Age Difference in Morphology and Immunohistology inthe Thymus and Spleen in Crl:CD (SD) Rats

We investigated chronological changes in immunohistochemical phenotyping in the thymus and spleen in Crl:CD rats up to the age of about one year. In the thymus, T cells increased markedly from 3 to 4 weeks of age. Proliferating cells also increased markedly at these points. B cells tended towards an increase with age. In the spleen, white pulp increased until 9 weeks of age and remained fairly stable thereafter. In the periarteriolar lymphoid sheath and marginal zone, T cells gradually increased until 9 weeks of age and became almost flat thereafter. In the lymph follicle, T cells increased with age. B cells tended towards an increase with age in all areas of the spleen. It was concluded that development of the thymus was most marked from 3 to 4 weeks of age and that both the thymus and spleen had matured by 9 weeks of age.     

Proteome analysis of cerebrospinal fluid in healthy beagles and canine encephalitis

We performed proteomics analysis of the cerebrospinal fluid (CSF) of healthy dogs and dogs with meningoencephalitis of unknown etiology (MUE). By comparing two-dimensional electrophoreses (2DE), an upregulated spot was found in MUE dogs. This protein was identified as a neuron-specific enolase (NSE) by analysis with MALDI-TOF mass spectrometry. In comparing dot blots using an antibody against NSE, the NSE levels in the CSF of MUE dogs was significantly higher than that of the controls. NSE is a diagnostic marker of neuroendocrine tumors, brain injury and spinal cord trauma in humans. It seems that the NSE concentration in the CSF is increased by cellular destruction in canine encephalitis. Though elevation of NSE may not be specific in canine encephalitis because the NSE level was increased in other CNS diseases, further study including measurement with serum is necessary.     

Parathyroid hormone (1-34) improves bone mineral density and glucose metabolism in Spontaneously Diabetic Torii-Lepr(fa) rats

The Spontaneously Diabetic Torii-Lepr(fa) (SDT-fa/fa) rat, a model of obese type 2 diabetes, shows obesity, hyperglycaemia and low bone mineral density (BMD). The objective of this study is to evaluate the effects of parathyroid hormone (1-34) [PTH(1-34)] on BMD and glucose metabolism in the SDT-fa/fa rat. SDT-fa/fa rats showed obesity with hyperglycaemia and decreased serum osteocalcin levels and the tibial BMD. A 4-week treatment of PTH(1-34) (20 μg/kg/day) increased the serum osteocalcin levels and the tibial BMD, and decreased the serum glucose levels without changing the serum insulin levels. These findings indicate that PTH(1-34) improved not only BMD but also glucose metabolism in SDT-fa/fa rats. This study suggests that PTH(1-34) is a novel agent for the treatment of diabetic osteoporosis.     

Proliferation capacity, neuronal differentiation potency and microstructures after the differentiation of canine bone marrow stromal cells into neurons

We examined the proliferation capacity and neuronal differentiation potency of canine bone marrow stromal cells (BMSCs). In addition, the microstructures of neuron-like cells after neuronal differentiation were observed under a scanning electron microscope. Canine BMSCs grew to confluency at 10.0 ± 2.5 days, and 3.8 ± 2.1 × 10(6) BMSCs were collected in one passage. Approximately 65% of canine BMSCs changed to neuron-like morphology after neuronal differentiation, and nearly all neuron-like cells stained positive against neuron-specific enolase. In addition, microstructures such as the cellular organelles, filaments and growth cones of these cells bore a close resemblance to those of the original mature neurons. These results suggested that canine BMSCs might be capable of differentiating into neurons.     

Factors Influencing the Frequency of Pregnancy Loss among Thoroughbred Mares in Hidaka,Japan

Pregnancy loss in mares is thought to be a main problem associated with reproductive efficiency. To clarify the situation of pregnancy loss in Thoroughbred mares in Japan, the occurrence of pregnancy loss before and after 35 days of gestation was investigated with 1,476 Thoroughbred mares in Hidaka, Japan, from 2007 to 2009. Pregnancy loss on days 17-35 was determined by ultrasound examination between 17 and 35 days after the last mating. Follow-up surveys were conducted between 35 days and foaling to determine pregnancy loss on day 35 until foaling in 843 of these mares. Using multiple logistic regression analysis, we assessed the influence of mare age, reproductive status, twin pregnancy reduction, body condition score (BCS), estrus type in foaling mares (foal heat or not), progesterone therapy, and endometrial cysts on pregnancy loss rates on days 17-35 and on day 35 until foaling in this population of mares. The pregnancy loss rates on days 17-35 and on day 35 until foaling were 5.8% and 8.7%, respectively. The overall pregnancy loss rate (day 17 until foaling, including parturient losses) was 14.7%. Risk factors for pregnancy loss included decrease in BCS between 17 and 35 days, <5 BCS at day 35, mating during foal heat, and endometrial cysts. In all, 14.7% of pregnancies were lost between day 17 and birth, contributing significantly to reduced reproductive efficiency in Thoroughbred mares in Japan. These observations indicate that mares should be maintained at high BCS and should be prevented from mating in foal heat to decrease the pregnancy loss rates.     

Expression of extracellular matrix metalloproteinase inducer (EMMPRIN) and its expected roles in the bovine endometrium during gestation

Extracellular matrix metalloproteinase inducer (EMMPRIN) and its induced matrix metalloproteinases (MMPs) play a crucial role in tissue remodeling during the peri-implantation period. However, the role of EMMPRIN in the bovine placenta is still unclear. We have postulated that EMMPRIN might play a regulatory role in trophoblastic cell functions during gestation by itself or through the regulation of MMP expression. In this study, EMMPRIN mRNA was detected in the bovine placentome and interplacentome throughout gestation, and its expression was significantly higher in the cotyledon during late gestation. In situ hybridization showed that EMMPRIN mRNA was expressed in the caruncular epithelium and the cotyledonary epithelium, including binucleate cells. Western blot analysis detected a band representing a protein of approximately 65 kDa in the caruncular and cotyledonary tissues, and the intensity of its expression was increased in both of these tissues during late gestation. The expression levels of MMP-2 and MMP-14 in the bovine placenta were higher during late gestation, as was observed for EMMPRIN. Therefore, EMMPRIN might regulate trophoblastic cell functions, especially those of binucleate cells, through MMP expression in the bovine placenta.     

Estimation of glomerular filtration rate in calves using the contrast medium iodixanol

To develop a simple procedure for estimating glomerular filtration rate (GFR) in calves, a three-sample method using iodixanol was first compared to that using the standard agent inulin. Iodixanol and inulin were co-administered intravenously to calves at 40 mg I/kg and 40 mg/kg, respectively, and blood was collected 30, 60, 120, and 180 min later. Serum iodixanol and inulin concentrations were separately determined by high performance liquid chromatography and colorimetry. Serum urea nitrogen (UN) and creatinine concentrations were also measured. GFR estimated by iodixanol was consistent with that using inulin in clinically healthy calves. Based on GFR estimations in healthy calves and those renal-loaded with iodixanol, it was found that the serum creatinine concentrations became elevated when GFR decreased to 60% of the reference value. In contrast, serum UN concentrations fluctuated widely, presumably due to extra-renal factors. When GFR was estimated using the three-sample method and compared with the single-blood-sample method, 62/69 (90%) of samples tested were within the agreement plots. The results demonstrated that the single-blood-sample method using iodixanol may be useful in monitoring GFR in calves.     

Production of bacteriocin by Leuconostoc mesenteroides 406 isolated from Mongolian fermented mare's milk,airag

The purification and characterization of a bacteriocin produced by Leuconostoc mesenteroides strain 406 that was isolated from traditional Mongolian fermented mare's milk, airag, were carried out. Leuconostoc mesenteroides strain 406 was identified on the basis of its morphological and biochemical characteristics and carbohydrate fermentation profile and by API 50 CH kit and 16S ribosomal DNA analyses. The neutral‐pH cell‐free supernatant of this bacterium inhibited the growth of several lactic acid bacteria and food spoilage and pathogenic organisms, including Listeria monocytogenes and Clostridium botulinum. The bacteriocin was heat‐stable and not sensitive to acid and alkaline conditions, but was sensitive to several proteolytic enzymes such as pepsin, pronase E, proteinase K, trypsin, and α‐chymotrypsin, but not catalase. Optimum bacteriocin production (4000 activity units/mL) was achieved when the strain was cultured at 25°C for 24–36 h in Man Rogosa Sharpe medium. The bacteriocin was partially purified by ammonium sulfate precipitation (80% saturation), dialysis (cut‐off MW: 1000), and gel filtration chromatography. Sodium dodecyl sulfate‐polyacrylamide gel electrophoresis revealed that the bacteriocin had a molecular weight of approximately 3.3 kDa. To our knowledge, this is the first report of the isolation of a bacteriocin‐producing Leuconostoc strain from airag. An application to fermented milks would be desired.     

Seasonal change and distribution of grass nutritive values and minerals in an open pasture surrounded by forest

The forage quality of pasture species is spatially heterogeneous. In this study, we evaluated the dynamics and spatial structure of forage quality and attempted to relate the observed variation to differences in light availability and soil properties. We compared grass forage quality in open grasslands with forest edge environments and established plots in open Japanese grasslands surrounded by secondary forests. We sampled Anthoxanthum odoratum L. and soil from each plot in May and September. These samples were analyzed to determine the crude protein (CP), neutral detergent fiber, and mineral concentrations. We calculated Moran’s I coefficients across discrete lag distance classes to construct autocorrelograms. The CP model showed a non-random pattern in May at a lag distance of 40 m, but a clustered pattern in September at a small scale. The mineral models produced similar clustered spatial patterns. We found no strong correlation between aboveground (plant) and belowground (soil) mineral contents. CP and some mineral element concentrations in the plants collected in September were significantly greater at the forest edge than in open grasslands. The forest edge therefore appears to be a valuable place to provide grazing animals with adequate protein and mineral intake during the later part of the growing season.     

Distribution of photoperiod-insensitive allele Ppd-A1a and its effect on heading time in Japanese wheat cultivars

The Ppd-A1 genotype of 240 Japanese wheat cultivars and 40 foreign cultivars was determined using a PCR-based method. Among Japanese cultivars, only 12 cultivars, all of which were Hokkaido winter wheat, carried the Ppd-A1a allele, while this allele was not found in Hokkaido spring wheat cultivars or Tohoku-Kyushu cultivars. Cultivars with a photoperiod-insensitive allele headed 6.9–9.8 days earlier in Kanto and 2.5 days earlier in Hokkaido than photoperiod-sensitive cultivars. The lower effect of photoperiod-insensitive alleles observed in Hokkaido could be due to the longer day-length at the spike formation stage compared with that in Kanto. Pedigree analysis showed that ‘Purple Straw’ and ‘Tohoku 118’ were donors of Ppd-A1a and Ppd-D1a in Hokkaido wheat cultivars, respectively. Wheat cultivars recently developed in Hokkaido carry photoperiod-insensitive alleles at a high frequency. For efficient utilization of Ppd-1 alleles in the Hokkaido wheat-breeding program, the effect of Ppd-1 on growth pattern and grain yield should be investigated. Ppd-A1a may be useful as a unique gene source for fine tuning the heading time in the Tohoku-Kyushu region since the effect of Ppd-A1a on photoperiod insensitivity appears to differ from the effect of Ppd-B1a and Ppd-D1a.