to term; however, fetal growth was affected by the culture duration or culture conditions during the initial phase of follicular growth.     

Effects of chicken thymic stromal cells on the growth and differentiation of thymocytes in vitro

We examined contact-mediated effects of chicken thymic stromal cells (TSC) on thymocyte differentiation by co-cultivation of these cell populations. The primary cultures of TSC isolated from thymus mainly have consisted of epithelial cells which were polygonal in shape, possessed long processes and expressed MHC class II antigen. When thymocytes were co-cultured with TSC, 60% to 70% of thymocytes attached to TSC and some of them engulfed underneath TSC. These attached thymocytes were CD4-CD8- and CD4+CD8+ subsets and expressed alpha/beta TCRhigh or gamma/delta TCRlow. Some of the thymocytes attaching to TSC showed an increase of intracellular and nuclear density, fragmentation of cytoplasm and nuclei, and DNA fragmentation. And also, thymocytes attaching to TSC contained a higher percentage of cycling (S and G2 + M phase) cells than nonattaching cells. These results indicate that specific subsets in thymocytes selectively bind to TSC and undergo apoptotic death or proliferation because of interaction with TSC. Chicken TSC may play an important role in thymic differentiation by direct contact within the thymus as in mammals.     

Effect of growth factors on the migration of equine oral and limb fibroblasts using an in vitro scratch assay

The objective of this study was to determine the effect of platelet derived growth factor BB (PDGF), epidermal growth factor (EGF), transforming growth factor β1 (TGFβ1), insulin like growth factor-1 (IGF-1) and fibroblast growth factor-2 (FGF-2) on the proliferation and migration of equine oral mucosa and leg skin fibroblast cell lines, using an in vitro scratch assay. Fibroblasts from the two sites were firstly grown to confluence and then an area of cells removed (cell void area). Cell migration alone (with the addition of the mitosis inhibitor mitomycin-C to the culture media) and proliferation and migration combined (without mitomycin-C) into the cell void area were observed at 0, 5, 10, 24 and 36h. The presence of mitomycin-C in the culture media significantly slowed the closure of the cell void area, as mitosis was inhibited. For the oral cells only, TGFβ1 significantly slowed both migration (with mitomycin-C) and proliferation and migration combined (without mitomycin-C). For the limb cells only, both PDGF and FGF-2 significantly increased fibroblast proliferation and migration combined (without mitomycin-C). For both cell types, EGF significantly reduced migration (with mitomycin-C). IGF-1 had no effect on any of the parameters measured. It was concluded that TGFβ1, PDGF and FGF-2 have differential effects on the proliferation and migration of equine oral and limb fibroblasts. These differences in fibroblast responses to growth factors may in part form the basis of the different clinical outcomes for oral and limb wounds.     

Effect of chronic infusion of placental lactogen on ovine fetal growth in late gestation

To test the hypothesis that placental lactogen (PL) is a Immoral regulator of fetal growth, six singleton sheep fetuses received a continuous intravenous fusion of 1.2 mg/d of purified ovine PL (oPL) for 14 d, beginning on Day 122 of gestation. The plasma concentration of oPL was approximately four-fold higher in infused fetuses than in six control fetuses that received a continuous infusion of saline. The circulating insulin-like growth factor 1 (IGF-1) concentration was also significantly elevated in PL-infused fetuses (43.1 ± 1.7 vs. 31.9 ± 4.l ng/ml; P < 0.05). Animals were slaughtered on Day 136, and the placenta and all major fetal tissues were dissected, weighed, and subsampled for chemical analysis. Fetal weight and crown-rump length were not significantly affected by treatment; however, the aggregate weight of the brain, liver, lungs, and heart tended to be larger (85.3 ± 2.1 vs. 79.9 ± 1.5 g/kg fetus; mean ± SE, P = 0.07) and the thyroid gland was smaller (0.18 ± 0.l vs. 0.26 ± 0.02 g/kg fetus; P < 0.05) in the PL-infused fetuses. The livers of the PL-infused fetuses had also accumulated additional glycogen (13.1 ± l.7 vs. 8.4 ± 0.7 g; P < 0.05). In late gestation, PL within the fetal compartment increases fetal plasma IGF-1 concentration and hepatic glycogen deposition and may affect the growth of several vital organs.     

Fetal growth of beef calves. II. Effect of sire on prenatal development of the calf and related placental characteristics

Fifteen Hereford and 47 crossbred heifers were allotted by breed and body weight to be artificially inseminated to one of two Angus sires selected for progeny birth weights (L = low; H = high). Forty-two of the heifers were randomly assigned to be slaughtered at 200, 215, 230, 245 or 260 d of gestation for measurement of fetal and placental characteristics. Twenty heifers were allowed to go to term and five calves from each sire group were randomly assigned to be euthanized and dissected within 24 h after birth. Sire differences in birth weight (BW) and dystocia score (32.9 vs 35.4 kg; 1.8 vs 3.1, L vs H sires, respectively) existed (P less than .01), and there was a sire effect (P less than .01) for fetal calf weights (FW) and eviscerated calf weights (EW). However, there was a sire X calf sex interaction for BW (P less than .05), EW (P less than .01), FW (P less than .01), femur length (P less than .05), heart weight (P less than .05), kidney weight (P less than .01) and pituitary weight (P less than .01). Weight differences suggested these interactions were a result of the relationship of the organ weights to fetal body weights and the interaction effects on calf weights resulted from limitations in the maternal environment which restricted growth of H-sired male calves in utero. Sire X fetal age interaction effects were all nonsignificant (P greater than .10) except for cerebrum weight. This finding indicates that fetus and calf growth rates were parallel for the L and H sires. A sire effect was found for biceps (P less than .01) and liver weights (P less than .01), but not for cerebrum weights (P greater than .10). Increasing weight due to fetal age was attributed to hypertrophy for the cerebrum (P less than .05) and liver (P approximately equal to .01), while the biceps increased (P less than .05) by both hypertrophy and hyperplasia, as determined from deoxyribonucleic acid and protein analyses. All measured fetal organ weights except heart, when expressed as a ratio with EW, decreased (P less than .05) with increasing fetal age. Brain (cerebrum + cerebellum):liver weight ratios were higher (P less than .01) in L-sired calves (.32 vs. .28) than in H-sired calves. Total placentome weight (b' = 91; P less than .01) and placental fluid volume (b' = .32; P less than .01) were highly associated with FW, accounting for 84% of the variation in FW.(ABSTRACT TRUNCATED AT 400 WORDS)     

Regulation of in vitro growth of preantral follicles by growth factors in goats

Goat preantral follicles were cultured to investigate the effects of insulin-like growth factor-I (IGF-I), epidermal growth factor (EGF) and basic fibroblast growth factor (bFGF) on the in vitro growth and viability of oocytes. Preantral follicles were isolated mechanically and enzymatically (using collagenase and DNase) from prepuberal goat ovaries. The working medium was composed of Defined Eagle's Minimum Essential Medium (DMEM) supplemented with HEPES (20 mM), 10% fetal calf serum (FCS), hypoxanthine (2 mM), dibutyryl cyclic adenosine 3',5'-monophosphate (dbcAMP) (2 mM), penicillin (75 ng/ml) and streptomycin (50 ng/ml). The culture medium consisted of the working medium with follicle stimulating hormone (FSH) (100 ng/ml) and hydrocortisone (40 ng/ml) added. In the experiment, goat preantral follicles were cultured for 9 days in the culture medium and in the culture medium supplemented with either IGF-I (100 ng/ml), EGF (50 ng/ml), bFGF (50 ng/ml) or IGF-I (100 ng/ml)+EGF (50 ng/ml). The results indicated that IGF-I (100 ng/ml) effectively maintained the survival of oocytes and promoted their growth; EGF (50 ng/ml) enhanced the survival rate of oocytes but had a negative effect on oocyte growth; bFGF (50 ng/ml) stimulated oocyte survival but had no obvious effect on their growth while IGF-I (100 ng/ml) and EGF (50 ng/ml) in combination had a greater effect on both survival and growth rate of oocytes than IGF-I or EGF alone. The supplementation of IGF-1 and EGF to the culture medium is recommended in the culture of goat preantral follicles.     

Expression of arginase by mouse myeloid leukemic cell differentiation in vitro induced with tumor necrosis factor.

Induction of arginase activity in mouse myeloid leukemic M1 cells by treatment with recombinant human tumor necrosis factor (rH-TNF) or TNF-elicited mouse serum (TNS) were examined in vitro. M1 cells differentiated into macrophage-like cells by addition of rH-TNF or TNS. The differentiated cells expressed phagocytic function and did not grow anymore. Cytolytic effect of rH-TNF or TNS was not observed. The differentiation of M1 cells into phagocytic cells by the TNS treatment was more rapidly than that by the rH-TNF treatment though TNS contained 25-time less amounts of TNF indicating species specificity of TNF action on myeloid leukemic cells or TNS containing other differentiation factor(s). 3H-ornithine formation from 3H-arginine is catalyzed by arginase (EC. 3.5.3.1). The enzyme product increased in the M1 cell culture medium by the treatment with rH-TNF or TNS. The arginase activity statistically correlated with the appearance percentage of differentiated cells with phagocytic function. These results suggest that in vitro differentiation of M1 cells is accompanied by induction of arginase activity.     

Effects of growth factors on development of fetal islet B-cells in vitro

To investigate the role of growth factors (epidermal growth factor [EGF], betacellulin, and activin A) in the development of islet B cells of rat fetal pancreatic explants in vitro, pancreases from rat fetuses at day 18 of gestation were cultured for 96 hr, with or without these growth factors. Culture medium was changed every 24 hr, and the level of insulin released in the culture medium was measured. After 72 hr of culture, pancreases were examined histologically. As a result, EGF promoted cell proliferation, but reduced B cell volume. Whereas, betacellulin and activin A inhibited cell division, but promoted increased B cell volume and insulin secretion, especially activin A, which stimulated insulin release in a time dependent manner. These results suggest that EGF, betacellulin, and activin A promote pancreatic cell proliferation, islet B-cell differentiation, and islet B-cell differentiation and functional maturation, respectively, and that EGF, betacellulin, and activin A, in this order, regulate islet B-cell neogenesis.     

Effect of recombinant porcine somatotropin on fetal and placental growth in gilts with reduced uterine capacity

Crowded uterine conditions were induced by unilateral hysterectomy-ovariectomy (UHO) in 42 gilts to determine the effect of recombinant porcine somatotropin on fetal and placental growth. Gilts were randomly assigned across three replicates to one of three treatments: Control (C; n = 14), daily injections of 1 mL saline from d 0 to 64 of gestation, Early (E; n = 12), 5 mg of rpST/d from d 0 to 30, followed by 1 mL saline from d 31 to 64, and Late (L; n = 16), 1 mL saline/d from d 0 to 29, followed by 5 mg of rpST/d from d 30 to 64 of gestation. Blood was collected from each gilt via jugular venipuncture at d 0 and every 15 d thereafter. Gilts were hysterectomized on d 65 of gestation. Length of placental attachment and fetal crown-rump length were measured. Placentas and fetuses were weighed. Placental length, wet weight, and dry weight were recorded. Treatment with rpST (either E or L) increased (P < 0.0001) maternal plasma IGF-I concentrations relative to controls. Treatment with rpST did not affect placental wet weight or placental DNA content. However, E and L treatments increased the percentage of placental protein (P = 0.01) and placental dry matter (P = 0.10) and increased contact area of uterine-placental interface (P = 0.01). Despite changes in placental composition and morphology, weights of fetuses collected from L-treated gilts did not differ from controls, whereas weights of fetuses collected from E-treated gilts tended to be less than controls (P < 0.06). Administration of rpST increased maternal IGF-I concentrations and placental surface area but failed to increase fetal growth in the UHO model. Therefore, mechanisms that are independent of maternal IGF-I or placental contact area may control early fetal growth under crowded uterine conditions.     

In vitro retinoid-induced growth inhibition and morphologic differentiation of canine osteosarcoma cells

OBJECTIVE: To determine differentiation and growth inhibition effects of retinoids on canine osteosarcoma cells. SAMPLE POPULATION: 3 osteosarcoma cell lines established from osteosarcomas in dogs. PROCEDURE: Osteosarcoma cells were incubated with various concentrations of all-trans-retinoic acid and 9-cis-retinoic acid or control medium, counted daily for 10 days, and evaluated for morphologic changes. Synthesis of DNA was measured by use of a cell proliferation ELISA. To analyze effect of retinoids on colony formation on plastic dishes, cells were cultured for 14 days, fixed, and stained; number of colonies was counted. RESULTS: In a dose-dependent manner, both retinoids induced morphologic differentiation and growth inhibition in the 3 osteosarcoma cell lines and inhibited each cell's ability to form anchorage-dependent colonies. CONCLUSION AND CLINICAL RELEVANCE: Retinoids induced differentiation of osteosarcoma cells of dogs, resulting in altered expression of their malignant phenotype. Induction of differentiation by retinoids may have potential as an adjunctive treatment for osteosarcoma in dogs.     

Production of mouse fibroblast growth factor 4 in E. coli and its application for isolation and maintenance of mouse trophoblast stem cells in vitro

Fibroblast growth factor 4 (FGF4) promotes isolation of trophoblast stem (TS) cells from mouse blastocysts and maintenance of TS cells in an undifferentiated state in vitro. To date, commercially available, bacterially expressed human FGF4 (RhFGF4) has been used generally for this purpose. In this study, HismFGF4, a 6x histidine-tagged mouse FGF4, was produced in E. coli and purified using heparin column chromatography. We demonstrated that HismFGF4 (25 ng/ml) more efficiently generates mouse TS cells from a single blastocyst than RhFGF4 (25 ng/ml) and that TS cells isolated and maintained with HismFGF4 retained their ability to differentiate into the trophoblast cell lineage in vitro. In addition, TS cells cultured with HismFGF4 (25 ng/ml) were maintained in an undifferentiated state better than with RhFGF4 (25 ng/ml). To the best of our knowledge, this is the first application of a mouse FGF4 derivative for isolation and maintenance of mouse TS cells.     

干旱胁迫对多年生落花生生长及溶质累积的影响

采用盆栽控水的方法,研究了不同土壤干旱胁迫程度与胁迫时间对多年生落花生Arachis duranensis生长及溶质累积的影响。结果表明:轻度的土壤干旱处理(60%~65%的田间持水量)有利于多年生落花生的生长,而中度及以上程度的土壤干旱(≤45%的田间持水量)影响它生长及其作为草坪草的观赏价值;其游离脯氨酸和可溶性糖含量在中度及以上程度土壤干旱胁迫下均有不同程度的上升,而且随着干旱胁迫时间的延长,在多年生落花生体内有明显的累积趋势。     

成纤维细胞生长因子信号通路在小鼠牙齿发育中的表达及其作用

小鼠牙齿发育是一个外胚层来源的上皮和神经嵴来源的间充质相互作用的过程.在这个相互作用过程中,成纤维细胞生长因子(FGFs)信号通路在牙齿发生位置的确定,牙齿发育的起始、细胞的增殖分化和牙齿的形态建成等方面都发挥着重要作用.该家族由23个配体(Fgf1-23)组成,通过4种特异性受体(FGFR1-4)来发挥作用.本文综述FGFs信号通路的配体和受体在小鼠牙齿发育中具体的表达模式及其作用.