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.     

MCT1, MCT4 and CD147 gene polymorphisms in healthy horses and horses with myopathy

Polymorphisms in human lactate transporter proteins (monocarboxylate transporters; MCTs), especially the MCT1 isoform, can affect lactate transport activity and cause signs of exercise-induced myopathy. Muscles express MCT1, MCT4 and CD147, an ancillary protein, indispensable for the activity of MCT1 and MCT4. We sequenced the coding sequence (cDNA) of horse MCT4 for the first time and examined polymorphisms in the cDNA of MCT1, MCT4 and CD147 of 16 healthy horses. To study whether signs of myopathy are linked to the polymorphisms, biopsy samples were taken from 26 horses with exercise-induced recurrent myopathy. Two polymorphisms that cause a change in amino acid sequence were found in MCT1 (Val₄₃₂Ile and Lys₄₅₇Gln) and one in CD147 (Met₁₂₅Val). All polymorphisms in MCT4 were silent. Mutations in MCT1 or CD147 in equine muscle were not associated with myopathy. In the future, a functional study design is needed to evaluate the physiological role of the polymorphisms found.     

MCT1, MCT4 and CD147 gene polymorphisms in healthy horses and horses with myopathy

Polymorphisms in human lactate transporter proteins (monocarboxylate transporters; MCTs), especially the MCT1 isoform, can affect lactate transport activity and cause signs of exercise-induced myopathy. Muscles express MCT1, MCT4 and CD147, an ancillary protein, indispensable for the activity of MCT1 and MCT4. We sequenced the coding sequence (cDNA) of horse MCT4 for the first time and examined polymorphisms in the cDNA of MCT1, MCT4 and CD147 of 16 healthy horses. To study whether signs of myopathy are linked to the polymorphisms, biopsy samples were taken from 26 horses with exercise-induced recurrent myopathy. Two polymorphisms that cause a change in amino acid sequence were found in MCT1 (Val₄₃₂Ile and Lys₄₅₇Gln) and one in CD147 (Met₁₂₅Val). All polymorphisms in MCT4 were silent. Mutations in MCT1 or CD147 in equine muscle were not associated with myopathy. In the future, a functional study design is needed to evaluate the physiological role of the polymorphisms found.     

Polymorphisms in MCT1, CD147, PDK4, and DMRT3 genes in Arabian and Quarter Horses

Monocarboxylate transporter isoform 1 (MCT1) and its ancillary protein, CD147 function to transport H⁺ and lactate ions, retarding systemic acidosis and fatigue. The PDK4 gene is involved in the adenosine triphosphate production, and the DMRT3 gene is involved in the locomotor system. This study investigated polymorphisms of MCT1, CD147, PDK4, and DMRT3 in Arabian and Quarter Horses to associate polymorphisms with performance. Arabian Horses were divided into high performance and untrained horses groups based on success in endurance competition, whereas Quarter Horses were separated by a speed index. Polymorphisms were analyzed by sequencing (MCT1 and CD147), ARMS-PCR (PDK4), and PCR-RFLP (DMRT3). To compare the frequencies of SNPs, the Fisher's exact test was performed in the software R at 5% of significance. The A alleles from the polymorphisms Lys457Gln:1573A>C of MCT1 and Ile51Val:168A>G of CD147 were essentially fixed in both breeds. Only two Arabian from the high performance and one from the untrained horses group appeared AC in MCT1. For the DMRT3 polymorphism (g.22999655C>A:ECA23), all the animals were CC, except one Arabian in the high and two Quarter Horses in the low performance group that were AC. For the PDK4 polymorphism (g.38973231A>G:ECA4), Arabian showed a significantly greater frequency of the G allele than Quarter Horses (P < .01). Considering the kind of exercise practiced by each breed, it is likely that the PDK4 polymorphism influences the pathway to energy production.     

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.     

Gentamicin tissue concentrations in equine small intestine and large colon

Gentamicin sulfate (2.2 mg/kg of body weight, IV) was given to anesthetized horses. Jejunal and large colon tissue samples (1 g), serum, and urine were collected over a 4-hour period. Maximum gentamicin concentrations in serum (10.06 +/- 2.85 micrograms/ml) occurred at 0.25 hours after injection. Maximum gentamicin concentrations in the large colon (4.13 +/- 1.80 micrograms/ml) and jejunum (2.26 +/- 1.35 micrograms/ml) occurred in horses at 0.5 and 0.33 hours, respectively. Tissue concentrations decreased in parallel with serum concentrations and were still detectable at the end of the 4-hour period. During the time that samples were collected, the total amount of gentamicin excreted in the urine ranged from 7.21 +/- 3.11 mg to 11.91 +/- 7.12 mg, with a mean urinary concentration of 57.01 +/- 5.37 micrograms/ml. Over the 4-hour collection period, the fraction of dose that was excreted unchanged in the urine was 4.8 +/- 1.9%. Pharmacokinetic analyses of the serum concentration-time data gave a serum half-life of 2.52 +/- 1.29 hours, volume of distribution of 227 +/- 83 ml/kg, and body clearance of 1.12 +/- 0.26 ml/min/kg. The half-lives of the antibiotic in the jejunum and large colon were 1.32 and 1.33 hours, respectively.     

Immunohistochemical detection of SWC3, CD2, CD3, CD4 and CD8 antigens in paraformaldehyde fixed and paraffin embedded porcine lymphoid tissue

Identification of the different cell types of the immune system is important for in situ studies on the pathogenesis of infectious diseases in various animals, including the pig. Unfortunately, many monoclonal anti-leukocyte antibodies are only useful for staining frozen tissue sections with inherent poor tissue morphology, and are not readily adapted to formaldehyde fixed and paraffin embedded tissue with well preserved morphology. Seven well characterised monoclonal antibodies against porcine leukocyte antigens were tested on neutral buffered paraformaldehyde fixed and paraffin embedded porcine tissue sections using the highly sensitive tyramide signal amplification system. Combining this method with different antigen retrieval techniques enabled us to detect CD2, CD3, CD4, CD8 and SWC3 antigen expressing cells in porcine lymphoid tissue. Thus, we describe herein methods for the detection of several major cell types of the porcine immune system in fixed tissue with optimal preservation of histological details.     

Lesions in the small intestine of newborn pigs inoculated with porcine, feline, and canine coronaviruses

The infectivity and pathogenicity to newborn pigs of antigenically related coronaviruses from pigs (transmissible gastroenteritis virus; TGEV), cats (feline infectious peritonitis virus; FIPV), and dogs (canine gastroenteritis virus; CGEV) were studied by light, scanning electron, and immunofluorescence microscopy. Hysterectomy-derived, 12-hour-old pigs were orally given tissue culture or frozen preparations of 6 coronavirus strains (3 porcine, 2 feline, and 1 canine). The pigs were killed at regular intervals between 24 and 144 hours after exposure. Virulent TGEV and virulent FIPV produced necrosis of villous epithelium, resulting in villous atrophy in the jejunum and the ileum. Similar, but less extensive and severe lesions, were produced by the 4 other viruses. Coronaviral antigens were identified by immunofluorescence in villous epithelial cells of pigs that had been inoculated with virulent TGEV, attenuated TGEV, virulent FIPV, and tissue culture-adapted FIPV. In contrast, coronaviral antigens were not induced by the small plaque variant TGEV and virulent CGEV in the villous epithelium, but rather in cells of the lamina propria and crypt epithelium.     

水通道蛋白在小鼠小肠中的表达和定位

探明水通道蛋白在小鼠小肠中的表达及定位,为研究小肠水转运过程中的细胞和分子机制奠定理论基础。本文运用RT-PCR和免疫印迹技术检测小鼠十二指肠、空肠和回肠中水通道蛋白的表达情况,结果显示,AQP3在小鼠空肠中存在基因和蛋白表达,AQP4在小鼠回肠中存在基因和蛋白表达。为进一步明确这两种水通道蛋白在小肠中的表达位置,通过免疫组织化学的方法进一步证实AQP3主要表达在小鼠空肠黏膜上皮细胞,而AQP4主要表达在小鼠回肠隐窝细胞基底膜。     

鼠伤寒沙门氏菌对鼠淋巴细胞MHC－1、MHC－2、ICAM－1和CD4表达的影响

应用流式细胞仪技术，检测经口灌服１０３个鼠伤寒沙门氏菌（Ｓａｌｍｏｎｅｌｌａｔｙｐｈｉｍｕｒｉｕｍ），４８ｈ后迫杀的Ｃ５７Ｂ１６／Ｈ２ｂ鼠外周淋巴结、肠系膜淋巴结、脾和外周血中淋巴细胞ＭＨＣ－１、ＭＨＣ－２、ＩＣＡＭ－１和ＣＤ４４种受体的变化。结果表明，鼠伤寒沙门氏菌使外周血淋巴细胞ＭＨＣ－１、ＭＨＣ－２和ＩＣＡＭ－１的表达显著升高（Ｐ＜０．００１），肠系膜淋巴结中淋巴细胞ＭＨＣ－２、ＣＤ４和外周血淋巴细胞ＣＤ４的表达也有升高。     

Differential Expression of Monocarboxylate Transporter 1 and Ancillary Protein CD147 in Red Blood Cells of Show Jumping Horses

We compare the expression levels of the lactate transporter complex consisting of the lactate transporter, monocarboxylate transporter 1 (MCT1), and its ancillary protein, cluster of differentiation 147 (CD147), in the membranes of red blood cells (RBCs) from two breeds of jumping horses and associate the expression levels of these proteins with their jumping ability. The expression levels of MCT1 and CD147 proteins on the membranes of RBCs collected from 30 show jumping horses of two different breeds were quantified: the Brazilian Sport Horses (n = 17) and the European Warmbloods (n = 13). The levels of MCT1 and CD147 in the RBC membranes were measured by western blot using horse-specific antibodies. Statistical analyses included unpaired Student t test and chi-squared test. According to the expression levels of MCT1 and CD147 proteins, 88% of the Brazilian Sport Horses were categorized as high lactate transporters (HTs) and the remaining 12% as low lactate transporters (LTs). The opposite was found for the European Warmbloods, where most animals (77%) were classified as LTs and the remaining animals (23%) were classified as HTs. Brazilian Sport Horses express statistically significantly higher levels of CD147 and MCT1 than European Warmbloods. The classification of horses considering the expression of proteins involved in the ability to transport lactate through the complex MCT1-CD147 seems to be breed dependent, with horses that are able to jump higher obstacles showing lower expression of the MCT1-CD147 complex in their RBCs.     

Small intestine and small colon neuropathy in equine dysautonomia (grass sickness)

The number of neurons in the coeliacomesenteric ganglia and the myenteric and submucosal plexuses of the jejunum, ileum and small colon, and the pathological changes induced in them, were studied in various types of equine dysautonomia. In all forms of dysautonomia, severe and extensive neuron loss and damage occurred in the ileum. In acute and subacute dysautonomia, jejunal neuron loss and damage were severe, but in chronic cases significantly less loss or damage occurred. The damage followed the same pattern in the small colon but it was always less obvious than in the jejunum. The distribution of the damage was uniform within a segment of the intestine. In fatal cases of dysautonomia, the clinical severity and duration of illness seems, in most instances, to be related to the amount of neuronal disruption occurring in the jejunum. Severe disruption results in acute/subacute dysautonomia, while milder damage leads to the chronic form.No case of dysautonomia was encountered in which enteric neuron loss and damage occurred without significant neuronal disruption also occurring in the coeliacomesenteric ganglia.Ileal neuronal damage and loss are not invariably worse than that in the jejunum, and the possible reasons for this, together with the relationship between neuronal damage and possible causes of dysautonomia, are discussed.Abbreviations H&E haematoxylin and eosin Deceased. Formerly of the Moredun Research Institute, 408 Gilmerton Road, Edinburgh, EH17 7JH, UK     

猪圆环病毒2型感染小鼠脾脏内CD4~+CD25~+调节性T细胞的动态变化

为分析猪圆环病毒2型(PCV2)感染小鼠后脾细胞中CD4+CD25+调节性T细胞(Tregs)占CD4+T细胞比例的动态变化,探讨Tregs与PCV2感染的关系,本研究选择清洁级昆明小鼠60只,随机分成实验组和对照组,实验组腹腔接种PCV2,分别在接种后第0 d、5 d、10 d、20 d、30 d和60 d取脾脏制备单细胞悬液,用FITC-CD4和PE-CD25单克隆抗体标记Tregs,采用流式细胞仪检测Tregs占总CD4+T细胞百分比的动态变化。结果表明感染组小鼠脾细胞中Tregs百分比从第5 d开始逐步上升,第20 d达峰值后逐渐下降,感染组Tregs百分比第10 d、20 d、30 d时显著高于对照组(p0.05),但第60 d两组间差异不显著(p0.05)。试验结果证明PCV2感染昆明小鼠后,可在小鼠脾脏内诱导明显的Tregs增殖,这些增殖的细胞可能在PCV2感染中发挥免疫抑制作用。     

猪圆环病毒2型感染对猪CD80和CD86 mRNA表达的影响

通过构建猪共刺激分子CD80和CD86的缺失cDNA竞争分子,用竞争PCR技术定量检测了猪圆环病毒2型(PCV2)感染后猪肺泡巨噬细胞(PAM)中CD80和CD86的mRNA水平,分析了PCV2感染对猪共刺激分子CD80和CD86的mRNA表达的影响.结果显示,PCV2感染后,PAM中CD80和CD86的mRNA水平变化趋势一致,只是CD86的升降幅度更大.在PCV2感染后3d,CD80与CD86的mRNA水平下降至最低,随后迅速上升,至14d达到高峰,尔后快速下降,21d及以后恢复正常.本研究结果表明,PCV2初期可明显抑制猪共刺激分子CD80和CD86的基因转录.     

Effects of indomethacin on Salmonella typhimurium- and cholera toxin-induced fluid accumulation in the porcine small intestine

The effect of the cyclooxygenase and prostaglandin E2 (PGE2) synthesis inhibitor, indomethacin, on the secretory responses induced by Salmonella serotype Typhimurium (ST) and cholera toxin (CT), in the porcine small intestine was investigated. ST (10(10) colony-forming units) and CT (56 micrograms) were instilled in tied-off intestinal loops in young anaesthetized pigs receiving intravenous indomethacin in a total dose of 7.5 mg/kg, or saline. The accumulated fluid in the loops and the luminal content of endogenous secretagogues PGE2 and 5-hydroxytryptamine (5-HT) were measured. ST induced fluid accumulation in the jejunum, whereas CT induced fluid accumulation in the jejunum and ileum. Indomethacin had no effect on the secretory responses. Indomethacin had a significant effect on the luminal content of PGE2 in jejunal ST and CT loops, whereas no effect of indomethacin was observed on the luminal content of 5-HT in ST and CT loops. In ST and CT loops, an increased content of PGE2 and 5-HT compared with test loops infused with Ringer's solution was observed. These results indicate that the porcine jejunal secretory response to ST and CT does not involve prostaglandins although indomethacin has an influence on the luminal release of PGE2 but not of 5-HT.