Quantitation of Sertoli cell-germ cell desmosome gap junctions in relation to meiotic divisions in the male rat.

Desmosome-gap (D-G) junctions were quantified in relation to germ cell meiosis in the male, specifically to test the hypothesis that the loss of these junctions is related to successful passage of cells through diplotene phase of Meiosis I and the two cytokineses that follow. Such a hypothesis has been proposed as the cause for the resumption of meiosis that occurs prior to ovulation in the female. D-G junctions were quantified in pachytene spermatocytes (stage XII), diplotene spermatocytes (stage XII), secondary spermatocytes (stage XIV) and step 1 spermatids (stage I). These were referred to as the cells of interest as compared with spermatocytes (zygotene spermatocytes, zygotene spermatocytes, pachytene spermatocytes, pachytene spermatocytes) in the same stages, respectively, that served as controls termed control cells. Since gap junctions are not easily recognized in the average sectioned profile of a desmosome-gap junction, only the desmosomal component was quantified. The data were expressed as both numbers and length of junctions per tubule, per cell profile and per unit lineal membrane length to overcome errors inherent in the methodologies utilized. There was no indication that numbers of junctions changed specifically in the cells of interest after passage through diplotene suggesting that these junctions do not have a comparable role in meiotic continuance in the male as proposed for the female. Interestingly, the control cells always showed greater numbers and length of junctions than the cells of interest suggesting that junction may relate more to the period of initiation of meiosis than to its continuance.     

Protection against acute paraquat toxicity by ambroxol

An expression vector for G-CSF, pASLB3-3, was constructed and introduced into Namalwa KJM-1 cells (Hosoi et al., 1988), and cells resistant to 100 nM of methotrexate (MTX) were obtained. Among them, the highest producer, clone SC57, was selected and the productivity of this clone was further characterized. The maximal production of G-CSF was at the most 1.8 g/ml/day using a 25 cm² tissue culture flask, even though the cell number was above 7×10⁵ cells/ml. The limiting factors at high density were analyzed as the deficiency of nutrients, such as glucose, cysteine and serine, and pH control. The depression of specific G-CSF productivity per cell under the batch culture conditions was overcome by using a perfusion culture system, Biofermenter^TM (Sato, 1983) with modifications of nutrients supplementation by a dialysis membrane and/or dissolved oxygen (DO) supplementation by microsilicone fibers. ITPSGF medium was modified to elevate concentrations of amino acids and glucose by 2.0- and 2.5-times, respectively. Under the control of pH at 7.4 and DO at 3 ppm, the specific G-CSF productivity was not depressed even at high cell density (above 1×10⁷ cells/ml), and the amount of G-CSF reached 41 g/ml. These results indicated the possibility of finding the optimum culture conditions for the production of recombinant proteins by Namalwa KJM-1 cells.Abbreviations ABTS 2,2-Azino-di-(3-ethylbenzothiazoline)-6-sulfonic acid - BSA Bovine Serum Albumin - BSA-PBS Phosphate-buffered Saline without Ca²⁺ and Mg²⁺ containing Bovine Serum Albumin - dhfr Dihydrofolate Reductase - DO Dissolved Oxygen - G-CSF Granulocyte Colony-stimulating Factor - HEPES 4-(2-Hydroxyethyl)-1-piperazineethansulfonic Acid - IFN Interferon - MTX Methotrexate - PBS(-) Phosphate-buffered saline without Ca²⁺ and Mg²⁺ - Tween-PBS Phosphate-buffered saline without Ca²⁺ and Mg²⁺ containing 0.05% of Tween 20     

Differentiation between the somatostatin inhibition and the post-somatostatin rebound observed on growth hormone secretion in vitro

A rebound in growth hormone secretion following somatostatin treatment has been shown in several systems where somatostatin suppresses secretion of the hormone. We have developed an in vitro system in which isolated and cultured pituitary cells were perfused after mild trypsinization. After washing, these cells retained their sensitivity and secreted growth hormone (GH) in response to physiological activators (norepinephrine, dopamine, serotonin) or inhibitors (somatostatin) as well as pharmacological activators (PGE2). The variation in GH secretion occurred within a minute after commencement of the infusion and was as rapidly reversible and repeatable minutes later. During somatostatin infusion the GH secretion was not totally suppressed (residual secretion (mean +/- S.D.) 34 +/- 7%). After the infusion a rapid rebound in GH secretion occurred, reaching levels in excess of the pretreatment value of 138 +/- 13%. This rebound effect occurred at doses higher than (10(-10)M) but not at lower doses, even when significant inhibition was observed. The inhibitory effect is of greater magnitude than the rebound effect (rebound = inhibition X 57 +/- 7% (mean +/- S.D.)). Furthermore, rebound was not enhanced by prolongation of somatostatin infusion. These latter results indicate that the rebound in secretion cannot be explained on the sole basis of storage of intracellular GH during somatostatin infusion and in fact suggest the involvement of a process of GH degradation and/or an inhibition of GH synthesis.     

Influence of clay minerals on sorption of bacteriolytic enzymes

Myxobacteria presumably produce extracellular bacteriolytic enzymes when they are growing in soil. In order to study their ecological significance, adsorption experiments were performed with lytic enzymes produced byMyxococcus virescens in casitone media. Different soils as well as montmorillonite and kaolinite can rapidly adsorb the bacteriolytic but not the proteolytic enzymes. About 1 gm of montmorillonite per liter of cell-free culture solution is enough for the adsorption of 97% of the bacteriolytic enzymes. The adsorption per unit weight is about 100 times greater on montmorillonite than on kaolinite. About 40% of the adsorbed enzymes can be eluted with solutions of high pH or high ionic strength. The only desorbed bacteriolytic enzyme is the alanyl-∈-N-lysine endopeptidase.     

JQ-1 ameliorates schistosomiasis liver granuloma in mice by suppressing male and female reproductive systems and egg development of Schistosoma japonicum

Schistosomiasis is a serious and widespread parasitic disease caused by infection with Schistosoma. Because the parasite’s eggs are primarily responsible for schistosomiasis dissemination and pathogenesis, inhibiting egg production is a potential approach to control the spread and severity of the disease. The bromodomain and extra-terminal (BET) proteins represent promising targets for the development of epigenetic drugs against Schistosoma. JQ-1 is a selective inhibitor of the BET protein family. In the present study, JQ-1 was applied to S. japonicum in vitro. By using laser confocal scanning microscopy and EdU incorporation assays, we showed that application of JQ-1 to worms in vitro affected egg laying and the development of both the male and female reproductive systems. JQ-1 also inhibited the expression of the reproductive-related genes SjPlk1 and SjNanos1 in S. japonicum. Mice infected with S. japonicum were treated with JQ-1 during egg granuloma formation. JQ-1 treatment significantly reduced the size of the liver granulomas and levels of serum alanine aminotransferase and aspartate aminotransferase in mice and suppressed both egg laying and the development of male and female S. japonicum reproductive systems in vivo. Moreover, the mRNA expression levels of some proinflammatory cytokines were decreased in the parasites. Our findings suggest that JQ-1 treatment attenuates S. japonicum egg–induced hepatic granuloma due at least in part to suppressing the development of the reproductive system and egg production of S. japonicum. These findings further suggest that JQ-1 or other BET inhibitors warrant additional study as a new approach for the treatment or prevention of schistosomiasis.     

Staining of Some Specific Regions of Human Chromosomes,particularly the Secondary Constriction of No. 9

SEVERAL procedures have been described recently which produce specific patterns of differential staining in human chromosomes^1–9. Techniques which involve DNA denaturation and reannealing reveal deeply stained areas on centromere and secondary constriction regions which have been equated with constitutive heterochromatin⁹.     

The mode of action of thidiazuron: auxins,indoleamines, and ion channels in the regeneration of <Emphasis Type="Italic">Echinacea purpurea</Emphasis> L.

The biochemical mechanisms underlying thidiazuron (TDZ)-induced regeneration in plant cells have not been clearly elucidated. Exposure of leaf explants of Echinacea purpurea to a medium containing TDZ results in undifferentiated cell proliferation and differentiated growth as mixed shoot organogenesis and somatic embryogenesis. The current studies were undertaken to determine the potential roles of auxin, indoleamines, and ion signaling in the dedifferentiation and redifferentiation of plant cells. E. purpurea leaf explants were found to contain auxin and the related indoleamine neurotransmitters, melatonin, and serotonin. The levels of these endogenous indoleamines were increased by exposure to TDZ associated with the induction of regeneration. The auxin-transport inhibitor 2,3,5-triiodobenzoic acid and auxin action inhibitor, p-chlorophenoxyisobutyric acid decreased the TDZ-induced regeneration but increased concentrations of endogenous serotonin and melatonin. As well, inhibitors of calcium and sodium transport significantly reduced TDZ-induced morphogenesis while increasing endogenous indoleamine content. These data indicate that TDZ-induced regeneration is the manifestation of a metabolic cascade that includes an initial signaling event, accumulation, and transport of endogenous plant signals such as auxin and melatonin, a system of secondary messengers, and a concurrent stress response.