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A synthesized mouse vasoactive intestinal contractor peptide, which belongs to a novel member of the endothelin family, induced a prolonged contraction in mouse ileum as well as porcine coronary artery in vitro. Studies comparing the effects of vasoactive intestinal contractor and endothelin on different tissues revealed that the maximum ileum contraction of vasoactive intestinal contractor was much higher than that of endothelin in both guinea pig and mouse systems, but that the vasoconstriction activity of vasoactive intestinal contractor was weaker than that of endothelin in porcine artery. These resuts show that vasoactive intestinal contractor might be a novel gastrointestinal hormone.

Peroxisome proliferator-activated receptor alpha (PPARalpha) is a nuclear receptor that regulates the expression of genes associated with lipid metabolism. Recent studies have suggested that the expression of PPARalpha-dependent fibroblast growth factor 21 (FGF21) plays important roles in adaptation to fasting, such as lipolysis and ketogenesis. We found that a nighttime injection of bezafibrate, a ligand of PPARalpha, effectively induced FGF21 expression, whereas a daytime injection did not affect it. Furthermore, bezafibrate-induced circadian FGF21 expression was abolished in PPARalpha-deficient mice. These observations suggest that bezafibrate-induced circadian FGF21 expression is due to circadian variations in the responsiveness of the PPARalpha system in the liver.

Mutations that abolish expression of an X-linked gene, FMR1 , result in the pathogenesis of fragile X syndrome, the most common form of inherited mental retardation [1, 2]. To understand the normal function of the FMR1 protein, we have produced fly strains bearing deletions in a Drosophila homolog of FMR1 ( dfmr1 ). Since fragile X patients show a number of abnormal behaviors including sleep problems [3, 4], we investigated whether a loss-of-function mutation of dfmr1 affect circadian behavior [5–8]. Here we show that under constant darkness (DD), a lack of dfmr1 expression causes arrhythmic locomotor activity, but in light:dark cycles, their behavioral rhythms appear normal. In addition, the clock-controlled eclosion rhythm is normal in DFMR1-deficient flies. These results suggest that DFMR1 plays a critical role in the circadian output pathway regulating locomotor activity in Drosophila .

We evaluated 107 patients who were diagnosed as benign prostatic hypertrophy preoperatively and underwent transurethral resection of the prostate. We obtained the maximum cross section of the adenoma by transabdominal scanning using a real-time large-sized convex scanner and a 3.5 MNz. transducer. Regarding the adenoma as an ellipsoid with revolution around its transverse axis, 2 diameters (breadth and height) were measured with transverse scan (a and b, respectively). We then calculated the values of the estimated size of the adenoma according to the formula for a spheroid. V = (pi/6) ab2. Thus the calculated weight were correlated with the resected weight of the specimen after a complete transurethral resection of the prostate. As a result, the sonographic estimation provided a high correlation coefficient: r = 0.956. The result suggested that our method of transabdominal ultrasonography in the preoperative evaluation of the prostatic size was rapid, simple and noninvasive, was well adapted for screening sufferers from prostatic diseases, and gave preoperative information for transurethral resection of the prostate and other surgical operation.

We have determined the complete organization of the mouse CH gene family, which is comprised of the 8 CH genes in the order 5’-JH-6.5kb-Cμ-4.5kb-Cδ-55kb-Cγ3-34kb-Cγ1-21kb-Cγ2b-15kb-Cγ2a-14kb-Ce-12kb-Cα-3’. The S regions, which contain characteristic tandemly repeated unit sequences, are located 5’ to each CH gene except for the Cδ gene. There are at least two types of repetitive sequences dispersed in this 200 kb region. No pseudogenes are present. The arrangements of the CH genes in BALB/c and C57BL mice are similar, but the lengths of the S regions vary. The basic structures of all the CH genes are similar in that coding sequences are interrupted at the junctions of the domains and the hinge regions. Comparison of the nucleotide sequences of the CH genes revealed that sequence segments have been exchanged among members of the CH gene family. Cloning and characterization of human Cγ genes, i.e. Cγ1, Cγ2, Cγ3, Cγ4 and φCγ, indicate that the human Cγ gene family evolved by dynamic DNA rearrangements, including gene duplication, exon duplication, and exon reassortment by unequal crossing-over. A human pseudo-epsilon gene (Ce3) is a processed gene that has completely spliced out introns. The presence of movable genetic elements surrounding the Ce3 gene suggests that the Ce gene evolved by a translocation mechanism. Although S-S recombination has been shown to take place in myelomas and hybridomas secreting a large amount of immunoglobulin, analyses of the CH gene organization in normal spleen B cells bearing immunoglobulin on their surface suggest that RNA splicing may be responsible for the first step in class switching, followed by S-S recombination. The nucleotide sequences of S regions contain short common sequences, TGGG(G) and (G)AGCT. Comparison of nucleotide sequences surrounding recombination sites revealed common sequences TGAG and TGGG. A sister chromatid exchange model was proposed to explain deletion of CH genes accompanying S-S recombination. We have found that the S region serves as a preferred recombination site in E.coli extracts.

Effects on [Ca2+]i levels of endothelin-1 (ET) and vasoactive intestinal contractor peptide (VIC), which is a novel member of the endothelin family, were examined in fura 2-loaded neuroblastoma NG108-15 cells. VIC was found to be a very effective stimulus for intracellular Ca2+ mobilization and to be more potent than ET. Intracellular calcium response to sequential addition of two stimulants exhibited the homologous desensitization of either ET or VIC, but no heterologous desensitization between ET and VIC. This indicates evidence suggesting that these two peptides act through distinct receptors.

We have cloned cDNA encoding the mouse interleukin-2 (IL-2) receptor from a murine T cell line, CTLL using human IL-2 receptor cDNA as probe. COS 7 cells transfected with the cDNA expressed the antigen recognized by the monoclonal antibody against the murine IL-2 receptor. The cDNA identified 4 species of mRNA (4.5, 3.5, 2.2 and 1.5 kb) of the mouse IL-2 receptor in CTLL cells. Difference in the length of mRNA seems to be ascribed to the variable length of the 3' untranslated sequence. Total nucleotide sequence (approximately 1400 bp) of this cDNA was determined and compared with the human receptor. The nucleotide and amino acid sequences of the IL-2 receptor are 70% and 60%, respectively, homologous in average between the two species. The comparison has revealed several conserved regions localized to particular exons such as transmembrane and cytoplasmic portions, suggesting that these regions are important for receptor function and its regulation.

Immunoglobulin genes are shown to undergo dynamic rearrangements during differentiation as well as evolution. We have demonstrated that a complete immunoglobulin heavy chain gene is formed by at least two types of DNA rearrangement during B cell differentiation. The first type of rearrangement is V-D-J recombination to complete a variable region sequence and the second type is S-S recombination to switch a constant region sequence. Both types of recombination are accompanied by deletion of the intervening DNA segment. Structure and organization of CH genes are elucidated by molecular cloning and nucleotide sequence determination. Organization of H chain genes is summarized as VH-(unknown distance)-JH-(6.5 kb)-C mu-(4.5 kb)-C delta-(unknown distance)-C gamma 3-(34 kb)-C gamma 1-(21 kb)-C gamma 2b-(15 kb)-C gamma 2a-(14.5 kb)-C epsilon-(12.5 kb)-C alpha. The S-S recombination takes place at the S region which is located at the 5' side of each CH gene. Nucleotide sequence of the S region comprises tandem repetition of closely related sequences. The S-S recombination seems to be mediated by short common sequences shared among S regions. A sister chromatid exchange model was proposed as a mechanism for S-S recombination. Comparison of nucleotide sequences of CH genes indicates that immunoglobulin genes have scrambled by intervening sequence-mediated domain transfer during their evolution.

The human interleukin-2 (IL-2) receptor was purified by affinity chromatography using the anti-Tac monoclonal antibody, and its N-terminal amino acid sequence was determined. Complementary DNA clones were isolated and sequenced to reveal the primary structure of the IL-2 receptor precursor, which has 272 amino acid residues. The receptor is separated into two domains by a putative 19-residue transmembrane region. Two mRNAs (1.4 and 3.5 kilobases) hybridizing to the cDNA clone were found in human T cells bearing the IL-2 receptor. The cDNA directed synthesis of the IL-2 receptor in COS cells.

We cloned cDNAs of the human and mouse IL-2 receptors. Comparison of their structures allowed us to identify several conserved regions localized to exons 2 and 4, the cytoplasmic portion and the transmembrane portion. These regions might be important for the functions of the IL-2 receptor. The human IL-2 receptor, which was expressed on an IL-2-dependent murine T-cell line, CTLL-2, by cDNA transfection, was shown to be functionally active by blocking the endogenous mouse IL-2 receptor with monoclonal antibodies. On the other hand, the human IL-2 receptors expressed on non-lymphoid cells were functionally inactive. They were unable to mediate the growth signal, were of low affinity species and aberrant in internalization. We postulated that the dysfunction of the IL-2 receptors in non-lymphoid cells would be due to the absence of the putative converter protein which is expressed specifically in lymphoid cells. Since the human IL-2 receptor is active in the murine T cell, the converter may interact with the receptor at the portions conserved between man and mouse. We proposed the affinity conversion model that explained the high affinity state of the receptor by the ternary complex formation between IL-2, the IL-2 receptor and the converter.

The nephron is composed of various types of differentiated cells, each of which is located at the right place at the right time to exert its specific function. During nephrogenesis and physiologic turnover of cells in mature nephron, the cell cycle of differentiated cells and its stem/progenitor cells should be orchestrated, otherwise the morphologic and functional integrity of the kidney would be deranged. Based on this concept, we hypothesized that circadian rhythm, which is maintained by an intrinsic time-keeping system regulating behavior and physiologic functions of organisms, might organize the proliferation of cells in the kidney. In fact, daily administration of 5-bromo-2′-deoxyuridine (BrdU) to pup or adult Sprague-Dawley rats at a specific clock time revealed that renal tubular epithelial cells in cortical and corticomedullary segments of the nephron proliferated rapidly in a synchronized manner but cells in the rest of segments and glomeruli were quiescent or proliferate slowly without apparent synchronization. We confirmed that the activity of cell cycle-related protein molecules such as extracellular signal-regulated protein kinase (ERK)1 and 2 oscillated with a synchronized 24-hour cycle. Restricted daytime feeding, a well-established manipulation to dissociate the circadian clock of peripheral organs from that of suprachiasmatic nucleus (SCN) (the master clock of the body), clearly shifted the phase of the synchronized cell cycle. It means that the clock not in SCN but in the kidney somehow releases a cue for the growth of tubular epithelial cells at the specific segments. Generally, the existence and oscillating phase of the circadian clock can be perceived by the oscillating expression of clock genes such as Per 1 and Per 2. Because Per 1 and Per 2 existed throughout the nephron, including glomeruli and oscillated with a 24-hour cycle, it is intriguing that cells only in the limited area of the kidney proliferate in accordance with the circadian rhythm.