Effect of pH on giant cell formation induced by human immunodeficiency virus ( HIV )

The rate of multinucleated giant cell formation by the fusion of HIV chronically infected human T-cells (MOLT-4/HIV) and HIV uninfected MOLT-4 cells was examined under various pH conditions. The number of giant cells formed under the different pH conditions was quantitatively monitored by "MULTISIZER". The rate of giant cell formation was significantly less at the pH lower than 6 but not influenced at higher pH. Plaque assay under various pH conditions revealed that inhibition of giant-cell formation at lower pH was not due to the influence over the recognition between gp120 of HIV and CD4 molecules.     

Mapping of a gene determining tuberous sclerosis to human chromosome 11q14-11q23

Tuberous sclerosis (TSC) is a dominantly inherited disorder characterized by hamartomas and hamartias in one or more organs, most often in skin, brain, and kidneys. Analysis of the basic genetic defect in tuberous sclerosis would be greatly expedited by definitive determination of the chromosomal location of the TSC gene or genes. We have carried out genetic linkage studies in 15 TSC families, using 34 polymorphic markers including protein markers and DNA markers. Pairwise lod scores were calculated using LIPED, and multipoint analyses were carried out using MENDEL. In the pairwise linkage analysis, using a penetrance value of 90%, a significant positive lod score was obtained with MCT128.1 (D11S144), 11q22-11q23, Zmax 3.26 at theta = 0.08. The tyrosinase probe TYR (11q14-11q22) gave a maximum lod score of 2.88 at theta = 0. In the multipoint analyses the most likely order is (TYR,TSC)-MCT128.1-HHH172. Homogeneity analysis was carried out using the USERM9 subprogram of MENDEL, which conducts the admixture test of C. Smith (1963, Ann. Hum. Genet. 27: 175-182). This test provided no evidence for genetic heterogeneity (that is, non-11-linked families) in this data set.     

In vivo and in vitro synergistic antitumor effect of interleukin-2-cultured tumor-bearer spleen cells and immune fresh spleen cells

Summary The synergistic antitumor effect of interleukin-2(IL-2)-cultured tumor-bearer spleen cells (cultured lymphocytes) and immune fresh spleen cells was examined. Tumor-bearer cultured lymphocytes were obtained by culturing BALB/c spleen cells from syngeneic MOPC104E-tumor-bearing mice for 11 days with crude IL-2 and a soluble tumor extract. These cultured lymphocytes had weak antitumor activity when transferred i.p. into tumor-bearing mice that had been inoculated i.p. with 10⁵ tumor cells 5 days previously. Immune fresh spleen cells, obtained from mice in complete remission after the treatment with cyclophosphamide, also had weak antitumor activity when transferred at the same schedule. The cultured cells and the fresh cells, mixed together before transfer, significantly augmented the therapeutic effect. At least 1×10⁷ tumor-bearer cultured lymphocytes and 4×10⁷ immune cells were needed for the synergistic effect. A tumor-specific combination was needed for both cultured and fresh cells. The effective subpopulation of tumor-bearer cultured lymphocytes was a cytotoxic one from an Lyt2⁺ precursor, and that of the immune fresh spleen cells was noncytotoxic, Lytl⁺ and Lyt2⁺ T-cells.A similar synergistic effect was also observed during in vitro coculture of tumor-bearer and immune cells. Cytotoxicity, as assessed by the ⁵¹Cr-release test, of tumor-bearer IL-2-cultured lymphocytes was maintained most effectively after 3 or 4 days of culture without IL-2 when the lymphocytes were cocultured with immune fresh spleen cells and tumor cells.     

Interconversion of polyamines in wild-type strains and mutants of yeasts and the effects of polyamines on their growth

Yeasts of wild-type strains, such as Saccharomyces cerevisiae, Schizosaccharomyces pombe and Candida albicans were shown to have the ability to form aminopropylcadaverine and aminopropylhomospermidine from cadaverine and homospermidine, respectively. A polyamine autotroph S. cerevisiae 179-5, which lacks ornithine decarboxylase, produced both aminopropylcadaverine and aminopropylhomospermidine, while another mutant S. cerevisiae Y 260 A, which lacks spermine synthase, formed only aminopropylcadaverine. Naturally-occurring triamines and tetraamines except norspermidine and norspermine stimulated the growth of S. cerevisiae 179-5. All the six aliphatic diamines with carbon chain length ranging from one to six were effective in activating the growth of S. cerevisiae 179-5, though all of them were not converted to either triamines or tetraamines.     

Hormonal induction of all stages of spermatogenesis in germ-somatic cell coculture from immature Japanese eel testis

In cultivated male eel, spermatogonia are the only germ cells present in testis. Our previous studies using an organ culture system have shown that gonadotropin and 11-ketotestosterone (11-KT, a potent androgen in teleost fishes) can induce all stages of spermatogenesis in vitro. for detailed investigation of the control mechanisms of spermatogenesis, especially of the interaction between germ cells and testicular somatic cells during 11-KT-induced spermatogenesis in vitro, we have established a new culture system in which germ cells and somatic cells are cocultured after they are aggregated into pellets by centrifugation. Germ cells (spermatogonia) and somatic cells (mainly Sertoli cells) were isolated from immature eel testis. Coculture of the isolated germ cells and somatic cells without forming aggregation did not induce spermatogenesis, even in the presence of 11-KT. In contrast, when isolated germ cells and somatic cells were formed into pellets by centrifugation and were then cultured with 11-KT for 30 days, the entire process of spermatogenesis from premitotic spermatogonia to spermatozoa was induced. However, in the absence of 11-KT in the culture medium spermatogenesis was not induced, even when germ cell and somatic cells were aggregated. These results demonstrate that physical contact of germ cells to Sertoli cells is required for inducing spermatogenesis in response to 11-KT.     

Promotion of Cocklebur Seed Germination by Allyl, Sulfur and Cyanogenic Compounds

The effects of allyl, sulfur and cyanogenic compounds on thegermination of upper cocklebur (Xanthium pennsylvanicum Wallr.)seeds were examined. Mercaptoethanol and methylmercaptan aswell as KCN, substrates for rßcyanoalanine synthase(CAS), and H₂S and thiocyanate, the products of the CAS catalyzingreaction, were effective in promoting germination, suggestingthe involvement of CAS in germination. Most of allyl compounds, especially allylthiourea, as well asethylene which activated CAS [Hasegawa et al. (1994) Physiol.Plant. 91: 141], promoted the germination in an abnormal typewhich occurred by the predominant growth of cotyledons as didC₂H₄ [Katoh and Esashi (1975) Plant Cell Physiol. 16: 687].However, they failed to activate CAS unlike ethylene, and toliberate free ethylene during an incubation period. It was thuspossible that an C₂H₄-like double bond within allyl compoundscan act to promote seed germination. (Received June 10, 1996; Accepted August 21, 1996)     

In situ hybridization to metaphase chromosomes in six species ofPhaseolus andVigna using ribosomal DNA as the probe

In situ hybridization with a biotin-labeled rice ribosomal DNA (rDNA) probe to the somatic metaphase chromosomes of six species ofPhaseolus andVigna (P. angularis, P. calcaratus, P. coccineus, P. vulgaris, V. sesquipedalis andV. sinensis) was done to determine the sites of rDNA. Hybridization signals were present in the terminal and subterminal chromosome regions of each of the six species. The number of rDNA sites was two inP. angularis andP. calcaratus, four inP. coccineus andP. vulgaris, and six inV. sesquipedalis andV. sinensis.     

W-heterochromatin of chicken; its unusual DNA components,late replication,and chromatin structure

About 65% of DNA in the chicken W chromosome has been shown to consist ofXhoI andEcoRI family repetitive sequences. These sequences showed remarkable delay in the electrophoretic mobility at low temperature on a polyacrylamide gel. Three dimensional structures of the 0.7-kbXhoI and the 1.2-kbEcoRI family repeating units were estimated to be irregular solenoids using a computer program based on wedge angles of all the 16 dinucleotide steps. Fluorescencein situ hybridization demonstrated that these two family sequences were localized in a major heterochromatic body in an interphase nucleus. Incorporation of bromodeoxyuridine into the W chromosome in the synchronous culture of MSB-1 cells occurred about 1 h later than the peak of S phase. The chromatin structure formed alongXhoI andEcoRI family sequences was suggested to be different from the total chromatin or chromatin containing the β-actin gene sequence in that the linker DNA lengths of the former were significantly longer. Fractionation of theHaeIII-digested MSB-1 nuclei yielded a chromatin fraction in whichXhoI family sequences were partially enriched. Several DNA-binding proteins showing higher affinity for theXhoI family sequence were present in this fraction.     

Stable transformation of cultured cells of the liverwortMarchantia polymorpha by particle bombardment

Suspension-cultured cells (A-18 line) of the liverwortMarchanta polymorpha were bombarded by a pneumatic particle gun with plasmid pCH harbouring the hygromycin phosphotransferase (HPT) gene (hpt) under the control of the cauliflower mosaic virus (CaMV) 35 S promoter and the nopaline synthase polyadenylation region. Nine weeks after bombardments, 128 hygromycin-resistant calluses were obtained from an approximate total of 7×10⁶ cells. Ten cell lines chosen randomly were analysed further. Southern blot analysis showed that all of the ten lines contain thehpt gene in the genome, demonstrating that these lines are transformants. An HPT enzyme activity assay confirmed the expression of the gene in all of the transformant lines.     

Successful expression in pollen of various plant species of in vitro synthesized mRNA introduced by particle bombardment

Gold particles coated with -glucuronidase (GUS) mRNA with a 5 cap structure that had been synthesized in vitro were introduced, by use of a pneumatic particle gun, into pollen grains of lily (Lilium longiflorum), freesia (Freesia refracta) and tulip (Tulipa gesneriana). A fluorometric assay for the GUS activity indicated that in vitro synthesized GUS mRNA introduced into these pollen cells by particle bombardment was successfully expressed. GUS activity in extracts of the bombarded lily pollen became detectable fluorometrically within 30 min after bombardment, peaked at 6 h, then gradually decreased. This activity changed as a function of the developmental stage of the pollen cell of lily.