Search

In this study, the application of medium- and high-entropy (MoTa)xNbTiZr alloys in biomedical implants was systematically analyzed. The alloy with the best combination of mechanical properties was selected and characterized for in vitro and in vivo response for the first time to examine its biomedical properties. A logarithmic increase in the hardness and the yield strength was observed as a function of the Mo and Ta content. Alloys with up to 0.4 mol fraction of Mo and Ta showed a plastic strain of more than 30 % under compression. The nanoindentation results showed that the addition of Mo and Ta increased the elastic modulus of the system linearly. It was surmised that the addition of Ta and Mo above a critical concentration (mole fraction = 0.4) was unfavorable from a biomedical perspective as it increased the brittleness and elastic modulus and decreased the ductility of the system. Therefore, the (MoTa)0.2NbTiZr alloy is a potential structural material for biomedical implants because of its excellent strength and ductility. The developed alloy was investigated for its corrosion properties and compared with commercial biomedical alloys. Furthermore, the biocompatibility of the alloy was examined using an in vivo examination. The alloy was implanted in the skeletal muscles of mice for four weeks and the histology of the surrounding tissue was studied. The alloy exhibited strong passive behavior in a phosphate buffer solution and non-toxic soft tissue response.

Worldwide, obesity has become a major risk factor associated with health risks such as diabetes, hypertension, hypercholesterolemia, cardiovascular disease, stroke, and certain forms of cancer. In this study, we estimated the anti-obesity effect of the bacterial strain Lactobacillus plantarum LB818 (designated as LB818) using male C57BL/6J mice, which were treated with high-fat diet (HFD) to induce obesity. Next, LB818 (109 colony-forming units [CFU]/mL) was orally administered for 8 weeks. The results showed that feeding HFD+LB818 (109 CFU/mL) ameliorated body weight gain and decreased total body fat by regulating fasting glucose levels in HFD-fed mice. LB818 treatment significantly lowered aspartate aminotransferase, alanine aminotransferase (ALT), total cholesterol (TC), triglyceride (TG), and elevated high-density lipoprotein levels in serum and decreased deposition of fat droplets in liver. LB818 treatment increased the respective abundances of essential bacteria, including Bacteroidetes, Akkermansia, Bifidobacterium, Lactobacillus, and increased the Bacteroidetes:Firmicutes ratio; however, it significantly decreased the levels of Firmicutes. Taken together, this study demonstrates that LB818 is effective in attenuating obesity and hepatic steatosis and regulated gut microbiota in HFD-fed obese mice.

본 연구의 목적은 12주간 짐볼 코어 안정화 운동 프로그램을 통해 고등부 축구선수들의 신체조성 및 8방향 균형능력에 미치는 영향을 알아보는 것이다. 연구 대상자는 S시 Y고등학교 축구부 선수 28명을 12주간의 짐볼 코어 안정화 운동프로그램 실시 후 신체조성 및 8방향 균형능력을 사전·사후 비교하였다. 본 연구의 결과는 다음과 같다. 첫째 고등부 축구선수의 신체조성에 있어, 짐볼 코어 안정화 프로그램을 통해 몸무게, 체지방, 골격근, BMI 모든 항목에서 유의한 차이가 나타나지 않았다(p

Helicobacter pylori are etiological agents in the development of gastritis, gastroduodenal ulcers, gastric cancer, and mucosa-associated lymphoid tumors. Our previous investigations demonstrated that standardized combined plants extracts (Rubus crataegifolius and Ulmus macrocarpa) inhibit the growth of H. pylori in in vitro experiments. Also, we demonstrated that Gardenia jasminoides is effective in preventing gastritis and gastric ulcers in animal experiments. In the present work, we tested the standardized combined three plant extract (RUG-com) on the mouse model of H. pylori infectious disease to examine the effects of RUG-com on both the prevention and curing on the stomachs of infected mice. After the final administrations, biopsy samples of gastric mucus were assayed for bacterial numbers, biochemical analysis, inflammatory scores, and histology. Treatment with standardized plants extracts, single or combined, reduced the H. pylori load compared with the control. Treatment also significantly (P

[Purpose] Strenuous exercise often induces skeletal muscle damage, which results in impaired performance. Sphingolipid metabolism contributes to various cellular processes, including apoptosis, stress response, and inflammation. However, the relationship between exercise-induced muscle damage and ceramide (a key component of sphingolipid metabolism), is rarely studied. The present study aimed to explore the regulatory role of sphingolipid metabolism in exercise-induced muscle damage. [Methods] Mice were subjected to strenuous exercise by treadmill running with gradual increase in intensity. The blood and gastrocnemius muscles (white and red portion) were collected immediately after and 24 h post exercise. For 3 days, imipramine was intraperitoneally injected 1 h prior to treadmill running. [Results] Interleukin 6 (IL-6) and serum creatine kinase (CK) levels were enhanced immediately after and 24 h post exercise (relative to those of resting), respectively. Acidic sphingomyelinase (A-SMase) protein expression in gastrocnemius muscles was significantly augmented by exercise, unlike, serine palmitoyltransferase-1 (SPT-1) and neutral sphingomyelinase (N-SMase) expressions. Furthermore, imipramine (a selective A-SMase inhibitor) treatment reduced the exercise-induced CK and IL-6 elevations, along with a decrease in cleaved caspase-3 (Cas-3) of gastrocnemius muscles. [Conclusion] We found the crucial role of A-SMase in exercise-induced muscle damage.

Background: Preterm birth is a known leading cause of neonatal mortality and morbidity. The underlying causes of pregnancy-associated complications are numerous, but infection and inflammation are the essential high-risk factors. However, there are no safe and effective preventive drugs that can be applied to pregnant women. Objective: The objectives of the study were to investigate a natural product, Abeliophyllum distichum leaf (ADL) extract, to examine the possibility of preventing preterm birth caused by inflammation. Methods: We used a mouse preterm birth model by intraperitoneally injecting lipopolysaccharides (LPS). ELISA, Western blot, real-time PCR and immunofluorescence staining analyses were performed to confirm the anti-inflammatory efficacy and related mechanisms of the ADL extracts. Cytotoxicity and cell death were measured using Cell Counting Kit-8 (CCK-8) analysis and flow cytometer. Results: A daily administration of ADL extract significantly reduced preterm birth, fetal loss, and fetal growth restriction after an intraperitoneal injection of LPS in mice. The ADL extract prevented the LPS-induced expression of TNF-&alpha, in maternal serum and amniotic fluid and attenuated the LPS-induced upregulation of placental proinflammatory genes, including IL-1&beta, IL-6, IL-12p40, and TNF-&alpha, and the chemokine gene CXCL-1, CCL-2, CCL3, and CCL-4. LPS-treated THP-1 cell-conditioned medium accelerated trophoblast cell death, and TNF-&alpha, played an essential role in this effect. The ADL extract reduced LPS-treated THP-1 cell-conditioned medium-induced trophoblast cell death by inhibiting MAPKs and the NF-&kappa, B pathway in macrophages. ADL extract prevented exogenous TNF-&alpha, induced increased trophoblast cell death and decreased cell viability. Conclusions: We have demonstrated that the inhibition of LPS-induced inflammation by ADL extract can prevent preterm birth, fetal loss, and fetal growth restriction.

Background: Preterm birth is a known leading cause of neonatal mortality and morbidity. The underlying causes of pregnancy-associated complications are numerous, but infection and inflammation are the essential high-risk factors. However, there are no safe and effective preventive drugs that can be applied to pregnant women. Objective: The objectives of the study were to investigate a natural product, Abeliophyllum distichum leaf (ADL) extract, to examine the possibility of preventing preterm birth caused by inflammation. Methods: We used a mouse preterm birth model by intraperitoneally injecting lipopolysaccharides (LPS). ELISA, Western blot, real-time PCR and immunofluorescence staining analyses were performed to confirm the anti-inflammatory efficacy and related mechanisms of the ADL extracts. Cytotoxicity and cell death were measured using Cell Counting Kit-8 (CCK-8) analysis and flow cytometer. Results: A daily administration of ADL extract significantly reduced preterm birth, fetal loss, and fetal growth restriction after an intraperitoneal injection of LPS in mice. The ADL extract prevented the LPS-induced expression of TNF-α in maternal serum and amniotic fluid and attenuated the LPS-induced upregulation of placental proinflammatory genes, including IL-1β, IL-6, IL-12p40, and TNF-α and the chemokine gene CXCL-1, CCL-2, CCL3, and CCL-4. LPS-treated THP-1 cell-conditioned medium accelerated trophoblast cell death, and TNF-α played an essential role in this effect. The ADL extract reduced LPS-treated THP-1 cell-conditioned medium-induced trophoblast cell death by inhibiting MAPKs and the NF-κB pathway in macrophages. ADL extract prevented exogenous TNF-α-induced increased trophoblast cell death and decreased cell viability. Conclusions: We have demonstrated that the inhibition of LPS-induced inflammation by ADL extract can prevent preterm birth, fetal loss, and fetal growth restriction.

Worldwide, obesity has become a major risk factor associated with health risks such as diabetes, hypertension, hypercholesterolemia, cardiovascular disease, stroke, and certain forms of cancer. In this study, we estimated the anti-obesity effect of the bacterial strain

Rubus crataegifolius (black raspberry, RF), Ulmus macrocarpa (elm, UL), and Gardenia jasminoides (cape jasmine, GJ) are well known for hundreds of years as folk medicines in China and Korea to treat various gastrointestinal disturbance. The present study evaluated the gastroprotective effects of these plants either single or in combination against HCl/EtOH-induced gastritis and indomethacin-induced ulcer in rat model. Stomach ulcer was induced by oral ingestions of HCl/EtOH or indomethacin. Treatment with RF, UL, and GJ separately or in combination was done 1 h before ulcer induction. On HCl/EtOH-induced gastritis RF, UL, and GJ at a dose of 150 mg/kg showed comparable antigastritis effect (less than 50% inhibition) with lesion index of 94.97±8.05, 108.48±11.51, and 79.10±9.77 mm compared to cimetidine (45.33±23.73 mm). However, the combination of RF, UL, and GJ at a dose of 150 mg/kg with a ratio of 50:50:50 showed remarkable antigastritis effect with 77% inhibition. The observed lesion index at a ratio of 50:50:50 was 23.34±9.11 mm similar to cimetidine (18.88±19.88 mm). On indomethacin-induced ulcer, RF and GJ showed 38.28% and 51.8% inhibition whereas UL showed around 17.73% inhibition at 150 mg/kg. Combination of RF, UL, and GJ at 150 mg/kg showed strong antigastritis effect with 83.71% inhibition. These findings suggest strong gastroprotective effect of combined extract. In addition, these plants showed significant antioxidant activity in DPPH scavenging assay and antilipid peroxidation activity. Combination of black raspberry, elm, and cape jasmine might be a significant systemic gastroprotective agent that could be utilized for the treatment and/or protection of gastritis and gastric ulcer.

Ulcerative colitis is a complex inflammatory bowel disorder disease that can induce rectal and colonic dysfunction. Although the prevalence of IBD in Western countries is almost 0.5% of the general population, genetic causes are still not fully understood. In a recent discovery, itaconate was found to function as an immune-modulating metabolite in mammalian immune cells, wherein it is synthesized as an antimicrobial compound from the citric acid cycle intermediate cis-aconitic acid. However, the association between the Acod1 (Aconitate decarboxylase 1)-itaconate axis and ulcerative colitis has rarely been studied. To elucidate this, we established a DSS-induced colitis model with Acod1-deficient mice and then measured the mouse body weights, colon lengths, histological changes, and cytokines/chemokines in the colon. We first confirmed the upregulation of Acod1 RNA and protein expression levels in DSS-induced colitis. Then, we found that colitis symptoms, including weight loss, the disease activity index, and colon shortening, were worsened by the depletion of Acod1. In addition, the extent of intestinal epithelial barrier breakdown, the extent of immune cell infiltration, and the expression of proinflammatory cytokines and chemokines in Acod1-deficient mice were higher than those in wild-type mice. Finally, we confirmed that 4-octyl itaconate (4-OI) alleviated DSS-induced colitis in Acod1-deficient mice and decreased the expression of inflammatory cytokines and chemokines. To our knowledge, this study is the first to elucidate the role of the Acod1-itaconate axis in colitis. Our data clearly showed that Acod1 deletion resulted in severe DSS-induced colitis and substantial increases in inflammatory cytokine and chemokine levels. Our results suggest that Acod1 may normally play an important regulatory role in the pathogenesis of colitis, demonstrating the potential for novel therapies using 4-OI.

Helicobacter pylori is one of the most widespread infections involved in the pathogenesis of chronic gastritis, peptic ulcer, and gastric cancer. Hence, there is an urgent need to develop medications against H. pylori. This study aimed to evaluate synergistic effect of Rubus crataegifolius (RF) and Ulmus macrocarpa Hance (UL) against H. pylori. Antibacterial susceptibility of each extract either separately or in combination was studied against two H. pylori standard strains and 11 clinical isolates using agar dilution method. The effect of the extracts on H. pylori inoculated Balb/c mice model was also studied using single dosing (100 mg/kg each) approach. The MIC50 of RF and UL were more than 100 and 200 µg/ml, respectively, against the tested strains. However, simultaneous treatment with RF and UL at 75 and 50 µg/ml, respectively, showed decreased viable cell number, MIC70, and at 75 µg/ml each showed synergic effect with MIC90. On H. pylori inoculated Balb/c mice model, RF and UL separately (100 mg/kg each) showed moderate anti-H. pylori effect, while simultaneous treatment of RF and UL with same dose showed significant synergistic anti-gastric effects in stomach. The results showed a significant synergistic effect of plants extract against H. pylori infection and eventually gastric mucosal damage. Our finding could be considered a valuable support in the treatment of H. pylori induced gastritis and may contribute to the development of new and safe combined herbal product as anti-H. pylori regimens.

[Purpose] The aim of this study was to develop a new handle holder by modifying the inclination of the existing handle holder to reduce load on the wrist joints. [Subjects and Methods] The subjects of this study consisted of 25 elderly people aged 65 years or older accustomed to a walker-assisted gait. Two types of handle holders ((1) standard handle holder (2) inclination handle holder) were applied to all subjects and their wrist joint movement and muscle activity were measured while they conducted 10 cycle walker-assisted gait. [Results] The use of an inclination handle holder during the walker-assisted gait decreased considerably the extensor carpi radialis longus activity and angles of the ulnar deviation and wrist extension. [Conclusion] Improvements in the overall structure of a walker may be a new tool for improving existing walker users but the replacement cost will be expensive. The inclination handle holder presented in this study decreases the burden on the wrist joints of walker users without any overall structural changes in the walker, thereby reducing the occurrence of musculoskeletal diseases of the wrist joint during the walker-assisted gait of elderly people.

We aimed to analyze the value of each criterion for clinically insignificant prostate cancer (PCa) in the selection of men for active surveillance (AS) of low-risk PCa. We identified 532 men who were treated with radical prostatectomy from 2006 to 2013 who met 4 or all 5 of the criteria for clinically insignificant PCa (clinical stage ≤ T1, prostate specific antigen [PSA] density ≤ 0.15, biopsy Gleason score ≤ 6, number of positive biopsy cores ≤ 2, and no core with > 50% involvement) and analyzed their pathologic and biochemical outcomes. Patients who met all 5 criteria for clinically insignificant PCa were designated as group A ( n = 172), and those who met 4 of 5 criteria were designated as group B ( n = 360). The association of each criterion with adverse pathologic features was assessed via logistic regression analyses. Comparison of group A and B and also logistic regression analyses showed that PSA density > 0.15 ng ml −1 and high (≥7) biopsy Gleason score were associated with adverse pathologic features. Higher (> T1c) clinical stage was not associated with any adverse pathologic features. Although ≤ 3 positive cores were not associated with any adverse pathology, ≥4 positive cores were associated with higher risk of extracapsular extension. Among potential candidates for AS, PSA density > 0.15 ng ml −1 and biopsy Gleason score > 6 pose significantly higher risks of harboring more aggressive disease. The eligibility criteria for AS may be expanded to include men with clinical stage T2 tumor and 3 positive cores.

In this research we have confirmed that Nand flash cell current increases by optimization of contact barrier metal (BM) deposition and plug implantation (IIP) doping condition, which result in improved contact resistance (Rc) and cell leakage. Plug IIP dopant, BF2, which has been applied to P-type Transistor (Tr) for lowering Rc sacrifices contact process margin in cell area of n-type occurred by cell leakage. We could dramatically improve P-type Contact Rc and contact process margin by lowering BM deposition process temperature and adopting plug IIP doping with phosphorous (Ph.) respectively

－ Foods of plant origin, especially fruits and vegetables, have attracted attention because of their potential benefits to human health. In this report, Rubi Fructus (RF), the dried unripe fruit of Rubus coreanus Miq (Rosaceae) and ellagic acid (EA) purified from RF were used to test their potential hepatoprotective effect against acetaminophen (AAP)-induced hepatotoxicity in rats. RF extract (RFext) and EA reduced the elevated levels of alanine aminotransferase (ALT), aspartate aminotransferase (AST) in serum and the content of lipid peroxide in liver by AAP administration, while the increment of the cellular glutathione (GSH) content and the induction of glutathione S-transferase (GST) and glutathione peroxidase (GSH-PX) which were decreased by AAP administration. RFext and EA from RFext did not affect the two major form of cytochrome P450s, cytochrome P450 2E1 (CYP2E1) and cytochrome P450 1A2 (CYP1A2), but down- regulated the cytochrome P450 3A4 (CYP3A4) related to the conversion of AAP to N-acetyl-P-benzoquinone imine (NAPQI). These results suggest that RFext and EA from RF exhibit a hepatoprotective effect not only by increasing antioxidant activities but also by down-regulating CYP3A4 in the AAP-intoxicated rat.Keywords: Hepatoprotective effect, Acetaminophen

Multifunctional activities of the hepatitis B virus X-protein (HBx) in cells have been largely implicated in the development of liver cancer; one of these activities is the loss of p53 function by sequestering p53 in the cytoplasm. We have previously found that doxorubicin increased the p53 levels in cells containing p53-binding HBx protein and restored the p53-mediated transcriptional activity that was suppressed by HBx. Here, we investigated the mechanism underlying p53 reactivation. We found that six phosphorylation sites of the Serine residues of p53 were efficiently phosphorylated in HBx-expressing ChangX-34 cells, suggesting that the binding of HBx to the p53 protein does not interfere with the phosphorylation of p53 by signaling kinases. In addition, doxorubicin caused a dramatic reduction of Hdm2 mRNA and protein levels in cells expressing HBx. Intriguingly, reactivation of p53 was accompanied with a nuclear accumulation of p53 and the phosphorylated p53 at Serine15 was only detected in nuclear fraction, but not in cytosolic fraction of doxorubicin-treated ChangX-34 cells. Functional restoration of the p53 protein in HBx-expressing cells occurs according to the dual effects of doxorubicin: a significant reduction of Hdm2 expression and a nuclear accumulation of the phosphorylated p53 protein. Thus, proper usage of doxorubicin as an effective antitumor agent may be reevaluated and can be extended to tumors primarily caused by infection of DNA tumor viruses.

Background Our hypothesis is that the location of the seminal vesicles near the base of the prostate, the more positive cores are detected in the base, the greater the risk of seminal vesicle invasion. Therefore we investigate the clinical outcomes of base dominant prostate cancer (BDPC) in transrectal ultrasound (TRUS) - guided biopsies compared with anteromiddle dominant prostate cancer (AMPC). Methods From November 2003 to June 2014, a total of 990 intermediate and high risk prostate cancer (PCa) patients who underwent radical prostatectomy (RP) were enrolled and stratified into two groups according to proportion of positive cores-BDPC group had >= 33.3% ratio of positive cores from the prostate base among all positive cores and AMPC group < 33.3% in systemic biopsy. Between two groups, we compared the rate of pathologic outcomes and biochemical recurrence (BCR). We performed multivariate logistic regression model to confirm the significance of BDPC to seminal vesicle invasion (SVI) and Cox proportional hazard analysis to BCR. Results Among these 990 PCa patients, the 487 patients in BDPC group had more advanced clinical stage (p< 0.001), a higher biopsy GS (p = 0.002), and a higher rate of extracapsular extension (ECE), SVI and BCR (all p< 0.001) than AMPC group. The patients in BDPC group had poor BCR free survival rate via Kaplan-meier analysis (p< 0.001). The ratio of the base positive cores was a significant predictor to SVI in multivariate analysis (p < 0.001) and significant predictor of BCR in multivariate Cox proportional analysis (hazard ratio: 1.466, p = 0.004). Conclusions BDPC in TRUS-guided prostate biopsies was significantly associated with SVI and BCR after adjusting for other clinical factors. Therefore, BDPC should be considered to be a more aggressive tumor despite an otherwise similar cancer profile.

visualized. However, these lesions might not be easy to localize intraoperatively. Thus, the aim of this feasibility study was to evaluate intraoperative detection of metastatic lesions using a gamma probe after injection of a radiolabelled PSMA-ligand in correlation with postoperative histological findings. METHODS: 9 consecutive PCa patients with rising PSA after curative treatment (Median: 1.3ng/ml, range: 0.2 e 4.7ng/ml) and evidence of recurrent disease within the pelvis on 68Ga-HBEDPSMA PET hybrid imaging were included in this analysis. 24 hours before surgery patients received an intravenous injection of 111Inlabelled PSMA-I&T. Intraoperatively, metastatic lesions were detected by gamma probe with acoustic and visual feedback. Using the freehand SPECT technique optical tracking of the gamma probe allowed augmenting the live video stream of the field of surgery with reconstructed 3D image showing the position of positive lesions. Results of ex vivo radioactive rating (positive vs. negative) of resected tissue were compared to findings of postoperative histological analysis. RESULTS: In total, 32 separate histological specimens were evaluated (Median: 2, range: 2 e 7) and 15 specimens contained metastatic lesions at histological analysis. Lesions declared radioactive-positive intraoperatively and exhibiting a positive signal by the gamma probe ex vivo corresponded to metastatic PCa lesions with a sensitivity of 93.3% and a specificity of 100%. Detection of lesions as well as complete resection of even small metastatic lymph nodes was facilitated by the use of the gamma probe. Compared to preoperative 68Ga-HBED-PSMA PET hybrid imaging the intraoperative use of the gamma probe might even be more accurate as it detected all positive lesions and even additional lesions as small as 2mm. CONCLUSIONS: In this initial series, PSMA-radioguided surgery proved to be of high value for intraoperative detection of even small metastatic lesions in PCa patients. However, adequate patient selection for this individual treatment concept is crucial. Furthermore, greater patient numbers including follow-up data are needed to determine its possible role in clinical routine.

The aim of this study is to characterize the role of ellagic acid, a flavonoid from a medicinal herb which blocks HBeAg secretion in a HBV infected cell line and in HBeAg transgenic mice, in immune tolerance in chronic HBV infection. Using the mouse strain C57ML/6, HBeAg-producing transgenic mice (HBeAg-Tg), under the control of metal ion-inducible promoter were generated. The effect on immune tolerance of HBeAg-Tg and the release of immune tolerance by the inhibitor of HBeAg secretion, ellagic acid, was tested using T/B cell proliferation, HBeAg/HBeAb production, cytotoxic T-lymphocyte (CTL) and cytokine assays. C57ML/6 based HBeAg-producing HBeAg-Tg mice were tolerant to HBeAg at the T and B-cell level, did not produce antibodies to HBeAg in vivo and in vitro, produced minimal levels of cytokines (IL-4 and IFN-gamma) and decreased CTL responses, while feeding mice with ellagic acid (5mg/kg body weight) blocked the immune tolerance caused by HBeAg. Our results suggest that host immune tolerance induced by HBeAg during HBV infection, a viral strategy to guarantee HBV infection, can be overcome by ellagic acid, thus it can be used as a therapeutic for HBV-carriers.

This study determined whether exercise training prevents pathological hypertrophy in the left ventricle by modulation of myocardial and apoptosis-associated genes. We used spontaneously hypertensive rats (n=15, non-exercise SHR), exercise-trained SHR (n=15, treadmill exercise for 12 weeks), and sedentary Wistar-Kyoto (WKY) rats (n=15). Exercise-trained SHR expressed adaptive changes such as reduced body weight, heart rate, blood pressures, left ventricle wall thickness, lipid profiles, and homocysteine level. The mRNA expression of angiotensin converting enzyme, endothelin-1, and brain natriuretic peptides in the heart was lower in the exercise-trained SHR and in the WKY than in the non-exercise SHR, whereas mRNA expression of caveolin-3 and eNOS in the heart was higher. Bcl-2 protein was higher in the exercise-trained SHR than in the WKY and the non-exercise SHR. In contrast, Bax protein levels were lower in the exercise-trained SHR and in the WKY than in the non-exercise SHR. Furthermore, the levels of the active forms of caspase-3 (20 kDa) were lower in the exercise-trained SHR and in the WKY than in the non-exercise SHR. These findings suggest that exercise training prevents pathological hypertrophy in the left ventricle by modulation of myocardial genes and that it interferes with a signal transduction pathway of apoptosis secondary to the pathological cardiac hypertrophy.

[Purpose] This study verified the leg muscle activities of elderly subjects performing leg cycle ergometer exercise. [Subjects] Forty-one elderly persons were the subjects of this study. [Methods] For the three distances corresponding to knee flexion angles of 15, 45, and 70, the muscle activities of the rectus femoris, biceps femoris, tibialis anterior and lateral gastrocnemius were measured while the subjects exercised on a cycle ergometer. [Results] The rectus femoris and biceps femoris showed statistically significant increases as the distance between the cycle ergometer and the body increased, and the lateral gastrocnemius muscle activation showed a statistically significant increase as the distance from the body to the cycle ergometer decreased. [Conclusion] When the elderly have limb muscle weakness, leg cycle ergometer distances should be adjusted.

Osteoclasts are bone resorbing cells of hematopoietic origin. The hematopoietic transcription factor PU.1 is critical for osteoclastogenesis; however, the molecular mechanisms of PU.1-regulated osteoclastogenesis have not been explored. Here, we present evidence that the receptor activator of nuclear factor kappaB (RANK) gene that has been shown to be crucial for osteoclastogenesis is a transcriptional target of PU.1. The PU.1-/- progenitor cells failed to express the RANK gene and reconstitution of PU.1 in these cells induced RANK expression. Treatment of the PU.1 reconstituted cells with M-CSF and RANKL further augmented the RANK gene expression. To explore the regulatory mechanism of the RANK gene expression by PU.1, we have cloned the human RANK promoter. Transient transfection assays have revealed that the 2.2-kb RANK promoter was functional in a monocyte line RAW264.7, whereas co-transfection of PU.1 transactivated the RANK promoter in HeLa cells. Taken together, these results suggest that PU.1 regulates the RANK gene transcription and this may represent one of the key roles of PU.1 in osteoclast differentiation.

A flavonoid molecule that showed a unique anti-HBV function was isolated from Phyllanthus urinaria. The molecular formula was determined as C14H6O8 based on FAM-MS analysis and the structure was determined by NMR. The identified flavonoid molecule, ellagic acid, showed unique anti-HBV functions. Ellagic acid did not inhibit either HBV polymerase activity, HBV replication or block HBsAg secretion. Rather, ellagic acid blocks effectively HBeAg secretion in HepG2 2.2.15 cells (IC50 = 0.07 μg/ml). Since HBeAg is involved in immune tolerance during HBV infection, ellagic acid, a newly identified functional anti-HBV compound, may be a new candidate therapeutic against immune tolerance in HBV-infected individuals.

The hepatitis B virus-X protein (HBx) regulates fundamental aspects of mitochondrial physiology. We show that HBx down-regulates mitochondrial enzymes involved in electron transport in oxidative phosphorylation (complexes I, III, IV, and V) and sensitizes the mitochondrial membrane potential in a hepatoma cell line. HBx also increases the level of mitochondrial reactive oxygen species and lipid peroxide production. HBx does not activate apoptotic signaling, although it sensitizes hepatoma cells to apoptotic signaling, which is dependent on reactive oxygen species. Increased intrahepatic lipid peroxidation in HBx transgenic mice demonstrated that oxidative injury occurs as a direct result of HBx expression. Therefore, we conclude that mitochondrial dysfunction is a crucial pathophysiological factor in HBx-expressing hepatoma cells and provides an experimental rationale in the investigation of mitochondrial function in rapidly renewed tissues, as in hepatocellular carcinomas.

[Purpose] This study aims to improve the safety for caregivers and avoid musculoskeletal diseases by examining the shoulder and trunk muscle activity depending on wheelchair handle height as the caregiver provides propulsion to a wheelchair for wheelchair-bound patients. [Subjects] The participants were caregivers who met the criteria for this study (n=30). [Methods] To determine the activity of the shoulder and trunk muscles of caregivers depending on the handle height, the muscle activity was measured as a wheelchair was being pushed under three height conditions. [Results] According to the study results, the deltoid middle fiber, serratus anterior muscle, rhomboid muscle, and erector spinalis of lumbar part showed significantly low muscle activity at a height that produced an elbow joint flexion angle of 30°. Caregivers are highly likely to develop musculoskeletal disease if the handle height is excessively low. In addition, the burden on the shoulder and back muscles can be reduced at heights that lead to flexion of 30°. [Conclusions] As a caregiver pushes a wheelchair, the handle height has a close relationship with the burden on the shoulder and back muscles. If the handle height is excessively low, the caregiver is highly likely to develop musculoskeletal disease.

The PTEN gene (phosphatase and tensin homologous on chromosome 10) is frequently mutated or deleted in a number of malignancies including human hepatocellular carcinoma (HCC). We reported previously that the hepatitis B virus X (HBx) protein, known to be a causative agent in the formation of HCC, activates insulin-like growth factor II (IGF-II) expression through Sp1 phosphorylation by protein kinase C (PKC) or mitogen-activated protein kinase (MAPK) signaling. In this report we demonstrate that the PTEN effect on HBx induced IGF-II activation in a hepatoma cell line. Expression of PTEN and IGF-II was inversely related in different hepatoma cell lines. PTEN expression induced decreased Sp1 DNA binding by dephosphorylating Sp1 and interfered with transcriptional transactivation of IGF-II by HBx in hepatoma cells. The protein phosphatase activity was involved in PTEN downregulation of IGF-II transcription through downregulation of MAPK, MAPK kinase phosphorylation and PKC translocation. Our data suggest that PTEN blocks Sp1 phosphorylation in response to HBx, by inactivating PKC, MAPK and MAPK kinase which eventually downregulate IGF-II expression, during the formation of HCC.

The aim of the present study was to investigate the intracellular mediators of the third base mutant of codon 249 in p53 gene (p53mt249) mutation that potentiate IGF-II dependent IGF-I receptor (IGF-IR) signaling. p53mt249 enhanced IGF-II dependent IGF-IR signaling in p53 negative Hep3B hepatoma cells which were specifically prevented by IGF-IR antibody, αIR3 and lovastin. p53mt249 increased the number of IGF-II binding sites with no change in the affinity of IGF-IR. Enhanced levels of IGF-IR expression and transcription were identified in p53mt249 transfected Hep3B cells. Pre-transfection of cultured hepatoma cells with p53mt249 resulted in a three to fourfold increase in IGF-IR phosphorylation and downstream mediator IRS-I phosphorylation but, enhanced more than 15-fold after IGF-II treatment, which coincides well with the cell growth and thymidine uptake results. Our results showed that p53mt249 modulate IGF-II dependent IGF-IR signaling by upregulating IGF-IR and potentiating IGF-IRs where IGF-IRs became more sensitive on treatment with IGF-II. We concluded that p53mt249 stimulates IGF-II dependent IGF-IR signaling by upregulating the expression of both ligand (IGF-II) (Oncogene 19 (2000) 3717) and receptor (IGF-IR) through an autocrine and/or paracrine loop and we outline the physiological significance of potentiation of IGF-IR by p53 mutation in the development of hepatocellular carcinoma (HCC).

[Purpose] The purpose of this study was to investigate the effect of handle type, which is a fundamental element determining rollator type and the part that directly connects the user to a rollator, on the plantar foot pressure of elderly people during gait, and to examine how the gait of elderly people is affected by variation of the rollator handle. [Subjects] The subjects of this study were 12 elderly women over the age of 65 and 10 young adults who were given sufficient explanation about the objectives and method of this study and volunteered to participate in the study. [Methods] The elderly women learned how to use different types of rollators and performed gait without a rollator and with two types of rollators in a random order on a course that was 10 m long. The young adults, who served as the reference for the plantar foot pressure during gait, performed gait in the same manner without a rollator. [Results] The plantar foot pressure of the elderly people was compared with that of the young adults. The results show that there was a significant difference at the rearfoot, lateral midfoot, lateral forefoot, and hallux regions. The plantar foot pressure at the rearfoot and hallux regions was highest in the young adults, while it was lowest when the elderly used a rollator with a transverse handle. The plantar foot pressure at the lateral midfoot and lateral forefoot regions was highest when the elderly group did not use a rollator. [Conclusion] The results suggest that selective use of the handle type according to the purpose is important even though the use of a rollator assists the gait of the elderly regardless of the handle type. Moreover, the high dependency on the transverse handle rollator means that there can be a greater risk of falling down when grip on the handle is lost, and the vertical handle is more appropriate for gait training since it reduces dependency on the rollator during gait.

The recombinant human hepatitis B virus-X protein (rhHBx) has been expressed as inclusion bodies in Escherichia coli and purified. By sequential dialysis of urea, rhHBx was folded into the native structure, which was demonstrated by both the efficacy of its transcriptional activation of the adenovirus major late promoter, fluorescence and circular dichroism (CD) analysis. The increase in CD values at 220 nm and a corresponding blue shift of the intrinsic fluorescence emission confirmed the ability of HBx to refold in lower concentrations of urea to produce the active protein. After purification and renaturation, the rhHBx protein was found to be phosphorylated by protein kinase C (PKC) and mitogen-activated protein kinase (MAPK). In vivo phosphorylation of HBx was also demonstrated. Although PKC and MAPK enhance the HBx phosphorylation in vitro, neither protein kinase A nor caseine kinase II (CKII) phosphorylate HBx protein, though there are possible substrate residues of both kinases in HBx protein. Phosphoamino acid analysis of the total acid hydrolyzed HBx showed that serine residues can be phosphorylated by PKC or MAPK.

The possibility that hepatitis C virus core gene product (HCV-core) acts as a transactivator in insulin-like growth factor II (IGF-II) gene transcription was tested. HCV-core protein increases endogenous IGF-II expression from promoter 4 (P4) of the IGF-II gene through two cis-acting elements: Sp1 and Egr1 binding sites. Sp1 and Egr1 both bind to IGF-II P4 and functionally cooperate in mediating the maximal activity of IGF-II P4. HCV-core protein induced the binding of Sp1 and Egr1 on its binding sites on IGF-II P4. In addition, Sp1 and Egr1 were stimulated to phosphorylate by HCV-core, and its DNA binding activity was up-regulated upon HCV-core transfection. Transfection with HCV-core in HepG2 cells stimulated the membrane translocation of protein kinase C (PKC) and the treatment of HCV-core transfected cells with calphostin C, a PKC inhibitor, blocked induction of Sp1 and Egr1 DNA binding activity, and eventually transcriptional transactivations of the IGF-II gene. Increasing the DNA binding activity of the phosphorylated form of Sp1 and Egr1 might be an important mechanism for regulating IGF-II gene expression and for promoting cell division during hepatic carcinogenesis. These results indicate that HCV-core functions as a positive regulator of IGF-II transcription through the PKC pathway and that Sp1 and Egr1 are direct targets of the transcriptional regulation of the IGF-II gene which plays an important role in hepatitis C virus pathogenesis during the formation of hepatocellular carcinoma (HCC).

Aflatoxin B1 (AFB1) induced mutation of the p53 gene at codon 249 (p53mt249) is critical during the formation of hepatocellular carcinoma (HCC) following hepatitis B virus (HBV) infection. p53mt249 markedly increases insulin-like growth factor II (IGF-II) transcription largely from promoter 4, accumulating the fetal form of IGF-II. Modulation of the transcription factor binding to IGF-II P4 by wild-type p53 and p53mt249 was identified. Wild-type p53 inhibited binding of transcription factors Sp1 and TBP on the P4 promoter, while p53mt249 enhanced the formation of transcriptional complexes through enhanced DNA-protein (Sp1 or TBP) and protein-protein (Sp1 and TBP) interactions. p53mt249 stimulates transcription factor Sp1 phosphorylation which might be a cause of increased transcription factor binding on the P4 promoter while wild-type p53 does not. Transfection of hepatocytes with p53mt249 impaired induction of apoptosis by the HBV-X protein and TNF-alpha. Therefore, the blocking of apoptosis through enhanced production of IGF-II should provide a favorable opportunity for the selection of transformed hepatocytes. These results explain the molecular basis for the genesis of HCC by p53mt249 which was found to be induced by a potent mutagen, AFB1.

Progression through the cell cycle is controlled by the induction of cyclins and activation of cognate cyclin-dependent kinases. The human hepatitis B virus-X (HBV-X) protein functions in gene expression alterations, in the sensitization of cells to apoptotic killing and deregulates cell growth arrest in certain cancer cell types. We have pursued the mechanism of growth arrest in Hep3B cells, a p53-mutant human hepatocellular carcinoma (HCC) cell line. In stable or transient HBV-X transformed Hep3B cells, HBV-X increased protein and mRNA levels of the cyclin-dependent kinase inhibitor (CDKI) p21(waf1/cip1) increased binding of p21(waf1/cip1) with cyclin-dependent kinase 2 (CDK2), markedly inhibited cyclin E and CDK2 associated phosphorylation of histone H1 and induced the activation of a p21 promoter reporter construct. By using p21 promoter deletion constructs, the HBV-X responsive element was mapped to a region between -1185 and -1482, relative to the transcription start site. Promoter mutation analysis indicated that the HBV-X responsive site coincides with the ets factor binding sites. These data indicate that in human hepatocellular carcinoma cells, HBV-X can circumvent the loss of p53 functions and induces critical downstream regulatory events leading to transcriptional activation of p21(waf1/cip1). As a consequence, there is an increased chance of acquisition of mutations which can enhance the genesis of hepatomas. Our results also emphasize the chemotherapeutic potential of p21(waf1/cip1) inhibitors, particularly in the HBV-X infected hepatoma which lacks functional p53.

Systematic secondary structure simulation of a target mRNA sequence is shown to be effective for locating a good anti-sense target site. Multiple selected anti-sense sequences were placed in a single molecule. The anti-sense oligonucleotide (oligo) was covalently closed to avoid exonuclease activities and was designated CMAS (covalently closed multiple anti-sense)-oligo. CMAS-oligo was found to be stable, largely preserving its structural integrity after 24 h of incubation in the presence of either exonuclease III or serum. When human c-myb mRNA was targeted by the c-myb CMAS-oligo, expression of the gene was completely abolished. Further, tumour cell growth was inhibited by 82±3% as determined by an MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-2H-tetrazolium bromide] assay and by 90±1% by [3H]thymidine incorporation. When a leukaemic cell line K562 was treated with CMAS-oligo, colony formation on soft agarose was also decreased by 93%. In contrast, treatment with a scrambled control oligo did not significantly inhibit leukaemic cell growth. These results suggest that a rational target site search is possible for an anti-sense oligo and that CMAS-oligo can be employed as an effective anti-sense agent with enhanced stability.

Fed-batch cultures were carried out to overproduce human insulin-like growth factor I (IGF-I) in Escherichia coli. The effects of carbon sources (glucose or glycerol) and induction time on cell growth and IGF-I production were investigated in more detail. Glycerol was a better carbon source than glucose for IGF-I production in fed-batch culture. Induction at the mid-exponential phase with glycerol as a carbon source in the pH-stat fed-batch culture was optimal for IGF-I production. Under this condition, 2.8 g L−1 of fusion IGF-I was produced as inclusion bodies. We have also developed downstream processing for preparative scale purification of IGF-I from the fusion protein produced by the fed-batch culture using glycerol as a carbon source. After the fusion protein expressed was solubilized in 8 M urea and cleaved with hydroxylamine, the released IGF-I was purified by cation exchange chromatography, refolding and preparative scale reverse phase HPLC (rp-HPLC) to give recombinant IGF-I of >98% purity. The biological activities of the purified IGF-I were measured and found to be identical to those of commercial IGF-I. Journal of Industrial Microbiology & Biotechnology (2000) 24, 94–99.

We studied the effects of antisense oligonucleotides (AS oligos) with a novel structure. The AS oligos were covalently closed to avoid exonuclease activities by enzymatic ligation of two identical molecules. The AS oligos of a ribbon type (RiAS oligos) consist of two loops containing multiple antisense sequences and a stem connecting the two loops. Three antisense sequences targeting different binding sites were placed in a loop that was designed to form a minimal secondary structure by itself. RiAS oligos were found to be stable because they largely preserved their structural integrity after 24 h incubation in the presence of either exonuclease III or serums. When a human promyelocytic cell line, HL-60, was treated with RiAS oligos to c-myb, c-myb expression was effectively ablated. Cell growth was inhibited by90% determined by both the 3-[4,5-dimethythiazol-2-yl]-2,5-diphenyltetrazolium bromide assay and [(3)H]thymidine incorporation. Further, when the leukemic cell line K562 was treated with c-myb RiAS oligos, colony formation on soft agarose was reduced by 92 +/- 2%. These results suggest that RiAS oligos may be employed for developing molecular antisense drugs as well as for the functional study of a gene.

We prepared human hepatoma cell lines, which expressed the human hepatitis B virus-X gene product. The plasmid pMAMneo-X, containing an HBV-X gene promoter, an enhancer and a structural gene was constructed. Transfected HBV-X gene integration and expression were detected by Southern and Northern blotting, as well as by chloramphenicol acetylase transferase (CAT) assay using various kinds of promoter-CAT reporter systems. HBV-X protein expression in stable transfectants was confirmed by immunofluorescence microscopy. Transfected cell lines showed permanent expression of HBV-X proteins. The HBV-X transfectant activated its target promoters in promoter-CAT constructs as reporters. The HBV-X transfectant enhanced AP-1 transcription factor binding to its target DNA. Therefore, X-transfectants are not only stable, but also have specific biological functions. Cell cycle analysis by flow cytometry showed that the majority of the transfectant cells are arrested in the G1 or G2 phase of the cell cycle. These cell lines may be useful in analyzing the biological functions of HBV-X and its functional role in the formation of hepatocellular carcinomas.

In an attempt to induce the secretion of human granulocyte-colony stimulating factor (hG-CSF) in Escherichia coli , the gene encoding mature hG-CSF was fused to the gene for the pectate lyase B (pelB) signal peptide of Erwinia caratovora , and fed-batch fermentation of the recombinant E. coli was performed. No secretion of hG-CSF and no processing of the signal peptide from the fusion protein was observed. Instead, the fusion protein, pelB-hG-CSF, at a concentration of ca. 1.7 g/ l was produced in an inclusion body form. Interestingly, the biological activity of the purified fusion protein was found to be identical to that of mature hG-CSF, indicating that the pelB signal peptide attached to the N-terminal of hG-CSF had no effect on its biological activity.