Full-term development of offspring using round spermatids produced ectopically from fetal male germ cells

The continuous production of mammalian sperm is maintained by the proliferation and differentiation of spermatogonial stem cells, which originate from primordial germ cells in the early embryo. Previously, we reported that the transplantation of fetal male gonadal tissue into the recipient testis was effective obtaining functional sperm. This transplantation technique is a promising new approach for the preservation of testicular function in a mutant animal with embryonic lethality. In the present study, we examined whether spermatogenesis from fetal male germ cells is induced under ectopic conditions in male and female recipients. Nine to 10 weeks after the transplantation of male gonads prepared from embryos at 12.5 or 16.5 days post gestation, male germ cell differentiation occurred under the skin of male and female recipient nude mice. Histological analyses revealed that grafted gonads contained haploid germ cells such as round or elongated spermatids. Furthermore, we succeeded in obtaining normal progeny by injecting the ectopically produced round spermatids into the cytoplasm of oocytes, even when the male germ cells had been generated in female recipients. These results indicate that the transplantation of fetal male gonads under the skin of recipient mice is a useful technique for obtaining functional male gametes.     

Detection of in vivo DNA damage induced by ethanol in multiple organs of pregnant mice using the alkaline single cell gel electrophoresis (Comet) assay

Ethanol is principal ingredient of alcohol beverage, but considered as human carcinogen, and has neurotoxicity. Alcohol consumption during pregnancy often causes fetal alcohol syndrome. The DNA damage is one of the important factors in carcinogenicity or teratogenicity. To detect the DNA damage induced by ethanol, we used an in vivo alkaline single cell gel electrophoresis (Comet) assay in pregnant mice organs and embryos. Pregnant ICR mice on Day 7 of gestation were treated with 2, 4 or 8 g/kg ethanol, and maternal organs/tissues and embryos were subjected to the Comet assay at 4, 8, 12 and 24 hr after ethanol treatment. Four and 8 g/kg ethanol induced DNA damage in brain, lung and embryos at 4 or 8 hr after the treatment. Two g/kg ethanol did not cause any DNA damage, and 8 g/kg ethanol only increased the duration of DNA damage without distinct increase in the degree of the damage. No significant DNA damage was observed in the liver. To detect the effect of acetaldehyde, disulfiram, acetaldehyde dehydrogenase inhibitor, was administered before 4 g/kg ethanol treatment. No significant increase of DNA damage was observed in the disulfiram pre-treated group. These data indicate that ethanol induces DNA damage, which might be related to ethanol toxicity. Since pre-treatment of disulfiram did not increase DNA damage, DNA damage observed in this study might not be the effect of acetaldehyde.     

Antiinflammatory effect of Japanese horse chestnut (Aesculus turbinata) seeds

The antiinflammatory effects of Japanese horse chestnut (Aesculus turbinata) seeds were examined in vivo and in vitro. The extract of this seed (HCSE) inhibited croton oil-induced swelling of the mouse concha. HCSE inhibited cyclooxygenase (COX) -1 and -2 activities, but had no effect on 15-lipoxygenase and phospholipase A2 activities. Inhibition of COX-2 occurred at a lower concentration of HCSE than for COX-1. Japanese horse chestnut seeds contain coumarins and saponins, but these chemicals did not inhibit COX activities. These results suggest that the antiinflammatory effect of Japanese horse chestnut seeds is caused, at least partly, by the inhibition of COX. The inhibitor of COX in this seed may be a chemical(s) other than coumarins and saponins.     

Sugar expressions on the vaginal epithelium in pregnant mice

Sugar expressions were examined on the epithelium of both the middle portion of the vagina and the vaginal portion of the cervical canal (CC) in pregnant mice to understand the pathogenesis of bacterial infection in the female reproductive organ by using a panel of lectins. As a result, N-acetylglucosamine was positive before pregnant day (P) 7 but negative after P10 and at diestrus on both the vagina and the CC. In addition, some differences in sugar expressions were seen between them. These results suggest that sugar expressions on the mucosal surface would change not only site-specifically but also time-dependently, and these sugar differences indicate the possibility of the alteration of the settled bacterial species on the vaginal mucosa in pregnancy.     

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.     

Bilateral segmental aplasia with unilateral uterine horn torsion in a pomeranian bitch

Bilateral segmental aplasia of the uterine horns with unilateral pyometra and uterine horn torsion were diagnosed in a Pomeranian bitch that presented with chronic abdominal distension and an acute onset of anorexia and lethargy. Because radiographic and ultrasonographic findings revealed the presence of markedly enlarged bilateral uterine horns filled with fluid in the caudal abdomen, a tentative diagnosis of either pyometra or hydrometra with uterine horn torsion was made. Exploratory laparotomy showed bilateral, segmentally distended uterine horns with unilateral uterine horn torsion. Ovariohysterectomy was performed, and bilateral segmental aplasia of the uterine horns with the development of unilateral uterine horn torsion was diagnosed histopathologically. To the authors' knowledge, this is the first report of uterine horn torsion in conjunction with segmental aplasia of the uterine horn in a bitch.     

Relation of reproductive performances and rectal palpation for luteum function of heifers 7 days after estrus

Diagnosis of corpus luteum (CL) function by rectal palpation (RP) has been widely used for recipient selection of embryo transfer (ET), a technology essential for genetic improvements in cattle. To examine the accuracy of RP diagnosis method, the relationship between RP‐based CL function and reproductive performance was compared in this study. In Experiment 1, CL of Holstein heifers on day 7 after estrus was classified into functional or hypoplastic by RP, and the results were compared with ultrasonographic (US) images and plasma progesterone (P4) levels. As a result, heifers with functional CL judged by RP had a mean maximum CL diameter of 20.1 ± 3.1 mm on US and a mean P4 concentration of 8.1 ± 2.3 ng/mL. These values were significantly greater than those of heifers with hypoplastic CL (12.4 ± 5.4 mm, 4.0 ± 2.8 ng/mL) (P < 0.001). In Experiment 2, the length of the estrus cycle was examined between functional CL and hypoplastic CL. The rate of heifers with a normal estrus cycle length with 18–25 days was significantly lower with hypoplastic CL than with functional CL (16/24 vs. 43/46, P < 0.01). In Experiment 3, 543 inseminated heifers were similarly classified by CL function by RP 7 days after estrus. The heifers with functional CL showed higher pregnancy rate compared with the heifers with hypoplastic CL (75.2 vs. 47.9%, P < 0.0001). Finally, the CL function of 66 heifers was examined by RP on day 7 post‐estrus, and ET was performed in 49 (74.2%) heifers with functional CL. As a result, 27 (55.1%) of them became pregnant. Taken together, these results reconfirm that RP on day 7 after estrus is useful for selection of heifers with functional CL.     

Seasonal variations in the incidence of pine wilt and infestation by its vector, Monochamus alternatus, near the northern limit of the disease in Japan

During the last few decades, pine wilt disease has spread to cool-climate regions in Japan and, more recently, the potential risk of it spreading into the European midwest has also become a concern. In a coastal pine stand (84.7?ha) in Akita, near the northern limit of pine wilt disease in Japan, we investigated seasonal variations in the incidence of damage caused by the disease to trees and oviposition by the disease’s insect vector, Monochamus alternatus, during a two-year period. Foliage discoloration occurred throughout each year, and its seasonal variation showed a bimodal pattern in Pinus thunbergii (a higher peak in May–June and a smaller peak in October) and a clear peak in June in P. densiflora, which differed from the patterns in seasonal variation seen for warm-climate regions. Oviposition scars by M. alternatus were found in 40–45% of the trees damaged each year. The percentage of trees that had oviposition scars was higher in P. thunbergii than in P. densiflora. This appeared to reflect the difference in seasonal discoloration pattern between the two species. Analysis of the oviposition risk showed that trees that exhibited discoloration starting between July and October had a significantly higher risk or significantly higher oviposition scar densities, particularly for those that became discolored between August and September (2.5–14.6-fold higher risk than during other months). Oviposition scar densities per damaged tree were similar within the period of higher oviposition risk. Considering both oviposition risks and scar densities, we concluded that trees with discoloration that become apparent between July and October are important targets for preventing the spread of pine wilt disease in Akita.