Ameliorative Effects of Pueraria lobata Extract on Postmenopausal Symptoms through Promoting Estrogenic Activity and Bone Markers in Ovariectomized Rats.

Pueraria lobata (Willd.) Ohwi, known as kudzu, is one of the most popular traditional medicines in Asian countries. It has been widely used as a natural alternative to hormone replacement therapy for treating postmenopausal symptoms. This study aimed to investigate the estrogenic effect of P. lobata extract (PE) against postmenopausal osteoporosis in ovariectomized (OVX) rats. OVX rats were treated with PE (25-1600 mg/kg) for 8 weeks. Biochemical parameters, estradiol, and bone turnover markers (e.g., osteocalcin, C-terminal telopeptide fragment of type I collagen, deoxypyridinoline, and pyridinoline) were measured in plasma samples. In addition, estrogen receptor-alpha (ER-α) protein expression and morphology of uterine were evaluated. Long-term treatment with PE did not cause liver damage in OVX rats. PE supplementation reduced body weight gain in obese rats with high lipid accumulation induced by ovariectomy. Furthermore, PE exhibited a protective effect against insulin resistance, hyperlipidemia, and hepatic lipid peroxidation. PE treatment increased uterine weight and thickness of the uterine layers in cases of uterus atrophy due to removal of ovaries. The levels of bone turnover markers, which were significantly increased in OVX rats, were decreased by PE treatment. Western blotting analysis showed that ER-α protein expression was upregulated in PE-treated rats compared with OVX rats. These results suggest that PE could be a promising alternative functional food for improving menopausal symptoms.

The Intertester Reliability of a Modified Movement System Impairment Classification System Used to Evaluate Individuals With Prolonged Sitting: A Preliminary Study.

OBJECTIVE: Prolonged sitting while working at a computer leads to poor sitting postures, which can aggravate low back pain in many individuals. We examined the intertester reliability of using the modified musculoskeletal impairment schema for classifying participants sitting at computers for prolonged times. METHODS: Fifty participants were examined independently by each therapist using a test-retest design. Each therapist assigned an musculoskeletal impairment classification upon completion of the examination. The agreement percentages and the kappa coefficient were used to evaluate intertester reliability in classifying participants with prolonged sitting. RESULTS: The percentage agreement between the 2 examiners for participants who maintained the sitting posture for prolonged times was 84%. The calculated kappa coefficient was 0.73, reflecting a substantial level of agreement. CONCLUSIONS: The present findings provide some evidence to support the classification of individuals who sit at computers for prolonged times and participants with rotation with flexion pattern would need to manage asymmetry pattern in a subclinical group.

Antibacterial activity and effects of Colla corii asini on Salmonella typhimurium invasion in vitro and in vivo.

BACKGROUND: Salmonella enterica serovar Typhimurium is a foodborne pathogen that triggers inflammatory responses in the intestines of humans and livestock. Colla corii asini is a traditional medicine used to treat gynecologic and chronic diseases in Korea and China. However, the antibacterial activity of Colla corii asini has been unknown. In this study, we investigated the antibacterial activity and effects of Colla corii asini extract on Salmonella typhimurium invasion. METHODS: To tested for antibacterial effects of Colla corii asini extracts, we confirmed the agar diffusion using Luria solid broth medium. Also, we determined the MIC (minimum inhibitory concentration) and the MBC (minimum bactericidal concentration) value of the Colla corii asini ethanol extract (CEE) by using two-fold serial dilution methods. We evaluated the expression of salmonella invasion proteins including SipA, SipB and SipC by using Western blot and qPCR at the concentration of CEE without inhibition of bacterial growth. In vitro and vivo, we determined the inhibitory effect of invasion of S. typhimurium on CEE by using gentamicin assay and S. typhimurium-infected mice. RESULTS: CEE significantly inhibited the growth of Salmonella typhimurium in an agar diffuse assay and had an MIC of 0.78 mg/ml and an MBC of 1.56 mg/ml. Additionally, CEE reduced Salmonella typhimurium cell invasion via the inhibition of Salmonella typhimurium invasion proteins, such as SipA, SipB and SipC. Furthermore, CEE significantly suppressed invasion in the small intestines (ilea) of mice injected with Salmonella typhimurium. CONCLUSION: These findings show that Colla corii asini exerts antibacterial activity and suppresses Salmonella typhimurium invasion in vitro and in vivo. Together, these findings demonstrate that Colla corii asini is a potentially useful therapeutic herbal medicine for treating salmonella-mediated diseases.

Fermented herbal formula KIOM-MA-128 protects against acute colitis induced by dextran sodium sulfate in mice.

BACKGROUND: Colitis is a well-known subtype of inflammatory bowel disease and is caused by diverse factors. Previous research has shown that KIOM-MA elicits anti-inflammatory and anti-allergic effects on various diseases. KIOM-MA-128, our novel herbal formula, was generated from KIOM-MA using probiotics to improve the therapeutic efficacy. We investigated whether KIOM-MA-128 has protective activity in a mouse model of acute colitis induced by dextran sodium sulfate (DSS). METHODS: Colitis was induced by DSS administered to ICR mice in drinking water. KIOM-MA-128 (125 or 250 mg/kg) was orally administered once per day. The body weights of the mice were measured daily, and colonic endoscopies were performed at 5 and 8 days. Colon length as well as histological and cytokine changes were observed at the end of drug administration. RESULTS: KIOM-MA-128 has pharmacological activity in an acute colitis model. KIOM-MA-128 reduced the loss of body weight and disease activity index (DAI) and inhibited the abnormally short colon lengths and the colonic damage in this mouse model of acute colitis. Moreover, KIOM-MA-128 suppressed pro-inflammatory cytokine expression and maintained the integrity of the tight junctions during DSS-induced colitis. CONCLUSION: The results indicated that KIOM-MA-128 protects against DSS-induced colitis in mice and suggested that this formula might be a candidate treatment for inflammatory bowel disease (IBD).

Leonurus japonicus Houtt Attenuates Nonalcoholic Fatty Liver Disease in Free Fatty Acid-Induced HepG2 Cells and Mice Fed a High-Fat Diet.

We investigated the effects of a Leonurus japonicus ethanol extract (LJE) on nonalcoholic fatty liver disease (NAFLD). An in vitro model of hepatic steatosis was treated with 1 mM free fatty acid (FFA) in HepG2 cells. An in vivo NAFLD model was established using C57BL/6 mice fed a high-fat diet (HFD) and administered LJE (100 or 200 mg/kg) orally for 14 weeks. LJE treatment suppressed lipid accumulation and intracellular triglyceride levels significantly in a concentration-dependent manner in HepG2 cells. Moreover, LJE significantly reduced the expression of sterol regulatory element binding protein 1-c, and its downstream genes, which are associated with lipogenesis, in HepG2 cells. In HFD-fed mice, LJE treatment decreased body weight significantly and decreased serum alanine transaminase levels to normal values, concurrent with a decrease in hepatic lipid accumulation. Furthermore, LJE supplementation ameliorated insulin sensitivity by decreasing serum glucose and insulin levels. LJE improved hepatic steatosis by increasing the expression of phosphorylated AMP-activated protein kinase and peroxisome proliferator-activated receptor-α in HFD-fed mice and FFA-treated HepG2 cells. The results suggested that LJE might be a potential therapeutic agent to treat NAFLD.

In vivo hair growth-stimulating effect of medicinal plant extract on BALB/c nude mice.

CONTEXT: Chrysanthemum zawadskii var. latilobum (Asteraceae) (CZ) and Polygonum multiflorum Thunb. (Polygonaceae) (PM) have been used traditionally to treat different systemic diseases and acclaimed for various biological activities including hair growth. OBJECTIVE: This study investigates the hair restoration efficacy of selected medicinal plant extracts on nude mice. MATERIALS AND METHODS: Nude mice genetically predisposed to pattern balding were used in this study. Topical methanol extracts of CZ and PM (10 mg/mouse/d) with standardized vehicle formulation, only vehicle (propylene glycol:ethanol:dimethyl sulfoxide, 67:30:3% v/v) and Minoxidil (2%) were applied daily for 40 consecutive days. RESULTS: In our study, the maximum hair score (2.5 ± 0.29) was obtained in the CZ-treated group. Histological observation revealed a significant increase (p < 0.001) in the number of hair follicles (HF) in CZ-treated mice (58.66 ± 3.72) and Minoxidil-treated mice (40 ± 2.71). Subsequently, immunohistochemical analysis also confirmed the follicular keratinocyte proliferation by detection of BrdU-labeling, S-phase cells in Minoxidil and CZ-treated mouse follicular bulb and outer root sheaths. CONCLUSION: Our study revealed the underlying mechanism of stimulating hair growth in athymic nude mice by repair the nu/nu follicular keratin differentiation defect. Thus, the topical application of CZ may represent a novel strategy for the management and therapy of certain forms of alopecia.

The multi-herbal formula Chong-Myung-Tang improves spatial memory and increases cell genesis in the dentate gyrus of aged mice.

Chong-Myung-Tang (CMT) is a multi-herbal formula that has been used to improve memory. However, the potential mechanism remains unknown. The present study investigated the effects of CMT (50, 100, and 200 mg/kg) on spatial memory of aged mice. The behavioral training tests indicated that 200 mg/kg CMT treatment can significantly improve spatial memory of aged mice in the Morris water maze. Moreover, cell survival was examined by injecting bromodeoxyuridine (BrdU) on the first three days. The result showed that 200 mg/kg CMT treatment significantly increased cell survival in the dentate gyrus. Cell proliferation was determined by injecting BrdU 2 h before the mice were killed. The result suggested that CMT treatments had no influence on cell proliferation in the dentate gyrus. Thus, an increase in cell survival in the dentate gyrus stimulated by CMT may be involved in the effect of CMT on spatial memory improvement.

Chrysanthemum zawadskii extract induces hair growth by stimulating the proliferation and differentiation of hair matrix.

Chrysanthemum zawadskii has been proven to possess hair growth activity and has been used as treatment for hair loss. The aim of this study was to provide a novel explanation of the mechanism by which Chrysanthemum zawadskii extracts (CZe) promote hair growth and to characterize the affected hair follicle (HF) regions and the progression of growth. The n-butanol and water fractions of CZe were used for hair growth induction by topical application to the backs of C57BL/6 mice for up to 30 days. To investigate cell development during HF morphogenesis, bromodeoxyuridine-labeled skin sections were detected using immunohistochemistry. The results showed that the water fraction of CZe promoted hair shaft production and induced premature entry of telogen HFs into the anagen. Subsequently, immunohistochemical studies indicated that the water fraction of CZe stimulated the differentiation and proliferation of pluripotent epidermal matrix cells in the matrix region and epithelial stem cells in the basal layer of the epidermis. Additionally, flavonoids were identified as effective constituents. Therefore, the findings of this study suggested that the water fraction of CZe may be developed as a therapeutic agent for the prevention of hair loss.

Long-term administration of ginsenoside Rh1 enhances learning and memory by promoting cell survival in the mouse hippocampus.

Ginsenosides, the secondary plant metabolites produced by Panax ginseng are responsible for the enhancing effects on learning observed following treatment with Panax ginseng. A number of studies have provided correlational evidence that cell proliferation and survival are closely associated with hippocampal-dependent learning tasks. In this study, to investigate the beneficial effects of ginsenoside Rh1 on hippocampal cells and learning, mice (6 months old) were administered ginsenoside Rh1 at a dose of 5 and 10 mg/kg/day for a period of 3 months. Saline-treated mice were used as controls. The enhancement of memory and learning in the mice was evaluated by hippocampal-dependent tasks (passive avoidance tests and Morris water maze tests) and the immunohistochemical marker of cell proliferation, bromodeoxyuridine (BrdU). In addition, the levels of brain-derived neurotrophic factor (BDNF) were measured following treatment. Based on our data, the Rh1-treated group (5 and 10 mg/kg) showed a significantly improved learning and memory ability in the passive avoidance tests compared with the control group; however, only treatment with 10 mg/kg ginsenoside Rh1 significantly promoted spatial learning ability in the Morris water maze test. Ginsenoside Rh1 significantly enhanced cell survival in the dentate gyrus of mice, although it did not enhance hippocampal cell proliferation. In addition, ginsenoside Rh1 upregulated the expression of BDNF. These findings address the potential therapeutic significance of ginsenoside Rh1 as a nutritional supplement in memory loss and neurodegenerative diseases.

Ginsenoside Rh2 improves learning and memory in mice.

A wide range of plant foods and dietary supplements are able to modify the functioning of the central nervous system. In the present study, we observed that oral administration of ginsenoside Rh2 (10 mg/mL) for 3 weeks significantly improved spatial learning and memory. Spatial memory and learning was evaluated in mice by hippocampus-dependent tasks (Morris water maze test) and immunohistochemical marker of cell genesis bromodeoxyuridine. Ginsenoside Rh2 treatment (30 days) promoted cell survival and genesis. Further, ginsenoside Rh2 treatment in enriched condition had no significant effects on cell survival compared with standard condition exposure. These results revealed that ginsenoside Rh2-mediated spatial learning and memory improvement was associated with cell genesis and survival and may be parallel to the mechanism of environmental enrichment. Therefore, ginsenoside Rh2 may have efficacy as a dietary supplement for spatial learning and memory improvement.

Ginsenoside Rb1 improves spatial learning and memory by regulation of cell genesis in the hippocampal subregions of rats.

Ginsenoside Rb1 (Rb1) is known to improve learning and memory in hippocampus-dependent tasks. However, the cellular mechanism remains unknown. Cell genesis in hippocampus is involved in spatial learning and memory. In the present study, Rb1 was orally administrated to adult rats for 30days. The behavioral training tests indicated that Rb1 improved spatial cognitive performance of rats in Morris water maze (MWM). Furthermore, we investigated the effects of Rb1 on cell genesis in adult rats' hippocampus, using thymidine analog bromodeoxyuridine (BrdU) as a marker for dividing cells. It has been shown that hippocampal cell genesis can be influenced by several factors such as learning and exercise. In order to avoid the effects of the interfering factors, only the rats treated with Rb1 without training in MWM were used to investigate cell genesis in hippocampus. When BrdU was given to the rats 30days prior to being killed, it was shown that oral administration of Rb1 significantly increased cell survival in dentate gyrus and hippocampal subregion CA3. However, when BrdU was injected 2h prior to sacrifice, the results indicated that Rb1 had no significant influence on cell proliferation in the hippocampal subregions. Thus, an increase of cell survival in hippocampus stimulated by Rb1 may be one of the mechanisms by which ginseng facilitates spatial learning and memory. Our study also indicates that Rb1 may be developed as a therapeutic agent for patients with memory impairment.

Neuroprotective effects of SG-168 against oxidative stress-induced apoptosis in PC12 cells.

Postmortem examinations of tissues of humans and rodents with a host of neurodegenerative conditions, including Alzheimer's and Parkinson's diseases, have identified oxidative damage in proteins, lipids, and DNA. The aim of this study was to better understand the cellular mechanisms of neuronal cell degeneration induced via oxidative stress and the protective roles of bioactive substance. In order to achieve this aim, we established a screening program to discover therapeutic agents that exhibit preferential neuroprotective activity in H(2)O(2)-treated PC12 cells. During the course of our screening program, we isolated an active compound, SG-168, from Dendrobium nobile Lindley and identified it as a neuroprotective agent. SG-168 was identified as a compound with an acetal skeleton, a prototypical compound, by electrospray ionization-mass spectrometry analysis and various nuclear magnetic resonance spectroscopic methods. The protective effect of SG-168 in PC12 cells with H(2)O(2)-induced oxidative damage was investigated by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide reduction assay. As expected, incubation with H(2)O(2) for 2 hours resulted in cell viability of 31.8% compared to the control, while pretreatment of SG-168 increased cell viability by 15-50% compared to the H(2)O(2)-stressed control cells. These results showed that SG-168 inhibits H(2)O(2)-induced apoptotic cell death. Interestingly, flow cytometric analysis showed that H(2)O(2)-treated PC12 cells incubated with SG-168 exhibited greatly suppressed apoptosis. In summation, the results of this study suggest that SG-168 has potential as a new antioxidant agent against neuronal diseases.

Anti-amnesic effect of Chong-Myung-Tang on scopolamine-induced memory impairments in mice.

AIM OF THE STUDY: Chong-Myung-Tang (CMT) consisted of Acorus gramineus Soland, Polygala tenuifolia Willdenow, and Poria cocos Wolf is one of the traditional Korean herbal medicines used for the therapy of learning and memory improvement. The present study was investigated the effect of CMT on learning and memory functions in SCOP-induced memory deficits mice. MATERIALS AND METHODS: The cognitive-enhancing effect of CMT on amnesic mice induced by SCOP was investigated by assessing the passive avoidance test and the Morris water maze test. In order to confirm the underlying mechanisms of memory enhancing effects of CMT, activities of AChE, choline acetyltransferase (ChAT), and antioxidant enzymes were measured. RESULTS: Administration of CMT significantly restored memory impairments induced by SCOP in the passive avoidance test and also reduced escape latency during trial sessions in the Morris water maze test. The increased AChE activity produced by SCOP was significantly inhibited by CMT. CMT significantly enhanced ChAT activity. Moreover, treatment with CMT to the amnesic mice induced by SCOP considerably decreased malondialdehyde levels and restored activities of superoxide dismutase and catalase to the control values. CONCLUSIONS: These results suggest that CMT may be useful for the cognitive improvement via regulation of cholinergic marker enzyme activities and the antioxidant defense system.

Chemical inhibitors destabilize HuR binding to the AU-rich element of TNF-alpha mRNA.

Hu protein R (HuR) binds to the AU-rich element (ARE) in the 3UTR to stabilize TNF-alpha mRNA. Here, we identified chemical inhibitors of the interaction between HuR and the ARE of TNF-alpha mRNA using RNA electrophoretic mobility gel shift assay (EMSA) and filter binding assay. Of 179 chemicals screened, we identified three with a half-maximal inhibitory concentration (IC(50)) below 10 microM. The IC(50) of quercetin, b-40, and b-41 were 1.4, 0.38, and 6.21 microM, respectively, for binding of HuR protein to TNF-alpha mRNA. Quercetin and b-40 did not inhibit binding of tristetraprolin to the ARE of TNF-alpha mRNA. When LPS-treated RAW264.7 cells were treated with quercetin and b-40, we observed decreased stability of TNF-alpha mRNA and decreased levels of secreted TNF-alpha. From these results, we could find inhibitors for the TNF-alpha mRNA stability, which might be used advantageously for both the study for post-transcriptional regulation and the discovery of new anti-inflammation drugs.

Simultaneous quantification of 19 ginsenosides in black ginseng developed from Panax ginseng by HPLC-ELSD.

A high-performance liquid chromatographic method with evaporative light scattering detection (HPLC-ELSD) has been developed to identify and quantify 19 ginsenosides (Rg(1), Re, Rf, Rb(1), Rc, Rb(2), Rd, F(4), Rg(6), Rk(3), Rh(4), 20(S)-, 20(R)-Rg(3), 20(S)-, 20(R)-Rs(3), Rk(1), Rg(5), Rs(4), and Rs(5)) in black ginseng (BG, Korean white ginseng that was subjected to nine cycles of steam treatment). Ultrasonication is employed for sample preparation, and the analysis is achieved on a Discovery C(18) column using gradient elution of CH(3)CN-H(2)O-CH(3)COOH without buffer in 40min. The method was validated by linearity (r(2)> or =0.9994), precision (92.0-107.5%), intra- and inter-day accuracy (R.S.D.<3.21%), and limit of detection (LOD< or =93ng). The quantification method was applied to analyze the composition of ginsenosides in Korean white, red, and black ginsengs. During the preparatory process of BG, ginsenosides transform into constituents of low polarity by hydrolysis, isomerization, and dehydration at C-20, and hydrolysis also occurs at C-3 or C-6. The validated HPLC method is expected to provide the basis for the quality assessment of ginseng products.

Methanolic extracts of Uncaria rhynchophylla induce cytotoxicity and apoptosis in HT-29 human colon carcinoma cells.

In this paper, we report the anticancer activities of Uncaria rhynchophylla extracts, a Rubiaceae plant native to China. Traditionally, Uncaria rhynchophylla has been used in the prevention and treatment of neurotoxicity. However, the cytotoxic activity of Uncaria rhynchophylla against human colon carcinoma cells has not, until now, been elucidated. We found that the methanolic extract of Uncaria rhynchophylla (URE) have cytotoxic effects on HT-29 cells. The URE showed highly cytotoxic effects via the MTT reduction assay, LDH release assay, and colony formation assay. As expected, URE inhibited the growth of HT-29 cells in a dose-dependent manner. In particular, the methanolic URE of the 500 microg/ml showed 15.8% inhibition against growth of HT-29 cells. It induced characteristic apoptotic effects in HT-29 cells, including chromatin condensation and sharking occurring 24 h when the cells were treated at a concentration of the 500 microg/ml. The activation of caspase-3 and the specific proteolytic cleavage of poly (ADP-ribose) polymerase were detected over the course of apoptosis induction. These results indicate that URE contains bioactive materials with strong activity, and is a potential chemotherapeutic agent candidate against HT-29 human colon carcinoma cells.

Comparative analysis of effects of cytokine gene adjuvants on DNA vaccination against Mycobacterium tuberculosis heat shock protein 65.

DNA-based vaccines generate potent cellular immunity as well as humoral immunity. It seems evident that cytokines play a crucial role in generation of effector T cell subsets and in determining the magnitude of the response by DNA vaccines. In this study, we compared the effects of several TH1 cytokine genes as adjuvant in DNA vaccination using mycobacterial Hsp65 as a model antigen. Our results demonstrated that although the overall immune response to Hsp65 was enhanced by co-injection of Hsp65 DNA with cytokine genes, each cytokine gene was shown to affect different immune response elements. Co-injection of Hsp65 DNA with IL-12 or GM-CSF led to an increase in IFN-gamma production and represented potent protections against Mycobacterium tuberculosis challenge, while that with Eta-1, IL-12 or IL-18 gene led to an elevated IgG2a/IgG1 ratio. Interestingly, co-administration of Flt3L gene was shown to enhance the Ag-specific CTL response. These results show that the direction and magnitude of immune response in DNA vaccination against Hsp65 of M. tuberculosis could be modulated in different ways by co-injection of an appropriate cytokine gene as adjuvant.