Hypertrophy of rat plantaris muscle fibers after voluntary running with increasing loads

There have been no systematic comparisons ofskeletal muscle adaptations in response to voluntary wheel runningunder controlled loading conditions. To accomplish this, a voluntaryrunning wheel for rats and mice was developed in which a known load canbe controlled and monitored electronically. Five-week-old maleSprague-Dawley rats (10 rats/group) were assigned randomly to either a1) sedentary control group(Control); 2) voluntary exercisedwith no load (Run-No-Load) group; or3) voluntary exercised withadditional load (Run-Load) group for 8 wk. The load for the Run-Loadgroup was progressively increased to reach ~60% of body weightduring the last week of training. The proportions of fast glycolytic(FG), fast oxidative glycolytic (FOG), or slow oxidative (SO) fibers inthe plantaris were similar in all groups. The absolute and relativeplantaris weights were greater in the Run-Load group compared with theControl and Run-No-Load groups. The mean fiber cross-sectional areas of FG, FOG, and SO fibers were 20, 25, and 15% greater in the Run-Load than in Control rats. In addition, these fiber types were 16, 21, and12% larger in Run-Load than in Run-No-Load rats. The muscle weightsand mean cross-sectional areas of each fiber type were highlycorrelated with the average running distances and total work performedin the Run-Load, but not the Run-No-Load, group. The slope of therelationship between fiber size and running distance and total workperformed was significant for each fiber type but was higher for FG andFOG fibers compared with SO fibers. These data show that the load on arat running voluntarily can determine the magnitude of a hypertrophicresponse and the population of motor units that are recruited toperform at a given loading condition.

     

Reduced growth of Ehrlich ascites tumor cells in creatine depleted mice fed beta-guanidinopropionic acid

The effect of implantation of Ehrlich ascites tumor (EAT) cells on creatine distribution was investigated. It was also studied how depletion of creatine by feeding creatine-analogue beta-guanidinopropionic acid (beta-GPA) affects the growth of EAT cells in mice. Enhanced mobilization of creatine from host tissues to EAT cells against a greater concentration gradient was observed. The creatine (but not creatinine) level in blood plasma was lowered to 22% of the normal value by beta-GPA feeding alone and assimilation of 14C-creatine into EAT cells was inhibited. The growth of EAT cells was significantly reduced and the duration of survival of mice after implantation of EAT cells was extended when the creatine concentration was decreased. A decrease in daily food consumption and the degree of muscle atrophy after implantation of EAT cells was less in beta-GPA than control groups. In the creatine-depleted mice, the rate of increase in total EAT cell number and the volume of abdominal ascites were approximately half of the control values, and more dead EAT cells were observed. These results suggest that supplementation of beta-GPA inhibits creatine transfer to EAT cells and reduces the growth of cancer cells.     

Degradation of Ribulose-l,5-bisphosphate Carboxylase/Oxygenase in the Lysates of the Chloroplasts Isolated Mechanically from Wheat (Triticum aestivum L.) Leaves

Intact chloroplasts were isolated mechanically from the primaryleaves of 8- to 12-day old seedlings of wheat (Triticum aestivumL.) and purified by Percoll gradient centrifugation. The chloroplastswere lyzed by osmotic shock and the reaction mixtures containingthe lysates were incubated in the pH range of 5.3 to 9.4 at37°C. The degradation of ribulose-l,5-bisphosphate carboxylase/oxygenase(RuBisCO, EC 4.1.1.39 [EC] ) and its degradation products in the mixtureswere examined by using SDS-polyacrylamide gel electrophoresis.RuBisCO-hydrolase activity in the lysates was very weak, andit was difficult to assess the activity by measuring the lossof the amount of the large subunit of RuBisCO on the gels afterstaining with Coomassie Brilliant Blue. By using immunoblottingmethod, however, degradation products of RuBisCO could be detectedin the reaction mixtures. The hydrolase activity was pronouncedin the presence of 0.1 % (w/v) of SDS in the reaction mixtures.Among the products, the 35 kDa fragment was conspicuous andfound in the wide range of pHs. This degradation of RuBisCOwas inhibited in the presence of leupeptin and N-ethylmaleimide. (Received October 3, 1988; Accepted November 25, 1988)     

Effects of space flight on GLUT-4 content in rat plantaris muscle

 The effects of 14 days of space flight on the glucose transporter protein (GLUT-4) were studied in the plantaris muscle of growing 9-week-old, male Sprague Dawley rats. The rats were randomly separated into five groups: pre-flight vivarium ground controls (PF-VC) sacrificed approximately 2 h after launch; flight groups sacrificed either approximately 5 h (F-R0) or 9 days (F-R9) after the return from space; and synchronous ground controls (SC-R0 and SC-R9) sacrificed at the same time as the respective flight groups. The flight groups F-R0 and F-R9 were exposed to micro-gravity for 14 days in the Spacelab module located in the cargo bay of the shuttle transport system – 58 of the manned Space Shuttle for the NASA mission named ”Spacelab Life Sciences 2”. Body weight and plantaris weight of SC-R0 and F-R0 were significantly higher than those of PF-VC. Neither body weight nor plantaris muscle weight in either group had changed 9 days after the return from space. As a result, body weight and plantaris muscle weight did not differ between the flight and synchronous control groups at any of the time points investigated. The GLUT-4 content (cpm/μg membrane protein) in the plantaris muscle did not show any significant change in response to 14 days of space flight or 9 days after return. Similarly, citrate synthase activity did not change during the course of the space flight or the recovery period. These results suggest that 14 days of space flight does not affect muscle mass or GLUT-4 content of the fast-twitch plantaris muscle in the rat. Received: 25 March 1997 / Accepted: 18 August 1997     

Metabolism of S-methylcysteine and pectin esterification in Chinese cabbage, Brassica pekinensis Rupr.

S-Methyl-L-cysteine was actively metabolized in Chinese cabbageand carbon from its methyl group was distributed into both thesoluble and insoluble fractions. The high incorporation of ¹⁴Cfrom the methyl group into the insoluble fraction after administeringof S-methyl-L-cysteine-¹⁴CH₃, and our previous results thatS-methyl-L-cysteine is demethylated to give cysteine, suggestthat S-methyl-L-cysteine might act as a methyl donor in Chinesecabbage. To obtain evidence for this possibility, incorporationof the methyl-¹⁴C of S-methyl-L-cysteine into methyl estersof pectic substances was investigated. Most of the ¹⁴C incorporatedinto pectic substances was liberated by treatment with dilutealkali and pectin esterase. The results show that S-methyl-L-cysteineacts as a methyl donor to form pectin ester. (Received October 12, 1971; )     

Cloning and characterization of a molt-inhibiting hormone-like peptide from the prawn Marsupenaeus japonicus

Recently, it was demonstrated by PCR amplification that an additional molt-inhibiting hormone (MIH)-like peptide was present in the kuruma prawn Marsupenaeus japonicus. In this study, a cDNA encoding this peptide designated Pej-MIH-B was cloned. The Pej-MIH-B gene was expressed strongly in the nerve cord, and weakly in the eyestalk. It was possible to isolate Pej-MIH-B from the sinus glands in the eyestalks. The recombinant Pej-MIH-B expressed in Escherichia coli showed low molt-inhibiting activity, but did not exhibit hyperglycemic activity. These results suggest that Pej-MIH-B does not function as MIH or CHH intrinsically, but may have some unknown functions.     

Structure-activity relationship of crustacean molt-inhibiting hormone from the kuruma prawn Marsupenaeus japonicus

In crustaceans, molt-inhibiting hormone (MIH) controls molting by suppressing the synthesis and/or secretion of molting hormone. In our previous study, which determined the solution structure of MIH by NMR, we hypothesized that the peptide's functional site spanned the region encompassing the N-terminal alpha-helix and a portion of the C-terminus, both of which are located sterically close to each other [Katayama et al. (2003) J. Biol. Chem. 278, 9620-9623]. To confirm this hypothesis, various mutants of MIH were prepared and their molt-inhibiting activities were assessed. All peptides mutated at the putative functional site exhibited circular dichroism spectra similar to the natural MIH, suggesting that the mutants retained their natural conformation regardless of the mutations. As expected, a majority of the mutants, except for Delta12 (a deletion mutant of Gly(12)) and Delta75-77 (a deletion mutant of the last three residues of the C-terminus), were less active than the natural MIH. In particular, I72G exhibited no molt-inhibiting activity even at 200 nM, while N13A and S71Y exhibited low activity at the same concentration. In contrast, the natural and recombinant MIHs exhibited full inhibitory activity at 20 nM. All these results indicate that the functional site of MIH is located in the region containing the C-terminal ends of the N- and C-terminal alpha-helices, and that Asn(13), Ser(71), and Ile(72) are especially significant for conferring molt-inhibiting activity. Furthermore, these findings agree with the results and the proposed hypothesis presented in previous studies on the structure-activity relationship of MIH and its related peptides.     

Identification of a novel Japanese flounder (<Emphasis Type="Italic">Paralichthys olivaceus</Emphasis>) CC chemokine gene and an analysis of its function

A cDNA of Japanese flounder (Paralichthys olivaceus) CC chemokine designated as Paol-SCYA104 was cloned and sequenced. The cDNA contains an opening reading frame of 315 nucleotides encoding 104 amino acid residues. The full gene was cloned and sequenced from a BAC library. It has a length of approximately 750 bp from the start codon to the stop codon and is composed of four exons and three introns. Four cysteine residues are conserved in the same positions as those of mammalian and fish CC chemokines. Paol-SCYA104 gene was expressed in several organs, including peripheral blood leukocytes (PBLs), head kidney, trunk kidney, and spleen. The recombinant Paol-SCYA104 was expressed in Escherichia coli and the expressed protein was partially purified. The recombinant Paol-SCYA104 was able to attract Japanese flounder PBLs in a microchemotaxis chamber. On the other hand, a negative control, the fraction of the control cells carrying an expression vector lacking the Paol-SCYA104 cDNA, did not show chemotactic activity. These results indicate that Paol-SCYA104 probably acts as a CC chemokine.     

Two different types of hepcidins from the Japanese flounder Paralichthys olivaceus

The cysteine-rich peptide hepcidin is known to be an antimicrobial peptide and iron transport regulator that has been found in both fish and mammals. Recently, we found two different types (designated Hep-JF1 and Hep-JF2) of hepcidin cDNA in the Japanese flounder, Paralichthys olivaceus, by expressed sequence tag analysis. The identity of amino acid sequences between Hep-JF1 and Hep-JF2 was 51%. The Hep-JF1 and Hep-JF2 genes both consist of three exons and two introns, and both exist as single copies in the genome. The predicted mature regions of Hep-JF1 and Hep-JF2 have six and eight Cys residues, respectively. The first Cys residue of Hep-JF1 was deleted and the second was replaced with Gly. The number and positions of Cys residues in Hep-JF2 are the same as they are in human Hep. Hep-JF1 is specifically expressed in liver while the expression of Hep-JF2 was detected from gill, liver, heart, kidney, peripheral blood leucocytes, spleen and stomach. Gene expression of Hep-JF1 in liver decreased during experimental iron (iron-dextran) overload. Expression of Hep-JF1 in liver was decreased by injecting fish with iron-dextran and increased by injecting lipopolysaccharide. Iron overload did not significantly affect expression of Hep-JF2 in liver but it did increase expression of Hep-JF2 in kidney. Lipopolysaccharide injection increased expression of Hep-JF2 in both liver and kidney. In liver, some cells expressed both Hep-JF1 and Hep-JF2 while some other cells expressed just one of them. Synthesized Hep-JF2 peptide showed antimicrobial activity, while synthesized Hep-JF1 peptide did not against several bacteria including fish-pathogenic bacteria used in this study.