ORIGINAL ARTICLE: Inulin‐coated butyrate increases ileal MCT1 expression and affects mucosal morphology in the porcine ileum by reduced apoptosis

Carbohydrates, which were not digested in the jejunum, will be fermented by micro‐organisms to short chain fatty acids. These are transported by the monocarboxylate transporter 1 (MCT1) through the gut wall and serve as fuels for colonic cells. To deliver butyrate to the distal part of the intestine, inulin with a low precaecal digestibility was chosen as a coating material. Approximately 150 g of inulin‐coated butyrate (containing 81 g butyrate) per day was fed to pigs (mean weight: 97 kg) over a period of 6 days after an adaptation period of 6 days with linear increasing amounts of butyrate. The following observations compared to controls were observed: (1) coating was digested microbially in the ileum; (2) MCT1‐mRNA showed a higher expression in the ileum; (3) apoptosis was reduced in the ileum but mitosis was not changed; and (4) length of villi increased by approximately 25% in the ileum. Feeding inulin‐coated butyrate resulted in an increased ileal surface. Delivery of butyrate to the colon requires a more resistant inulin‐coating.     

成年猪结肠肥大细胞的分布

成年猪结肠按常规制备石蜡切片,用甲苯胺蓝改良法显示肥大细胞.结果粘膜型肥大细胞(MMCs)主要分布于固有膜深层,不同部位结肠粘膜中MMCs的数量存在极显著差异(P＞0.01),结缔组织型肥大细胞(CTMCs)主要分布于粘膜下层且不存在数量上的区域性差异(P＞0.05).结论猪结肠MMCs的分布类型具有区域异质性特征.     

Influence of weaning age on the villous height and disaccharidase activities in the porcine small intestine

Body weight gain after weaning is correlated with villous height and disaccharidase activity. This evidence suggests that the maintenance of the small intestinal structure and function after weaning is important for the growth of piglets. We demonstrated that the influence of weaning age was obtained by disaccharidase activities and villous height in eight sections of the porcine small intestine. Therefore, we designed three weaning ages (weaned at the ages of 14, 21 or 28 days) and the piglets were slaughtered after 7 or 14 days post‐weaning. The remaining suckling piglets were slaughtered at the age of 1, 7, 14, 21 and 28 days. Four piglets were slaughtered at each event; therefore, 44 piglets were used in this study. Villous height and disaccharidase activities were measured in each section of the small intestine. Early weaning such as that at 14 days had severe influence on villous and disaccharidase activities. In particular, weaning of 14‐day‐olds did not result in maltase activity at least 2 weeks post‐weaning. Accordingly, the weaning age of crossbred piglets is recommended to be at least 21 days or more on the basis of villous height and disaccharidase activity analyses.     

利用大规模F_2及F_3群体研究猪小肠长度及其生长发育性状的相关性

小肠是猪的主要消化吸收器官,对个体的生长发育非常重要,但到目前为止,国内外对猪小肠的研究还很少。为探讨其长度与生长发育性状之间的关系,本试验测定了白色杜洛克×二花脸资源家系F2和F3群体共计1 508头个体的240日龄小肠长度、出生重、21日龄体重、46日龄体重、0~46日龄平均日增重、46~240日龄平均日增重、240日龄平均日增重、240日龄宰前活重及屠宰后胴体直长、胴体斜长等10项生长性状指标,并对其与小肠长度的相关性进行了分析。结果显示猪240日龄小肠长度与这些生长性状都存在极显著相关(P0.001),表明小肠可能通过影响个体对营养成分的消化吸收而影响个体的生长发育。据我们所知,本试验是首次在大规模样本群体中研究猪小肠长度与生长发育性状的相关性,其结果为小肠和个体发育研究提供了重要参考。     

山羊乳腺上皮细胞的生长特性

用组织块培养法获得山羊乳腺细胞原代培养物。根据山羊乳腺成纤维细胞与上皮细胞对胰蛋白酶的敏感性不同将二者分离纯化，对细胞生长特性进行了光镜观察。结果如下：细胞可形成闭合型细胞群和开放型细胞群。乳腺上皮细胞与成纤维细胞混生时，细胞之间形成许多腔状结构。纯化的乳腺上皮细胞通过单细胞悬浮后传代，部分细胞形成岛屿状聚集，部分细胞以贴壁的自由单个细胞散在形式存在。上皮细胞增殖可形成圆顶型结构，呈乳头状，称之为乳球体；可产生乳腺上皮细胞并分泌乳汁。山羊乳腺上皮细胞含不同的细胞类型．大多数上皮细胞呈短梭形或多角形。细胞之间紧密相靠。互相衔接。连接成片，呈蜂窝状；细胞核呈圆形或椭圆形．核仁2～4枚；部分细胞呈圆饼状，体积较大；出现部分长形细胞；接触抑制的上皮细胞形态不均一。纯化的山羊乳腺上皮细胞传代至第15代时其生长仍正常，经透射电镜观察发现细胞表面微绒毛极为发达，细胞质中线粒体和粗面内质网丰富，细胞质内有大量脂滴及小泡，表明第15代山羊乳腺上皮细胞增殖活力旺盛。染色体数目分析表明．该细胞系稳定，在离体培养条件下细胞不发生转化。山羊乳腺上皮细胞系的细胞染色体数目为60．染色体组型为2n=60。     

Investigations on the conjunctival goblet cells and on the characteristics of glands associated with the eye in the guinea pig

Objective To investigate the distribution and density of conjunctival goblet cells (GC) and to study the anatomy and microscopic characteristics of glands associated with the eye in the guinea pig. Procedures Twenty‐five guinea pigs were used. Meibomian gland openings were counted using biomicroscopy. Conjunctiva, eyelids and glands were embedded in glycol methacrylate and paraffin. Sections were stained with hematoxylin and eosin (H&E), periodic acid Schiff’s reaction (PAS) and Alcian blue (AB). Results Highest GC densities were found in the bulbar and palpebral region of the nasal conjunctiva (GC index: 13.7–16.4%). Lowest GC densities (GC index: 0.0–1.0%) were found in 3/4 limbal regions (nasal and temporal upper eyelid, temporal lower eyelid). Guinea pigs have 27.1 ± 3.0 (mean ± SD) meibomian gland openings in the upper lid and 25.7 ± 2.3 in the lower lid. Difference between upper and lower lid was significant (P = 0.037). Two subconjunctival sebaceous glands occur temporal to each eye. The Harderian gland is very large. In the lacrimal gland three different cell types were distinguished both according to the cell structure and histochemical staining. Conclusions Goblet cell densities are lower in guinea pigs than in dogs and horses. Positive staining with PAS and AB could be an indication that mucins are produced in the lacrimal gland. If so, they may contribute to the mucin layer of the tear film. Both the extraordinarily large Harderian gland and the subconjunctival sebaceous glands produce lipids and may contribute to the lipid layer of the tear film.     

Comparison of effects of leptin and ghrelin on porcine ovarian granulosa cells

The aim of our studies was to compare the roles of leptin and ghrelin in the direct control of proliferation, apoptosis, and secretory activity by porcine ovarian cells. In our in vitro experiments, we analyzed the effects of leptin and ghrelin treatments (at 0, 1, 10, or 100 ng/mL medium) on the accumulation of proliferation-related peptides (PCNA, cyclin B1, MAP kinase [MAPK]) and apoptosis-associated peptides (Bax, caspase 3, p53), and on progesterone secretion by cultured porcine granulosa cells, using immunocytochemistry, SDS PAGE-Western immunoblotting, and radioimmunoassay (RIA). Leptin stimulated proliferation (PCNA, cyclin B1, MAPK), apoptosis (Bax, p53), and progesterone secretion. Ghrelin promoted proliferation (PCNA, cyclin B1, MAPK) and progesterone secretion but suppressed apoptosis (Bax, caspase 3, p53). These observations suggest that both leptin and ghrelin directly control proliferation, apoptosis, and secretory activity by porcine ovarian cells. At the level of the ovary, in contrast to the hypothalamo-hypophysial system, leptin and ghrelin may have similar action in promoting granulosa cell proliferation and progesterone secretion, but they may be antagonistic to one another (leptin, stimulator; ghrelin, inhibitor) in controlling apoptosis.     

Stimulation of butyrate production through the metabolic interaction among lactic acid bacteria, Lactobacillus acidophilus, and lactic acid-utilizing bacteria, Megasphaera elsdenii, in porcine cecal digesta

In this study, we examined the potential of Megasphaera elsdenii as a probiotic agent in combination with sodium gluconate. We also examined the combination of M. elsdenii with probiotic lactic acid bacteria (LAB), Lactobacillus acidophilus, to enhance butyrate production in the large intestine using a pig cecal in vitro model under the presence of sodium gluconate. Compared to the uninoculated control culture (with only diluted cecal digesta), the addition of M. elsdenii and LAB to the culture stimulated significantly the production of n‐butyrate (60% increase) and n‐valerate (50% increase) after 24 h incubation. n‐Butyrate is regarded as the most important short‐chain fatty acid in the large intestine because it stimulates the proliferation of epithelial cells, mucus release and water and mineral absorption. Therefore, lactate‐utilizing butyrate producers such as M. elsdenii play a significant role in supporting the health‐beneficial effects of lactogenic prebiotics and probiotics LAB. The potential of M. elsdenii as a probiotic was also suggested.     

年龄差异对家兔CD4+T细胞在胃肠分布的影响

为深入了解家兔消化道中CD4⁺T细胞的组织学定位以及在消化道不同解剖学部位的数量变化,本试验运用免疫组织化学方法,探讨了家兔胃(胃底腺区、幽门腺区)和小肠及大肠不同解剖学部位中的CD4⁺T细胞定位特点和数量变化规律。结果表明,在胃肠中,CD4⁺T细胞主要分布于固有层。胃底腺区阳性细胞数目要明显比幽门腺区少。小肠、大肠各段的阳性细胞分布数量未见明显差异,由十二指肠到回肠表现为小幅度递增,由盲肠到直肠表现为小幅度递减。胃肠中CD4⁺T细胞在各部位的数量随年龄增长表现出先增后减的变化趋势,老年组和幼年组CD4⁺T细胞数量在大肠中未见明显差异。结果提示,CD4⁺T细胞在青年家兔胃肠中的分布最为丰富。     

贵州香猪睾丸发育中支持细胞和生精细胞数量变化观察

为了解香猪睾丸发育过程中生殖细胞和支持细胞数变化规律,用手术取出30,40,50,70,90和110日龄(每个年龄组n=3～4)香猪右侧睾丸,经中性多聚甲醛固定,石蜡包埋,组织切片采用免疫组化SP法,用单克隆抗体GATA-4检测睾丸支持细胞的特异生长转录因子-4,经DAB显色、苏木素复染。光镜下核呈棕色者为支持细胞,核呈蓝色者则为生殖细胞;经显微照相并用Scion image软件测量生精小管及管壁面积。结果:30～110日龄睾丸支持细胞数维持在稳定水平(P>0.05),而生殖细胞数随日龄增加而增多,70日龄生殖细胞数快速增多(P<0.05),持续到110日龄。同样,从70日龄开始睾丸生精小管和管壁面积显著性增大(P<0.05)。香猪睾丸支持细胞快速增殖发生在30日龄前,而生殖细胞数随着日龄的增长而增多。     

Effects of 7S and 11S on the intestine of weaned piglets after injection and oral administration of soybean antigen protein

Soybeans are used increasingly in food products because of their health benefits. In this study, we investigated the effect of soybean antigen protein on weaned piglet intestine. Seventy piglets were randomly divided into seven groups with 10 piglets each. At 7 and 14 days of age, groups A–C were injected with saline, and D–G were intramuscularly injected with or orally administered 7S or 11S. Groups B–G were artificially sensitized by dietary 7S or 11S. At 27 days, the small intestinal tissues were collected to determine levels of histamine, sIgA protein, and IgA mRNA. Histamine in B–G was significantly decreased in the duodenum and ileum. Moreover, sIgA expression was higher in all groups than in A, with B/C>D–G and F/G>D/E; the trend in IgA expression was similar. Collectively, these results indicated that soybean antigen protein‐immunizing agents decrease sIgA and IgA levels. Additionally, the effect of injection immunization occurred prior to that of oral immunization.     

撒坝猪成纤维细胞系的建立及其生物学特性分析

实验采用撒坝猪耳缘组织块贴壁培养法首次对撒坝猪成纤维细胞进行体外培养,通过原代培养、传代培养、冷冻保存建立了撒坝猪耳缘成纤维细胞系;并对其细胞形态、细胞活率、细胞核型等生物学特性进行分析;还利用透射电镜观察了耳缘组织细胞。结果表明:细胞倍增时间为48 h,生长曲线为"S"型;染色体中正常二倍体染色体(2n=38)所占比例为92.56%;细菌、真菌、病毒和支原体等微生物污染检测均显示为阴性。透射电镜观察显示,成纤维细胞呈长梭型。     

Effects of fermentable carbohydrates and low dietary phosphorus supply on the chemical composition of faecal bacteria and microbial metabolites in the gastrointestinal tract of pigs

The objective was to determine the effects of fermentable carbohydrates on phosphorus (P) metabolism, the chemical composition of the faecal mixed bacterial mass (MBM) and the microbial activity in the large intestine (LI) of pigs. Eight barrows (mean BW 35.9 ± 0.9 kg), fitted with simple T‐cannulas at the terminal ileum, were either fed a low‐P corn–soybean meal‐based control diet or 75% of the control diet supplemented with 25% cellulose, starch or pectin according to a 4 × 3 Latin Square design. Both pectin and cellulose caused higher faecal than ileal P recoveries. Ileal volatile fatty acids (VFA) levels were more pronounced for the starch (p < 0.05) rather than the cellulose and pectin treatments, whereas pectin resulted in a higher faecal VFA concentration in comparison to starch and cellulose (p < 0.05). The differences in faecal VFA concentrations corresponded to the pH values obtained in faeces. The N content of MBM was higher (p < 0.05) when cellulose was supplemented. Pectin caused a decrease in the P content of the MBM compared to the control (p < 0.05). As a result, the N:P ratio was significantly higher for the pectin (N:P = 4.33) than for the control treatment (N:P = 2.63), while the Ca:P ratio remained constant for all treatments, suggesting changes in the accumulation of N, P and Ca in MBM, probably due to changes in the species composition and activity of the microflora.     

猪骨骼肌卫星细胞中miR-24功能的初步研究及靶基因分析

为了探讨miR-24对猪骨骼肌发育的影响,对体外培养1日龄纯种长白公猪骨骼肌卫星细胞分别转染miR-24过表达载体和干扰片段,发现过表达miR-24后,成肌分化基因MyoD、Myogenin和Myf5表达量显著升高,骨骼肌卫星细胞分化趋势增强;干扰miR-24后,Myogenin的表达量显著降低;同时,靶基因预测结果显示Myogenin可能是miR-24的靶基因。     

Influences of protein ingestion on glucagon‐like peptide (GLP)‐1‐immunoreactive endocrine cells in the chicken ileum

Influences of a specific dietary nutrient on glucagon‐like peptide (GLP)‐1‐containing cells in the chicken intestine are not yet clear. Significance of dietary protein level on GLP‐1‐containing cells in the chicken ileum was investigated. Chickens fed control or experimental diets of varying protein levels were examined using immunohistochemical and morphometrical techniques. We show that the protein ingestion had an impact on the activities of GLP‐1‐immunoreactive cells in the chicken ileum. Weight gains declined with decreasing dietary crude protein (CP) levels, but no significant differences were detected in the daily feed intake and villous height. GLP‐1‐immunoreactive cells with a round or oval shape were frequently observed in the lower CP level groups (4.5% and 0%). Frequencies of occurrence of GLP‐1‐immunoreactive cells were 41.1 ± 4.1, 38.5 ± 4, 34.8 ± 3.1 and 34.3 ± 3.7 (cells/mm², mean ± SD) for dietary CP level of 18%, 9%, 4.5% and 0% groups, respectively and significant differences were recognized between the control and lower CP level groups (P < 0.05). Multiple regression analysis indicated a significant correlation between the daily protein intake and frequencies of occurrence of GLP‐1‐immunoreactive cells. The protein ingestion is one of the signals that influence GLP‐1‐containing cells in the chicken small intestine.     

Associations of the variation in the porcine myogenin gene with muscle fibre characteristics, lean meat production and meat quality traits

Pig breeding is aimed at improving lean meat production ability as well as meat quality, and muscle fibre characteristics may be important for enhancing these traits. Therefore, new molecular markers have been demanded for selecting lean meat production ability and meat quality in live animals. Myogenin belongs to the MyoD gene family, and is a candidate gene responsible for muscle fibre characteristics. We identified a new single nucleotide polymorphism (SNP) site in the 5' upstream region of the myogenin gene (nucleotides C and T). A total of 252 pigs of three breeds were genotyped by polymerase chain reaction–restriction fragment length polymorphism using BspCNI . Additionally, they were genotyped for the previously detected Msp I site in the 3'-flanking region (alleles A and B). The CCBB diplotype had the highest frequency over breeds, followed by TCBB and CCAB. The other diplotypes were not found in studied pigs. Association analysis performed for the markers found that the TCBB diplotype has desirable effects on the total number of fibres (p   < 0.002), fibre cross-sectional area (p   < 0.06), and loin eye area (p   < 0.001) than the other diplotypes. Moreover, the diplotype had the highest muscle pH value (p   < 0.07) and all meat quality traits were near the upper limit of the normal range as a reddish pink, firm and non-exudative (RFN) pork. Therefore, we suggest that selection for the myogenin diplotypes could improve total muscle fibre number, size and lean meat production ability with good meat quality.     

Generation of embryonic stem‐like cells from in vivo‐derived porcine blastocysts at a low concentration of basic fibroblast growth factor

Although basic fibroblast growth factor (bFGF) is an essential factor supporting the maintenance of porcine embryonic stem (ES) cell self‐renewal and pluripotency, its high cost has limited previous studies, and the development of a low‐cost culture system is required. For these systems, in vivo blastocysts were progressively cultured under various conditions consisting of different culture mediums and/or different feeder cell numbers at a low concentration of bFGF. As the results, the sequential culture of in vivo‐derived porcine blastocysts on 5.0 × 10⁵ mouse embryonic fibroblast (MEF) feeder cells in alpha minimum essential medium‐based medium for primary culture, on 2.5 × 10⁵ MEF feeder cells in Mixture medium for the 1st subpassage, and on 2.5 × 10⁵ MEF feeder cells in DMEM/Ham's F10‐based medium for the post‐2nd subpassage could support the establishment and maintenance of porcine ES‐like cells at the low concentration of bFGF. The established porcine ES‐like cells showed ES cell‐specific characteristics such as self‐renewal and pluripotency. We confirmed that porcine ES‐like cells could be generated from in vivo‐derived porcine blastocysts at a low concentration of bFGF.     

Effects of leptin and leptin peptide amide on the release of luteinizing hormone, growth hormone and prolactin from cultured porcine anterior pituitary cells

The present study was carried out to determine whether leptin or leptin (116–130) peptide amide (lep (116–130)), an active fragment of the native protein in rats, is able to stimulate the release of luteinizing hormone (LH), growth hormone (GH) or prolactin (PRL) from cultured porcine anterior pituitary (AP) cells in vitro. The AP cells were obtained from 6 month‐old pigs and were incubated for 3 h with 10^?11?10^?7 mol/L leptin or lep (116–130) after being cultured in Dulbecco's modified Eagle's medium for 3–4 days. Leptin significantly increased the concentration of LH and GH in the culture medium at concentrations of 10^?8 and 10^?7 mol/L, respectively, compared with the controls (P < 0.05). Leptin did not increase the concentration of PRL in the culture medium. In contrast to these results, no effects of lep (116–130) on the release of LH, GH or PRL were seen in the cultured cells. These results suggest that leptin stimulates the release of LH and GH by acting directly on porcine AP cells, and that a fragment of leptin protein comprising amino acids 116–130 is not associated with the secretion of hormones in pigs.