Human-type ABO blood groups as genetic markers for the management of a squirrel monkey (Saimiri sciureus) breeding colony

The human-type ABO blood groups were determined for 94 families of the squirrel monkey which included 151 animals. Four phenotypes of ABO blood groups (A, B, AB, and O) were detected. Family analysis revealed that the human-type ABO blood groups in this species were governed by three alleles, codominantA andB and silentO. There were intraspecific differences in the distribution of phenotypes and gene frequency among three populations imported by different routes at different times. The usefulness of ABO blood groups for defining the genetic variability of a squirrel monkey breeding colony through successive generations is discussed on the basis of the difference in distribution of ABO blood groups between wild-originated parental and its first colony-born populations.     

Full-term development of in vitro-matured, vitrified and fertilized bovine oocytes

In vitro-matured bovine oocytes were vitrified in a mixture of 2 M-dimethyl sulphoxide (DMSO), 1 M-acetamide and 3 M-propylene glycol dissolved in mTCM199. After vitrification and thawing, the oocytes were exposed to 2-0.1M-sucrose solution in 1 or 12 steps to remove the cryoprotectants. Then the oocytes were fertilized in vitro and co-cultured with a monolayer of cumulus cells for 7 days. Nine of 88 inseminated oocytes developed to the blastocyst stage. Three blastocysts were transferred to 3 recipients, resulting in 2 pregnancies.     

Developmental changes in the incidence of chromosome anomalies of bovine embryos fertilized in vitro.

In total, 196 two- to 32-cell bovine embryo and 104 blastocysts were obtained by the in vitro fertilization of follicular oocytes matured in vitro, and 15 blastocysts fertilized in vivo were used. Chromosomal anomalies in these embryos and the inner cell mass (ICM) separated immunologically were investigated. Chromosomal anomalies were observed in 12.1% (5/41) of 2-cell embryos, 20.0-36.4% of 4- to 16-cell embryos, 7.1% (1/14) of 17- to 32-cell embryos, 44.2% (15/34) of blastocysts, and 18.6% (13/70) of ICM cells derived from in vitro fertilization. These anomalies were mainly 3N and 4N at 2-cell stage, 1N and 1N/2N at 4- to 32-cell stages, and 2N/4N in blastocysts and in their ICM cells. Chromosomal anomalies of blastocysts from in vivo fertilization and their ICM were observed in 20.0% (1/5) of blastocysts and 33.3% (3/9) of ICM cells and these compositions were mainly 2N/4N. These results indicate that the abnormalities at early and blastocyst stages of embryos derived from in vitro fertilization were caused by abnormal fertilization in vitro and abnormal cleavage, respectively. Furthermore, a definite location of the chromosomal anomalies was observed in the trophectoderm of blastocysts derived from in vitro fertilization.     

Induction of apoptosis of T cells by infecting mice with murine cytomegalovirus.

Cytomegalovirus (CMV) is associated with several lymphocyte dysfunctions, but the precise mechanisms of the dysfunctions are still unclear. To elucidate the mechanisms, a cell cycle-DNA content analysis was performed on splenic T cells of murine CMV (MCMV)-infected BALB/c mice. T cells from mice infected with 3 x 10(3) PFU of MCMV contained a higher percentage of hypodiploid nuclei after 12 or 24 h of culture than those from naive mice. T cells from infected mice also contained a larger amount of fragmented DNA. Taken together, these results suggested that infection with MCMV induced the apoptotic cell death of T cells. This induction of apoptosis accounted for the dysfunction of lymphocytes, at least partially. Flow cytometric analysis showed that T cells as well as B cells from MCMV-infected mice expressed an augmented level of Fas antigen, an apoptosis-associated cell surface molecule, which might be the cause of the apoptosis of cells. T cells from MCMV-infected C57BL/6-lpr/lpr mice with mutations at the lpr/fas locus, however, also showed a substantial level of apoptosis, which was reproducibly lower than that seen in C57BL/6 mice. Therefore, it was suggested that the Fas-mediated pathway contributed to but was not sufficient for the induction of apoptosis and that mechanisms other than the Fas-associated pathway were also involved in the induction of apoptosis.     

Expression of calcium-binding protein regucalcin mRNA in fetal rat liver is stimulated by calcium administration

The expression of hepatic calcium-binding protein regucalcin mRNA in fetal rats was investigated. The alteration in regucalcin mRNA levels was analyzed by Northern blotting using liver regucalcin cDNA (0.9 kb with complete open reading frame). Hepatic regucalcin mRNA levels were progressively increased with fetal development; the mRNA was clearly expressed at 15 and 21 days of pregnancy but only slightly at the 8 days. Meanwhile, -actin mRNA levels in the fetal liver were remarkable at 8 and 15 days of pregnancy. The fetal liver regucalcin mRNA levels at 15 days of pregnancy were significantly decreased by overnight-fasting of maternal rats. The oral administration of calcium chloride (50 mg Ca/100 g body weight) to maternal rats at 15 days of pregnancy caused a remarkable elevation (about 2 fold) of regucalcin mRNA levels in the fetal liver; this increase was seen 60 and 180 min after the calcium administration. After birth, regucalcin mRNA was increasingly expressed in the livers of newborn and weanling rats, while hepatic -actin mRNA expression was not appreciably altered with increasing ages. These findings demonstrate that the expression of hepatic regucalcin mRNA is increased with fetal development, and that the gene expression may be stimulated by the ingestion of dietary calcium.     

Gender preselection in cattle with intracytoplasmically injected, flow cytometrically sorted sperm heads

We investigated the development to the blastocyst and subsequent live-offspring stages of in vitro-matured bovine oocytes intracytoplasmically injected with flow cytometrically sorted bull sperm heads. Bull sperm heads, prepared by ultrasound sonication, were distinguished and sorted on the basis of their relative DNA contents using a flow cytometer/cell sorter modified for sorting sperm. By fluorescence in situ hybridization, the proportion of sperm confirmed as having Y specific DNA in the fraction sorted for the Y sperm was 82%. Injection with single sorted sperm heads of in vitro-matured oocytes (cultured for 24 h) resulted in 46.6% cleavage and 6.9% blastocyst development rates. Embryo transfer of 48 blastocysts (Days 7-8) to recipients (one per recipient) resulted in 20.8% pregnancy and 20.8% normal live offspring production rates. The birth of 8 male and 2 female calves represents an 80% sex preselection accuracy rate.     

Disulfide formation in bovine zona pellucida glycoproteins during fertilization: evidence for the involvement of cystine cross-linkages in hardening of the zona pellucida

The time for solubilization of the bovine zona pellucida in a hypotonic buffer containing 5% (v/v) beta-mercaptoethanol and 7 mol urea l-1 increased by 10% after fertilization. Coupling with a specific fluorescent thiol probe, monobromobimane (mBBr), was markedly greater in the zona pellucida of ovarian eggs compared with fertilized eggs, indicating that the cysteine residues in the zona pellucida of unfertilized eggs are oxidized to cystines during fertilization. After endo-beta-galactosidase digestion to remove N-acetyllactosamine repeats of the carbohydrate chains, three zona pellucida glycoproteins (ZPA, ZPB and ZPC) coupled with the fluorescent bimane groups were fractionated efficiently by reverse-phase HPLC. Estimation of bimane groups in the three components and SDS-PAGE revealed that intramolecular disulfide bonds in ZPA and intra- and intermolecular disulfide bonds in ZPB were formed during fertilization, but oxidation of cysteine residues in ZPC was low. Specific proteolysis of ZPA during fertilization was also observed. These results indicate that the formation of disulfide linkages together with specific proteolysis result in the construction of a rigid zona pellucida structure, which is responsible for hardening of the zona pellucida.     

Involvement of abscisic acid in bulb dormancy of Allium wakegi Araki. II. A comparison between dormant and nondormant cultivars

The content of abscisic acid (ABA) in bulbs of two Allium wakegi Araki cultivars, Kiharabansei No. 1 (dormant type) and Ginoza (nondormant type), was similar and changed similarly during the development and storage of the bulbs. It increased during bulb development, reached a maximum shortly after bulb harvesting, and gradually decreased during bulb storage. The bulbs of Kiharabansei No. 1 showed dormancy correlated with the change in ABA content, but those of Ginoza did not show significant dormancy throughout the experimental period. The ABA content in the buds of dormant bulbs of Kiharabansei No. 1 did not change after planting of bulbs, but that of nondormant bulbs of Ginoza planted on the same day rapidly decreased after planting. Application of ABA to bulbs delayed sprouting of both cultivars, but dormant bulbs of Kiharabansei No. 1 had higher sensitivity to ABA than the bulbs of Ginoza or the bulbs of Kiharabansei No. 1 partly released from dormancy. These results suggest that the decrease in the ABA content after planting (watering) and low sensitivity to ABA are correlated with the nondormancy of Ginoza.     

Correction to: Rhamphocottus nagaakii (Cottoidea: Rhamphocottidae), a new species of grunt sculpin from the northwestern Pacific,with notes on the phylogeography of the genus Rhamphocottus

A new species of grunt sculpin, Rhamphocottus nagaakii inhabiting the northwestern Pacific, previously identified as Rhamphocottus richardsonii Günther 1874, is described based on genetic evidence and morphological differences. The new species can be distinguished based on morphometric characters related to the head, including head length [45.3–54.6% of standard length (SL)], postorbital head length (18.8–25.5% SL) and the length of pectoral-fin base (15.8–20.7% SL), which are smaller than in R. richardsonii (53.6–60.5% SL, 26.2–31.7% SL, and 19.5–25.2% SL, respectively). Genetic differences between two species markedly exceed levels for intra-specific differences. Rhamphocottus nagaakii is considered to have arisen from a common ancestor of the two species, which probably inhabited somewhere the North Pacific Rim around the Aleutian Archipelago. During a period of cooling in the Pliocene or the Miocene, R. nagaakii and R. richardsonii became separated to the southern regions of the northwestern and northeastern Pacific, and subsequently underwent speciation.