Branched-chain amino acid supplementation attenuates muscle soreness, muscle damage and inflammation during an intensive training program.

AIM: The aim of this paper was to assess the effects of branched-chain amino acid (BCAA) supplementation on muscle soreness, muscle damage and inflammation during an intensive training program. METHODS: Twelve long-distance runners (20 + or - 1 year-old) participated in a double-blinded crossover designed study conducted during two intensive training periods (three-day). The subjects were provided either a drink containing BCAA (0.8% BCAA in a 3.5% carbohydrate solution; 2,500 mL/day) or an isocaloric placebo drink during each training period. All subjects completed the same training program (total running distance: males: 86 km, females: 64 km), and ate the same meals during the training period. Whole body muscle soreness and fatigue sensation were measured in the morning before and during the training period by Visual Analogue Scale method. Plasma creatine kinase (CK), lactate dehydrogenase (LDH), and granulocyte elastase (GEL) levels were measured as indicators of muscle damage and inflammation before and after the training period. RESULTS: Muscle soreness and fatigue sensation during the training period in the BCAA trial were lower than those in the placebo trial (-32% and -24%, respectively; P<0.05). The plasma CK, LDH, and GEL levels after the training program in the BCAA trial were lower than those in the placebo trial (-21%, -6%, and -15%, respectively; P<0.05). CONCLUSIONS: These results demonstrate that BCAA supplementation during an intensive training program effectively reduces the muscle soreness and fatigue sensation, and that the perceived changes could be attributed to the attenuation of muscle damage and inflammation.

Branched-chain amino acids supplementation attenuates the accumulation of blood lactate dehydrogenase during distance running.

AIM: We investigated the effect of branched-chain amino acids (BCAA) supplementation on tissue damage during distance running. METHODS: Eight male distance runners (mean +/- standard deviation; age: 20.4+/-1.2 years, body weight: 58.4+/-4.2 kg) participated in a double blinded cross over designed study conducted during training camp. During each intervention period, the subjects were asked to participate in a 25-km run, and the blood BCAA and lactate dehydrogenase (LDH) level, an index of tissue damage, were measured pre- and post-run. Either a drink containing BCAA (0.4% BCAA in a 4% carbohydrate solution) or an iso-calorie placebo drink was provided to the subjects 5 times during the run without any restriction in the volume. RESULTS: The total volume of the drink consumed by the subjects did not differ substantially between the trials: 591+/-188 (2.36 g BCAA) vs 516+/-169 mL in BCAA and placebo trial, respectively. During the run, the blood BCAA concentration was maintained in the BCAA trial. However, the blood BCAA concentration level tended to decrease in the placebo trial (P<0.1). The extent of the blood LDH increase in the BCAA trial was significantly less than that of the placebo trail (48% vs 58%, P<0.05). CONCLUSION: Maintaining the blood BCAA level throughout a long distance run contributes to a reduction in the LDH release and, therefore, the effect of BCAA supplementation is suggested to reduce the degree of muscle damage.

Analysis of pharmacological effect and molecular mechanisms of a traditional herbal medicine by global gene expression analysis: an exploratory study.

BACKGROUND AND OBJECTIVES: Although traditional herbal medicines have been used for more than 2000 years, there are few studies on their molecular mechanisms of action. We examined the pharmacological effect and mechanism of action of a traditional herbal medicine (Kososan) with global gene expression analysis using a DNA chip. METHODS: Fourteen healthy subjects were given the herbal medicine (Kososan) for 2 weeks and interviewed. Peripheral blood was collected before and after 2-week medication. Based on the outcomes of the interview, the subjects were divided into responders and non-responders. We analysed the blood samples from responder and non-responder groups, respectively, using a DNA chip. RESULTS: In the Kososan responder group, 70 genes were over-expressed (2-fold or more), and their over-expression was normalized by Kososan (0.5-fold or less), whereas 24 genes were under-expressed (0.5-fold or less), and the under-expression was normalized by Kososan (2-fold or more). CONCLUSION: This exploratory study suggests that gene expression profiling is a possible approach for studying the effects of complex herbal remedies. Some of the genes studied seem to have functions related to the pharmacological effects of Kososan, which have been known for a long time.

In vitro formation of corticospinal synapses in an organotypic slice co-culture.

In order to study biological properties of the corticospinal tract, we have reconstructed this system in an in vitro slice culture preparation. Motor cortex and spinal cord slices, prepared from newborn rats, were co-cultured on pored membranes for 16-24 days. Anterograde labeling with biocytin showed that substantial neural connections had formed between the cortex and spinal cord slices. Retrograde labeling with horseradish peroxidase or 1,1'-dioctadecyl-3,3,3',3'-tetramethylindocarbocyanine perchlorate demonstrated that the parent cells were located primarily in the deeper layer of the cortex, as is found in vivo. Stimulation of the deep layer of the cortex elicited extracellular postsynaptic responses and intracellular excitatory postsynaptic potentials (EPSPs) in the co-cultured spinal cord that were mediated by the 1-amino-3-hydroxy-5-methyl-4-isoxazolepropionate/ kainate-type glutamate receptor. The intracellular injection of biocytin after EPSPs were recorded showed that one-third of these cells were large stellate cells, which are thought to be motoneurons, while a large portion of the remaining labeled cells were bipolar cells of smaller sizes. Using this reconstructed in vitro preparation, we recorded field EPSPs (fEPSPs) along a 100-microm-interval lattice in the spinal gray matter, which allowed the quantitative evaluation of synapse formation. The fEPSP amplitudes were more than two-fold larger when the forelimb cortex was co-cultured with cervical cord rather than lumbar cord. However, hindlimb cortex did not show this preference. The fEPSP amplitudes were more than twice as large when the dorsal side of the spinal cord was adjacent to the cortex than the ventral side. In summary, we have reconstructed the corticospinal projection and synapses in vitro using cortical and spinal explants. This system allows for an efficient quantitative evaluation of synapse formation and for studies of postsynaptic cells. Our results suggest that synapse formation shows preferences along and perpendicular to the neuraxis of the spinal cord.

Hyperbaric oxygen therapy given 30 minutes after spinal cord ischemia attenuates selective motor neuron death in rabbits.

OBJECTIVE: Spinal cord ischemia sometimes causes paraplegia because the spinal motor neuron cells are vulnerable to ischemia. Although various protective remedies for spinal cord injury have been reported, there have been few established clinical methods. Although hyperbaric oxygen (HBO) has been used clinically as a treatment for ischemia, the reason for its effectiveness is still uncertain because sufficient experimental data are lacking. DESIGN: Prospective, randomized, controlled study. SETTING: Experimental animal research laboratory in a university research center. SUBJECTS: Twenty-three Japanese white rabbits, weighing 2-3 kg. INTERVENTIONS: A modified rabbit spinal cord ischemia model of infrarenal aortic occlusion for 15 mins was employed. Rabbits were randomly assigned to four groups; the rabbits in group A did not undergo ischemic insults (n = 5). The rabbits in groups B and C underwent ischemic insult for 15 mins, followed by 1 hr of HBO treatment at 3 atm absolute with 100% oxygen at 30 mins (n = 6) or 6 hrs (n = 7) after reperfusion, respectively. The rabbits in group D underwent ischemic insult for 15 mins without HBO treatment (n = 5). MEASUREMENTS AND MAIN RESULTS: We observed neurologic functions for 14 days. The sections of the spinal cords were stained with hematoxylin and eosin, and the number of spinal motor neurons in ventral region was counted by light microscopy. All rabbits in groups A and B could stand, whereas all rabbits in groups C and D showed irreversible paraplegia on days 2 and 14 after reperfusion. Spinal motor neurons in ventral gray matter in groups C and D decreased significantly compared with those in groups A and B. CONCLUSIONS: HBO therapy shortly after ischemic insult had protective effects against ischemic spinal cord damage. However, delayed treatment with HBO did not change the prognosis.

Adenovirus-mediated glial cell line-derived neurotrophic factor gene delivery reduces motor neuron injury after transient spinal cord ischemia in rabbits.

OBJECTIVE: Glial cell line-derived neurotrophic factor (GDNF) has protective effects on various injuries involving the central and peripheral nervous systems in vitro and vivo. However, the possible protective effect of GDNF on spinal cord ischemia and the exact mechanism involved in the ameliorative effect of GDNF on ischemic spinal cord injuries are not fully understood. Therefore, we investigated the possible protective effect of the adenovirus-mediated GDNF gene delivery on transient spinal cord ischemia in rabbits. METHODS: The adenoviral vector (lacZ gene as a control or GDNF gene contained) was injected directly into the lumbar spinal cord via a needle inserted into the dorsal spine 2 days before the animal was subjected to 15 minutes of spinal cord ischemia induced by infrarenal aortic occlusion. In situ terminal deoxynucleotidyl transferase (TdT)-mediated dUTP-biotin nick-end labeling (TUNEL staining) was performed, and temporal profiles of the GDNF and caspase-3 (caspase-3 is the marker of apoptotic change) immunoreactivity were investigated. RESULTS: In the control rabbit, the majority of motor neurons showed selective cell death at 7 days of reperfusion. Immunocytochemistry showed that in situ TUNEL staining was selectively detected at 2 days of reperfusion in motor neuron nuclei. GDNF and caspase-3 were selectively induced in the motor neuron cells at 8 hours of reperfusion. In the GDNF-treated group, a large population of motor neuron cells was still surviving at 7 days after having been subjected to 15 minutes of ischemia. Unlike the control group, the GDNF-treated group expressed GDNF persistently. Induction of TUNEL staining and immunoreactivity for caspase-3 were greatly reduced by the GDNF treatment. CONCLUSION: These results suggest that the reduction in motor neuron death by GDNF was greatly associated with a reduction in DNA fragmentation and apoptotic signals of the caspase-3 cascade; they further suggest a great potential for gene therapy for paraplegic patients in the future.

Orally administered decoction of Kampo (Japanese herbal) medicine, "Juzen-Taiho-To" modulates cytokine secretion and induces NKT cells in mouse liver.

The effects of orally administered decoction of Juzen-Taiho-To (JTT; Si-Quan-Da-Bu-Tang in Chinese) on cytokine production in hepatic lymphocytes were studied in mice. JTT was found to increase interferon gamma (IFN-gamma), as well as interleukin-4 (IL-4), IL-5 and IL-6 secretion from stimulated hepatic lymphocytes, whereas IL-2 secretion was reduced. The number of IFN-gamma- and IL-4-spot forming cells (SFC) were not changed by administration of JTT. These results suggest that modulation of cytokine secretion by JTT might not be due to changes in the number of cytokine secreting cells within liver lymphocytes. CD4/CD8 ratio and alphabeta/gammadelta T cell receptor (TCR) ratio in hepatic lymphocytes were not changed. However, flow cytometric analysis revealed that the population of CD3 positive intermediate cells in NK positive cells (NKT cells) was increased after oral administration of JTT. The population of CD3int IL-2Rbeta+ cells was also increased. The induction of NKT cells by JTT was reduced by injection of 2-chloroadenosine. JTT enhanced transcription of IL-12 mRNA in liver. From these results, it may be concluded that a rise in NKT cell population contributes, at least partially, to the modulating effect of JTT on cytokine production in liver lymphocytes, and macrophages. The production of IL-12 in liver may also contribute to this NKT induction.

Restoration of shoulder movement in quadriplegic and hemiplegic patients by functional electrical stimulation using percutaneous multiple electrodes.

The purpose of this study is to restore the motion of the paralyzed shoulder caused by upper motor neuron disorders using functional electrical stimulation (FES). Percutaneous wire electrodes were implanted into twelve muscles of the shoulder in six patients with stroke or cervical spinal cord injury. The motion of the paralyzed shoulder was controlled by a portable FES computer system, with the three standard stimulation patterns for restoring motion of 90 degrees flexion to 90 degrees horizontal abduction, 90 degrees flexion to 20 degrees horizontal adduction, and 90 degrees abduction to 90 degrees horizontal adduction. Shoulder movements were repeatedly controlled according to the created stimulation patterns in five of the patients. The two dimensional motion analyzer also confirmed shoulder control over a satisfactorily broad range of excursion. One hemiplegic patient, who was a signboard painter, had his paretic left upper extremity improved by FES, and he drew a large picture on a board with his normal right hand and, with his affected left arm against the wall, to support his trunk. This may be a world first case of producing shoulder motion through FES.

B-cell proliferation activity of pectic polysaccharide from a medicinal herb, the roots of Bupleurum falcatum L. and its structural requirement.

Pectic polysaccharide fraction (BR-2) containing pharmacologically active pectic polysaccharide, bupleuran 2IIc, which was prepared from a medicinal herb, the roots of Bupleurum falcatum L., was administered orally to C3H/HeJ mice for 7 consecutive days. Proliferative responses of spleen cells were enhanced in the presence of the purified pectic polysaccharide, bupleuran 2IIc, but another B-cell mitogen, lipopolysaccharide (LPS) did not give a similar effect. In vitro studies using spleen cells showed that bupleuran 2IIc also stimulated lymphocytes, depleted of adherent cells or T cells. Bupleuran 2IIc treatment increased subpopulation of CD25+ and surface immunoglobulin M-positive (sIgM+) lymphocytes. Non-specific immunoglobulin secretion of spleen cells treated with bupleuran 2IIc was increased according to the culture time, and coexistence of interleukin-6 (IL-6) enhanced the secretion more than that of bupleuran 2IIc alone. These results suggest that bupleuran 2IIc proliferates B cells in the absence of macrophages, and the resulting activated B cells are then induced into antibody-forming cells in the presence of IL-6. Among the structural region of bupleuran 2IIc, ramified region (PG-1), which consists of rhamnogalacturonan core rich in neutral sugar chain, showed the potent mitogenic activity suggesting it to be an active site. Mitogenic activity of bupleuran 2IIc was reduced in the presence of antipolysaccharide antibody (antibupleuran 2IIc/PG-1-IgG), which recognizes the ramified region of bupleuran 2IIc as the antigenic epitope. Mitogenic activity of bupleuran 2IIc was also reduced by the addition of beta-d-GlcpA-(1-->6)-beta-d-Galp-(1-->6)-d-Galp or beta-d-GlcpA-(1-->6)-d-Galp, which are a part of the epitopes of antibupleuran 2IIc/PG-1-IgG. These results suggest that the epitopes in bupleuran 2IIc act as active sites of the polysaccharide during mitogenic activity.

Characterization of antigenic epitopes in anti-ulcer pectic polysaccharides from Bupleurum falcatum L. using several carbohydrases.

A polyclonal antibody (anti-bupleuran 2IIc/PG-1-IgG) against the "ramified" region (PG-1) of an anti-ulcer pectic polysaccharide was prepared and its antigenic epitopes were analyzed by using several carbohydrases. Enzymatic removal of arabinosyl residues from PG-1 by endo-(1-->5)-alpha-L-arabinanase (from Aspergillus niger) did not reduce the binding ability of anti-bupleuran 2IIc/PG-1-IgG to PG-1. When the endo-(1-->5)-alpha-L-arabinanase-resistant fraction (EA-1) was digested with rhamnogalacturonase A (rRGase A from A. aculeatus), a high-molecular-mass fragment fraction (RA-1) and an oligosaccharide fraction (RA-3) were obtained. RA-3 contained at least four kinds of oligosaccharides liberated from the rhamnogalacturonan core. This partial removal of the rhamnogalacturonan core in EA-1 also did not reduce the binding of the antibody to the polysaccharide. Further digestion of RA-1 with exo-(1-->3)-beta-D-galactanase (from Irpex lacteus), gave a high-molecular-mass fragment (EXG-1) and a trace of oligosaccharides (EXG-3). Methylation and FABMS analyses indicated that EXG-3 contained mono- and di-galactosyl oligosaccharides possessing terminal GlcA or GlcA4Me. Removal of the EXG-3 fraction from RA-1 by exo-(1-->3)-beta-D-galactanase significantly reduced the ability of the binding of the antibody to the polysaccharide. When PG-1 was digested with endo-(1-->6)-beta-D-galactanase (from Trichoderma viride) or beta-D-glucuronidase (from A. niger), the reactivities of both enzyme-resistant fractions to the antibody were decreased in comparison with that of PG-1. Both radish arabinogalactan (containing GlcA4Me) and beta-D-GlcpA-(1-->6)-beta-D-Galp-(1-->6)-D-Galp were shown to inhibit the reactivity of PG-1 to the antibody by competitive ELISA. These results suggest that 6-linked galactosyl chains containing terminal GlcA or GlcA4Me attached to (1-->3)-beta-D-galactosyl chains, are important sugar residues in the antigenic epitopes of the "ramified" region of bupleuran 2IIc.

Suppressive effect of the angiogenesis inhibitor TNP-470 on the development of carcinogen-induced hepatic nodules in rats.

Tumor metastasis can be prevented by inhibiting angiogenesis. In the present study, we have demonstrated that the angiogenesis inhibitor TNP-470 also suppresses the development of primary hepatic nodules. Hepatocarcinogenesis was performed by the feeding of 2-acetylaminofluorene to hepatectomized rats during 8-14 weeks of age. Predominantly arterial-to-portal circulation and sinusoidal capillarization were determined by the staining of nodules with arterially infused ink and immunostaining for factor VIII-related antigen, respectively. Intraperitoneal administration of 30 mg/kg b.w. of TNP-470 twice a week significantly reduced the number of hepatic nodules. Among the nodules, hyperplastic nodules stained with ink, atypical hyperplastic nodules and hepatocellular carcinoma, all of which possess structurally altered sinusoidal endothelial cells or capillary-type endothelial cells, were dramatically decreased in number. Suppression was observed equally in nodules of all sizes. TNP-470 was more effective when administered during 8-20 weeks than during 14-26 weeks. In contrast, ink-non-stained hyperplastic nodules, which have normal sinusoidal endothelial cells, were not affected at all. The present results indicate that TNP-470 suppresses the development of primary hepatic nodules whose microvessels are capillaries or transitional forms from sinusoids to capillaries.

Salvage therapy for relapsed or refractory childhood acute lymphocytic leukemia by alternative administration a lymphoid- and myeloid-directed chemotherapeutic regimen consisting of dual modulation of ara-C, hydroxyurea, and etoposide.

Risk-directed chemotherapeutic regimens in recent use have improved the prognosis of children with acute lymphocytic leukemia (ALL). However, many patients relapse during or shortly after cessation of the initial continuation chemotherapy. Since achievement of a second complete remission (CR) is the initial step in successful retreatment effort, it is important to develop salvage protocols for children with relapsed or refractory ALL. In the present study, we developed a new salvage protocol (MLL-93) and applied the concept of dual chemical modulation of cytarabine, hydroxyurea, and etoposide with the alternative administration of high doses of myeloid- and lymphoid-directed agents. We also planned to perform allogeneic bone marrow transplantation (BMT) following a CR if patients had HLA-identical donor(s). The six patients treated with the MLL-93 protocol achieved a second CR. One patients in CR died of interstitial pneumonia after an unrelated allogeneic BMT. The other five patients have been in CR for 12-41 months. We suggest that the concepts of alternative administration of lymphoid- and myeloid-directed drugs and biochemical modulation are useful in the treatment of children with relapsed or refractory ALL.

Isolation and identification of hematopoietic stem cell-stimulating substances from Kampo (Japanese herbal) medicine, Juzen-taiho-to.

We have previously found that TJ-48 has the capacity to accelerate recovery from hematopoietic injury induced by radiation and the anti-cancer drug mitomycin C (MMC). The effects are found to be due to its stimulation of spleen colony-forming unit (CFU-S) counts on day 14. In the present study, we attempt to isolate and purify the active components in TJ-48 extracts using a new in vitro hematopoietic stem cell (HSC) assay method. n-Hexane extract from TJ-48 shows a significant stimulatory activity. The extract is further fractionated by silica gel chromatography and HPLC in order to identify its active components. 1H-NMR and GC-EI-MS indicate that the active fraction is composed of free fatty acids (oleic acid and linolenic acid). When 27 kinds of free fatty acids (commercially available) are tested using the HSC proliferating assay, oleic acid, elaidic acid, and linolenic acid are found to have potent activity. The administration of oleic acid to MMC-treated mice enhances CFU-S counts on days 8 and 14 to twice the control group. These findings strongly suggest that fatty acids contained in TJ-48 actively promote the proliferation of HSCs. Although many mechanisms seem to be involved in the stimulation of HSC proliferation, we speculate that at least one of the signals is mediated by stromal cells, rather than any direct interaction with the HSCs.

[Bone metabolism and phosphorus metabolism in patients with prostate cancer: paracrine and endocrine effects produced by prostate neoplasm].

We examined whether paracrine factors produced by prostate cancer cells can modulate bone metabolism in proportion to the volume of cancer cells in bone metastasis. Endocrine factors produced by prostate cancer cells affect both phosphate and 1,25-dihydroxyvitamin D metabolisms. Levels of urine pyridinoline (U-Pyr) excretion and serum carboxy-terminal propeptide of type 1 procollagen (P1CP) in patients with bone metastasis were significantly higher than those in patients without bone metastasis (P < 0.05). In patients with bone metastasis (n = 17), serum prostate-specific antigen (PSA) levels were significantly correlated with the levels of U-Pyr and urine deoxypyridinoline (U-dPyr) excretion, serum cross-linked carboxyterminal telopeptide of type 1 collagen (1CTP), and P1CP levels (p < 0.05). However, serum PSA levels were not correlated with U-Pyr, U-dPyr excretions, serum 1CTP and P1CP levels in patients without bone metastasis. Therefore, prostate cancer cells appear to have some paracrine effects on bone cells. In controls (n = 15), serum 1,25-dihydroxyvitamin D levels (1,25-(OH)2D) were inversely correlated with serum phosphorus levels (P < 0.01). In prostate cancer patients with bone metastasis, the ability to regulate the serum 1,25-(OH)2D levels in response to serum phosphorus levels is lost. These results suggest that endocrine factors produced by prostate cancer cells disturb the regulation of serum 1,25-(OH)2D in response to serum phosphorus levels.

Reduced resuscitation fluid volume for second-degree experimental burns with delayed initiation of vitamin C therapy (beginning 6 h after injury).

We studied the hemodynamic effects of delayed initiation (6 h postburn) of antioxidant therapy with high-dose vitamin C in second-degree thermal injuries. Seventy percent body surface area burns were produced by subxiphoid immersion of 12 guinea pigs into 100 degrees C water for 3 s. The animals were resuscitated with Ringer's lactate solution (R/L) according to the Parkland formula (4 ml/kg/% burn during the first 24 h) from 6 h postburn, after which the resuscitation fluid volume was reduced to 25% of the Parkland formula volume. Animals were divided into two groups. The vitamin C group (n = 6) received R/L to which vitamin C (340 mg/kg/24 h) was added after 6 h postburn. The control group (n = 6) received R/L only. Both groups received identical resuscitation volumes. Heart rates, mean arterial blood pressure, cardiac output, hematocrit level, and water content of burned and unburned tissue were measured before injury and at intervals thereafter. No animals died. There were no significant differences in heart rates or blood pressures between the two groups throughout the 24-h study period. The vitamin C group showed significantly (P < 0.05) lower hematocrits 8 and 24 h postburn, and higher cardiac outputs after 7 h postburn. At 24 h postburn, the burned skin in the vitamin C group had a significantly (P < 0.05) lower water content (73.1 +/- 1.1) than that of the control group (76.0 +/- 0.8). In conclusion, delayed initiation of high-dose vitamin C therapy beginning 6 h postburn with 25% of the Parkland formula volume significantly reduced edema formation in burned tissue, while maintaining stable hemodynamics.

Effect of a single bout of exercise and beta-carotene supplementation on the urinary excretion of 8-hydroxy-deoxyguanosine in humans.

We investigated the effects of acute exhaustive exercise and beta-carotene supplementation on urinary 8-hydroxy-deoxyguanosine (8-OHdG) excretion in healthy nonsmoking men. Fourteen untrained male (19-22 years old) volunteers participated in a double blind design. The subjects were randomly assigned to either the beta-carotene or placebo supplement group. Eight subjects were given 30 mg of beta-carotene per day for 1 month, while six subjects were given a placebo for the same period. All subjects performed incremental exercise to exhaustion on a bicycle ergometer both before and after the 1-month beta-carotene supplementation period. The blood lactate and pyruvate concentrations significantly increased immediately after exercise in both groups. The baseline plasma beta-carotene concentration was significantly 17-fold higher after beta-carotene supplementation. The plasma beta-carotene decreased immediately after both trials of exercise, suggesting that beta-carotene may contribute to the protection of the increasing oxidative stress during exercise. Both plasma hypoxanthine and xanthine increased immediately after exercise before and after supplementation. This thus suggests that both trials of exercise might enhance the oxidative stress. The 24-h urinary excretion of 8-OHdG was unchanged for 3 days after exercise before and after supplementation in both groups. However, the baseline urinary excretion of 8-OHdG before exercise tended to be lower after beta-carotene supplementation. These results thus suggest that a single bout of incremental exercise does not induce the oxidative DNA damage, while beta-carotene supplementation may attenuate it.

Detection and tissue distribution of anti-ulcer pectic polysaccharides from Bupleurum falcatum by polyclonal antibody.

Anti-sera against the "ramified" region (PG-1) of an anti-ulcer polysaccharide (bupleuran 2IIc), which was purified from the roots of Bupleurum falcatum L, were obtained by immunization of rabbits, and a polyclonal anti-bupleuran 2IIc/PG-1-antibody of the IgG class was purified by Protein G and "ramified" region (PG-1) immobilized affinity chromatographies. The antigenic specificity of anti-bupleuran 2IIc/PG-1-IgG was examined by a two-site sandwich ELISA which was developed as an improved method for microanalysis of bupleuran 2IIc using a biotinylated antibody. Another pectin from B. falcatum and anti-complementary pectins from Angelica acutilaba and Glycyrrhiza uralensis also showed significant reactivity to anti-bupleuran 2IIc/PG-1-IgG, although these reactivities were lower than that of bupleuran 2IIc. Other polysaccharides tested such as apple pectin, araban, yeast mannan, pullulan, etc., had negligible reactivity. The KDO-containing region and oligogalacturonides, which were obtained by endo-alpha-(1-->4)-polygalacturonase digestion of bupleuran 2IIc, were also not significantly recognized by anti-bupleuran 2IIc/PG-1-IgG. When bupleuran 2IIc was administered to the mice i.v., the polysaccharide disappeared from the circulation within 24 h and was mainly detected in the liver by the two-site sandwich ELISA. However the clearance of bupleuran 2IIc from the circulation was delayed by pretreatment with iota-carrageenan. When the crude polysaccharide fraction (BR-2), containing mainly bupleurans 2IIb and 2IIc from B.falcatum, was administrated orally to the mice, the polysaccharides were detected in the liver and Peyer's patch.

Effect of capacitively coupled electric fields on an experimental model of delayed union of fracture.

To evaluate the effect of capacitively coupled electric fields (CCEF) on delayed union of fractures, an experimental model of delayed union was produced in the radius of rabbits, and the process of healing was investigated by radiography, bone mineral density (BMD) measured with dual energy x-ray absorptiometry, and histological survey. It was confirmed radiographically and histologically that callus formation was enhanced in the group treated with CCEF. After stimulation, the average BMD increased more than 18% compared with the controls. Our experiment on a delayed union model suggests that CCEF is effective for the treatment of delayed union of fractures.