Kyu-Min Cha

Pectinase-treated Panax ginseng ameliorates hydrogen peroxide-induced oxidative stress in GC-2 sperm cells and modulates testicular gene expression in aged rats

Background To investigate the effect of pectinase-treated Panax ginseng (GINST) in cellular and male subfertility animal models. Methods Hydrogen peroxide (H2O2)-induced mouse spermatocyte GC-2spd cells were used as an in vitro model. Cell viability was measured using MTT assay. For the in vivo study, GINST (200 mg/kg) mixed with a regular pellet diet was administered orally for 4 mo, and the changes in the mRNA and protein expression level of antioxidative and spermatogenic genes in young and aged control rats were compared using real-time reverse transcription polymerase chain reaction and western blotting. Results GINST treatment (50 μg/mL, 100 μg/mL, and 200 μg/mL) significantly (p < 0.05) inhibited the H2O2-induced (200 μM) cytotoxicity in GC-2spd cells. Furthermore, GINST (50 μg/mL and 100 μg/mL) significantly (p < 0.05) ameliorated the H2O2-induced decrease in the expression level of antioxidant enzymes (peroxiredoxin 3 and 4, glutathione S-transferase m5, and glutathione peroxidase 4), spermatogenesis-related protein such as inhibin-α, and specific sex hormone receptors (androgen receptor, luteinizing hormone receptor, and follicle-stimulating hormone receptor) in GC-2spd cells. Similarly, the altered expression level of the above mentioned genes and of spermatogenesis-related nectin-2 and cAMP response element-binding protein in aged rat testes was ameliorated with GINST (200 mg/kg) treatment. Taken together, GINST attenuated H2O2-induced oxidative stress in GC-2 cells and modulated the expression of antioxidant-related genes and of spermatogenic-related proteins and sex hormone receptors in aged rats. Conclusion GINST may be a potential natural agent for the protection against or treatment of oxidative stress-induced male subfertility and aging-induced male subfertility.

Korean red ginseng extract rejuvenates testicular ineffectiveness and sperm maturation process in aged rats by regulating redox proteins and oxidative defense mechanisms

Distortion of intracellular oxidant and antioxidant balances appears to be a common feature that underlies in age-related male sexual impairment. Therefore regulating oxidative defense mechanisms might be an ideal approach in improving male sexual dysfunctions. In the present study, the effect of Korean red ginseng aqueous extract (KRG) on age-induced testicular dysfunction in rats was investigated. KRG (200mg/kg) mixed with regular pellet diet was administered orally for six months and the morphological, spermatogenic and antioxidant enzyme status in testis of aged rats (18months) were evaluated. Data indicated a significant change in morphology and decrease in spermatogenesis-related parameters in aged rats (AC) compared with young rats (YC). Sperm number, germ cell count, Sertoli cell count and Sertoli cell index were significantly (p<0.05) restored in KRG-treated aged rat groups (G-AC). Further the increased lipid peroxidation as measured by malondialdehyde (p<0.05), and altered enzymatic (superoxide dismutase, glutathione peroxidase, glutathione S-transferase, glutathione reductase and catalase) and non-enzymatic (reduced glutathione, ascorbic acid and α-tocopherol) antioxidants (p<0.05) were attenuated by KRG treatment in aged rats to near normal levels as in YC groups. Furthermore, proteomic analysis demonstrated differential expression of selected proteins such as phosphatidylinositol transfer protein, fatty acid binding protein-9, triosephosphate isomerase-1 and aldehyde (aldose) reductase-1in aged rats was significantly (p<0.05) protected by KRG treatment. In conclusion, long-term administration of KRG restored aging-induced testicular ineffectiveness in rats by modulating redox proteins and oxidative defense mechanisms.

Pectinase-treated Panax ginseng protects against chronic intermittent heat stress-induced testicular damage by modulating hormonal and spermatogenesis-related molecular expression in rats

Background Elevated testicular temperature disrupts spermatogenesis and causes infertility. In the present study, the protective effect of enzymatically biotransformed Panax ginseng Meyer by pectinase (GINST) against chronic intermittent heat stress-induced testicular damage in rats was investigated. Methods Male Sprague–Dawley rats (4 wk old, 60–70 g) were divided into four groups: normal control (NC), heat-stress control (HC), heat-stress plus GINST-100 mg/kg (HG100), and heat-stress plus GINST-200 mg/kg (HG200) treatment groups. Each dose of GINST (100 mg/kg and 200 mg/kg) was mixed separately with a regular pellet diet and was administered orally for 24 wk. For inducing heat stress, rats in the NC group were maintained at 25°C, whereas rats in the HC, HG100, and HG200 groups were exposed to 32 ± 1°C for 2 h daily for 6 mo. At week 25, the testes and serum from each animal were analyzed for various parameters. Results Significant (p < 0.01) changes in the sperm kinematic values and blood chemistry panels were observed in the HC group. Furthermore, spermatogenesis-related molecules, sex hormone receptors, and selected antioxidant enzyme expression levels were also altered in the HC group compared to those in the NC group. GINST (HS100 and HS200) administration significantly (p < 0.05) restored these changes when compared with the HC group. For most of the parameters tested, the HG200 group exhibited potent effects compared with those exhibited by the HG100 group. Conclusion GINST may be categorized as an important medicinal herb and a potential therapeutic for the treatment of male subfertility or infertility caused by hyperthermia.

A new validated analytical method for the quality control of red ginseng products

The main active components of Panax ginseng are ginsenosides. Ginsenoside Rb1 and Rg1 are accepted as marker substances for quality control worldwide. The analytical methods currently used to detect these two compounds unfairly penalize steamed and dried (red) P. ginseng preparations, because it has a lower content of those ginsenosides than white ginseng. To manufacture red ginseng products from fresh ginseng, the ginseng roots are exposed to high temperatures for many hours. This heating process converts the naturally occurring ginsenoside Rb1 and Rg1 into artifact ginsenosides such as ginsenoside Rg3, Rg5, Rh1, and Rh2, among others. This study highlights the absurdity of the current analytical practice by investigating the time-dependent changes in the crude saponin and the major natural and artifact ginsenosides contents during simmering. The results lead us to recommend (20S)- and (20R)-ginsenoside Rg3 as new reference materials to complement the current P. ginseng preparation reference materials ginsenoside Rb1 and Rg1. An attempt has also been made to establish validated qualitative and quantitative analytical procedures for these four compounds that meet International Conference of Harmonization (ICH) guidelines for specificity, linearity, range, accuracy, precision, detection limit, quantitation limit, robustness and system suitability. Based on these results, we suggest a validated analytical procedure which conforms to ICH guidelines and equally values the contents of ginsenosides in white and red ginseng preparations.

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.

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.

Korean red ginseng improves testicular ineffectiveness in aging rats by modulating spermatogenesis-related molecules

Abstract Korean red ginseng (Panax ginseng Meyer) is known to rejuvenate testicular effectiveness and the sperm maturation process by regulating redox proteins in aged rats. This study was performed to investigate the effect of Korean red ginseng water extract (KRG‐WE) on the expression level of spermatogenesis‐related key biomolecules and sex hormone receptors as well as enzymes regulating oxidation, histone deacetylation, and growth‐related activities in aged rat testis. KRG‐WE (200 mg/kg) mixed with a regular pellet diet was administered to 12‐month‐old rats for 6 months (KRG‐AC), whereas the young (YC, 2 months) and aged (AC, 12 months) controls received the vehicle only. The results showed that the expression levels of spermatogenesis‐related key biomolecules (inhibin‐&agr;, nectin‐2, and cyclic adenosine monophosphate [cAMP] responsive element binding protein [CREB]‐1), sex hormone receptors (androgen, luteinizing‐ and follicle‐stimulating hormone receptors [AR, LHR, and FSHR, respectively]), and antioxidant enzymes (glutathione S‐transferase mu [GSTm]‐5, glutathione peroxidase [GPx]‐4, peroxiredoxin [PRx]‐3), as well as histone deactylation (silent mating type information regulation 2 homolog 1, SIRT1) and growth‐related (mammalian target of rapamycin complex 1, mTORC1) molecules were significantly altered in the AC group rat testes compared with those of the YC group. However, KRG‐WE treatment of the AC group significantly (p < 0.05) attenuated these molecular changes. From these results, it can be concluded that long‐term administration of KRG‐WE significantly delayed the aging‐induced testicular dysfunction. HighlightsKorean red ginseng (KRG) improves testicular dysfunction in aged rats.KRG attenuated the altered expression levels of sex hormone receptors in aged rats.KRG restored the expression of antioxidant enzyme status in aged rats.KRG attenuates the aging‐induced changes in spermatogenesis‐related molecules.

Canola oil is an excellent vehicle for eliminating pesticide residues in aqueous ginseng extract

Background We previously reported that two-phase partition chromatography between ginseng water extract and soybean oil efficiently eliminated pesticide residues. However, an undesirable odor and an unpalatable taste unique to soybean oil were two major disadvantages of the method. This study was carried out to find an alternative vegetable oil that is cost effective, labor effective, and efficient without leaving an undesirable taste and smell. Methods We employed six vegetable oils that were available at a grocery store. A 1-mL sample of the corresponding oil containing a total of 32 pesticides, representing four categories, was mixed with 10% aqueous ginseng extract (20 mL) and equivalent vegetable oil (7 mL) in Falcon tubes. The final concentration of the pesticides in the mixture (28 mL) was adjusted to approximately 2 ppm. In addition, pesticides for spiking were clustered depending on the analytical equipment (GC/HPLC), detection mode (electron capture detector/nitrogen–phosphorus detector), or retention time used. Samples were harvested and subjected to quantitative analysis of the pesticides. Results Soybean oil demonstrated the highest efficiency in partitioning pesticide residues in the ginseng extract to the oil phase. However, canola oil gave the best result in an organoleptic test due to the lack of undesirable odor and unpalatable taste. Furthermore, the qualitative and quantitative changes of ginsenosides evaluated by TLC and HPLC, respectively, revealed no notable change before or after canola oil treatment. Conclusion We suggest that canola oil is an excellent vehicle with respect to its organoleptic property, cost-effectiveness and efficiency of eliminating pesticide residues in ginseng extract.

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.