Apigenin Isolated from Carduus crispus Protects against H2O2-Induced Oxidative Damage and Spermatogenic Expression Changes in GC-2spd Sperm Cells.

Testicular oxidative stress is one of the most common factors underlying male infertility. Welted thistle, Carduus crispus Linn., and its bioactive principles are attracting scientific interest in treating male reproductive dysfunctions. Here, the protective effects of apigenin isolated from C. crispus against oxidative damage induced by hydrogen peroxide (H2O2) and dysregulation in spermatogenesis associated parameters in testicular sperm cells was investigated. Cell viabilities, ROS scavenging effects, and spermatogenic associated molecular expressions were measured by MTT, DCF-DA, Western blotting and real-time RT-PCR, respectively. A single peak with 100% purity of apigenin was obtained in HPLC conditions. Apigenin treated alone (2.5, 5, 10 and 20 µM) did not exhibit cytotoxicity, but inhibited the H2O2-induced cellular damage and elevated ROS levels significantly (p < 0.05 at 5, 10 and 20 µM) and dose-dependently. Further, H2O2-induced down-regulation of antioxidant (glutathione S-transferases m5, glutathione peroxidase 4, and peroxiredoxin 3) and spermatogenesis-associated (nectin-2 and phosphorylated-cAMP response element-binding protein) molecular expression in GC-2spd cells were attenuated by apigenin at both protein and mRNA levels (p < 0.05). In conclusion, our study showed that apigenin isolated from C. crispus might be an effective agent that can protect ROS-induced testicular dysfunctions.

Cordycepin mitigates spermatogenic and redox related expression in H2O2-exposed Leydig cells and regulates testicular oxidative apoptotic signalling in aged rats.

CONTEXT: Cordycepin (COR), from Cordyceps militaris L., (Cordycipitaceae), is a valuable agent with immense health benefits. OBJECTIVE: The protective effects of COR in ageing-associated oxidative and apoptosis events in vivo and hydrogen peroxide (H2O2)-exposed spermatogenesis gene alterations in TM3 Leydig cells was investigated. MATERIALS AND METHODS: Male Sprague-Dawley rats were divided into young control (YC), aged control (AC) and COR treated (COR-20) aged groups. COR-20 group received daily doses of COR (20 mg/kg) for 6 months. Cell viability and hormone levels were analysed by MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] and enzyme immunoassay kits with COR treated at 1, 5, and 10 µg/mL. Oxidative enzymes, spermatogenic, and apoptotic expression in testis tissues were evaluated by Western blotting and real-time RT-PCR. RESULTS: COR treatment (1, 5, and 10 µg/mL) significantly (p < 0.05 ∼ p < 0.001) inhibited the H2O2-induced decrease in the percentage of viable cells (from 63.27% to 71.25%, 85.67% and 93.97%, respectively), and reduced the malondialdehyde (MDA) content (from 4.28 to 3.98, 3.14 and 1.78 nM MDA/mg protein, respectively). Further, the decreased antioxidant enzymes (glutathione-S-transferase mu5, glutathione peroxidase 4 and peroxiredoxin 3), spermatogenesis-related factors (nectin-2 and inhibin-α) and testosterone levels in H2O2-exposed TM3 cells were significantly (p < 0.05 ∼ p < 0.001) ameliorated by COR. In aged rats, COR (20 mg/kg) restored the altered enzymatic and non-enzymatic antioxidative status and attenuated the apoptotic p53 and Bax/Bcl-2 expression significantly (p < 0.05). CONCLUSION: COR might be developed as a potential agent against ageing-associated and oxidative stress-induced male infertility.

Pectinase-treated Panax ginseng protects heat stress-induced testicular damage in rats.

Testicular hyperthermia is well studied to cause impaired spermatogenesis. In the present study, the protective effect of enzymatically modified (pectinase-treated) Panax ginseng (GINST) against intermittent sub-chronic heat stress-induced testicular damage in rats was investigated. Male Sprague-Dawley rats were divided into four groups: normal control (NC), heat-stressed control (HC), heat-stressed plus GINST-100 mg/kg/day (HG100) and heat-stressed plus GINST-200 mg/kg/day (HG200) treatment groups. GINST (100 and 200 mg/kg/day) was mixed separately with a regular pellet diet and was administered orally for 8 weeks starting from 1 week before heat exposure. Parameters such as organ weight, blood chemistry, sperm kinetic values, expression of antioxidant enzymes, spermatogenesis molecules and sex hormone receptors levels were measured. Data revealed that kidney and epididymis weight were significantly (P < 0.05) decreased with heat stress and recovered by GINST treatment. Further, the altered levels of blood chemistry panels and sperm kinetic values in heat stress-induced rats were attenuated when GINST was administered (P < 0.05). Furthermore, the expression levels of antioxidant-related enzymes (GSTM5 and GPX4), spermatogenesis-related proteins (CREB1 and INHA) and sex hormone receptors (androgen receptor, luteinizing hormone receptor and follicle-stimulating hormone receptor) were reduced by heat stress; however, GINST treatment effectively ameliorated these changes. In conclusion, GINST was effective in reducing heat-induced damage in various male fertility factors in vivo and has considerable potential to be developed as a useful supplement in improving male fertility.

Lactobacillus vespulae sp. nov., isolated from gut of a queen wasp (Vespula vulgaris).

A Gram-stain-positive, oxidase- and catalase-negative, rod-shaped, facultatively anaerobic bacterial strain, DCY75T, was isolated from a queen wasp (Vespula vulgaris). Growth occurred at 4­37 °C (optimum, 30 °C), at pH 3.5­8.0 (optimum, pH 5.0­6.0) and with ≤ 7.0 % (w/v) NaCl. Strain DCY75T produced gas during growth on glucose. Phylogenetic analysis based on 16S rRNA gene sequences indicated that strain DCY75T belonged to the genus Lactobacillus and was closely related to Lactobacillus sanfranciscensis ATCC 27651T and Lactobacillus lindneri DSM 20690T at sequence similarities of 96.7 and 96.4 %, respectively. A comparison of two housekeeping genes, pheS and rpoA, revealed that strain DCT75T was well separated from other species of the genus Lactobacillus. Strain DCY75T produced d- and l-lactic acid isomers in a ratio of 22.5 : 77.5 (v/v). The major fatty acids were summed feature 8 (comprising C18 : 1ω7c and/or C18 : 1ω6c), C16 : 0, C18 : 1ω9c and C18 : 0.The peptidoglycan structure was of the A4α (l-Lys­d-Asp) type. Cell-wall sugars were glucose, galactose and ribose. The DNA G+C content was 35.5 ± 1.3 mol%. Based on phenotypic and genotypic properties, strain DCY75T represents a novel species of the genus Lactobacillus, for which the name Lactobacillus vespulae sp. nov. is proposed. The type strain is DCY75T ( = KCTC 21023T = JCM 19742T).

Protection of apigenin against kainate-induced excitotoxicity by anti-oxidative effects.

Apigenin (5,7,4'-trihydroxyflavone) is a principal ingredient of Cirsium japonicum. These experiments were performed to determine whether apigenin has neuroprotective effects against kainic acid (KA)-induced excitotoxicity in vitro and in vivo. Intraperitoneal (i.p.) administration of apigenin (25, 50 mg/kg) decreased the seizure scores induced by KA injection (40 mg/kg, i.p.) in mice. In addition, the convulsion onset time was significantly delayed by apigenin administration. Moreover, we found that apigenin blocked KA-induced seizure-form electroencephalogram (EEG) discharge activity in the brain cortex. In hippocampal cells, apigenin inhibited KA-induced excitotoxicity in a dose-dependent manner as measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. To study the possible mechanisms underlying the in vitro neuroprotective effects of apigenin against KA-induced cytotoxicity, we also examined the effect of apigenin on intracellular reactive oxygen species (ROS) elevations in cultured hippocampal neurons and found that apigenin treatment dose-dependently inhibited intracellular ROS elevation. The remarkable reduction of glutathione (GSH) levels induced by KA in hippocampal tissues was reversed by apigenin in a dose-dependent manner. In addition, similar results were obtained after pretreatment with free radical scavengers such as trolox and dimethylthiourea (DMTU). Finally, after confirming the protective effect of apigenin in hippocampal CA3 region, we found apigenin is an active compound in KA-induced neuroprotection. These results collectively indicate that apigenin alleviates KA-induced excitotoxicity by quenching ROS as well as inhibiting GSH depletion in hippocampal neurons.

Effect of long-term administration of cordycepin from Cordyceps militaris on testicular function in middle-aged rats.

This study was carried out to examine the potential beneficial effect of cordycepin on the decline of testicular function induced with age. A total of 30 male Sprague-Dawley rats (twenty-four 12-month-olds and six 2-month-olds) were divided into five groups. The young control (YC) and middle-aged control (MC) groups received vehicle only. Cordycepin-treated groups were administered daily doses of oral cordycepin at 5, 10, and 20 mg/kg body weight for 4 months. As a result, the MC group exhibited epididymal weight loss, decreased sperm motility, and reduced spermatogenesis compared to the young control group. Interestingly, the epididymal weights of middle-aged rats were dose-dependently increased by treatment with cordycepin. Cordycepin also improved calcium levels and decreased urea and nitrogen, uric acid, and creatinine in the blood of middle-aged rats. In addition, cordycepin significantly increased sperm motility and the progressiveness of sperm movement. All cordycepin-treated groups showed well-arranged spermatogonia, densely packed cellular material, and increased numbers of mature spermatozoa in the seminiferous lumen compared to the middle-aged control group. These results indicate that long-term administration of cordycepin can counteract the decline of testicular function in middle-aged rats.

Cordycepin from Medicinal Fungi Cordyceps militaris Mitigates Inflammaging-Associated Testicular Damage via Regulating NF-κB/MAPKs Signaling in Naturally Aged Rats.

Inflammaging in male reproductive organs covers a wide variety of problems, including sexual dysfunction and infertility. In this study, the beneficial effects of cordycepin (COR), isolated from potential medicinal fungi Cordyceps militaris, in aging-associated testicular inflammation and serum biochemical changes in naturally aged rats were investigated. Male Sprague Dawley rats were divided into young control (YC), aged control (AC), and COR (5, 10, and 20 mg/kg) treated aged rat groups. Aging-associated serum biochemical changes and inflammatory parameters were analyzed by biochemical assay kits, Western blotting, and real-time RT-PCR. Results showed a significant (p < 0.05) alteration in the total blood cell count, lipid metabolism, and liver functional parameters in AC group when compared with YC group. However, COR-treated aged rats ameliorated the altered biochemical parameters significantly (p < 0.05 and p < 0.01 at 5, 10, and 20 mg/kg, respectively). Furthermore, the increase in the expression of inflammatory mediators (COX-2, interleukin (IL)-6, IL-1ß, and tissue necrosis factor-alpha) in aged rat testis was significant (p < 0.05) when compared with YC group. Treatment with COR at 20 mg/kg to aged rats attenuated the increased expression of inflammatory mediators significantly (p < 0.05). Mechanistic studies revealed that the potential attenuating effects exhibited by COR in aged rats was mediated by regulation of NF-κB activation and MAPKs (c-Jun N-terminal kinase, extracellular signal-regulated kinase 1/2, and p38) signaling. In conclusion, COR restored the altered serum biochemical parameters in aged rats and ameliorated the aging-associated testicular inflammation proving the therapeutic benefits of COR targeting inflammaging-associated male sexual dysfunctions.

Korean Red Ginseng aqueous extract improves markers of mucociliary clearance by stimulating chloride secretion.

BACKGROUND: Abnormal chloride (Cl-) transport has a detrimental impact on mucociliary clearance in both cystic fibrosis (CF) and non-CF chronic rhinosinusitis. Ginseng is a medicinal plant noted to have anti-inflammatory and antimicrobial properties. The present study aims to assess the capability of red ginseng aqueous extract (RGAE) to promote transepithelial Cl- secretion in nasal epithelium. METHODS: Primary murine nasal septal epithelial (MNSE) [wild-type (WT) and transgenic CFTR-/-], fisher-rat-thyroid (FRT) cells expressing human WT CFTR, and TMEM16A-expressing human embryonic kidney cultures were utilized for the present experiments. Ciliary beat frequency (CBF) and airway surface liquid (ASL) depth measurements were performed using micro-optical coherence tomography (µOCT). Mechanisms underlying transepithelial Cl- transport were determined using pharmacologic manipulation in Ussing chambers and whole-cell patch clamp analysis. RESULTS: RGAE (at 30µg/mL of ginsenosides) significantly increased Cl- transport [measured as change in short-circuit current (ΔISC = µA/cm2)] when compared with control in WT and CFTR-/- MNSE (WT vs control = 49.8±2.6 vs 0.1+/-0.2, CFTR-/- = 33.5±1.5 vs 0.2±0.3, p < 0.0001). In FRT cells, the CFTR-mediated ΔISC attributed to RGAE was small (6.8 ± 2.5 vs control, 0.03 ± 0.01, p < 0.05). In patch clamp, TMEM16A-mediated currents were markedly improved with co-administration of RGAE and uridine 5-triphosphate (8406.3 +/- 807.7 pA) over uridine 5-triphosphate (3524.1 +/- 292.4 pA) or RGAE alone (465.2 +/- 90.7 pA) (p < 0.0001). ASL and CBF were significantly greater with RGAE (6.2+/-0.3 µm vs control, 3.9+/-0.09 µm; 10.4+/-0.3 Hz vs control, 7.3 ± 0.2 Hz; p < 0.0001) in MNSE. CONCLUSION: RGAE augments ASL depth and CBF by stimulating Cl- secretion through CaCC, which suggests therapeutic potential in both CF and non-CF chronic rhinosinusitis.

Korean Red Ginseng alleviates neuroinflammation and promotes cell survival in the intermittent heat stress-induced rat brain by suppressing oxidative stress via estrogen receptor beta and brain-derived neurotrophic factor upregulation.

BACKGROUND: Heat stress orchestrates neurodegenerative disorders and results in the formation of reactive oxygen species that leads to cell death. Although the immunomodulatory effects of ginseng are well studied, the mechanism by which ginseng alleviates heat stress in the brain remains elusive. METHODS: Rats were exposed to intermittent heat stress for 6 months, and brain samples were examined to elucidate survival and antiinflammatory effect after Korean Red Ginseng (KRG) treatment. RESULTS: Intermittent long-term heat stress (ILTHS) upregulated the expression of cyclooxygenase 2 and inducible nitric oxide synthase, increasing infiltration of inflammatory cells (hematoxylin and eosin staining) and the level of proinflammatory cytokines [tumor necrosis factor α, interferon gamma (IFN-γ), interleukin (IL)-1ß, IL-6], leading to cell death (terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling assay) and elevated markers of oxidative stress damage (myeloperoxidase and malondialdehyde), resulting in the downregulation of antiapoptotic markers (Bcl-2 and Bcl-xL) and expression of estrogen receptor beta and brain-derived neurotrophic factor, key factors in regulating neuronal cell survival. In contrast, KRG mitigated ILTHS-induced release of proinflammatory mediators, upregulated the mRNA level of the antiinflammatory cytokine IL-10, and increased myeloperoxidase and malondialdehyde levels. In addition, KRG significantly decreased the expression of the proapoptotic marker (Bax), did not affect caspase-3 expression, but increased the expression of antiapoptotic markers (Bcl-2 and Bcl-xL). Furthermore, KRG significantly activated the expression of both estrogen receptor beta and brain-derived neurotrophic factor. CONCLUSION: ILTHS induced oxidative stress responses and inflammatory molecules, which can lead to impaired neurogenesis and ultimately neuronal death, whereas, KRG, being the antioxidant, inhibited neuronal damage and increased cell viability.

Compound K improves skin barrier function by increasing SPINK5 expression.

BACKGROUND: The skin acts as a barrier to protect organisms against harmful exogenous agents. Compound K (CK) is an active metabolite of ginsenoside Rb1, Rb2 and Rc, and researchers have focused on its skin protective efficacy. In this study, we hypothesized that increased expression of the serine protease inhibitor Kazal type-5 (SPINK5) may improve skin barrier function. METHODS: We screened several ginsenosides to increase SPINK5 gene promoter activity using a transactivation assay and found that CK can increase SPINK5 expression. To investigate the protective effect of CK on the skin barrier, RT-PCR and Western blotting were performed to investigate the expression levels of SPINK5, kallikrein 5 (KLK5), KLK7 and PAR2 in UVB-irradiated HaCaT cells. Measurement of transepidermal water loss (TEWL) and histological changes associated with the skin barrier were performed in a UVB-irradiated mouse model and a 1-chloro-2,4-dinitrobenzene (DNCB)-induced atopic dermatitis-like model. RESULTS: CK treatment increased the expression of SPINK5 and decreased the expression of its downstream genes, such as KLKs and PAR2. In the UVB-irradiated mouse model and the DNCB-induced atopic dermatitis model, CK restored increased TEWL and decreased hydration and epidermal hyperplasia. In addition, CK normalized the reduced SPINK5 expression caused by UVB or DNCB, thereby restoring the expression of the proteins involved in desquamation to a level similar to normal. CONCLUSIONS: Our data showed that CK contributes to improving skin-barrier function in UVB-irradiated and DNCB-induced atopic dermatitis-like models through SPINK5. These results suggest that therapeutic attempts with CK might be useful in treating barrier-disrupted diseases.

Cordycepin causes p21WAF1-mediated G2/M cell-cycle arrest by regulating c-Jun N-terminal kinase activation in human bladder cancer cells.

Cordycepin (3'-deoxyadenosine), a bioactive compound of Cordyceps militaris, has many pharmacological activities. The present study reveals novel molecular mechanisms for the anti-tumor effects of cordycepin in two different bladder cancer cell lines, 5637 and T-24 cells. Cordycepin treatment, at a dose of 200 microM (IC(50)) during cell-cycle progression resulted in significant and dose-dependent growth inhibition, which was largely due to G2/M-phase arrest, and resulted in an up-regulation of p21WAF1 expression, independent of the p53 pathway. Moreover, treatment with cordycepin-induced phosphorylation of JNK (c-Jun N-terminal kinases). Blockade of JNK function using SP6001259 (JNK-specific inhibitor) and small interfering RNA (si-JNK1) rescued cordycepin-dependent p21WAF1 expression, inhibited cell growth, and decreased cell cycle proteins. These results suggest that cordycepin could be an effective treatment for bladder cancer.

Korean Red Ginseng (Panax ginseng Meyer) with enriched Rg3 ameliorates chronic intermittent heat stress-induced testicular damage in rats via multifunctional approach.

BACKGROUND: Panax ginseng Meyer, known as Korean Red Ginseng (KRG), is one of the important age-old traditional herbs used in boosting libido and improving male fertility. In this study, the effects of Rg3-enriched KRG extract (KGC04P) on heat stress-induced testicular damage in experimental rats was evaluated. METHODS: Male rats (Sprague-Dawley) were divided into four groups (n = 10): normal control (NC), heat-stressed control (HC), heat-stressed plus KGC04P-100 mg/kg (HK100), and heat-stressed plus KGC04P-200 mg/kg (HK200) groups. Starting 1 week prior to heat stress, animals were administered orally with KGC04P (100 and 200 mg/kg) mixed with a regular pellet diet and continued for 25 weeks. Heat stress was induced to HC, HK100, and HK200 groups by intermittently exposing the animals to high temperatures (32 ± 1°C, 2 h/day). After 6 months, animals were euthanized under general anesthesia with carbon dioxide and evaluated for various parameters in serum and testicular tissue by using Western blotting, biochemical kits, and reverse transcription-polymerase chain reaction. RESULTS: Significant (p < 0.05) alterations in several parameters, such as body/organ weight, sperm kinematics, and lipid metabolism marker levels, in the serum and testis of rats were observed. Further, the expression of testicular antioxidant enzymes, inflammatory cytokines, sex hormonal receptors, and spermatogenesis-related genes were also affected significantly (p < 0.05) in the heat-stressed group. However, KGC04P prevented the heat stress-induced changes in rats significantly (p < 0.05) at both concentrations. CONCLUSION: KGC04P attenuated heat stress-induced testicular damage by a multifunctional approach and can be developed as an excellent therapeutic agent for hyperthermia-mediated male infertility.

Cordycepin, an Active Constituent of Nutrient Powerhouse and Potential Medicinal Mushroom Cordyceps militaris Linn., Ameliorates Age-Related Testicular Dysfunction in Rats.

Age-related male sexual dysfunction covers a wide variety of issues, together with spermatogenic and testicular impairment. In the present work, the effects of cordycepin (COR), an active constituent of a nutrient powerhouse Cordyceps militaris Linn, on senile testicular dysfunction in rats was investigated. The sperm kinematics, antioxidant enzymes, spermatogenic factors, sex hormone receptors, histone deacetylating sirtuin 1 (SIRT1), and autophagy-related mammalian target of rapamycin complex 1 (mTORC1) expression in aged rat testes were evaluated. Sprague Dawley rats were divided into young control (2-month-old; YC), aged control (12-month-old; AC), and aged plus COR-treated groups (5 (COR-5), 10 (COR-10), and 20 (COR-20) mg/kg). The AC group showed reduced sperm kinematics and altered testicular histomorphology compared with the YC group (p < 0.05). However, compared with the AC group, the COR-treated group exhibited improved sperm motility, progressiveness, and average path/straight line velocity (p < 0.05-0.01). Alterations in spermatogenesis-related protein and mRNA expression were significantly ameliorated (p < 0.05) in the COR-20 group compared with the AC group. The altered histone deacetylating SIRT1 and autophagy-related mTORC1 molecular expression in aged rats were restored in the COR-20 group (p < 0.05). In conclusion, the results suggest that COR holds immense nutritional potential and therapeutic value in ameliorating age-related male sexual dysfunctions.

Protective effects of Korean Red Ginseng against sub-acute immobilization stress-induced testicular damage in experimental rats.

BACKGROUND: Excessive stress causes varied physiological and psychological disorders including male reproductive problems. Here, we attempted to investigate the protective effects of Korean Red Ginseng (Panax ginseng Meyer; KRG) against sub-acute immobilization stress-induced testicular damage in experimental rats. METHODS: Male rats (age, 4 wk; weight, 60-70 g) were divided into four groups (n = 8 in each group): normal control group, immobilization control group, immobilization group treated with 100 mg/kg of KRG daily, and immobilization group treated with 200 mg/kg of KRG daily. Normal control and immobilization control groups received vehicle only. KRG (100 mg/kg and 200 mg/kg) was mixed in the standard diet powder and fed daily for 6 mo. Parameters such as organ weight, blood chemistry, sperm kinematic values, and expression levels of testicular-related molecules were measured using commercially available kits, Western blotting, and reverse transcription polymerase chain reaction. RESULTS: Data revealed that KRG restored the altered testis and epididymis weight in immobilization stress-induced rats significantly (p < 0.05). Further, KRG ameliorated the altered blood chemistry and sperm kinematic values when compared with the immobilization control group and attenuated the altered expression levels of spermatogenesis-related proteins (nectin-2, cAMP responsive element binding protein 1, and inhibin-⍺), sex hormone receptors (androgen receptor, luteinizing hormone receptor, and follicle-stimulating hormone receptor), and antioxidant-related enzymes (glutathione S-transferase m5, peroxiredoxin-4, and glutathione peroxidase 4) significantly in the testes of immobilization stress-induced rats. CONCLUSION: KRG protected immobilization stress-induced testicular damage and fertility factors in rats, thereby indicating its potential in the treatment of stress-related male sterility.

Sanguinarine-induced G1-phase arrest of the cell cycle results from increased p27KIP1 expression mediated via activation of the Ras/ERK signaling pathway in vascular smooth muscle cells.

The present study identified a novel mechanism for the effects of sanguinarine in vascular smooth muscle cells (VSMC). Sanguinarine treatment of VSMC resulted in significant growth inhibition as a result of G1-phase cell-cycle arrest mediated by induction of p27KIP1 expression, and resulted in a down-regulation of the expression of cyclins and CDKs in VSMC. Moreover, sanguinarine-induced inhibition of cell growth appeared to be linked to activation of Ras/ERK through p27KIP1-mediated G1-phase cell-cycle arrest. Overall, the unexpected effects of sanguinarine treatment in VSMC provide a theoretical basis for clinical use of therapeutic agents in the treatment of atherosclerosis.

Effect of 2,3,7,8-tetrachlorodibenzo-p-dioxin on testicular spermatogenesis-related panels and serum sex hormone levels in rats.

OBJECTIVES: To determine the detrimental role of tetrachlorodibenzo-p-dioxin (TCDD) in testicular histology, spermatogenesis-related panels and proteome, and serum sex hormone levels. MATERIALS AND METHODS: In all, 40 male rats were divided into equal groups: a normal control (NC) group that received vehicle and saline, and a TCDD-treated (TT) group injected intraperitoneally with TCDD (one dose, 50 microg/kg body weight). The rats were killed 4 weeks after TCDD exposure and testicular weight, histopathology, proteome and variables related to spermatogenesis, and serum sex hormone levels were investigated. RESULTS: TCDD induced a significant decrease in testicular weight, Johnsen's score, seminiferous tubular size, percentage of tubules containing sperm, sperm counts, germ cell counts and Sertoli cell index. In addition, there was a significant decrease in serum testosterone level (P < 0.01) and a remarkable increase in oestradiol (P < 0.01), follicle-stimulating hormone (P < 0.05) and luteinizing hormone (P < 0.05) levels in the TT group. The expression of six testicular proteins including testis-specific heat shock protein (Hsp70), protein disulphide isomerase A3 precursor, 3-phosphoglycerate dehydrogenase, nonmuscle myosin heavy-chain type B-like protein, and superoxide dismutase 1 were significantly up-regulated (P < 0.05-0.01). Interestingly, fertility protein SP22 and phosphatidylethanolamine-binding protein were down-regulated but this was only significant for fertility protein SP22 (P < 0.05). CONCLUSION: TCDD induces marked histological changes in the testis, impairs variables related to spermatogenesis, and increases serum oestradiol levels but decreases testosterone levels. In particular, TCDD disturbs testicular proteome profiles in rats.

Activation of matrix metalloproteinase-9 by TNF-alpha in human urinary bladder cancer HT1376 cells: the role of MAP kinase signaling pathways.

The expression of matrix metalloproteinase-9 (MMP-9) has been implicated in tumor invasion and metastasis. In this study, the factors and signaling pathways that are involved in the regulation of the MMP-9 expression were examined in urinary bladder cancer HT1376 cells. Tumor necrosis factor-alpha (TNF-alpha) stimulated the secretion of MMP-9 in HT1376 cells, as shown by zymography and immunoblot analysis. At the level of transcription, TNF-alpha also stimulated 5'-flanking promoter activity of MMP-9. Transcription factor NF-kappaB, AP-1 and Sp-1 binding sites were identified by a gel shift assay to be cis-elements for TNF-alpha activation of the MMP-9 promoter. TNF-alpha activates multiple signaling pathways in HT1376 cells, including the extracellular signal-regulated kinase (ERK1/2), p38 MAP kinase and JNK pathways. Chemical inhibitors, which specifically inhibit each of these TNF-alpha-activated pathways, were used to examine the signaling pathways involved in TNF-alpha-mediated MMP-9 expression. The ERK1/2 inhibitor, U0126 and the p38 MAP kinase inhibitor, SB203580, significantly down-regulated TNF-alpha-induced MMP-9 expression and promoter activity. The transactivation of TNF-alpha-stimulated NF-kappaB, AP-1 and Sp-1 were inhibited by U0126 and SB203580 treatment. In conclusion, the findings of the present study indicate that TNF-alpha induces MMP-9 expression in HT1376 cells by activating transcription factors, which are involved in the ERK1/2- and p38 MAP kinase-mediated control of MMP-9 regulation, namely, NF-kappaB, AP-1 and Sp-1.

Microeukaryotic diversity in marine environments, an analysis of surface layer sediments from the East Sea.

Molecular techniques, based on clone library of 18S rRNA gene, were employed to ascertain the diversity of microeukaryotic organisms in sediments from the East Sea. A total of 261 clones were recovered from surface sediments. Most of the clone sequences (90%) were affiliated with protists, dominated by Ciliates (18%) and Dinoflagellates (19%) of Alveolates, phototrophic Stramenopiles (11%), and Cercozoa (20%). Many of the clones were related to uncultivated eukaryotes clones retrieved from anoxic environments with several highly divergent 18S rRNA gene sequences. However, no clones were related to cultivated obligate anaerobic protists. Protistan communities between subsurface layers of 1 and 9 cm shared 23% of total phylotypes which comprised 64% of total clones retrieved. Analysis of diversity indices and rarefaction curve showed that the protistan community within the 1 cm layer exhibited higher diversity than the 9 cm layer. Our results imply that diverse protists remain to be uncovered within marine benthic environments.

Korean red ginseng attenuates hypercholesterolemia-enhanced platelet aggregation through suppression of diacylglycerol liberation in high-cholesterol-diet-fed rabbits.

Intake of Korean red ginseng (KRG, ginseng Radix rubra), rich in glycosylated saponins (ginsenosides), has been known to inhibit platelet aggregation in the normocholesterolemic condition. However, it is unclear whether KRG can attenuate hypercholesterolemia-enhanced platelet aggregation. This study examined whether the daily consumption of a KRG-water extract (WE) could prevent the hypercholesterolemia-enhanced platelet aggregation and progression of hypercholesterolemic atherosclerosis. KRG-WE administration (200 mg/kg/day) for 8 weeks potently inhibited the platelet aggregation induced by low doses of agonists (0.5 microg/mL collagen and 0.025 unit/mL thrombin), whereas it weakly reduced the blood-cholesterol levels and formation of atheromatous lesions. In further investigation, KRG-WE significantly suppressed collagen-induced 1,2-diacylglycerol liberation, but had no significant effect on arachidonic acid liberation. Taken together, it can be suggested that the antiplatelet effect of KRG-WE may, at least partly, be due to the inhibition of 1,2-diacylglycerol generation rather than regulation of blood lipid levels. In conclusion, daily consumption of KRG-WE could be a useful alternative measure for the prevention of thrombus and atheroma formation in hypercholesterolemia.

Effects of ginseng on two main sex steroid hormone receptors: estrogen and androgen receptors.

Ginseng has been used in China for at least two millennia and is now popular in over 35 countries. It is one of the world's popular herbs for complementary and alternative medicine and has been shown to have helpful effects on cognition and blood circulation, as well as anti-aging, anti-cancer, and anti-diabetic effects, among many others. The pharmacological activities of ginseng are dependent mainly on ginsenosides. Ginsenosides have a cholesterol-like four trans-ring steroid skeleton with a variety of sugar moieties. Nuclear receptors are one of the most important molecular targets of ginseng, and reports have shown that members of the nuclear receptor superfamily are regulated by a variety of ginsenosides. Here, we review the published literature on the effects of ginseng and its constituents on two main sex steroid hormone receptors: estrogen and androgen receptors. Furthermore, we discuss applications for sex steroid hormone receptor modulation and their therapeutic efficacy.