The Androgenic Alopecia Protective Effects of Forsythiaside-A and the Molecular Regulation in a Mouse Model.

This study examined the inhibitory effect of forsythiaside-A, a natural substance derived from Forsythia suspensa (F. suspensa), on entry into catagen induced by dihydrotestosterone (DHT) in an androgenic alopecia mouse model. In vitro experiment comparing finasteride with forsythiaside-A showed that forsythiaside-A treatment resulted in a 30% greater inhibition of DHT-induced apoptosis in human hair dermal papilla cell (HHDPCs) and human keratinocytes (HaCaTs). In vivo experiment showed that mouse hair density and thickness were increased by 50% and 30%, respectively, in the forsythiaside-A-treated group when compared to a DHT group. Tissue histological results revealed that the forsythiaside-A-treated group had an increase in size and shape of the hair follicles and a 1.5 times increase in the follicle anagen/telogen ratio when compared to the finasteride group. Western blot examination of TGF-ß2 expression related to apoptosis signaling in mouse skin verified that forsythiaside-A reduced the expression of TGF-ß2 by 75% and suppressed apoptosis by reducing the expression of caspase-9 by 40%, and caspase-3 by 53%, which play an roles up-regulator in the apoptosis signal. The forsythiaside-A group also showed a 60% increase in the Bcl-2/Bax ratio, which is a factor related to mitochondrial apoptosis. Our results indicated that forsythiaside-A prevents apoptosis by similar mechanism with finasteride, but forsythiaside-A is more effective than finasteride. In summary, forsythiaside-A controlled the apoptosis of hair cells and retarded the entry into the catagen phase and therefore represents a natural product with much potential for use as a treatment for androgenic alopecia.

Ginsenoside F2 reduces hair loss by controlling apoptosis through the sterol regulatory element-binding protein cleavage activating protein and transforming growth factor-ß pathways in a dihydrotestosterone-induced mouse model.

This study was conducted to test whether ginsenoside F2 can reduce hair loss by influencing sterol regulatory element-binding protein (SREBP) cleavage-activating protein (SCAP) and the transforming growth factor beta (TGF-ß) pathway of apoptosis in dihydrotestosterone (DHT)-treated hair cells and in a DHT-induced hair loss model in mice. Results for ginsenoside F2 were compared with finasteride. DHT inhibits proliferation of hair cells and induces androgenetic alopecia and was shown to activate an apoptosis signal pathway both in vitro and in vivo. The cell-based 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay showed that the proliferation rates of DHT-treated human hair dermal papilla cells (HHDPCs) and HaCaTs increased by 48% in the ginsenoside F2-treated group and by 12% in the finasteride-treated group. Western blot analysis showed that ginsenoside F2 decreased expression of TGF-ß2 related factors involved in hair loss. The present study suggested a hair loss related pathway by changing SCAP related apoptosis pathway, which has been known to control cholesterol metabolism. SCAP, sterol regulatory element-binding protein (SREBP) and caspase-12 expression in the ginsenoside F2-treated group were decreased compared to the DHT and finasteride-treated group. C57BL/6 mice were also prepared by injection with DHT and then treated with ginsenoside F2 or finasteride. Hair growth rate, density, thickness measurements and tissue histotological analysis in these groups suggested that ginsenoside F2 suppressed hair cell apoptosis and premature entry to catagen more effectively than finasteride. Our results indicated that ginsenoside F2 decreased the expression of TGF-ß2 and SCAP proteins, which have been suggested to be involved in apoptosis and entry into catagen. This study provides evidence those factors in the SCAP pathway could be targets for hair loss prevention drugs.

Lactobacillus yonginensis sp. nov., a lactic acid bacterium with ginsenoside converting activity isolated from Kimchi.

A Gram-reaction-positive, non-motile, non-spore-forming, catalase-negative, facultatively anaerobic, rod-shaped, ß-glucosidase-producing lactic acid bacterium, designated strain THK-V8(T), was isolated from the Korean fermented food, Kimchi, and its taxonomic position was investigated by using a polyphasic approach. Strain THK-V8(T) was able to grow at 4-40 °C (optimum, 30 °C) and pH 4.0-7.0 (optimum, pH 6.0). Strain THK-V8(T) had the ability to transform ginsenoside Rb1 to Rd. On the basis of 16S rRNA gene sequence similarity data, strain THK-V8(T) was shown to belong to the genus Lactobacillus. Strain THK-V8(T) was related to Lactobacillus koreensis DCY50(T) (98.8% sequence similarity), Lactobacillus parabrevis LMG 11984(T) (97.7%), Lactobacillus senmaizukei L13(T) (97.5%), Lactobacillus hammesii TMW1.1236(T) (97.3%) and Lactobacillus brevis ATCC 14687(T) (97.2%). Subsequently, sequence analysis of the RNA polymerase alpha subunit gene (rpoA) confirmed that strain THK-V8(T) showed a maximum rpoA gene sequence similarity value of 93% with Lactobacillus brevis LMG 6906(T). The G+C content of the genomic DNA was 47.8 mol%. The DNA-DNA hybridization values between strain THK-V8(T) and Lactobacillus parabrevis DCY50(T) and Lactobacillus parabrevis LMG 11984(T) were 46.1 ± 4.9% and 10.6 ± 2.9%, respectively. The major fatty acids were summed feature 7 (comprised of C(19:0) cyclo ω10c/19ω6), C(14:0), C(16:0) and C(18:1)ω9c. The cell wall peptidoglycan was of the A4α L-Lys-D-Asp type. The phenotypic and molecular properties indicated that strain THK-V8(T) represents a novel species within the genus Lactobacillus, for which the name Lactobacillus yonginensis sp. nov. is proposed. The type strain is THK-V8(T) ( =KACC 16236(T) =JCM 18023(T)).

Pedobacter ginsenosidimutans sp. nov., with ginsenoside-converting activity.

A Gram-reaction-negative, strictly aerobic, non-motile, non-spore-forming and rod-shaped bacterial strain, designated THG-45(T), was isolated from soil of a ginseng field of Pocheon province in the Republic of Korea and its taxonomic position was investigated by a polyphasic approach. Growth occurred at 4-30 °C, at pH 5.5-9.0 and with 0-2 % (w/v) NaCl on nutrient agar. On the basis of 16S rRNA gene sequence similarity, strain THG-45(T) was shown to belong to the genus Pedobacter and was related to Pedobacter borealis G-1(T) (98.8 %), P. alluvionis NWER-II11(T) (97.9 %), P. agri PB92(T) (97.9 %), P. terrae DS-57(T) (97.5 %), P. suwonensis 15-52(T) (97.4 %), P. sandarakinus DS-27(T) (97.0 %) and P. soli 15-51(T) (97.0 %), but DNA relatedness between strain THG-45(T) and these strains was below 36 %. The G+C content of the genomic DNA was 39 mol%. The only isoprenoid quinone detected in strain THG-45(T) was menaquinone-7 (MK-7). The predominant fatty acids were iso-C15 : 0, summed feature 3 (comprising C16 : 1ω6c and/or C16 : 1ω7c) and iso-C17 : 0 3-OH, and the major polar lipids were phosphatidylethanolamine and an unidentified aminophosphoglycolipid. Phenotypic data and phylogenetic inference supported the affiliation of strain THG-45(T) to the genus Pedobacter, and a number of biochemical tests differentiated strain THG-45(T) from the recognized species of the genus Pedobacter. Therefore, the novel isolate represents a novel species, for which the name Pedobacter ginsenosidimutans sp. nov. is proposed, with THG-45(T) as the type strain ( = KACC 14530(T) = JCM 16721(T)).

Hair growth activity of Crataegus pinnatifida on C57BL/6 mouse model.

Crataegus pinnatifida has a long history of use in traditional oriental herbal medicine to stimulating digestion and improving blood circulation. Based on nutrition of hair, the present study was conducted to assess the effect of C. pinnatifida extract on hair growth using mouse model and its mechanisms of action. The C. pinnatifida extract containing the contents of total polyphenol of 5.88□0.82 g gallic acid/100 g extract and proanthocyanidin of 9.15□1.58 mg cyaniding chloride/100 g extract was orally administered daily at a dosage of 50 mg/kg weight to the 7-week-old C57BL/6 mice in telogen. The C. pinnatifida extract promoted hair growth by inducing anagen phase in mice in telogen, reflected by color of skin, thickness of hair shaft, and density of hair. The ratio of anagento telogen was determined by shape of hair follicles in vertically sectioned slide and increased by oral administration of C. pinnatifida extract. The number and the size of hair follicles were also enlarged, indicating anagen phase induction. The proliferation of human dermal papilla cells (hDPC) was accelerated by addition of C. pinnatifida extract, which activated the signaling of mitogen-activated protein kinases (Erk, p-38, and JNK) and Akt. Moreover, the ratio of Bcl-2/Bax as the determinant of cell fate was also raised in skin. These results suggest that the C. pinnatifida extract promotes hair growth by inducing anagen phase, which might be mediated by the activation of cellular signalings that enhance the survival of cultured hDPC and the increase of the ratio of Bcl-2 to Bax that protects cells against cell death.

Effects of Galla chinensis extracts on UVB-irradiated MMP-1 production in hairless mice.

Galla chinensis (GAC) is a natural traditional Chinese medicine that has been widely used in folk medicine. Although GAC compounds (mainly gallic acid and methyl gallate) possess strong antiviral, antibacterial, anticancer, and antioxidant activities, there is no report regarding topical or oral administration of GAC compounds on UVB irradiation-induced photoaging in hairless mice (SKH: HR-1). In the present study, we examined cell viability, intracellular reactive oxygen species (ROS), matrix metalloproteinase-1 (MMP-1), and interleukin-6 (IL-6) in skin fibroblasts and keratinocytes induced by UVB in vitro. We also studied skin damage by measuring skin thickness, elasticity, wrinkling and levels of protein MMP-1, elastin, procollagen type I, and transforming growth factor-ß1 (TGF-ß1) in hairless mouse skin chronically irradiated by UVB in vivo. GAC treatment significantly prevented skin photoaging by reducing the levels of ROS, MMP-1, and IL-6 and promoting production of elastin, procollagen type I, and TGF-ß1. According to the results of H&E staining and Masson's trichrome staining, GAC reduced skin thickness and wrinkle formation while it increased skin elasticity. The effects of GAC on UVB-induced skin photoaging may be due to suppressed MMP-1 expression. These findings could be referenced for the development of new agents that target UVB-induced photoaging.

Vigna angularis water extracts protect against ultraviolet b-exposed skin aging in vitro and in vivo.

Exposure to ultraviolet (UV) radiation induces various pathological changes, such as thickened skin and wrinkle formation. In particular, UVB irradiation increases matrix metalloproteinase (MMP)-1 production and collagen degradation, leading to premature aging, termed photoaging. The azuki bean (Vigna angularis; VA) has been widely used as a food product as well as a traditional medicine. However, its activity needs additional study to confirm its functional application in foods and cosmetics for protecting skin. In this study, hot-water extract from VA (VAE) and its active component, rutin, were investigated to determine their antiphotoaging effects. VAE was found to have antioxidant activity. In UVB-exposed normal human dermal fibroblasts cells with VAE and rutin treatments, MMP-1 production was significantly suppressed (90% and 47%, respectively). The effects of both topical and oral administration of VAE were tested in UVB-irradiated hairless mice. VAE suppressed wrinkle formation and skin thickness by promoting elastin, procollagen type I, and TGF-ß1 expression (118%, 156%, and 136%, respectively) and by diminishing MMP-1 production. These results suggest that VAE may be effective for preventing skin photoaging accelerated by UVB radiation.

The inductive effect of ginsenoside F2 on hair growth by altering the WNT signal pathway in telogen mouse skin.

This study was conducted to confirm the possibility of using minor ginseng saponin F2 by oral administration on hair anagen induction effects. The signaling pathway and anagen induction effect of ginsenoside F2 were investigated and compared with finasteride on the effect of hair growth induction. The cell-based MTT assay results indicated that the proliferation rates of HHDPC and HaCaT treated with F2 significantly increased by 30% compared with the finasteride-treated group. A western blot study showed that the expression of ß-catenin Lef-1 and DKK-1 increased by 140, 200% and decreased by 40% in the F2-treated group, respectively compared to that of finasteride-treated group. C57BL/6 mice were subjected to the same treatments. The hair growth promotion rates were compared with groups treated with finasteride, which was 20% higher in the F2-treated group. Tissue histological analysis results showed the number of hair follicles, thickness of the epidermis, and follicles of the anagen phase which increased in the F2-treated group, compared with the finasteride-treated groups. Moreover, the effect of F2 on hair growth was confirmed through the immunofluorescence (IF) methods indicating the expression aspect of Wnt signal pathway-related factors in the tissue of C57BL/6 mouse. Our results considered the expression increase in ß-catenin, Lef-1 which was suggested as a major factor related to the development and growth of hair follicle and the decrease in DKK-1 when entering catagen by F2. As the data showed, F2 might be a potential new therapeutic source for anagen induction and hair growth through the Wnt signal pathway.

The bone regenerative effects of fucosterol in in vitro and in vivo models of postmenopausal osteoporosis.

SCOPE: The aim of this study was to investigate the bone regenerative effects of fucosterol in estrogen-deficient ovariectomized (OVX) rats. METHODS AND RESULTS: Bone regeneration was assessed in fucosterol-treated MG63 cells in vitro via assays for osteoblast proliferation, alkaline phosphatase, and osteoclast differentiation. Osteoblast proliferation rates, alkaline phosphatase activity, and mineralization were increased in the fucosterol-treated group. Moreover, differentiation of osteoclasts was decreased in the fucosterol-treated group. In the in vivo assay, female rats were OVX. Twelve weeks after ovariectomy, rats were divided into seven groups, each oral administrate everyday for 7 weeks. The bone mineral density of femoral bones was higher in fucosterol groups than in OVX control, and body weight was lower in fucosterol groups. Among bone-quality parameters, bone volume/total volume increased and trabecular separation decreased in fucosterol groups relative to the OVX control. Bone formation and resorption were evaluated using the serum biomarkers osteocalcin and CTx. Fucosterol tripled the level of serum osteocalcin relative to the OVX group and reduced the serum level of CTx. CONCLUSION: These results suggest that fucosterol has the dual potentials to activate osteoblasts to stimulate bone formation and suppress differentiation of osteoclasts so as to reduce bone resorption.

The protective effects of fucosterol against skin damage in UVB-irradiated human dermal fibroblasts.

Exposure to ultraviolet (UV) light causes matrix metalloproteinase (MMP) overexpression and extracellular matrix depletion, leading to skin photoaging. The activation of MMP is related to increased interlukin-6 (IL-6) and type I procollagen production, which is regulated by transforming growth factor-ß1 (TGF-ß1). Activator protein-1 (AP-1) activation induces MMP-1 production and reduces type I procollagen secretion. Fucosterol, which is extracted and purified from the brown algae Hizikia fusiformis, is a phytosterol. We assessed the effects of fucosterol on photodamage and investigated its molecular mechanism of action in UVB-irradiated normal human dermal fibroblasts by using enzyme-linked immunosorbent assay, Western blot analysis, and reverse transcription-polymerase chain reaction. Our results showed that fucosterol significantly decreased the UVB-induced expression of MMP-1, IL-6, p-c-Jun, and p-c-Fos. Additionally, fucosterol markedly increased the UVB-induced production of type I procollagen and TGF-ß1. Our results indicate that fucosterol regulates MMP-1 and type I procollagen expression by modulating AP-1 and TGF-ß1 signaling and that MMP-1 activation is correlated with IL-6. These data suggest that fucosterol is a promising botanical agent to protect against skin photodamage.

Immunostimulatory effect of fermented red ginseng in the mouse model.

In this study, Woongjin fermented red ginseng extract (WFRG) was evaluated for its potential ability to act as an adjuvant for the immune response of mice. For the in vitro study, macrophages were treated with serial concentrations (1 µg/mL, 10 µg/mL, and 100 µg/mL) of WFRG. For in vivo studies, mice were administered different concentrations (10 mg/kg/day, 100 mg/kg/day, and 200 mg/kg/day) of WFRG orally for 21 days. In vitro, the production of nitric oxide and TNF-α by RAW 264.7 cells increased in a dose-dependent manner. In vivo, WFRG enhanced the proliferation of splenocytes induced by two mitogens (i.e., concanavalin A and lipopolysaccharide [LPS]) and increased LPS-induced production of TNF-α and IL-6, but not IL-1ß. In conclusion, WFRG has the potential to modulate immune function and should be further investigated as an immunostimulatory agent.

Sphingobacterium ginsenosidimutans sp. nov., a bacterium with ginsenoside-converting activity isolated from the soil of a ginseng field.

A Gram-staining-negative, aerobic, non-motile, non-spore-forming, and rod-shaped bacterium designated THG 07(T) was isolated from the soil of a ginseng field of Pocheon in South Korea, and its taxonomic position was investigated by using a polyphasic study. Strain THG 07(T) grew optimally at 25-30°C and at pH 6.5-7.0 and in the absence of NaCl on nutrient agar. Strain THG-T17(T) displayed ß-glucosidase activity that was responsible for its ability to transform ginsenoside Rb1 (one of the dominant ginsenosides of ginseng) to compound C-K. On the basis of 16S rRNA gene sequence similarity, strain THG 07(T) was shown to belong to the family Sphingobacteriaceae and was related to Sphingobacterium canadense CR11(T) (98.7%), S. cladoniae No.6(T) (98.1%), S. detergens 6.2S(T) (98.0%), S. multivorum IAM14316(T) (97.9%), S. siyangense SY1(T) (97.8%) and S. thalpophilum DSM11723(T) (96.9%). The G+C content of the genomic DNA was 40.6 mol%. The major menaquinone, MK-7, and major fatty acids, iso-C15:0 and C16:1 ω7c and/or ω6c, supported the affiliation of strain THG 07(T) to the genus Sphingobacterium. The DNA-DNA relatedness values between strain THG 07(T) and its closest phylogenetic neighbors were below 26.9%. The results of physiological and biochemical tests enabled strain THG 07(T) to be differentiated phenotypically from the recognized species of the genus Sphingobacterium. Therefore the isolate represents a novel species, for which the name Sphingobacterium ginsenosidimutans sp. nov. is proposed, with the type strain THG 07(T) (=KACC 14526(T)=JCM 16722(T)).

Enzyme-processed Korean Red Ginseng extracts protects against skin damage induced by UVB irradiation in hairless mice.

UV irradiation is the main factor contributing to skin damages that are associated with an excessive production of matrix-degrading metalloproteinase (MMP)-1 and a deficient expression of collagens. To date, red ginseng has been revealed to possess many biomedical effects, such as anti-aging, anti-oxidation, and anti-inflammatory. In this study, we prepared the Korean Red Ginseng extracts treated with enzyme (KRGE) and investigated the effects of dietary KRGE on the formation of wrinkles generated by UVB irradiation in hairless mice. It was found that KRGE inhibited the UVB-induced formation of wrinkles, epidermal thickness, and skin dryness in hairless mice. Further results also showed that KRGE attenuated UVB-induced MMP-1 level, while accelerated procollagen type I, transforming growth factor-ß1 secretion. Interestingly, the expression of profilaggrin and filaggrin in both the epidermis and dermis were decreased due to UVB exposure and reversed by KRGE. The KRGE 0.06% was prior to KRGE 0.24%. In view of these results, which indicated that KRGE protected skin from UVB-induced photodamages, which may not only mediated by regulating of MMP-1 and procollagen type I, but also by increasing the production of profilaggrin and filaggrin. In conclusion, our results suggest that KRGE may be a promising agent for the treatment of skin photodamages. The challenge of KRGE will be expected as cosmeceuticals and nutraceuticals in order to intervene in aging-related degenerative skin changes.

Anti-Cancer Effect of Ginsenoside F2 against Glioblastoma Multiforme in Xenograft Model in SD Rats.

The glioblastoma multiforme (GBM) is the most common malignant brain tumor in adults. Despite combination treatments of radiation and chemotherapy, the survival periods are very short. Therefore, this study was conducted to assess the potential of ginsenoside F2 (F2) to treat GBM. In in vitro experiments with glioblastoma cells U373MG, F2 showed the cytotoxic effect with IC50 of 50 µg/mL through apoptosis, confirmed by DNA condensation and fragmentation. The cell population of cell cycle sub-G1 as indicative of apoptosis was also increased. In xenograft model in SD rats, F2 at dosage of 35 mg/kg weight was intravenously injected every two days. This reduced the tumor growth in magnetic resonance imaging images. The immunohistochemistry revealed that the anticancer activity might be mediated through inhibition of proliferation judged by Ki67 and apoptosis induced by activation of caspase-3 and -8. And the lowered expression of CD31 showed the reduction in blood vessel densities. The expression of matrix metalloproteinase-9 for invasion of cancer was also inhibited. The cell populations with cancer stem cell markers of CD133 and nestin were reduced. The results of this study suggested that F2 could be a new potential chemotherapeutic drug for GBM treatment by inhibiting the growth and invasion of cancer.

Hijikia fusiforme protects against ovariectomy-induced bone loss in rats.

The prophylactic effects of Hijikia fusiforme on bone metabolism were examined using in vitro indices of bone formation and resorption. As the indices of bone formation, osteoblast proliferation and differentiation were measured through mitochondrial enzyme activity [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay] and bone marker alkaline phosphatase (ALP) activity. The aqueous extract of H. fusiforme stimulated the proliferation of the human osteoblast-like cell line MG63 and the ALP activity of the mouse osteoblast-like cell line MC3T3-E1. Moreover, extracellular matrix mineralization was accelerated by the addition of H. fusiforme. As the indices of bone resorption, differentiation of the murine macrophage/osteoclast precursor cell line RAW 264.7 by receptor activator of nuclear factor-κB ligand (RANKL) was measured as tartrate-resistant acid phosphatase-positive multinucleated cells, which were suppressed by H. fusiforme. Additionally, H. fusiforme lowered the secreted amount of RANKL that is required for the osteoclastic differentiation and activation, but the amount of osteoprotegerin as a decoy receptor for RANKL was not affected. The bone-protective effects of H. fusiforme in estrogen-deficient ovariectomized rats were also investigated. Osteoporosis was induced in female Sprague-Dawley rats by ovariectomy for 15 weeks, and then H. fusiforme was orally administered at a dose of 100 mg/kg of body weight every day for 6 weeks. Bone mineral density in the group orally administered H. fusiforme was increased, compared with ovariectomized rats, but not significantly (P>.05). Oral administration of H. fusiforme improved microarchitecture of bone in terms of bone volume (bone volume/total volume ratio) and trabecular separation (P<.05). The amounts of osteocalcin and C-telopeptide type I collagen in serum were measured as the biomarkers for bone formation and resorption. The level of osteocalcin in serum was increased, but not significantly. However, the level of C-telopeptide type I collagen in serum was significantly decreased (P<.05). When the results are taken together, the present study indicates that H. fusiforme might be useful in the treatment of osteoporosis.