Sensorimotor Modulation Differs with Load Type during Constant Finger Force or Position

During submaximal isometric contraction, there are two different load types: production of a constant force against a rigid restraint (force task), and maintenance of position against a constant load (position task). Previous studies reported that the time to task failure during a fatigue task was twice as long in the force task compared with the position task. Sensory feedback processing may contribute to these differences. The purpose of the current study was to determine the influence of load types during static muscle contraction tasks on the gating effect, i.e., attenuation of somatosensory-evoked potentials (SEPs) and the cortical silent period (cSP). Ten healthy subjects contracted their right first dorsal interosseus muscle by abducting their index finger for 90 s, to produce a constant force against a rigid restraint that was 20% of the maximum voluntary contraction (force task), or to maintain a constant position with 10° abduction of the metacarpophalangeal joint against the same load (position task). Somatosensory evoked potentials (SEPs) were recorded from C3′ by stimulating either the right ulnar or median nerve at the wrist while maintaining contraction. The cortical silent period (cSP) was also elicited by transcranial magnetic stimulation. Reduction of the amplitude of the P45 component of SEPs was significantly larger during the position task than during the force task and under control rest conditions when the ulnar nerve, but not the median nerve, was stimulated. The position task had a significantly shorter cSP duration than the force task. These results suggest the need for more proprioceptive information during the position task than the force task. The shorter duration of the cSP during the position task may be attributable to larger amplitude of heteronymous short latency reflexes. Sensorimotor modulations may differ with load type during constant finger force or position tasks.     

Successful Treatment of Pulmonary Mucormycosis Caused by <Emphasis Type="Italic">Cunninghamella bertholletiae</Emphasis> with High-Dose Liposomal Amphotericin B (10 mg/kg/day) Followed by a Lobectomy in Cord Blood Transplant Recipients

Infection caused by Cunninghamella bertholletiae carries one of the highest mortality rates among mucormycosis, and there are no reported cases that survived from the infection in allogeneic hematopoietic stem cell transplantation recipients occurring before neutrophil engraftment. Here, we present two cases of pulmonary mucormycosis caused by C. bertholletiae occurring before neutrophil engraftment after cord blood transplantation. Both were successfully treated with high-dose liposomal amphotericin B (10 mg/kg/day) combined with micafungin, which was then followed by neutrophil recovery, reduction in immunosuppressive agents, and a subsequent lobectomy. The intensive antifungal therapy immediately administered upon suspicion of mucormycosis greatly suppressed the infection in its early stage and was well tolerated despite its prolonged administration and simultaneous use of nephrotoxic agents after transplantation. Although the synergic effect of micafungin remains unclear, these cases highlight the importance of prompt administration of high-dose lipid polyene when suspecting mucormycosis in highly immunocompromised patients, which enables subsequent diagnostic and therapeutic interventions, resulting in a favorable outcome.     

Improvement of Automatic In-Gel Digestion by in Situ Alkylation of Proteins

We have recently improved the automation of an in-gel digestion system, DigestPro 96, using in situ alkylation of proteins with acrylamide, conducted during one-dimensional (1D) SDS-PAGE. The improved method included the processes of destaining, dehydration, trypsin digestion, and extraction but excluded the reduction and alkylation steps following staining of proteins with CBB. The extracted peptide mixtures were directly loaded onto a micro C18 LC column of the mass spectrometer. The resultant spectra were processed with “Mascot” search engine to estimate the sequence coverage of the bovine serum albumin (BSA). The original method, designed for Laemmli 1D SDS gel applications, consisted of reduction and post-alkylation with iodoacetamide, which produced carboxyamidemethyl (CAM; –S–CH₂CONH₂) derivatives. The original method also included a desalting step essential for mass spectrometry, especially matrix-assisted laser desorption/ionization time-of-flight mass spectrometry. We compared the original and improved methods using BSA (3 pmol loaded to the gel, one third of digested peptide mixture injected into LC-MS). The original method yielded both CAM and propionicamide (PAM; –S–CH₂CH₂CONH₂) derivatives. The source of PAM derivatives is the unpolymerized acrylamide formed during electrophoresis. The sequence coverage of CAM derivatives of BSA by the original method was 10% with desalting and 19% without desalting. The sequence coverage of PAM derivative by the improved method was 32%. Our results clearly show the advantage of our improved automated in-gel digestion method for in situ PAM alkylated protein with respect to peptide recovery, compared with the original method with CAM post-alkylation.     

Intranasal administration of His-D-Trp-Ala-Trp-D-Phe-LysNH2 (growth hormone releasing peptide) increased plasma growth hormone and insulin-like growth factor-I levels in normal men

The effects of intranasal and iv administration of His-D-Trp-Ala-Trp-D-Phe-LysNH2 (GHRP) on plasma GH, PRL, LH, FSH, TSH, cortisol, insulin, IGF-I as well as GHRH-like immunoreactivity (LI) levels were examined in 6 healthy male subjects. An iv bolus injection of GHRP(1 micrograms/kg BW) caused a remarkable increase in plasma GH levels with a mean (+/- SE) peak of 54.9 +/- 4.2-micrograms/L. In addition an intranasal administration of GHRP resulted in a significant, dose-related increase in plasma GH with peaks of 39.6 +/- 15.3 micrograms/L at a dose of 30 micrograms/kg BW, 14.1 +/- 5.0 micrograms/L at 15 micrograms/kg BW and 7.5 +/- 5.7 micrograms/L at 5 microgram/kg BW. Plasma PRL and cortisol levels were slightly but significantly increased after iv administration of GHRP, whereas GHRP totally failed to affect plasma TSH, LH, FSH, insulin, blood sugar and GHRH-LI levels. Seven consecutive, intranasal administrations of 15 micrograms/kg BW GHRP every 8h were well tolerated in all subjects examined. During this treatment, GH responsiveness to GHRP was not attenuated by desensitization and plasma IGF-I was increased from 94.5 +/- 5.8 micrograms/L before GHRP to 125.8 +/- 6.0 micrograms/L after repeated GHRP administration. These findings indicate that intranasal administration of GHRP stimulates GH secretion and consequently enhances IGF-I production in normal subjects. If GHRP is demonstrated to be beneficial in the treatment of some patients with GH deficiency, the intranasal route of administration may be more useful than the painful injection because a prolonged period is required for the treatment.     

Nitrated and oxidized products of a single tryptophan residue in human Cu,Zn-superoxide dismutase treated with either peroxynitrite-carbon dioxide or myeloperoxidase-hydrogen peroxide-nitrite

We reported previously that a single tryptophan residue, Trp32, in human Cu,Zn-superoxide dismutase is specifically modified by peroxynitrite-CO2 [Yamakura et al. (2001) Biochim. Biophys. Acta 1548, 38-46]. In this study, we modified Cu,Zn-superoxide dismutase by using a combination of myeloperoxidase, hydrogen peroxide, and nitrite. The modified enzyme showed no loss of copper and zinc, and 15% less enzymatic activity. Trp32 was the only significant amino acid lost. After trypsin digestion of the modified SOD with peroxynitrite-CO2 and the myeloperoxidase system, six newly appearing peptides containing tryptophan derivatives were observed on microLC-ESI-Q-TOF mass analyses and HPLC with a photodiode-array detector. The derivatives of the tryptophan residue exhibiting mass increases of 4, 16 (2 peaks), 32, 45 (major), and 45 Da (minor) were identified as kynurenine, oxindole-3-alanine and its derivatives, dihydroxytryptophan, 6-nitrotryptophan and 5-nitrotryptophan, respectively. We further identified 6-nitrotryptophan from the 1H-NMR spectrum for the pronase-digested product and calculated the yield of 6-nitrotryptophan as being about 30% for each of the modification methods. The tryptophan residue in the modified human Cu,Zn-superoxide dismutase gave the same spectra for the products including 6-nitrotryptophan as the major nitrated product with the two different modification systems.     

Hypoxia followed by reoxygenation induces secretion of cyclophilin A from cultured rat cardiac myocytes

We previously reported that hypoxia followed by reoxygenation (hypoxia/reoxygenation) rapidly activated intracellular signaling such as mitogen-activated protein kinases (MAPKs) including extracellular signal-regulated protein kinase (ERK) 1/2, p38MAPK, and stress-activated protein kinases (SAPKs). To investigate the humoral factors which mediate cardiac response to hypoxia/reoxygenation, we analyzed the conditioned media from cardiac myocytes subjected to hypoxia/reoxygenation by two-dimensional electrophoresis and mass spectrometry. We identified cyclophilin A (CyPA) as one of the proteins secreted from cardiac myocytes in response to hypoxia/reoxygenation. Hypoxia/reoxygenation induced the expression of CyPA and its cell surface receptor CD147 on cardiac myocytes in vitro. This was also confirmed by ischemia/reperfusion in vivo. Recombinant human (rh) CyPA activated ERK1/2, p38MAPK, SAPKs, and Akt in cultured cardiac myocytes. Furthermore, CyPA significantly increased Bcl-2 in cardiac myocytes. These data strongly suggested that CyPA is released from cardiac myocytes in response to hypoxia/reoxygenation and may protect cardiac myocytes from oxidative stress-induced apoptosis.     

Doppel-induced Purkinje cell death is stoichiometrically abrogated by prion protein

Activin A can induce erythroid differentiation, whereas basic fibroblast growth factor (bFGF) can maintain the undifferentiated status of erythroid progenitors. How these two factors together can affect the regulation of erythroid differentiation in hematopoietic cells has not been elucidated. This study demonstrates that bFGF antagonizes activin A-mediated growth inhibition and hemoglobin (Hb) synthesis in K562 cells. Analyses of mitogen-activated protein kinases revealed that activin A-induced p38 phosphorylation and inhibited ERK1/2 phosphorylation. In contrast, bFGF worked antagonistically to induce ERK1/2 phosphorylation and inhibited p38 phosphorylation in K562 cells. Furthermore, co-treatment of cells with activin A and bFGF decreased p38 phosphorylation and increased ERK1/2 phosphorylation. SB203580 inhibition of p38 activity eliminated activin A-mediated growth inhibition and Hb synthesis, whereas U0126 inhibition of ERK1/2 activity augmented the effects of activin A on K562 cells. These results suggest that bFGF can negatively modulate p38 and positively modulate ERK1/2 to antagonize activin A-mediated growth inhibition and Hb synthesis in K562 cells.