Comparison of renal biomarkers with glomerular filtration rate in susceptibility to the detection of gentamicin-induced acute kidney injury in dogs.

Fourteen renal biomarkers were compared with measurement of glomerular filtration rate (GFR) in detecting acute kidney injury (AKI) in beagle dogs given gentamicin (40 mg/kg/day by subcutaneous injection) for 7 consecutive days. Serum and urinary biomarkers were measured before administration of gentamicin and then on days 4 and 8 after starting administration. GFR was derived by use of a simplified equation. Increased urinary cystatin C and decreased GFR occurred from day 4 and were detected before increases in blood urea nitrogen (BUN) and serum creatinine concentrations and changes in other urinary parameters. The closest correlation was between urinary cystatin C and GFR. At termination, microscopical examination revealed extensive necrosis of proximal tubular epithelium with hyaline casts in the kidney of treated dogs. These data indicate that urinary cystatin C is the most sensitive index of kidney injury and GFR reflects the kidney functional mass.

Silent period and H reflex from soleus muscle as an index in a neuro-muscular function after reconstruction of anterior cruciate ligament.

To clarify that the mixed nerve silent period (SP) and H reflex from soleus muscle may become an index expressed neuro-muscular function after reconstruction of anterior cruciate ligament (ACL), we studied the alteration of SP and H reflex from the soleus muscle in the patient with ACL reconstruction. Subjects were three patients with after ACL reconstruction, who were two male athletes (case A and B) and a female aerobics instructor (case C). Recording of SP and H reflex was performed longitudinally from one month post operation to six months post operation. As a result of this study, in case A, following findings were appeared; shortening of duration of SEP increasing the coefficient of variation of SP and increasing the amplitude ratio of H/M in non-operative side at the same time when his overusing activity of non-operative side. With case B, an appearance of long latency reflex (LLR) and an increase of coefficient of variation of SP in the operative side were verified in the period when many assists were required in order to correct motion image in rehabilitation. Concretely, from after the reconstruction over 5 months, LLR was appeared in SP of operative side, and increases of coefficient of variation was most remarkably in 4 months after the reconstruction, amplitude H/M ratio of operative side showed high value in 4 months after the reconstruction. With case C, remarkable problematical point was not found at the time of activity of daily life (ADL) acquisition and occupation return. So, with case C, feature findings of both SP and H reflex were not appeared. The results suggested that SP and H reflex from soleus muscle have become an index expressed neuro-muscular function of lower extremity and supra-spinal function in the patient after reconstruction of ACL.

Isolation of microorganisms and substances inhibitory to aflatoxin production.

Aflatoxins are toxic and carcinogenic secondary metabolites produced by Aspergillus flavus and A. parasiticus. The contamination of crops, feeds, and foods with aflatoxins can have serious effects on the health of humans and animals. Although many studies have been done to develop aflatoxin-control strategies, most are limited in their effectiveness. As part of an effort to develop control procedures, we have devised simple and safe methods that are useful for identifying microorganisms that effectively inhibit aflatoxin production by fungi. These include the microtitre agar plate assay using norsolorinic acid-accumulating mutant fungi, the ultraviolet light photography method using an instant film, the tip culture method, a convenient RNA extraction method for reverse transcription-polymerase chain reaction (RT-PCR) analysis, and other methods. Results of a recent trial have shown that Achromobacter xylosoxidans significantly inhibited aflatoxin production by A. parsiticus, and that the main inhibitory substance produced by the bacterium was cyclo(L-leucyl-L-prolyl). This result confirms that the methods described herein are useful for identifying microorganisms that inhibit aflatoxin production by fungi and could contribute to the development of methods to reduce aflatoxin contamination in commodities.

Neuromuscular function after reconstruction of anterior cruciate ligament--a case study using evoked electromyography.

To clarify the neuro-muscular function of a lower extremity after reconstruction of anterior cruciate ligament (ACL), we studied the alteration of H reflex and Silent period (SP) from the soleus muscle in a patient with ACL reconstruction. Subject was an eighteen year-old male and high school basketball player after a reconstruction of right ACL. Recording of H reflex and SP was started at one month after operation, and ended at six months post operation with a test at every month. As a result of this study, the amplitude ratio of H/M(max) was increased at post four months after ACL reconstruction on operative side. Little variation of SP was acquired in non-operative side. However, that in operative side was increased compared with non-operative side and gradually decreased until post six months. Post six months, the variation of SP in operative side was recovered as that of non-operative side. And, on the operation side, long-latency reflex (LLR) was appeared during SP from one month to five months after, especially its appearance pattern was most markedly post four months. However, post six months, LLR disappeared during SP. The result of H reflex, SP and LLR appearance suggested a following hypothesis; until post six months after ACL reconstruction, even the small and simple task (i.e. an ankle planter flexion with a slight effort) needs to an intervention of a various supra-spinal function. In this case, autonomy of central nervous system related to an out-put mechanism in lower-extremity was acquired post six months.

Implications for using H-max/M-max ratio in H-reflex parameters for elderly subjects compared with young subjects.

The H-max/M-max ratio has long been used to understand motoneuron excitability induced by the Ia fibers. However, we contend that when comparing young and elderly subjects, this measurement can be controversial. We compared the modulation of the soleus H-max and M-max at five different muscle lengths between elderly and young subjects. The young subjects demonstrated a significant modulation of both H-max and M-max between dorsiflexion (DF) and plantarflexion (PF) positions. In contrast, the elderly subjects demonstrated no difference in H-max and M-max between DF and PF positions. However, it was more interesting to note that those elderly subjects who had similar H-max/M-max ratios to the young subjects at 0 deg showed identical modulation of H-max and M-max to that of the young In this paper, we discuss that H-reflex comparisons between young and elderly subjects may be confounded by the initial H-max/M-max ratio. This finding has implications for both the statistical analysis of this data as well as the theoretical interpretation of H-reflex measurements.

Gastric myoelectrical activity increases after moderate-intensity exercise with no meals under suppressed vagal nerve activity.

Postprandial gastric myoelectrical activity recorded by electrogastrogram (EGG) with the subject in a supine position has shown to be enhanced after moderate-intensity pedaling exercise in an upright seated position, despite the suppression of vagal nerve activity. However, it is still unknown whether the effect is due to the exercise itself and/or a meal or how the position change has influenced the effects. To address this, we used a position-controllable cycle ergometer to examine the effects of the moderate-intensity exercise on EGG activity and the high-frequency (HF) component of heart rate variability (HRV), an index of vagal nerve activity. To eliminate the effect of position change, we carried out the exercise and the EGG recording in the supine position. The peak amplitude of the EGG was enhanced by prior moderate-intensity exercise with a reduced HF component of HRV, which did not differ for postexercise conditions with or without a meal. The small amount of meal itself, however, enhanced both the peak amplitude of the EGG and the HF component of HRV. The peak frequency of EGG was reduced and the instability coefficient of EGG was increased only after the exercise itself. Taken together, these results suggest that the enhanced amplitude of gastric myoelectrical activity can be induced by moderate-intensity exercise itself, even with suppressed vagal nerve activity, and that the mechanism underlying the exercise effects would differ from that underlying the effect of a meal alone.

Early pathophysiologic feature of arthropathy in juvenile dogs induced by ofloxacin, a quinolone antimicrobial agent.

Arthropathy in dogs induced by ofloxacin, a quinolone antimicrobial agent, was pathophysiologically investigated. In the in vivo studies, ofloxacin was administered orally once or twice at 20 mg/kg/day to male juvenile (3-month-old, n=3) or adult (36-month-old, n=2) dogs, and the humeral and femoral heads were examined pathologically. Unlike adult dogs, fluid-filled vesicles were macroscopically observed on the articular surfaces of one juvenile dog 24 hours after a single treatment with ofloxacin. These lesions were seen in all juvenile dogs by twice dosing. Microscopically, fissures or cavity formations in the middle zone of the articular cartilage were noted only in juvenile dogs. Furthermore, the cartilage matrix from the abnormal area to the articular surface showed a decreased safranin-O staining intensity, suggesting proteoglycan depletion. Ultrastructurally, chondrocytes in the middle zone of juvenile dogs displayed dilatation of the cisternae in the rough endoplasmic reticulum as an initial hallmark. In the in vitro studies, chondrocytes isolated from the articular cartilage of naive juvenile dogs were exposed to ofloxacin at 6.3-100 microg/ml for 24 hours. Although no changes were noted in the deoxyribonucleic acid synthesis, protein synthesis, or proteoglycan release at concentrations of up to 100 microg/ml, the proteoglycan synthesis was evidently decreased in a dose-dependent manner from 12.5 microg/ml. The results obtained suggest that the inhibitory action of ofloxacin on proteoglycan syntheses in the chondrocytes may largely contribute to the early morphologic features in the articular cartilage of the juvenile dog.

Enzyme reactions and genes in aflatoxin biosynthesis.

Aflatoxins are highly toxic and carcinogenic substances mainly produced by Aspergillus flavus and Aspergillus parasiticus. Sterigmatocystin is a penultimate precursor of aflatoxins and also a toxic and carcinogenic substance produced by many species, including Aspergillus nidulans. Recently, the majority of the enzyme reactions involved in aflatoxin/sterigmatocystin biosynthesis have been clarified, and the genes encoding the enzymes have been isolated. Most of the genes constitute a large gene cluster in the fungal genome, and their expression is mostly regulated by a product of the regulatory gene aflR. This review will summarize the enzymatic steps and the genes in aflatoxin/sterigmatocystin biosynthesis.

Effects of high versus low-intensity resistance training on bone mineral density in young males.

The purpose of this study was to investigate the effect of high-intensity and low-intensity resistance training upon bone mineral density (BMD) by comparing the BMD of young male powerlifters (n = 5), recreational trainees (n = 5), and controls (n = 5). Lumbar spine (L2-L4), proximal femur, and whole body BMDs were measured using dual-energy X-ray absorptiometry (DXA). The high-intensity group showed a significantly greater BMD when the whole body and trochanter regions were measured than the low-intensity and control group. The BMD of the lumbar spine, femoral neck, and Ward's triangle was greater in the high-intensity group compared with the control group. There was no significant BMD difference between the low-intensity and control group except at the trochanter region. These results suggest that high-intensity resistance training is effective for increasing BMD, but low-intensity resistance training is not.

A non-arthropathic dose and its disposition following repeated oral administration of ofloxacin, a new quinolone antimicrobial agent, to juvenile dogs.

A non-arthropathic dose and disposition of ofloxacin, a potent new quinolone antimicrobial agent, were assessed in male juvenile (3-month-old) dogs, when administered orally at 5, 10 and 20 mg/kg/day once daily for 8 consecutive days. Ofloxacin concentrations in sera and articular cartilages were analyzed by high-performance liquid chromatography (HPLC). Macroscopically, arthropathy characterized by fluid-filled vesicles in articular surface of the humerus and femur was observed in animals receiving 10 and 20 mg/kg/day of ofloxacin, but not in those given 5 mg/kg/day. At 20 mg/kg/day, arthropathy of comparable severity also occurred on day 2. Microscopically, the cavity formation in the middle zone of the articular cartilage was first identified and then necrotic chondrocytes were found numerous around the cavity, followed by appearance of chondrocyte clusters. In pharmacokinetics, peak serum concentration (Cmax) and area under the concentrations (AUC0-24) were increased in a dose-dependent manner. However, no remarkable differences in these two parameters were noted between a single and repeated treatments, suggesting no accumulation of the drug. The articular ofloxacin concentration 2 hr after treatment was approximately 1.8 (day 2) to 2.0 times (day 8) higher than the serum concentration. Based on these results, a non-arthropathic dose of ofloxacin in male juvenile dogs following an 8-day treatment is considered to be 5 mg/kg/ day, and its Cmax, AUC0-24 and articular cartilage concentrations 2 hr after treatment were 3.4 microg/ml, 35.1 microg-hr/m/ and 7.0 microg/g, respectively, under these experimental conditions. Thus, arthropathy due to ofloxacin may be predicted by monitoring serum drug concentration.

Absence of aflatoxin biosynthesis in koji mold (Aspergillus sojae).

Ten strains isolated from industrial soy sauce producing koji mold were identified as Aspergillus sojae and distinguished from Aspergillus parasiticus morphologically and physiologically. There was no detectable aflatoxin in any culture extracts of A. sojae strains. Strain 477 was chosen as a representative strain of industrial A. sojae for further molecular analysis. All enzymatic activities associated with the aflatoxin biosynthesis were not detected or negligible in strain 477 compared with that of the A. parasiticus strain. Southern analysis suggested that the genomic DNA of strain 477 contained aflatoxin biosynthetic pathway genes. In contrast, all industrial strains lacked detectable transcripts of aflR, the main regulatory gene for aflatoxin biosynthesis, under the aflatoxin-inducing condition. Our data suggest that defects in aflR expression cause the lack of expression of aflatoxin-related genes which results in the absence of aflatoxin biosynthesis in A. sojae strains.

Automatic postural response systems in individuals with congenital total blindness.

This study examined the effects of the absence of vision from birth on automatic postural responses to platform displacements during stance. Postural responses were induced by producing randomly four types of perturbations which consisted of forward and backward translations, and toe up and down rotations. Nine congenitally totally blind and nine sighted adults served as subjects. EMG signals were recorded from four muscles in the right leg, and reaction time to somatosensory stimuli generated by platform displacements was measured by pushing a hand-held button. To assess the ability to control postural balance, the root mean square (RMS) values for lateral and antero-posterior sway before, during, and after perturbations were calculated. The EMG amplitude in the gastrocnemius muscle of a blind subject was smaller than that of a sighted subject with eyes closed. No significant differences were found between blind and sighted subjects in EMG latencies of the lower extremity muscles in response to perturbations. The blind subjects had significantly faster reaction times to somatosensory stimuli triggered by platform displacements, but in toe down rotations no significant difference was found between blind and sighted subjects. The difference in the EMG latencies and reaction times between the two groups suggests that blindness from birth may not affect the spinal stretch reflex, but may affect a volitional act mediated through the motor cortex. There were also no significant differences in the RMS values for postural sway between blind and sighted subjects with eyes open or closed, although blind subjects swayed more after backward translations than did sighted subjects with eyes open. Results suggest that the ability to control postural balance during perturbations was not affected by vision loss from birth. Our findings suggest that the automatic postural response systems of humans are unaffected by the absence of vision from birth and are rather hard wired.

Quantitative imaging of tyrosine hydroxylase and calmodulin in the human brain.

The distributions of tyrosine hydroxylase and calmodulin in adult normal postmortem human brain were analyzed quantitatively. Consecutive coronal sections were obtained from the anterior area of the right hemisphere and were stained immunohistochemically for tyrosine hydroxylase and calmodulin. Stained sections were divided into approximately 3 million microareas at 50 microm intervals, and the immunohistochemical fluorescence intensity in each area was measured by a human brain mapping analyzer, which is a microphotometry system for analysis of the distribution of neurochemicals in a large tissue slice. Immunoreactive staining of tyrosine hydroxylase and calmodulin was observed in almost all brain regions, but its intensity varied. Relatively high levels of calmodulin were observed in brain regions with high levels of tyrosine hydroxylase, though high levels of tyrosine hydroxylase were not always observed in brain regions where high levels of calmodulin were distributed. In particular, high levels of both of tyrosine hydroxylase and calmodulin were distributed in the caudate nucleus and putamen. Previously it was shown that tyrosine hydroxylase was activated and dopamine synthesis was enhanced in the neostriatum region in mice and rats by the intracerebroventricular administration of calcium through a calmodulin-dependent system. The present results combined with these previous findings suggest that the activity of tyrosine hydroxylase in the caudate nucleus and putamen of humans may also be regulated by a calcium/calmodulin-dependent system.

Role of cardiac renin-angiotensin system in sarcoplasmic reticulum function and gene expression in the ischemic-reperfused heart.

The aim of this study was to explore the possible participation of cardiac renin-angiotensin system (RAS) in the ischemia-reperfusion induced changes in heart function as well as Ca2+-handling activities and gene expression of cardiac sarcoplasmic reticulum (SR) proteins. The isolated rat hearts, treated for 10 min without and with 30 microM captopril or 100 microM losartan, were subjected to 30 min ischemia followed by reperfusion for 60 min and processed for the measurement of SR function and gene expression. Attenuated recovery of the left ventricular developed pressure (LVDP) upon reperfusion of the ischemic heart was accompanied by a marked reduction in SR Ca2+-pump ATPase, Ca2+-uptake and Ca2+-release activities. Northern blot analysis revealed that mRNA levels for SR Ca2+-handling proteins such as Ca2+-pump ATPase (SERCA2a), ryanodine receptor, calsequestrin and phospholamban were decreased in the ischemia-reperfused heart as compared with the non-ischemic control. Treatment with captopril improved the recovery of LVDP as well as SR Ca2+-pump ATPase and Ca2+-uptake activities in the postischemic hearts but had no effect on changes in Ca2+-release activity due to ischemic-reperfusion. Losartan neither affected the changes in contractile function nor modified alterations in SR Ca2+-handling activities. The ischemia-reperfusion induced decrease in mRNA levels for SR Ca2+-handling proteins were not affected by treatment with captopril or losartan. The results suggest that the improvement of cardiac function in the ischemic-reperfused heart by captopril is associated with the preservation of SR Ca2+-pump activities; however, it is unlikely that this action of captopril is mediated through the modification of cardiac RAS. Furthermore, cardiac RAS does not appear to contribute towards the ischemia-reperfusion induced changes in gene expression for SR Ca2+-handling proteins.

Quantitative maps of GAbAergic and glutamatergic neuronal systems in the human brain.

GABAergic and glutamatergic neuronal systems in adult normal human brains were shown quantitatively and in detail through the distributions of glutamate decarboxylase (GAD) and glutamate dehydrogenase (GDH), respectively. Consecutive coronal sections containing part of the striatum and the substantia nigra were obtained from the right hemisphere of three deceased persons with no history of neurological or psychiatric diseases and were stained immunohistochemically for GAD and GDH. Each stained section was divided into approximately 3 million microareas and the immunohistochemical fluorescence intensity in each area was measured by a human brain mapping analyzer, which is a microphotometry system for analysis of the distribution of neurochemicals in a large tissue slice. In the analyzed brain regions, conspicuously intense GAD-like immunoreactivity was observed in the substantia nigra, globus pallidus, and hypothalamus. GDH was widely and rather evenly distributed in the gray matter compared to GAD, although intense GDH-like immunoreactivity was observed in the lateral geniculate nucleus and substantia nigra. Within the substantia nigra, the globus pallidus, and other regions, characteristic distributions of GAD- and GDH-like immunoreactivity were found. We believe that the analysis of the human brain by this novel technique can help to understand the functional distribution of neuronal systems in the normal human brain and may be able to identify abnormal changes in the diseased human brain. It can also provide basic data to help in the interpretation of functional magnetic resonance imaging or positron emission tomography.

Effect of Zena F-III, a liquid nutritive and tonic drug, on the neurochemical changes elicited by physical fatigue in mice.

The effects of a liquid nutritive and tonic drug (NTD) on the neurochemical changes elicited by physical fatigue in mice were investigated in terms of the calcium-dependent dopamine synthesizing function of the brain. In this study, Zena F-III (Taisho Pharmaceutical Co., Ltd., Japan), one of the most popular NTDs in Japan, containing 15 crude drug extracts together with taurine, caffeine, and vitamins, and formulated based on the precepts of traditional Chinese medicine, was used. Male mice were forced to walk for 0-6 h at a speed of 3 m/min using a programmed motor-driven wheel cage. The serum and brain calcium levels in the mice were significantly increased following forced walking. The increase in brain calcium level began later and was more gradual than that in the serum calcium level, and reached its maximum value following forced walking for 3 h. The neostriatal dopamine level was also significantly increased, and locomotor activity significantly decreased following forced walking for 3 h. Prior oral administration of F-III (10 ml/kg) attenuated the increases in the serum and brain calcium levels, the increase in the brain dopamine levels, and the decrease in locomotor activity induced by forced walking. Taking into consideration these findings with our previous reports, it is suggested that physical fatigue leads to an increase in dopamine synthesis in the brain through a calcium/calmodulin-dependent system, thereby inducing behavioral changes, and that F-III inhibits this pathway and may alleviate overwork-induced physical fatigue.

Physical aging by soft ultrasonic wave enhances ethanol metabolism: metabolic process of wine as followed by 400 mhz 1H-NMR spectroscopy.

In natural aging of spirits or wine, the dynamic structure of ethanol-water clusters changes to a smaller and more uniform state. Through experience we know that naturally aged ones have higher metabolism than the non-aged ones. Also, the same effect as natural aging can be obtained in various types of spirits or wines by the treatment for a period of time with soft ultrasonic wave (US). In this study, we compared ethanol metabolism in human subjects dosed with non-treated white wine (control = CON) and with US treated wine. Ethanol levels in human sera were followed by 400 MHz 1H-NMR spectroscopy after administration of wine doses. Experimental results indicated that ethanol metabolism was enhanced 18% in subjects when US treated wine was used rather than when non-treated (CON) was used. Other experiments using rabbits showed that a 20% ethanol-aqueous solution was absorbed 18% more rapidly by the group dosed with US wine than by the CON group. From these experimental facts, it was theorized that ethanol metabolism depends on the rapidity of ethanol absorption in the human body. And it can be concluded that US treatment brings about the same effect on spirits or wines as natural aging.

Mechanomyographic investigation of muscle contractile properties in preadolescent boys.

The aim of the present study was to elucidate the muscle contractile properties of preadolescent boys using the mechanomyogram (MMG) and electromyogram (EMG). In 8 preadolescent boys and 10 male young adults, the EMG and MMG were recorded from quadriceps muscles during isometric knee extension contractions at levels of 10 to 80% MVC. The relationship of EMG and MMG to absolute force was approximated by a regression line (r > 0.8). The regression line of the EMG in preadolescent was located above that of the adults, whereas the MMG in both subject groups was fitted by virtually the same regression line. Thus, the MMG seems to be indicative of absolute force in preadolescents as well as young adults. Both groups showed a non-linear increase in the EMG and MMG with relative force (%MVC). The EMG and %MVC relationship was not significantly different between the two groups, whereas the MMG in preadolescents was smaller than that of adults over the range of force studied. The MMG increment with %MVC was more remarkable above 30 or 40% MVC in both groups; however, the rate of the increment in preadolescents was smaller than that in adults. The MMG and %MVC relationship suggests that the muscle contractile properties in preadolescent boys are characterized by immaturity of the fast twitch fibers.

A role of PKC in the improvement of energy metabolism in preconditioned heart.

OBJECTIVE: A possible link between activation of PKC and improvement of energy metabolism during reperfusion in ischemic preconditioning hearts was examined. METHODS: Isolated perfused rat hearts were preconditioned by 5-min ischemia and 5-min reperfusion in the presence and absence of a PKC inhibitor polymyxin B (50 microM) and then subjected to 40-min sustained ischemia and subsequent 30-min reperfusion. In another set of experiments, the hearts pretreated with and without a PKC activator PMA (15 pmol/5 min) were subjected to the sustained ischemia and reperfusion. Myocardial high-energy phosphates, glycolytic intermediates and mitochondrial oxygen consumption capacity were determined at appropriate experimental sequences. RESULTS: Preconditioning enhanced the recovery of cardiac function such as left ventricular developed pressure, heart rate and rate-pressure product of the reperfused heart, suppressed the release of creatine kinase, enhanced the reperfusion-induced restoration of myocardial high-energy phosphates, attenuated the reperfusion-induced accumulation in glucose 6-phosphate and fructose 6-phosphate contents, abolished the ischemia-induced increase in tissue lactate content and prevented the ischemia-induced decrease in mitochondrial oxygen consumption capacity. Treatment of the perfused heart with PMA mimicked the effects of preconditioning on post-ischemic contractile function, enzyme release, levels of myocardial energy store, glycolytic intermediates and lactate, and mitochondrial function. Polymyxin B-treatment abolished the preconditioning-induced recovery of post-ischemic contractile function, the suppression of the release of CK, the restoration of myocardial energy store, and the preservation of mitochondrial function, whereas it did not cancel the improvement of glycolytic intermediate levels and the reduction in tissue lactate accumulation. Post-ischemic contractile function was closely related to restoration of high-energy phosphates and mitochondrial oxygen consumption capacity in all hearts subjected to ischemia/reperfusion. CONCLUSION: The results suggest that activation of PKC and preservation of mitochondrial function are closely linked with each other in the preconditioned heart, which may lead to the improvement of post-ischemic contractile function.