Cell-free extract from porcine induced pluripotent stem cells can affect porcine somatic cell nuclear reprogramming

Pretreatment of somatic cells with undifferentiated cell extracts, such as embryonic stem cells and mammalian oocytes, is an attractive alternative method for reprogramming control. The properties of induced pluripotent stem cells (iPSCs) are similar to those of embryonic stem cells; however, no studies have reported somatic cell nuclear reprogramming using iPSC extracts. Therefore, this study aimed to evaluate the effects of porcine iPSC extracts treatment on porcine ear fibroblasts and early development of porcine cloned embryos produced from porcine ear skin fibroblasts pretreated with the porcine iPSC extracts. The Chariot^TM reagent system was used to deliver the iPSC extracts into cultured porcine ear skin fibroblasts. The iPSC extracts-treated cells (iPSC-treated cells) were cultured for 3 days and used for analyzing histone modification and somatic cell nuclear transfer. Compared to the results for nontreated cells, the trimethylation status of histone H3 lysine residue 9 (H3K9) in the iPSC-treated cells significantly decreased. The expression of Jmjd2b, the H3K9 trimethylation-specific demethylase gene, significantly increased in the iPSC-treated cells; conversely, the expression of the proapoptotic genes, Bax and p53, significantly decreased. When the iPSC-treated cells were transferred into enucleated porcine oocytes, no differences were observed in blastocyst development and total cell number in blastocysts compared with the results for control cells. However, H3K9 trimethylation of pronuclear-stage-cloned embryos significantly decreased in the iPSC-treated cells. Additionally, Bax and p53 gene expression in the blastocysts was significantly lower in iPSC-treated cells than in control cells. To our knowledge, this study is the first to show that an extracts of porcine iPSCs can affect histone modification and gene expression in porcine ear skin fibroblasts and cloned embryos.     

初情期前母猪垂体细胞LH和FSH的释放及其对GnRH类似物的反应

利用垂体细胞单层培养模型研究了30，60，90，120和150日龄北京黑猪母猪垂体细胞LH和FSH的释放及其对LRH－A3反应能力。结果表明，猪垂体细胞LH和FSH释放对LRH－A3的反应呈S形剂量依赖型同线。LH基础释放量和LRH－A3刺激的最大释放量在不同日龄间没有显著差异，说明初情期前母猪垂体细胞已达最大LH释放和对GnRH反应的能力。FSH的基础释放量和LRH－A3刺激的最在释放量在30日龄与60日龄之间无差异，但90日龄以后垂体细胞FSH的释施量随日龄增大而降低，提出可能由于垂体在体内时受过抑制素的作用。     

不同培养液对体外培养的小鼠胸腺上皮细胞增殖的影响

分离纯化小鼠胸腺上皮细胞（mTEC）,以DMEM、DMEM/F12、RPMI 1640 3种基础培养液加10%胎牛血清完全培养液进行培养, MTT法和流式细胞术分别检测mTEC的增殖及细胞周期,筛选出最适合mTEC生长的培养液。结果表明,DMEM培养的mTEC增殖快,细胞增殖指数明显高于DMEM/F12、RPMI 1640培养液,表明DMEM是mTEC体外培养的最佳培养液。     

High‐level exogenous trans10, cis12 conjugated linoleic acid plays an anti‐lipogenesis role in bovine mammary epithelial cells

Primary bovine mammary epithelial cells (BMECs) were treated by 0, 37.5, 75, 112.5, 150 μmol/L trans10, cis12 conjugated linoleic acid (CLA) to evaluate the effects of different level trans10, cis12 CLA on lipogenesis in BMEC. Addition of 75–150 μmol/L trans10, cis12 CLA reduced significantly the triacylglycerol (TAG) content (P < 0.05), but did not have inhibiting action on cell proliferation (P > 0.05). Treatment with 150 μmol/L trans10, cis12 CLA for 48 h resulted in a 17.1% reduction (P < 0.0001) of medium chain fatty acids (MCFA, C14 < C < C16), a 26.5% reduction (P < 0.0001) of unsaturated fatty acids (UFA) and a corresponding reduction of the mRNA abundance of acetyl coenzyme A (acetylCoA) carboxylase (ACC) (P = 0.046), fatty acid synthase (FAS) (P = 0.017) and stearoylCoA desaturase1 (SCD1) (P = 0.002). Another finding was that trans10, cis12 CLA elevated expression of diacylglycerol acyltransferase2 (DGAT2) (P = 0.020) and long chain acylCoA synthetases (ACSL) (P = 0.032). In conclusion, higher trans10, cis12 CLA, not low trans10, cis12 CLA, inhibited milk fat synthesis and changed fatty acid composition by regulating the expression of FAS, ACC, SCD1, DGAT2 and ACSL.     

Localization of putative pituitary stem/progenitor cells in female dairy cattle

Research on sex-determining region Y-box 2 (SOX2)-positive pituitary stem/progenitor cells, as a source of hormone-producing cells, is progressing rapidly in rodents. However, the stem/progenitor cells supplying hormone-producing cells that are essential for growth, reproduction, and lactation in bovines have not yet been identified. In this study, we characterized SOX2-positive cells in the pituitary gland of dairy cattle (Holstein heifers) after sexual maturity. Immunofluorescence analysis revealed that the localization pattern of SOX2-positive cells in the dairy cattle pituitary gland was similar to that observed in the rodent pituitary gland; the marginal cell layer (MCL), dense cell clusters, and single cells scattered in the parenchyma of the anterior lobe. Furthermore, most of the SOX2-positive cells were positive for the pituitary stem/progenitor cell niche markers E-cadherin and cytokeratin 8+18, which have been reported in rodents. In addition, in the MCL of the anterior lobe, there was a subpopulation of SOX2-positive cells positive for paired-related homeobox 1 and 2, whereas negative for S100β. Moreover, in the parenchyma of the anterior lobe, co-localization of SOX2 and pituitary hormones was infrequent. In summary, this study reveals the localization of putative pituitary stem/progenitor cells positive for SOX2 in dairy cattle. These results provide valuable information to support further investigation of cell supply in the dairy cattle pituitary gland.     

摩杂一代水牛脑垂体前叶的超微结构研究

利用超薄切片对摩杂一代水牛脑垂体前叶的腺细胞进行透射电镜观察,发现5种腺细胞各有其超微结构特征：促性腺激素细胞胞体呈圆形,直径为7.6～9.0 μm;分泌颗粒分为明显的大小2种颗粒,且电子密度高。促甲状腺激素细胞的直径为6.0～7.5 μm;分泌颗粒少而最小,且大小无显著差异,多沿质膜分布;细胞核小,多位于中央。促肾上腺皮质激素细胞多在邻近垂体神经部的腺体前叶中,细胞核不规则,常偏向一侧,分泌颗粒常于细胞周边或一端;胞体直径为7.1～8.5 μm。促生长激素细胞数量多,核大而圆,分泌颗粒多且均一,充满胞质。催乳素细胞胞体最大,直径为11.2～13.4 μm,数量多,核小,常偏于一侧;分泌颗粒多而大小不等,充满胞质。旨在为研究摩杂一代水牛生殖泌乳的内分泌调控机制提供形态学基础资料。     

不同苜蓿品种在青岛地区的适应性

2007-2010年,在青岛地区对引进的23个国内外苜蓿品种进行了膜侧种植、无灌溉、无施肥模式的田间试验,观测分析了各品种产草量、株高、茎叶比等指标。结果表明,23个苜蓿品种在青岛普遍生长良好,国外品种生产性能优于国内品种;播种当年干草产量膜侧种植组明显高于直接播种组,干草产量增幅(除甘农4号、新疆大叶和牧歌401外)均在50%以上;干草产量与年均日照时数、年均温的灰色关联度分别达到0.795 5和0.878 9,而与年均降水量关联度仅为0.569 1;株高最高的是WL323为87.8 cm,最低的是敖汉苜蓿,仅74.7 cm;供试23个品种的干草产量与株高均存在显著性差异,株高与草产量相关系数达到0.65以上,具有很高的正相关性;茎叶比最低的是牧歌401,仅为1.22,绝大多数苜蓿品种的茎叶比有逐年增加的趋势。综合评价认为供试23个苜蓿品种中的WL323、牧歌401、驯鹿3个品种综合生产性能较好,适宜在青岛地区辅以膜侧种植模式进行大面积推广。     

Radiosensitivity of canine osteosarcoma cells transfected with wild‐type p53 in vitro*

The p53 gene is one of the important tumour suppressor genes that are involved with the cell survival signal pathway. One of the major functions of the p53 protein is to organize cell cycle regulation and induction of apoptosis for cellular genetic stability. It has been documented that more than 50% of all human cancers include a p53 mutation. We evaluated the difference in radiosensitivity between upregulating the expression of canine wild‐type p53 (cp53) in cultured osteosarcoma (D17) cells and naive D17 cells in vitro. We found that upregulating transfected cp53 D17 cells increased their radiation sensitivity in vitro, and there was a significant decrease (P < 0.009) in survival between cp53‐transfected D17 cells and naive D17 cells. In this experiment, a p53 enhancement ratio (p53ER) reached approximately 3.0 at high doses. The transfected cp53 D17 cells were significantly more radiosensitive at all doses evaluated than naive D17 cells, except at 1 Gy where too few data points were available. The p53ER increased rapidly at doses less than 4 Gy, achieving a maximum of about 3.0 for doses of 4 Gy and above. This study shows the enhanced radiosensitivity of the transfected p53 at clinically relevant doses.