Expression of monocarboxylate transporter 1 (MCT1) in the dog intestine

In this study, the expression and distribution of monocarboxyolate transporter 1 (MCT1) along the intestines (duodenum, jejunum, ileum, cecum, colon and rectum) of dogs were investigated at both the mRNA and protein levels. The expression of MCT1 protein and its distribution were confirmed by Western blotting and immunohistochemical staining using the antibody for MCT1. We identified mRNA coding for MCT1 and a 43-kDa band of MCT1 protein in all regions from the duodenum to the rectum. Immunoreactive staining for MCT1 was also observed in epithelial cells throughout the intestines. MCT1 immunoreactivity was greater in the large intestine than in the small intestine. MCT1 protein was predominantly expressed on the basolateral membranes along intestinal epithelial cells, suggesting that MCT1 may play an important role in lactate efflux and transport of short-chain fatty acids (SCFAs) to the bloodstream across the basolateral membranes of the dog intestine.     

Expression of mRNA for sodium-glucose transporter 1 and fatty acid translocase in the ruminant gastrointestinal tract before and after weaning

The mRNA expression of sodium‐glucose transporter 1 (SGLT1) and fatty acid translocase (CD36) in the gastrointestinal tract of Holstein cattle and Saanen goats before and after weaning was investigated. Before weaning, the expression of both SGLT1 and CD36 was highest in the jejunum, relative to the other parts of the gastrointestinal tract in both species. The expression of SGLT1 and CD36 in the duodenum was second highest in the goats. After weaning, SGLT1 and CD36 expression in the small intestine significantly decreased in both species. The expression of both types of transporters was also detected in the forestomach. From these results, it was concluded that the jejunum is probably the major absorption site for glucose and long‐chain fatty acids before weaning, and that the expression of both types of transporters decreases after weaning in cattle and goats.     

GHSR—1a在奶山羊胃肠道的分布与表达

试验采用免疫组织化学、Real—timePCR和Western blotting方法测定ghrelin的功能性受体GHSR-1a（Growth hormone seeretagogue receptor-1a,GHSR-1a）在奶山羊胃肠道的分布和表达。免疫组织化学结果显示,GHSR—1a免疫阳性细胞广泛分布于奶山羊胃肠道。在皱胃主要定位于黏膜层和肌层;瘤胃、网胃和瓣胃黏膜层及肌层中也可见GHSR-1a免疫阳性细胞;在小肠主要位于十二指肠、空肠和回肠的黏膜层、黏膜下层和肌层;在结肠、盲肠和直肠GHSR—1a免疫阳性细胞也有广泛分布;GHSR—1a主要表达于内在神经丛神经细胞、胃底腺上皮细胞、肠腺上皮细胞、复层鳞状上皮细胞、平滑肌细胞中。real—timePCR和Westernblotting结果显示,皱胃、十二指肠、空肠、回肠、盲肠、结肠和直肠GHSR—1a的表达水平相对较高,显著高于瘤胃、网胃和瓣胃的表达（P〈0．05）。结果表明,ghrelin可能通过GHsR-1a对奶山羊胃肠功能具有重要的调节作用。     

Expression of KA1 kainate receptor subunit in the substantia gelatinosa of the trigeminal subnucleus caudalis in mice

The KA1 kainate receptor (KAR) subunit in the substantia gelatinosa (SG) of the trigeminal subnucleus caudalis (Vc) has been implicated in the processing of nociceptive information from the orofacial region. This study compared the expression of the KA1 KAR subunit in the SG of the Vc in juvenile, prepubescent and adult mice. RT-PCR, Western blot and immunohistochemistry analyses were used to examine the expression level in SG area. The expression levels of the KA1 KAR subunit mRNA and protein were higher in juvenile mice than in prepubescent or adult mice. Quantitative data revealed that the KA1 KAR subunit mRNA and protein were expressed at levels approximately two and three times higher, respectively, in juvenile mice than in adult mice. A similar expression pattern of the KA1 KAR subunit was observed in an immunohistochemical study that showed higher expression in the juvenile (59%) than those of adult (35%) mice. These results show that the KA1 KAR subunits are expressed in the SG of the Vc in mice and that the expression level of the KA1 KAR subunit decreases gradually with postnatal development. These findings suggest that age-dependent KA1 KAR subunit expression can be a potential mechanism of age-dependent pain perception.     

Evaluation of safety and efficacy of DNA vaccines against bovine herpesvirus-1 (BoHV-1) in calves

Four DNA vaccines against BoHV-1 were evaluated for their efficacy in calves. Twelve animals were divided into four groups which were injected with four different DNA vaccines: pVAX-tgD (Vaccine A); pVAX-tgD co-immunised with pVAX-48CpG (Vaccine B); pVAX-UbiLacI-tgD-L (Vaccine C); pVAX-UbiLacI-tgD-L co-immunised with pVAX-48CpG (Vaccine D). Three additional calves were given the plasmid vector and served as controls. Ninety days after the first vaccination all calves were challenge infected with BoHV-1.All animals developed a severe form of infections bovine rhinotracheitis. Only the calves given the pVAX-tgD co-immunised with pVAX-48CpG (Vaccine B) developed humoral antibodies against BoHV-1 between 56 and 90 days after the first vaccination, whereas in calves of other groups and in the controls, antibodies appeared only after the infection. In the calves vaccinated with either pVAX-tgD (Vaccine A) or pVAX-tgD combined with pVAX-48CpG (Vaccine B), BoHV-1-specific IFN-γ secreting cells were detected in PBMCs 90 days after the first vaccination and their number increased after challenge exposure. In the other groups the IFN-γ secreting cells were detected after virus infection and at low values.     

Expression of CD147 and monocarboxylate transporters MCT1, MCT2 and MCT4 in porcine small intestine and colon

Lactate, formed mainly in the stomach and small intestines, and short-chain fatty acids (SCFAs) formed in the colon, are ionised and require transporter proteins such as monocarboxylate transporters (MCTs) for absorption. The amounts of MCT1, MCT2, MCT4 and CD147, an ancillary protein for MCT1 and MCT4, were measured by immunoblotting the small intestine and colon of 40 pigs (Landrace, Yorkshire and LandracexYorkshire). MCT1 and MCT4 were found in both small intestine and colon, but MCT2 only in the small intestine. In both small intestine and colon, Yorkshire pigs had more CD147 than Landrace pigs, while no interbreed differences were found in MCT isoforms. Since CD147 is essential for the activity of MCT1 and MCT4, the breed difference suggests that MCT activity is higher in Yorkshire than in Landrace pigs. The absence of MCT2 in the colon suggests that it is mainly a lactate transporter, while MCT1 and MCT4 facilitate the transport of both lactate and SCFA.     

MCT1在犊牛消化道的分布以及SCFA对MCT1表达的影响

短链脂肪酸(SCFAs)是由纤维素性饮食在瘤胃内发酵产生的乙酸、丙酸、丁酸以及戊酸等组成的混合物。SCFA对维持反刍动物的能量平衡至关重要,目前对SCFA的吸收机制和影响因素至今尚不清楚,本试验主要研究一元羧酸转运蛋白(monocarboxylate/proton cotransporter isoform 1,MCT1)在犊牛消化道内的分布及SCFA对MCT1表达的影响。运用免疫印迹Western blot和实时荧光定量PCR检测MCT1在犊牛消化道内的分布和表达;在体外试验中,体外添加一定比例的SCFA(乙酸、丙酸和丁酸的混合物)处理犊牛原代瘤胃上皮细胞,检测SCFA对MCT1表达水平的影响。结果显示:MCT1在整个消化道内均有分布,并且在瘤胃内的表达水平显著高于其他部位(P<0.01);体外处理的瘤胃上皮细胞,正常组MCT1的表达水平显著高于(P<0.05)SARA组。结果表明:正常水平的SCFA能够促进MCT1的表达和转运能力,而过量的SCFA抑制MCT1的表达和转运能力。     

堆型艾美球虫3-1E基因在大肠杆菌中的可溶性表达及鉴定

将堆型艾美球虫（E．acervulina）3-1E基因克隆至pET-32a（＋）载体中。转化大肠杆菌BL21，在37℃下，用终浓度为1.0mmol／L的IPTG诱导表达，得到相对分子质量约为38500的重组蛋白。Western blot检测证实，该重组蛋白可以与特异性抗血清结合。在20℃条件下，以终浓度分别为2．0、1．0、0．5mmol／L的IPTG进行诱导．当IPTG终浓度为0．5mmol／L时，以可溶性形式存在的目的蛋白含量最高。     

扁穗冰草脱水素基因的克隆和表达特性分析

扁穗冰草(Agropyron cristatum(L.)Gaertn)生长于干旱、半干旱地区,具有耐旱、耐寒、抗病等特性。采用RT-PCR技术从扁穗冰草叶片中克隆3个脱水素基因Acwcor410,Acwzy2和Acwcs120)和1个肌动蛋白β-actin基因(Acβ-actin)。半定量RT-PCR分析表明,Acwcor410基因对温度敏感,对干旱胁迫不敏感;Acwzy2基因对温度不敏感,而对干旱胁迫敏感;Acwcs120基因则对冷胁迫及干旱胁迫均有响应。Western blot表明,扁穗冰草含有40kD脱水素,该蛋白的表达随冷胁迫和干旱胁迫程度的变化而变化,对干旱胁迫的响应比冷胁迫更为明显,推测该脱水素属于干旱敏感型。     

Natural (auto)antibodies in calves are affected by age and diet

Background: Natural autoantibodies (N(a)ab) were found in every species tested so far, and are likely important in maintaining homeostasis.

Objectives: (1) To determine N(a)ab in Bos taurus calves, (2) evaluate effects of diet and age on N(a)ab binding repertoires in calves, and (3) delineate bovine liver cell lysate (BLL) antigens related with variation in rumen score and body weight.

Animals and methods: Effects of age and diet on staining of BLL fragments by IgM and IgG antibodies in serum samples collected at 20 or at 26 weeks of age from bull calves either fed a restricted or ad libitum diet were analyzed using quantitative Western blotting. Correlations between fragments stained and grouping of calves were done by Principal Component Analysis (PCA). Redundancy analysis (RDA) was done to relate rumen score and body weight variation at slaughter at 27 weeks of age with stained BLL fragments.

Results: In sera from all calves IgM and IgG antibodies binding BLL antigens were found. Corresponding fragments were stained, but quantitative differences in staining intensities were related to diet and age for both IgM and IgG. PCA revealed that age had a greater influence than diet on BLL fragment staining. RDA suggested that staining by IgM or IgG of specific BLL fragments was related with variation in rumen score and body weight.

Conclusions and clinical importance: Analyses of N(a)ab in serum could be a potential tool to estimate the health status of cattle, and be used to evaluate effects of husbandry practices.     

GHSR-1a在奶山羊胃肠道的分布与表达

试验采用免疫组织化学、Real-time PCR和Western blotting方法测定ghrelin的功能性受体GHSR-1a(Growth hormone secretagogue receptor-1a,GHSR-1a)在奶山羊胃肠道的分布和表达。免疫组织化学结果显示,GHSR-1a免疫阳性细胞广泛分布于奶山羊胃肠道。在皱胃主要定位于黏膜层和肌层;瘤胃、网胃和瓣胃黏膜层及肌层中也可见GHSR-1a免疫阳性细胞;在小肠主要位于十二指肠、空肠和回肠的黏膜层、黏膜下层和肌层;在结肠、盲肠和直肠GHSR-1a免疫阳性细胞也有广泛分布;GHSR-1a主要表达于内在神经丛神经细胞、胃底腺上皮细胞、肠腺上皮细胞、复层鳞状上皮细胞、平滑肌细胞中。real-time PCR和Western blotting结果显示,皱胃、十二指肠、空肠、回肠、盲肠、结肠和直肠GHSR-1a的表达水平相对较高,显著高于瘤胃、网胃和瓣胃的表达(P<0.05)。结果表明,ghrelin可能通过GHSR-1a对奶山羊胃肠功能具有重要的调节作用。     

Expression of Mycoplasma bovis variable surface membrane proteins in the respiratory tract of calves after experimental infection with a clonal variant of Mycoplasma bovis type strain PG45

The pathomorphological findings and the expression and distribution of variable surface protein antigens (Vsp) of Mycoplasma (M.) bovis were characterised immunohistochemically in lungs of eight calves following inoculation with a Vsp A-expressing clonal variant of M. bovis type strain PG45. Within 48 h post inoculation (p.i.) an innate immune response dominated by macrophages and neutrophils develops. The monoclonal antibodies (mAbs) 1A1 and 1E5 detected M. bovis Vsp antigens in paraffin tissue sections of seven calves. Vsp antigens were widely distributed and were already present at day two p.i. within macrophages and other lung compartments. Taken together, the results demonstrate that the bovine is unable to eliminate M. bovis during the time period examined. Based on the different immunohistochemical labelling patterns obtained with the mAbs, the results also support the speculation that the in vivo variability of Vsps together with immunological factors may contribute to the chronicity of pulmonary disease.     

马身猪和大白猪不同组织DECR1基因的表达

旨在研究DECR1基因在猪不同组织和品种中mRNA和蛋白水平的表达规律,探讨该基因与脂肪代谢的关系。以山西马身猪与大白猪为试验材料,提取肝脏、心脏、肾脏、脾脏、肺脏、胃、小肠、皮下脂肪和背最长肌组织的总RNA和总蛋白,应用实时荧光定量PCR技术检测DECR1基因在2个品种各组织中mRNA的相对表达量,采用Western blot技术对各组织中DECR1蛋白进行半定量分析。实时荧光定量PCR结果显示:DECR1基因在各组织中均有表达,组织之间的表达量存在极显著差异(P<0.01),DECR1基因在肝脏、皮下脂肪与心脏中为高丰度表达;在不同品种的皮下脂肪组织中,大白猪DECR1的mRNA表达极显著高于马身猪(P<0.01)。Western blot检测结果显示:DECR1在各组织中均有表达,不同组织的表达量存在极显著差异(P<0.01),在皮下脂肪、肝脏与小肠中高表达;不同品种的皮下脂肪组织中,大白猪DECR1蛋白的表达显著高于马身猪(P<0.05)。猪DECR1基因在不同组织的表达差异可能与脂肪代谢和脂肪沉积有关。     

MCT1抑制剂p-CMB对乙酰胆碱诱导的大鼠胰液分泌的影响

采用在体试验方法研究单羧酸转运蛋白第1亚型（monocarboxylate transporter 1,MCT1）抑制剂（p-chloromercuribenzene sulfonate,p-CMB）对乙酰胆碱（acetylcholine,Ach）诱导的大鼠胰液分泌的影响。Spra-gue-Dwley（SD）雄性大鼠,体重180g-210g,分6组,每组5只,试验前禁食24h,自由饮水。动物麻醉后经外科手术后,每隔10min收集一次胰液,采用Lowry法测定胰液蛋白量,Bernfeld法测定胰酶。结果对照组（-p-CMB）和实验组（＋p-CMB）,在注射1、5、10μg／kg的Ach后对胰液分泌量、胰蛋白分泌量以及胰淀粉酶分泌量都呈现增加作用,注射后1h内达到峰值,之后逐渐降低。然而,试验组的胰液分泌各项指标比对照组的显著减少（P〈0．01）。表明MCT1抑制剂p-CMB减弱了Ach增加大鼠胰液分泌的作用。     

Pathogenicity of an Indian isolate of bovine viral diarrhea virus 1b in experimentally infected calves

The aim of this study was to determine the pathogenicity of an Indian bovine viral diarrhea virus (BVDV) 1b isolate in 7-9-months-old male calves. Infected (four) and control (two) calves were bled at three days interval for hematological, virological and serological studies until day 27. All infected calves developed respiratory illness, biphasic pyrexia, mild diarrhea, leucopenia and mild thrombocytopenia. Viraemia was demonstrated between 3 and 15dpi and the infected calves seroconverted by 15dpi. Prominent kidney lesions were endothelial cell swelling, proliferation of mesangial cells and podocytes leading to glomerular space obliteration. Degeneration and desquamation of cells lining seminiferous tubules were observed in two infected calves. Consolidation of lungs with interstitial pneumonia, mild gastroenteritis and systemic spread were also evident. It was concluded that Indian BVDV isolate induced moderate clinical disease in calves and glomerulonephritis resulting from acute BVDV infection was observed for the first time.     

Immunohistochemical analysis of MCT1 and CD147 in equine skeletal muscle fibres

Monocarboxylate transporter 1 (MCT1) and its ancillary protein CD147 facilitate efflux of lactate from the muscle. Expression of MCT1 and CD147 were studied with immunohistochemistry in type I, IIA, IIAB and IIB fibres of equine gluteal muscle. Staining intensity of MCT1 in the cytoplasm as well as in the membranes of fibre types decreased in the order I = IIA > IIAB > IIB and correlated with the oxidative capacity. Capillaries were pronounced in the MCT1 staining. CD147 antibody stained plasma membranes of all fibre types evenly, whereas the staining in the cytoplasm followed that of MCT1. In the middle gluteal muscle the expression of MCT1 follows the oxidative capacity of muscle fibres, but the expression of CD147 in sarcolemma does not vary among fibre types. The use of horse specific MCT1 and CD147 antibodies can in future studies help to evaluate lactate efflux from different muscle fibre types.     

Recovery of intact IgG in the gastrointestinal tract of the growing rat following ingestion of an ovine serum immunoglobulin

The aim of this study was to determine whether orally ingested ovine serum IgG partly resists digestion in the growing rat. Fifteen Sprague‐Dawley male rats were allocated to one of three diets for a 3‐week study: a control diet (CON) and two test diets containing either freeze‐dried ovine serum immunoglobulin (FDOI) or inactivated ovine serum immunoglobulin (IOI). Samples of stomach chyme and intestinal digesta from the ad libitum‐fed rats were subjected to ELISA and Western blot analysis. Amounts of intact ovine IgG for the FDOI diet were found to be 13.9, 20.0, 34.1, 13.0 and 36.9 μg in the total wet digesta from the stomach chyme, duodenal, jejunal, ileal and colonic digesta respectively. Qualitative detection by Western blot revealed the presence of intact ovine serum IgG with a ~150 kDa MW. This was detected in all of the gut segments (stomach chyme, duodenal, jejunal, ileal and colonic digesta) for growing rats fed the FDOI diet. No ovine IgG was detected in the chyme or digesta from rats fed the CON or the IOI diets. Ovine serum IgG partly resisted digestion in the growing rat fed the FDOI diet and was found throughout the digestive tract. These results provide a basis to explain the reported biological effects of orally administered immunoglobulin.