Mapping of fish myosin light chains by two-dimensional gel electrophoresis

1. Myosins were prepared from the ordinary muscle of 16 fish species as well as from rabbit fast muscle, and light chain subunits [alkali light chains A1, A2 and DTNB (5,5'-dithio-bis-2-nitrobenzoate) light chain] were separated on two-dimensional gel electrophoresis in combination with isoelectric focusing and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. 2. A1 light chains showed mol. wts ranging from 21,000 to 22,900 and isoelectric points ranging from 4.51 to 4.62. DTNB light chains were spotted in a narrow area, with a mol. wt range of 16,800-17,600 and an isoelectric point range of 4.48-4.55. On the other hand, A2 light chains were most species-specific, with a mol. wt range of 14,000-19,500 and an isoelectric point range of 4.31-4.46. 3. It was suggested that the lower species-specificity in A1 as opposed to A2 is accounted for by the addition of an N-terminal peptide ("difference peptide") in the former. The properties and possible role of this peptide are discussed.     

Physiological polyspermy: Selection of a sperm nucleus for the development of diploid genomes in amphibians

The union between a sperm and an egg nucleus in egg fertilization is necessary to mix genetic materials to create a new diploid genome for the next generation. In most animals, only one sperm is incorporated into the egg (monospermy), but several animals exhibit physiological polyspermy in which several sperms enter the egg during normal fertilization. However, only one sperm nucleus forms the zygote nucleus with the egg nucleus, even in a polyspermic egg. The cellular and molecular mechanisms involved in the selection of sperm nuclei in the egg cytoplasm have been well investigated in urodele amphibians. The principal sperm nucleus develops a larger sperm aster and contacts the egg nucleus to form a zygote nucleus, whereas other accessory sperm nuclei are unable to approach the egg nucleus. The diploid zygote nucleus induces cleavage and participates in embryonic development, whereas the accessory sperm nuclei undergo pyknosis and degenerate. We propose several models to account for the mechanisms of the selection of one sperm nucleus and the degeneration of accessory sperm nuclei. The roles of physiological polyspermy in animal reproduction are discussed by comparison with other polyspermic species.     

Increased stearoyl-CoA desaturase index and triglyceride content in the liver of rats after a single bout of swimming exercise

Up-regulation of stearoyl-CoA desaturase (SCD) is closely related to improved insulin resistance. We investigated whether the SCD indices in tissues were influenced by a single-endurance exercise and low content of dietary medium-chain fatty acid (FA). Wistar rats were fed a long-chain (S) or medium- and long-chain FA (M) diet for 2 weeks. At the end of the experiment, the rats were further assigned to two sub-groups (sedentary, Sed; exercise, Ex). These groups were defined as S-Sed, S-Ex, M-Sed, and M-Ex. The rats in the exercise groups were subjected to swimming exercise for 4 h, and tissue samples were obtained. The exercise significantly increased the triglyceride (TG) content and SCD index only in the liver. In contrast, no such changes were apparent by intake of the M diet. A single bout of endurance exercise increased the hepatic TG content and SCD index which might be effective in protecting against insulin resistance.     

Role of MAPK phosphatase-1 in the induction of monocyte chemoattractant protein-1 during the course of adipocyte hypertrophy

Monocyte chemoattractant protein-1 (MCP-1), an important chemokine whose expression is increased during the course of obesity, plays a role in macrophage infiltration into obese adipose tissue. This study was designed to elucidate the role of mitogen-activated protein kinase (MAPK) phosphatase-1 (MKP-1) in the induction of MCP-1 during the course of adipocyte hypertrophy. We examined the time course of MKP-1 and MCP-1 mRNA expression and extracellular signal-regulated kinase (ERK) phosphorylation in the adipose tissue from mice rendered mildly obese by a short term high fat diet. We also studied the role of MKP-1 in the induction of MCP-1 in 3T3-L1 adipocytes during the course of adipocyte hypertrophy. MCP-1 mRNA expression was increased, followed by ERK activation and down-regulation of MKP-1, an inducible dual specificity phosphatase to inactivate ERK, in the adipose tissue at the early stage of obesity induced by a short term high fat diet, when macrophages are not infiltrated. Down-regulation of MKP-1 preceded ERK activation and increased production of MCP-1 in 3T3-L1 adipocytes in vitro during the course of adipocyte hypertrophy. Adenovirus-mediated restoration of MKP-1 in hypertrophied 3T3-L1 adipocytes reduced the otherwise increased ERK phosphorylation, thereby leading to the significant reduction of MCP-1 mRNA expression. This study provides evidence that the down-regulation of MKP-1 is critical for increased production of MCP-1 during the course of adipocyte hypertrophy.     

A novel gene, CRR9, which was up-regulated in CDDP-resistant ovarian tumor cell line, was associated with apoptosis

In the screening for cisplatin (CDDP)-resistance related genes by a mRNA differential display method, we detected some increased bands in CDDP resistant ovarian tumor cell line 2008/C13*5.25. One of them, named DD9, was a positive fragment on Northern blot analysis. We cloned it as a full length cDNA by 5'RACE and found a novel gene, CRR9 (Cisplatin Resistance Related gene 9). The CRR9 gene was transcribed into a 2.0 kb mRNA, encoding 512 amino acids. The putative protein had transmembrane-like domains and well conserved on C terminus with human CLPTM1 and the homologs found in Drosophila and C. elegans. Transfection assay showed that the CDDP-sensitive strain 2008 with CRR9 was more sensitive to CDDP, indicating that CRR9 was not associated with the CDDP-resistance, but the CDDP-induced apoptosis.     

Highly potent PDE4 inhibitors with therapeutic potential

Based on the hypothesis that the dose-limiting side effects of PDE4 inhibitors could be mediated via the central nervous system (CNS), design and synthesis of a hydrophilic analogue is considered to be one approach to improving the side-effect profile of Ariflo 1. Water-soluble piperidine derivatives were found to possess therapeutic potential.