Berberine impairs embryonic development in vitro and in vivo through oxidative stress‐mediated apoptotic processes

Berberine, an isoquinoline alkaloid isolated from several traditional Chinese herbal medicines, has been shown to suppress growth and induce apoptosis in some tumor cell lines. However, berberine has also been reported to attenuate H₂O₂‐induced oxidative injury and apoptosis. The basis for these ambiguous effects of berberine—triggering or preventing apoptosis—has not been well characterized to date. In the current investigation, we examined whether berberine exerts cytotoxic effects on mouse embryos at the blastocyst stage and affects subsequent embryonic development in vitro and in vivo. Treatment of blastocysts with berberine (2.5‐10 μM) induced a significant increase in apoptosis and a corresponding decrease in trophectoderm cell number. Moreover, the implantation success rate of blastocysts pretreated with berberine was lower than that of their control counterparts. Pretreatment with berberine was also associated with increased resorption of postimplantation embryos and decreased fetal weight. In an animal model, intravenous injection of berberine (2, 4, or 6 mg/kg body weight/d) for 4 days resulted in apoptosis of blastocyst cells and early embryonic developmental injury. Berberine‐induced injury of mouse blastocysts appeared to be attributable to oxidative stress‐triggered intrinsic apoptotic signaling processes that impaired preimplantation and postimplantation embryonic development. Taken together, our results clearly demonstrate that berberine induces apoptosis and retards early preimplantation and postimplantation development of mouse embryos, both in vitro and in vivo.     

Effects of ochratoxin a on mouse oocyte maturation and fertilization,and apoptosis during fetal development

We previously reported that ochratoxin A (OTA), a mycotoxin found in many foods worldwide, causes nephrotoxicity, hepatotoxicity, and immunotoxicity, and is a risk factor for abnormal embryonic development. More specifically, OTA triggers apoptotic processes in the inner cell mass of mouse blastocysts, decreasing cell viability and embryonic development. In the current study, we investigated the deleterious effects of OTA on mouse oocyte maturation, in vitro fertilization (IVF), and subsequent pre‐ and postimplantation development both in vitro and in vivo. Notably, OTA significantly impaired mouse oocyte maturation, decreased IVF rates, and inhibited subsequent embryonic development in vitro. Preincubation of oocytes with OTA during in vitro maturation increased postimplantation embryonic resorption and decreased mouse fetal weight. In an in vivo animal model, provision of 1–10 μM OTA in the drinking water or intravenous injection of 1 or 2 mg/kg body weight of OTA decreased oocyte maturation and IVF, and had deleterious effects on early embryonic development. Importantly, preincubation of oocytes with a caspase‐3‐specific inhibitor effectively blocked these OTA‐triggered deleterious effects, suggesting that the embryonic injury induced by OTA is mediated via a caspase‐dependent apoptotic mechanism. Furthermore, OTA upregulated the levels of p53 and p21 in blastocyst cells derived from OTA‐pretreated oocytes, indicating that such cells undergo apoptosis via p53‐, p21‐, and caspase‐3‐dependent regulatory mechanisms. This could have deleterious effects on embryonic implantation and fetal survival rates, as seen in our animal models. © 2014 Wiley Periodicals, Inc. Environ Toxicol 31: 724–735, 2016.     

Oxidative stresses‐mediated apoptotic effects of ginsenoside Rb1 on pre‐ and post‐implantation mouse embryos in vitro and in vivo

Ginsenoside Rb1, the major saponin component of ginseng root, has a wide range of therapeutic application. Previous studies have established that ginsenoside Rb1 inhibits the cell cycle and induces apoptosis. However, its side‐effects, particularly those on embryonic development, have not been well characterized to date. In the current study, we examined whether ginsenoside Rb1 exerts a cytotoxic effect on mouse embryos at the blastocyst stage, and affects subsequent embryonic development in vitro and in vivo . Blastocysts treated with 25–100 μg mL^?1 ginsenoside Rb1 exhibited significantly increased apoptosis and a corresponding decrease in total cell number. Notably, the implantation success rate of blastocysts pretreated with ginsenoside Rb1 was lower than that of their control counterparts. Moreover, in vitro treatment with 25–100 μg mL^?1 ginsenoside Rb1 was associated with increased resorption of post‐implantation embryos and decreased fetal weight. In an in vivo model, intravenous injection with ginsenoside Rb1 (1, 3, 5 mg kg^?1 body weight/day) for 4 days resulted in apoptosis of blastocyst stage embryos and early embryonic developmental injury. In addition, ginsenoside Rb1 appeared to induce injury in mouse blastocysts through oxidative stresses‐triggered intrinsic apoptotic signaling processes to impair sequent embryonic development. The collective results strongly indicate that ginsenoside Rb1 induces apoptosis and retards early pre‐ and post‐implantation development of mouse embryos, both in vitro and in vivo . © 2016 Wiley Periodicals, Inc. Environ Toxicol 32: 1990–2003, 2017.     

Hazardous effects of sanguinarine on maturation of mouse oocytes,fertilization, and fetal development through apoptotic processes

Previously, we reported that sanguinarine, a phytoalexin with antimicrobial, anti‐oxidant, anti‐inflammatory and pro‐apoptotic effects, is a risk factor for normal embryonic development that triggers apoptotic processes in the inner cell mass of mouse blastocysts, causing decreased embryonic development and cell viability. In the current study, we investigated the deleterious effects of sanguinarine on mouse oocyte maturation, in vitro fertilization (IVF), and subsequent pre‐ and postimplantation development both in vitro and in vivo. Notably, sanguinarine significantly impaired mouse oocyte maturation, decreased IVF rates, and inhibited subsequent embryonic development in vitro. Preincubation of oocytes with sanguinarine during in vitro maturation induced an increase in postimplantation embryo resorption and a decrease in mouse fetal weight. In an in vivo animal model, 1 to 5 μM sanguinarine, provided in drinking water, caused a decrease in oocyte maturation and IVF, and led to deleterious effects on early embryonic development. Importantly, preincubation of oocytes with a caspase‐3‐specific inhibitor effectively blocked sanguinarine‐triggered deleterious effects, clearly implying that embryonic injury induced by sanguinarine is mediated by a caspase‐dependent apoptotic mechanism. © 2014 Wiley Periodicals, Inc. Environ Toxicol 30: 946–955, 2015.     

Prevention of ochratoxin A‐induced oxidative stress‐mediated apoptotic processes and impairment of embryonic development in mouse blastocysts by liquiritigenin

Ochratoxin A (OTA), a mycotoxin constituent of a range of food commodities, including coffee, wine, beer, grains, and spices, exerts toxicological and pathological effects in vivo, such as nephrotoxicity, hepatotoxicity, and immunotoxicity. In a previous report, we highlighted the potential of OTA to induce apoptosis via reactive oxygen species (ROS) generation in mouse blastocysts that led to impaired preimplantation and postimplantation embryo development in vitro and in vivo. Here, we have shown that liquiritigenin (LQ), a type of flavonoid isolated from Glycyrrhiza radix, effectively protects against OTA‐mediated apoptosis and inhibition of cell proliferation in mouse blastocysts. Preincubation of blastocysts with LQ clearly prevented OTA‐triggered impairment of preimplantation and postimplantation embryonic development and fetal weight loss, both in vitro and in vivo. Detailed investigation of regulatory mechanisms revealed that OTA mediated apoptosis and embryotoxicity through ROS generation, loss of mitochondrial membrane potential (MMP), and activation of caspase‐9 and caspase‐3, which were effectively prevented by LQ. The embryotoxic effects of OTA were further validated in an animal model in vivo. Intravenous injection of dams with OTA (3 mg/kg/day) led to apoptosis of blastocysts, impairment of embryonic development from zygote to blastocyst stage and decrease in day 18 fetal weight. Notably, preinjection of dams with LQ (5 mg/kg/day) effectively prevented OTA‐induced apoptosis and toxic effects on embryo development. Our collective results clearly demonstrate that OTA exposure via injection has the potential to damage preimplantation and postimplantation embryonic development against which LQ has a protective effect.     

Injury effects of ginkgolide B on maturation of mouse oocytes,fertilization, and fetal development in vitro and in vivo

Ginkgolide B (GKB), the major active component of Ginkgo biloba extracts, exerts both stimulatory and inhibitory effects on apoptotic signaling. Previous studies by our group demonstrated that ginkgolide treatment of mouse blastocysts induces apoptosis, decreases cell number, hinders early postimplantation blastocyst development, and increases early-stage blastocyst death. Here, we further investigate the effects of GKB on oocyte maturation, and subsequent pre- and postimplantation development in vitro and in vivo. In our experiments, GKB induced a significant reduction in the rate of oocyte maturation, fertilization, and in vitro embryonic development. Treatment of oocytes with 1–6 μM GKB during in vitro maturation (IVM) led to increased resorption of postimplantation embryos and decreased placental and fetal weights. Data obtained using an in vivo mouse model further disclosed that consumption of drinking water containing 3–6 μM GKB led to decreased oocyte maturation and in vitro fertilization, as well as early embryo developmental injury, specifically, inhibition of development to the blastocyst stage in vivo. To our knowledge, this is the first study to investigate the impact of GKB on maturation of mouse oocytes, fertilization, and sequential embryonic development.     

Resveratrol protects against methylglyoxal‐induced apoptosis and disruption of embryonic development in mouse blastocysts

Methylglyoxal (MG) is a glucose metabolite. Diabetic patients have increased serum levels of MG, and MG is also implicated in tissue injury during embryonic development. In the present work, we show that MG induces apoptosis in the inner cell mass of mouse blastocysts and inhibits cell proliferation. Both effects are suppressed by resveratrol, a grape‐derived phytoalexin with known antioxidant and anti‐inflammatory properties. MG‐treated blastocysts displayed lower levels of implantation (compared to controls) when plated on culture dishes in vitro and a reduced ability to proceed to later stages of embryonic development. Pretreatment with resveratrol prevented MG‐induced disruption of embryonic development, both in vitro and in vivo. Further investigation of these processes revealed that MG directly promotes reactive oxygen species (ROS) generation, loss of mitochondrial membrane potential (MMP), and activation of caspase‐3, whereas resveratrol effectively blocks MG‐induced ROS production and the accompanying apoptotic biochemical changes. Our results collectively imply that MG triggers the mitochondrion‐dependent apoptotic pathway via ROS generation, and the antioxidant activity of resveratrol prevents MG‐induced toxicity. © 2011 Wiley Periodicals, Inc. Environ Toxicol 28: 431–441, 2013.     

Retinoic acid influences the embryoid body formation in mouse embryonic stem cells by induction of caspase and p38 MAPK/JNK‐mediated apoptosis

Although all‐trans retinoic acid (RA), the oxidative metabolite of vitamin A, is essential for normal development, high levels are teratogenic in many species. RA results in immediate effects on the preimplantation embryo and on blastocyst development in vitro and in vivo. To further elucidate the cellular mechanisms of early postimplantation embryo development induced by RA, we present an embryonic cell line, B5, as a candidate system for the investigation of these processes. We used undifferentiated ES cells as the model, which is from the undifferentiated status to differentiated status [embryoid body (EB) formation] mimicking postimplantation embryo development (egg‐cylinder stage of embryo formation) to clarify the cellular mechanism of action of RA in the implanted blastocysts and cell apoptosis following the series of exposures to differing RA concentrations. Using an in vitro model, we identified the impact of RA on undifferentiated embryonic stem (ES) cells, including inhibition of cell proliferation and induction of cell apoptosis. JNK, P‐38 and caspase activation were shown in the nature of RA‐triggered apoptotic signaling in ES cells. The carry‐on influences of RA on the ES cell were shown in the formation of EB from the pretreated ES cells. RA resulted in apparent impact on undifferentiated ES cells in vitro, with increased numbers of apoptotic cells initially and inhibited cell proliferation, which led to decreased size of EB. The process of EB formation (mimicking the early postimplantation embryo development) is regulated by RA‐induced apoptosis through the activation of caspase and P38 MAPK/JNK pathway. © 2011 Wiley Periodicals, Inc. Environ Toxicol, 2013.     

Hazardous impacts of silver nanoparticles on mouse oocyte maturation and fertilization and fetal development through induction of apoptotic processes

Silver nanoparticles (AgNPs) are antibacterial materials widely used in numerous products and medical supplies. Previously, we showed that AgNPs trigger apoptotic processes in mouse blastocysts, leading to a decrease in cell viability and impairment of preimplantation and postimplantation embryonic development in vitro and in vivo. In the present study, we further investigated the hazardous effects of AgNPs on mouse oocyte maturation, in vitro fertilization (IVF), and subsequent preimplantation and postimplantation development in vitro and in vivo. Data from in vitro experiments revealed that AgNPs impair mouse oocyte maturation, decrease IVF rates, and induce injury effects on subsequent embryonic development to a significant extent. In an animal model, intravenous injection of AgNPs (5 mg/kg body weight) led to a significant decrease in mouse oocyte maturation and IVF concomitant with impairment of early embryonic development in vivo. Importantly, pretreatment with N‐acetylcysteine effectively prevented AgNP‐triggered reactive oxygen species (ROS) production and apoptosis, clearly suggesting a critical role of ROS as an upstream initiator or key regulator of AgNP‐induced hazardous effects on oocyte maturation and sequent embryonic development. Furthermore, preincubation of oocytes with Ac‐DEVD‐cho, a caspase‐3‐specific inhibitor, effectively prevented hazardous effects, highlighting the potential involvement of caspase‐dependent apoptotic signaling cascades in AgNP‐mediated events. Expression levels of p53 and p21 of blastocysts were upregulated upon preincubation of mouse oocytes with AgNPs. Our collective results imply that cell apoptosis in mouse blastocysts derived from the AgNP‐pretreated oocytes via intracellular ROS generation, which is further mediated through p53‐, p21‐, and caspase‐3‐dependent regulatory mechanisms.     

Ginsenoside Rg1 protects primary cultured rat hippocampal neurons from cell apoptosis induced by β-amyloid protein

Objectives: Estrogen is known to prominently benefit neuronal syndromes and neurodegenerative diseases. Ginsenoside Rg1, an active ingredient found in a Chinese plant, ginseng root, was previously demonstrated to exert estrogen-like activity. This study was performed to assess the neuroprotective effect of ginsenoside Rg1 against apoptosis induced by β-amyloid protein 25–35 (Aβ_25–35) in primary cultured rat hippocampal neuronal cells as well as in the underlying mechanisms.

Methods: We first measured cell viability and lactate dehydrogenase (LDH) release from primary cultured rat hippocampal neurons. After that, the inhibition effects of ginsenoside Rg1 on neuronal cell apoptosis were evaluated with flow cytometric analysis. Furthermore, western blot analysis was used for detecting the expression of apoptosis-related proteins Bcl-2, Bax, and active caspase 3.

Results: The results show that ginsenoside Rg1 could increase neuronal viability and reduce LDH release; rescue cell apoptosis induced by Aβ_25–35; decrease the expression of caspase 3, increase the ratio of Bcl-2/Bax at the protein levels compared with the cells only treated with Aβ_25–35.

Conclusions: Taken together, our results indicate that the apoptosis induced by Aβ_25–35 could be reversed by ginsenoside Rg1. Furthermore, this neuroprotective effect is probably mediated by up-regulating the ratio of Bcl-2/Bax that activates caspase 3.     

Embryonic toxicity of sanguinarine through apoptotic processes in mouse blastocysts

In this study, we examined the cytotoxic effects of sanguinarine, a phytoalexin with antimicrobial, anti-oxidant, anti-inflammatory and pro-apoptotic effects, on the blastocyst stage of mouse embryos, subsequent embryonic attachment and outgrowth in vitro and in vivo implantation via embryo transfer. Blastocysts treated with 0.5-2 μM sanguinarine exhibited significantly increased apoptosis and a corresponding decrease in total cell number. Notably, the implantation success rates of blastocysts pretreated with sanguinarine were lower than that of their control counterparts. Moreover, in vitro treatment with 0.5-2 μM sanguinarine was associated with increased resorption of post-implantation embryos and decreased fetal weight. Our results collectively indicate that sanguinarine induces apoptosis and retards early post-implantation development in vitro and in vivo. In addition, sanguinarine induces apoptotic injury effects on mouse blastocysts through intrinsic and extrinsic apoptotic signaling processes to impair sequent embryonic development. However, the extent to which sanguinarine exerts teratogenic effects on early human development is not known at present, and further studies are required to establish effective protection strategies against its cytotoxic effects.     

Effect of genistein on mouse blastocyst development in vitro

AIM: To examine the cytotoxic effects of genistein, an isoflavone compound, on early postimplantation embryonic development in vitro. METHODS: Mouse blastocysts were incubated in medium with or without genistein (25 or 50 micromol/L) or daidzein (50 micromol/L) for 24 h. Cell proliferation and growth was investigated by dual differential staining, apoptosis was analyzed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling (TUNEL) assay, and apoptotic or necrotic cells were visualized by Annexin-V and propidium iodide (PI) staining. Implantation and postimplantation development of embryos were measured by in vitro development analysis. RESULTS: TUNEL staining and Annexin-V/PI staining showed that genistein dose-dependently increased apoptosis in mouse blastocysts, while daidzein, another soy isoflavone, had no such effect. The pretreatment of the blastocysts with genistein caused fewer cells than the control group and this effect was primary in the inner cell mass. The genistein-pretreated blastocysts showed normal levels of implantation on culture dishes in vitro, but significantly fewer genistein-pretreated embryos reached the later stages of embryonic development versus the controls, with many of the former embryos dying at relatively early stages of development. In addition, genistein treatment decreased the development of morulas into blastocysts, and dietary genistein was found to induce cell apoptosis and decrease cell proliferation in an animal assay model of embryogenesis. CONCLUSIONS: Our results collectively indicate that genistein treatment of mouse blastocysts induces apoptosis, decreases cell numbers, retards early postimplantation blastocyst development, and increases early-stage blastocyst death in vitro, while dietary genistein appears to negatively affect mouse embryonic development in vivo by inducing cell apoptosis and inhibiting cell proliferation. These novel findings provide important new insights into the effect of genistein on mouse blastocysts.     

Dosage‐related beneficial and deleterious effects of ginsenoside Rb1 on mouse oocyte maturation and fertilization and fetal development

Ginsenoside Rb1 (GRb1), the major saponin component of ginseng root, has a wide range of therapeutic applications for various diseases. Previously, our group showed that GRb1 triggers ROS‐mediated apoptotic cascades in mouse blastocysts, leading to decreased cell viability and impairment of pre‐ and postimplantation embryonic development, both in vitro and in vivo. In this study, we further found that GRb1 exerted dose‐dependent effects on oocyte maturation and sequent development in vitro. Oocytes preincubated with 25 μg/mL GRB1 displayed significantly enhanced maturation and in vitro fertilization (IVF) rates, along with progression of subsequent embryonic development. In contrast, treatment with 50 and 100 μg/mL GRB1 led to impairment of mouse oocyte maturation, decreased IVF rates, and injurious effects on subsequent embryonic development. In vivo, intravenous injection of 1 mg/kg body weight GRb1 significantly promoted mouse oocyte maturation, IVF, and early‐stage embryo development after fertilization while administration of 5 mg/kg body weight GRb1 led to a marked decrease in oocyte maturation and IVF rates concomitant with impairment of early embryonic development in our animal model. In terms of the mechanisms underlying the regulatory effects of GRb1 demonstrated increased intracellular reactive oxygen species (ROS) production and apoptosis in the 100 μg/mL GRb1 treatment group. However, we observed a significant decrease in total intracellular ROS content and inhibition of apoptosis events in the 25 μg/mL GRb1 treatment group, signifying that the intracellular ROS content serves as a key upstream regulator of GRb1 that influences its dose‐dependent beneficial or deleterious effects on oocyte maturation and sequent embryonic development. For further clarification of the mechanisms underlying GRb1‐triggered injurious effects, oocytes were pretreated with Ac‐DEVD‐CHO, a caspase‐3‐specific inhibitor, which effectively blocked injury to oocyte maturation, fertilization, and sequent development. In sum, study findings highlight the potential involvement of p53‐, p21‐, and caspase‐3‐dependent regulatory signaling cascades in GRb1‐mediated apoptotic processes.     

Apoptotic effects on maturation of mouse oocytes,fertilization and fetal development by puerarin

Previously we identified puerarin, an isoflavone compound, as a risk factor for normal embryonic development that triggers apoptotic processes in the inner cell mass of mouse blastocysts, leading to retardation of embryonic development and cell viability. In the current study, we investigated whether puerarin exerts deleterious effects on mouse oocyte maturation, in vitro fertilization (IVF) and subsequent pre- and post-implantation development, both in vitro and in vivo. Notably, puerarin caused significant impairment of these processes in vitro. Pre-incubation of oocytes with puerarin during in vitro maturation led to increased post-implantation embryo resorption and decreased mouse fetal weight. In an in vivo animal model, intravenous injection with or without puerarin (1, 3 and 5?mg/kg body weight/day) for 4 days caused a decrease in oocyte maturation and IVF, and led to deleterious effects on early embryonic development. Importantly, pre-incubation of oocytes with a caspase-3-specific inhibitor effectively blocked puerarin-triggered deleterious effects, clearly implying that embryonic injury induced by puerarin is mediated by a caspase-dependent apoptotic mechanism. These results clearly demonstrate that puerarin has deleterious effects on mouse oocyte maturation, fertilization and subsequent embryonic development in vitro and in vivo.     

Apoptotic effects of dillapiole on maturation of mouse oocytes,fertilization and fetal development

Previously, we reported that dillapiole, a phenylpropanoid with antileishmanial, anti-inflammatory, antifungal and acaricidal activities, is a risk factor for normal embryonic development that triggers apoptotic processes in the inner cell mass of mouse blastocysts, leading to impaired embryonic development and cell viability. In the current study, we investigated the deleterious effects of dillapiole on mouse oocyte maturation, in vitro fertilization (IVF) and subsequent pre- and post-implantation development, both in vitro and in vivo. Notably, dillapiole induced significant impairment of mouse oocyte maturation, decrease in the IVF rate and inhibition of subsequent embryonic development in vitro. Pre-incubation of oocytes with dillapiole during in vitro maturation led to an increase in post-implantation embryo resorption and decrease in mouse fetal weight. In an in vivo animal model, 2.5, 5 or 10?μM dillapiole provided in drinking water caused a decrease in oocyte maturation and IVF, and led to deleterious effects on early embryonic development. Importantly, pre-incubation of oocytes with a caspase-3-specific inhibitor effectively blocked dillapiole-triggered deleterious effects, clearly implying that embryonic injury induced by dillapiole is mediated via a caspase-dependent apoptotic mechanism. To the best of our knowledge, this is the first study to establish the impact of dillapiole on maturation of mouse oocytes, fertilization and sequential embryonic development.     

Ginsenoside Rg1 attenuates the inflammatory response in DSS-induced mice colitis

Ginsenoside Rg1 is a major active constituent of Panax ginseng and possesses anti-inflammatory effects. It has been reported to have therapeutic effects on various diseases. In the present study, we investigated the role of ginsenoside Rg1 in dextran sodium sulfate (DSS)-induced mouse colitis. Our results showed that ginsenoside Rg1 markedly reduces proinflammatory cytokines release upon DSS stimulation of mouse dendritic cells, that ginsenoside Rg1 suppresses IL-1β (Interleukin 1 beta) and TNF-α (Tumor necrosis factor alpha) release via up-regulation of NLRP12 (NACHT, LRR and PYD domains-containing protein 12) expression, and that ginsenoside Rg1 significantly decreases the inflammatory response to DSS-induced mouse colitis, as evidenced by increased body weight, reduced colonic damage scores and disease activity index (DAI), and lowered proinflammatory cytokines levels. These results highlight the potential therapeutic use of ginsenoside Rg1 as an anti-inflammatory agent in the treatment of colitis.     

Induction of cytotoxicity and apoptosis in mouse blastocysts by silver nanoparticles

Silver nanoparticles (nanoAg) are antibacterial materials widely used in various products and medical supplies. In this report, we examined the cytotoxic effects of nanoAg on mouse embryos at the blastocyst stage, subsequent embryonic attachment and outgrowth in vitro, and in vivo implantation by embryo transfer. Blastocysts treated with 50 μM nanoAg exhibited significantly increased apoptosis and a corresponding decrease in total cell number. Importantly, the implantation success rate of blastocysts pretreated with nanoAg was lower than that of their control counterparts. Moreover, in vitro treatment with 50 μM nanoAg was associated with increased resorption of post-implantation embryos and decreased fetal weight. Our results collectively indicate that in vitro exposure to nanoAg induces apoptosis and retards early post-implantation development after transfer to host mice. However, nanoAg-stimulated embryonic cytotoxicity appeared lower than that induced by the Ag⁺ ion. The results collectively show that nanoAg has the potential to induce embryo cytotoxicity. Further studies are required to establish effective protection strategies against the cytotoxic effects of these nanoparticles.     

Induction of apoptosis by ginsenoside Rk1 in SK-MEL-2-human melanoma

Ginsenosides are active compounds isolated from Panax ginseng Meyer. Among these ginsenosides, less polar ginsenosides such as ginsenoside Rg3 and ginsenoside Rh2 have been demonstrated to have tumor inhibitory effects because of their cytotoxicity. In this study, we evaluated the apoptotic effects of ginsenoside Rk1 in SK-MEL-2 human melanoma. Ginsenoside Rk1 isolated from red ginseng is one of the novel ginsenosides that shows strong cytotoxicity compared to ginsenoside Rg3 in dose- and time-dependent manners. The results of DNA fragmentation, 4′,6-diamidino-2-phenylindole staining, and flow cytometric analysis are corroborated that ginsenoside Rk1 induced apoptosis in SK-MEL-2 cells. Western blot analysis revealed up-regulation of Fas, FasL, and Bax protein expression and down-regulation of procaspase-8, procaspase-3, mutant p53 and Bcl-2 protein expression. These findings suggest that ginsenoside Rk1 might be a promising compound to induce apoptosis through both extrinsic and intrinsic pathways in SK-MEL-2 cells.     

Enhanced oral bioavailability and anti‐tumour effect of paclitaxel by 20(s)‐ginsenoside Rg3 in vivo

The purpose of this study was to investigate the effect of paclitaxel in combination with 20(s)‐ginsenoside Rg3 on its anti‐tumour effect in nude mice. In the Caco‐2 transport assay, the apparent permeability from the apical side to the basal side (P_app) (A‐B) and P_app (B‐A) of paclitaxel were measured when co‐incubated with different concentrations of 20(s)‐ginsenoside Rg3. The results indicated that the penetration of paclitaxel through the Caco‐2 monolayer from the apical side to the basal side was facilitated by 20(s)‐ginsenoside Rg3 in a concentration‐dependent manner. Meanwhile, 20(s)‐ginsenoside Rg3 inhibited P‐glycoprotein (P‐gp), and the maximum inhibition was achieved at 80 µ m (p < 0.05). The pharmacokinetic parameters of paclitaxel after oral co‐administration of paclitaxel (40 mg/kg) with various doses of 20(s)‐ginsenoside Rg3 in rats were investigated by an in vivo pharmacokinetic experiment. The results showed that the AUC of paclitaxel co‐administered with 20(s)‐ginsenoside Rg3 was significantly higher (p < 0.001 at 10 mg/kg) compared with the control. The relative bioavailability (RB) % of paclitaxel with 20(s)‐ginsenoside Rg3 was 3.4‐fold (10 mg/kg) higher than that of the control. The effect of paclitaxel orally co‐administered with 20(s)‐ginsenoside Rg3 against human tumour MCF‐7 xenografts in nude mice was also evaluated. Paclitaxel (20 mg/kg) co‐administered with 20(s)‐ginsenoside Rg3 (10 mg/kg) exhibited an effective anti‐tumour activity with the relative tumor growth rate (T/C) values of 39.36% (p <0.05). The results showed that 20(s)‐ginsenoside Rg3 enhanced the oral bioavailability of paclitaxel in rats and improved the anti‐tumour activity in nude mice, indicating that oral co‐administration of paclitaxel with 20(s)‐ginsenoside Rg3 could provide an effective strategy in addition to the established i.v. route. Copyright © 2012 John Wiley & Sons, Ltd.