Prolonged survival of mouse epididymal spermatozoa stored at room temperature

Summary: The viability and fertility of isolated mouse epididymal spermatozoa kept for up to 7 days at various temperatures (4°C, 22°C, and 37°C) were determined. Spermatozoa kept for 3 days at 22°C were still active, while those kept at 37°C or 4°C exhibited great reduction in motility within 2 days after isolation. In vitro fertilizing abilities of spermatozoa left for 0, 1, 2, and 3 days at 22°C were 69.2, 32.5, 9.5, and 4.9%, respectively, when the cleavage rate to two‐cell stage was examined. Transfer of two‐cell embryos produced in vitro with spermatozoa left for 1, 2, and 3 days at 22°C resulted in production of fetuses with efficiencies of respectively 30.2, 11.5, and 16.7%, which were lower (63.3%) than that of embryos derived from in vitro fertilization with fresh spermatozoa. These findings indicate that spermatozoa kept for up to 3 days at 22°C can fertilize oocytes, although at relatively low efficiency. genesis 31:147–155, 2001. © 2001 Wiley‐Liss, Inc.     

Detection of quantitative trait loci causing abnormal spermatogenesis and reduced testis weight in the small testis (Smt) mutant mouse.

The small testis (Smt) mutant mouse is characterized by a small testis of one third to one half the size of a normal testis, and its spermatogenesis is mostly arrested at early stages of meiosis, although a small number of spermatocytes at the late prophase of meiosis and a few spermatids can sometimes be seen. We performed quantitative trait locus (QTL) analysis of these spermatogenic traits and testis weight using 221 F2 males obtained from a cross between Smt and MOM (Mus musculus molossinus) mice. At the genome-wide 5% level, we detected two QTLs affecting meiosis on chromosomes 4 and 13, and two QTLs for paired testis weight as a percentage of body weight on chromosomes 4 and X. In addition, we found several QTLs for degenerated germ cells and multinuclear giant cells on chromosomes 4, 7 and 13. Interestingly, for cell degeneration, the QTL on chromosome 13 interacted epistatically with the QTL on chromosome 4. These results reveal polygenic participation in the abnormal spermatogenesis and small testis size in the Smt mutant.     

Three-Dimensional Electron Microscopy of the Photosystem II Core Complex

A three-dimensional image of the spinach photosystem II core complex composed of CP47, D1, D2, cytochromeb-559, andpsbI gene product was reconstructed at 20-Å resolution from the two-dimensional crystals negatively stained with phosphotungstate. Confirming the previous proposal, the crystal had ap22₁2₁symmetry. One PSII core complex was measured to be 80 × 80 Å in the membrane plane and 88 Å normal to it. The mass distribution was asymmetric about the lipid bilayer, consistent with predictions from the amino acid sequences. The lumenal mass consisted of three domains forming a characteristic triangular platform with another domain on top of it. Three stromal domains were smaller and linearly arranged. Due to strong stain exclusion in the hydrophobic core part of the lipid bilayer, the transmembrane region appeared to be imaged with a reversed contrast. Inverting the contrast resulted in a reasonable density distribution for that part. Thus, though the information on the transmembrane region is limited, the domain structure of the PSII core complex was revealed and allowed us to propose a model for the arrangement of subunits in the PSII core complex.     

Immunohistochemical distribution of epimorphin in human and mouse tissues

A novel protein epimorphin has been identified as a mesenchymal signal factor. We reported previously ubiquitous expression of epimorphin in normal skin and a significant increased expression in diseased human skin. The present immunofluorescence study was conducted to determine systematically the distribution of epimorphin in adult human organs with an anti-epimorphin monoclonal antibody. Epimorphin was found to be widely distributed in all human organs examined. It was present in the connective tissue adjacent to or around various epithelial tissues, muscles and vessels. In particular, strong staining was present on the endomysium of muscles, the adventitia of blood vessels, along the sinusoidal lining of hepatocytes and connective tissue around epithelial cells, exocrine and endocrine glands. The results suggest that epimorphin may play a key role in maintaining normal tissue structure and interaction between mesenchymal tissue and epithelial tissue in vivo. ©; 1998 Chapman & Hall     

Concluding remarks of Autonomous Biological Systems (ABS) experiments.

Team efforts for analysis on the Autonomous Biological Systems (ABS) space experiments are summarized here to conclude scientific findings, and to scope the extended studies in future. From the three experiments on Space Shuttle and Space Station Mir, a closed ecological system modeled by the ABS was verified to be capable of sustaining its members of animals and plants under space environment for a period of several months. The animals successfully completed their life cycle in space during the course of these experiments, this was the first time that the life cycle of higher organisms had been completed in space and ecological system. Importance of gravity for ecology was proven at the same time. Gravity is a dominant factor for ecology by formulating spatial patterns and distribution of members of ecological system. Under microgravity, the fate of ecological system was found highly sensitive against the variation of environmental factor, such as light illumination cycle.     

The role of major histocompatibility complex expression on head and neck cancer cells in the induction of autologous cytotoxic T lymphocytes

Using head and neck tumors, we studied the role of HLA class I and DR antigens on tumor cells in cytotoxic T lymphocyte (CTL) induction. Expression of major histocompatibility complex (MHC) antigens was investigated by two-color flow cytometry analysis and for this study we used the tumor cells, over 50% of which expressed both HLA class I and DR antigens on their surface. In seven cases, tumor cells were divided into three groups according to the specificity of monoclonal antibodies (mAb) to MHC to study the role of MHC antigens on tumor cells in CTL induction: one was not blocked (MHC double-positive tumor), a second was blocked by anti-class I mAb (class-Ingative DR-positive tumor) and third was blocked by anti-DR mAb (class-I-positive DR-negative tumor). Subsequently, these tumors were used to stimulate an autologous mixed lymphocyte/tumor cell culture for 5 days (MLTC) followed by further cultivation with interleukin-2 for 12 days. The induced autologous tumor killer cells were most cytotoxic when non-treated tumors, which consist mainly of cells that are both HLA-class I and DR-positive, were used as stimulator cells. When the tumor cells blocked by anti-DR mAb were used as stimulators, autologous tumor killer activity was lower than that induced by tumor cells blocked by anti-class-I mAb. Moreover, cytolysis by autologous tumor killer cells induced by stimulation of non-treated tumor cells was blocked during the effector phase, 26.6%–42.3% and 32.7%–53.8% by anti-class-I and anti-DR mAb respectively, suggesting that majority of the autologous tumor killer cells are MHC-restricted CD8⁺ or CD4⁺ CTL. These results suggest that both MHC class I and class II antigens on head and neck tumor cells play a critical role in inducing CTL.     

Establishment of somatic hybrid cell lines between Zea mays L. (maize) and Triticum sect,trititrigia MacKey (trititrigia)

Summary Somatic hybrid cell lines were constructed by the fusion of protoplasts isolated from cell suspensions of Zea mays L. (maize, 2n = 20) and Triticum sect, trititrigia MacKey (trititrigia, 2n = 35), a perennial hybrid of T. durum Desf. and Elytrigia intermedium (Host) Nevski. Iodoacetamide-inactivated protoplasts of maize were fused with trititrigia protoplasts, which were sensitive to the PEG/DMSO fusion treatment at high pH and high calcium. Based on physiological complementation, approximately 0.002% of the total protoplasts cultured following fusion treatment developed into cell colonies, and 79 lines of them, almost a half, were singled out and subcultured. Among the subcultured lines three were, in comparison with the parents, identified as somatic hybrids by their coupled XbaI restriction patterns of total DNAs probed with the ribosomal DNA of rice. Southern analysis of the digested total DNAs with a mitochondrial gene, atpA., from pea, or a chloroplast gene, trnK, from rice, revealed that all the hybrids carried only the organellar DNAs of trititrigia, which excluded the possibilities of a chimeric callus or any DNA contamination. Cytogenetically, one hybrid was mixoploid with a 2n of 46–67 in which chromosomal endoreduplication, characterized by the appearance of diplochromosomes, was occasionally observed. Its hybridity was reconfirmed by the fact that it bore the satellite chromosomes of both maize and trititrigia, which were distinguishable from each other by size. In contrast, the other two hybrids were aneuploids. The potential of gene transfer between Zea and Triticum species was thus conclusively established.     

Some ultrastructural observations on the mode of association of microtubules with the plasmalemma in epidermal tendon cells of the river crab

Summary— The ultrastructural aspects of the association of microtubules (MTs) with the plasmalemma in epidermal tendon cells of the river crab, Polamon dehaani, were studied by thin-section electron microscopy combined with detergent treatment. In the tendon cell, MTs were linked laterally by anchoring filaments to the plasmalemma via a submembranous electron-dense layer called the plasmalemmal undercoat. To further clarify how such anchoring filaments are spatially related to the plasmalemma through the undercoat, we carefully examined and compared thin-section images obtained from various specimen preparations using saponin and Triton X-100. When the tissues were treated with saponin or Triton, electron-dense materials in the undercoat were extracted in varying degrees to expose internal substructures. The undercoat appeared to show a two-layer organization, the inner and outer layers. In more extracted samples, filamentous networks became prominent in the outer layer. Anchoring filaments were seen to attach to such filamentous networks, which in turn were linked to the plasmalemma proper. Thus, it may be reasonable to consider that the filamentous network constitutes the core structure of the plasmalemmal undercoat which is structurally reinforced by extractable electron-dense materials.     

A novel growth-inducible gene that encodes a protein with a conserved cold-shock domain.

We have isolated a cDNA that encodes a novel member of the Y-box binding protein family, termed as RYB-a (Rat Y-box Binding protein-a). RYB-a is a 31 kDa protein that contains a conserved cold-shock domain and an amino acid alignment similar to those of charge zipper proteins. Expression of RYB-a mRNA was highly abundant in the skeletal muscle, spleen, and fetal liver. The expression is very low in new-born and adult livers, suggesting its expression is under developmental regulation. In addition, the expression of RYB-a mRNA was induced in the liver during regeneration and by stimulation of quiescent fibroblast cells with serum. Induction in the fibroblasts was inhibited by treating the cell with a specific tyrosine kinase inhibitor, genistein or by detachment of cell-adhesion. Since both treatments are known to inhibit G1 cells to enter S phase, RYB-a gene is thought to be a member of growth-inducible genes.     

The 5'-terminal sequence of TMV RNA. Question on the polymorphism found in vulgare strain

The complete nucleotide sequence of TMV RNA (common strain) reported in [Proc. Natl. Acad. Sci. USA (1982) 79, 5818] its 5'-end to be represented by two variants which differed in length. We have tested that result and sequenced the 5'-terminal regions of two strains of TMV RNA (common strain OM and tomato strain L) using cloned cDNA copies. The results showed that the 5'-terminal region of the TMV genome is not polymorphic and that one of the two variants cited above represents a tomato strain but not the common strain.