Upregulation of interferon-stimulated genes and interleukin-10 in peripheral blood immune cells during early pregnancy in dairy cows

In cows, interferon-tau (IFNT) regulates maternal recognition around days 15-19 after artificial insemination (AI). The present study hypothesized that if key target genes of IFNT are clearly upregulated in earlier stages of pregnancy, these genes could be use as indices of future pregnancy in cows. Therefore, we determined the expression of these genes in peripheral blood mononuclear leukocytes (PBMCs) and polymorphonuclear granulocytes (PMNs) during the maternal recognition period (MRP). Twenty multiparous Holstein cows were subjected to AI on day 0 and categorized into the following groups: pregnancy (Preg, n = 9), embryonic death (ED, n = 5) and non-pregnancy (NP, n = 6). Progesterone levels in the Preg group were higher than those in the NP group on days 12-21. ISG15 and OAS-1 (IFN-stimulated genes: ISGs) mRNA in PBMCs on day 8 was higher in the Preg group than in the NP group, and these mRNAs in PMNs was higher in the Preg group on day 5 than in the NP and ED groups. Interleukin-10 (IL-10, Th2 cytokine) mRNA expression increased on day 8 in the PBMCs of pregnant cows. Tumor necrosis factor α (TNFα, Th1 cytokine) mRNA expression was stable in all groups. In an in vitro cell culture experiment, IFNT stimulated mRNA expression of ISGs in both PBMCs and PMNs. IFNT stimulated IL-10 mRNA expression in PBMCs, whereas IFNT increased TNFα mRNA levels in PBMCs in vitro. The results suggest that ISGs and IL-10 could be responsive to IFNT before the MRP in peripheral blood immune cells and may be useful target genes for reliable indices of pregnancy before the MRP.     

Interspecies transmission of equine influenza virus (H3N8) to dogs by close contact with experimentally infected horses

In horse populations, influenza A virus subtype H3N8 (equine influenza virus, EIV) is a very important pathogen that leads to acute respiratory disease. Recently, EIV has emerged in dogs, and has become widespread among the canine population in the United States. The interspecies transmission route had thus far remained unclear. Here, we tested whether the interspecies transmission of EIV to dogs could occur as a result of close contact with experimentally EIV-infected horses. Three pairs consisting of an EIV-infected horse and a healthy dog were kept together in individual stalls for 15 consecutive days. A subsequent hemagglutination inhibition test revealed that all three dogs exhibited seroconversion. Moreover, two of the three dogs exhibited virus shedding. However, the dogs exhibited no clinical signs throughout the course of the study. These data suggest that the interspecies transmission of EIV to dogs could occur as a result of close contact with EIV-infected horses without clinical symptoms. Although the interspecies transmission of EIV is unlikely to become an immediate threat to canine hygiene, close contact between EIV-infected horses and dogs should be avoided during an EI epidemic.     

Segmentation of the Pathophysiological Stages of Diabetic Changes in the db/db Mouse

The db/db mouse is one of the diabetes mellitus animal models and if the pathophysiological stages of diabetic changes in the mouse model could simulate the stages in human diabetes, the db/db mouse could be used to better evaluate drug candidates. Blood insulin, HbA1c levels and morphological features of pancreatic islets in db/db mice were evaluated to determine the pathophysiological stage. At 6 weeks of age, db/db mice showed the highest level of plasma insulin and lowest level of HbA1c, and histopathological examination revealed enlarged islets with a circular shape and hypertrophic islet cells. By 9 and 12 weeks of age, the plasma insulin levels had decreased to mid levels and HbA1c had increased to mid to high levels; histopathological examination at this time revealed two types of islets coexisting, enlarged circular islets and small irregular-shaped islets. By 15 and 22 weeks of age, plasma insulin had decreased further to low levels and HbA1c was at its highest level; the histopathological examination at this time revealed an increase in irregular-shaped and small islets. Based on blood insulin levels, HbA1c levels and histopathology findings in the db/db mice in this study, the clinical staging of diabetic changes were recognized. The pathophysiological stages of diabetes mellitus in this animal model were similar to the stages in humans.     

Changes in follicular vascularity during the first follicular wave in lactating cows

Increase in the blood supply to individual follicles appears to be associated with follicular growth rates and the ability to become the dominant follicle, while reduced thecal vascularity appears to be closely associated with follicular atresia. Therefore, this study aimed to determine the real-time changes in the vascularity of the follicle wall during the first follicular wave in cycling Holstein cows. Normally cycling and lactating cows (n=5) were examined by transrectal color Doppler ultrasonography (the sensitivity for velocity: > 2 mm/sec) to determine the changes in the vasculature of the follicle wall (presence or absence of blood flow) and the diameter of follicles. A new follicular wave and ovulation were induced by GnRH injection at 48 h after an injection of PGF2alpha analogue. The ovaries were scanned daily for 7 days after GnRH injection. Follicles >2.5 mm were classified into 3 groups by the changes in diameter as follows: 1) largest follicle, 2) second largest follicle, and 3) small follicles, which included all other follicles >2.5 mm. Before the follicle selection, there was no significant difference in the percentage of follicles with detectable blood flow between the subsequently determined largest and second largest follicles. After the follicle selection, the percentage of follicles with detectable blood flow significantly decreased among the second largest follicles. In addition, small follicles with detectable blood flow kept larger diameters than those without detectable blood flow from one day before the occurrence of follicle selection. It is likely that maintenance of follicle vasculature and appropriate blood supply to the larger follicles is essential for follicle dominance. In small follicles, the presence of blood flow within the wall also appears to be required for recruitment. Consequently, the data suggest that the change of the blood supply to an individual follicle closely relates to the dynamics of follicular growth in the first follicular wave in the cow.     

Effects of LH and PGF2alpha in equine dominant follicles observed by MDS

Microdialysis System (MDS) is a novel technique used for investigation of molecule secretion between different cell populations. Local hormonal secretion at follicular wall has been still unclear. This MDS study was used to determine progesterone (P4), androstenedione (A4), estradiol-17beta (E2) and Prostaglandin F2alpha (PGF2alpha) release in mare pre-ovulatory follicles. Follicles larger than 30 mm were isolated from the ovary and follicular fluid aspirated for hormone assay. Follicular fluid collected from small, middle and large follicles were analyzed by EIA. The concentrations of P4 and PGF2alpha were similar among the different sizes of follicles. The release of A4 was observed in middle and large follicles. E2 concentration was observed in middle follicles and was higher in large follicles compared with middle follicles. Follicular wall was cut and incubated for MDS and when LH was infused, there was an increase in P4 and A4 release. PGF2alpha release was considerably high after LH infusion compared to the control group. Infusion of PGF2alpha increased P4 and A4 release but there was no change in E2 release. This results suggest that in pre-ovulatory follicles, LH stimulates theca interna cells and also PGF2alpha seemed to have a mediator role to induce steroid hormone production and luteinization of follicular cells. The nature of the mechanisms involved in selection of large follicles is still a perplexing research problem in reproduction.     

Evaluation of antigen detection kits for diagnosis of equine influenza

In this study, we evaluated whether five rapid antigen detection kits for human influenza could be used for the diagnosis of equine influenza (EI). Limiting dilution analyses showed that Directigen Flu A+B and ESPLINE INFLUENZA A&B-N had the highest sensitivities to equine-2 influenza viruses (EIVs) among the kits investigated. From the results of virus detection in nasal swabs taken from horses infected with EIV, these two kits could produce positive results in reasonable agreement with those obtained by virus isolation or RT-PCR, suggesting that these kits could be useful for rapid diagnosis of EI in the field. However, from the viewpoint of specificity for EIV, Espline seems to be superior to Directigen.     

Complete mitochondrial genome sequence of Brassica oxyrrhina and comparative analysis of the region containing orf108, a male sterility gene for mustard (Brassica juncea), among B. oxyrrhina,Diplotaxis erucoides and Sinapis species

To date, several cytoplasms of wild species have been introduced to Brassica juncea, for inducing cytoplasmic male sterility (CMS). One of the causal genes of CMS is orf108, which is widely distributed in Brassicaceae including Brassica oxyrrhina. To further understand the origin of orf108, we assembled the complete mitochondrial genome sequence of B. oxyrrhina. We also determined the DNA sequences upstream of atp1 including orf108 for D. erucoides and five Sinapis species. The orf108 was definitively placed in the mitochondrial genome of B. oxyrrhina consisting of 247,936 bp. In S. alba, previously reported to possess orf108, the gene was not present, whereas Sinapis arvensis and Sinapis turgida contained orf108. The orf108 sequence in the two Sinapis species showed a novel nucleotide substitution compared to D. erucoides. Northern hybridization suggested, furthermore, that orf108 mRNA was processed in the two species similarly to that in B. oxyrrhina. The results clarified the interspecific differentiation of orf108‐apt1 region in Sinapis.     

弱光照射和无机营养供给对冷藏绿芦笋品质变化的影响

绿芦笋嫩茎基部切口插入含有N、P、K营养的琼脂凝胶中 ,在 (6± 0 2 )℃下贮藏。贮藏期间 ,分别以有效光量子束密度 (PPFD)为 2 μmol·m-2 ·s-1的红色发光二极管和白色荧光灯作为弱光源 ,每天 2 4h连续照射绿芦笋 2 1d。结果表明 ,红光和白光对冷藏绿芦笋品质的保持具有相似的效果。供给无机营养的处理在红光或白光照射下 ,色值 (a 和ΔE ab)变化率、粗纤维含量增长率和可溶性固形物减少率为最低 ,其次是无营养供给的红光或白光处理 ,以黑暗处理为最高 ;供给无机营养处理的芦笋嫩茎伸长率大于无营养处理的 ;光线照射与否和无机营养供给有无与芦笋失重率之间无明显相关性