Maackiain Protects the Kidneys of Type 2 Diabetic Rats via Modulating the Nrf2/HO-1 and TLR4/NF-κB/Caspase-3 Pathways.

BACKGROUND: Type 2 diabetes (T2D) is aglobal health burden that accounts for about 90% of all cases of diabetes. Injury to the kidneys is aserious complication of type 2 diabetes. Maackiain, apterocarpan extracted from roots of Sophora flavescens, has been traditionally used for various disease conditions. However, nothing is known about its possible potential effect on HFD/STZ-T2D-induced nephrotoxicity. METHODS: In this study, T2D rat model is created by high-fat diet (HFD) for 2 weeks with injection of asingle dose of streptozotocin (35mg/kg body weight). T2D rats were orally administered with maackiain (10 and 20mg/kg body weight) for 7 weeks. RESULTS: Maackiain suppressed T2D-induced alterations in metabolic parameters, lipid profile and kidney functionality markers. By administering 10 and 20mg/kg maackiain to T2D rats, it was able to reduce lipid peroxidation while improving antioxidant levels (SOD, CAT, and GSH). Furthermore, the present study demonstrated the molecular mechanisms through which maackiain attenuated T2D-induced oxidative stress (mRNA: Nrf2, Nqo-1, Ho-1, Gclc and Gpx-1; protein: NRF2, NQO-1, HO-1 and NOX-4), inflammation (mRNA: Tlr, Myd88, IκBα, Mcp-1, Tgf-ß, col4, Icam1, Vcam1 and E-selectin; Protein: TLR4, MYD88, NF-κB, IκBα, MCP-1; levels: TNF-α and MCP-1) and apoptosis (mRNA: Bcl-2, Bax, Bad, Apaf-1, Caspase-9 and Caspase-3; protein: Bcl-2, Bax, Caspase-3 and Caspase-9) mediated renal injury. Additionally, significant improvement in kidney architecture was observed after treatment of diabetic rats with 10 or 20mg/kg maackiain. CONCLUSION: Maackiain protects the kidney by decreasing oxidative stress, inflammation, and apoptosis to preserve normal renal function in type 2 diabetes.

Bioavailability, tissue distribution and excretion studies of a potential anti-osteoporotic agent, medicarpin, in female rats using validated LC-MS/MS method.

Medicarpin, one of the active constituents isolated from the extract of Butea monosperma, has been shown to have various pharmacological activities including potent anti-osteoporotic properties. The aim of this study was to investigate the oral pharmacokinetics, tissue distribution and excretion of medicarpin following single oral dose administration in female rats. Oral pharmacokinetics was explored at 5 and 20 mg/kg while tissue distribution, urinary and fecal excretion were studied following 20 mg/kg oral dose. Medicarpin was quantified in rat plasma, urine, feces and tissue samples using a validated LC-MS/MS method following reverse-phase HPLC separation on RP18 column (4.6 mm × 50 mm, 5.0 µm) using methanol and 10 mM ammonium acetate (pH 4.0) as mobile phase in the ratio of 80:20 (v/v) at a flow rate of 0.8 mL/min. The oral bioavailability of medicarpin was found to be low with low systemic levels. The concentration in tissues was significantly higher than plasma. Highest tissue concentrations were found in the liver followed by bone marrow. Urinary and fecal excretion of medicarpin was < 1 %. In conclusion, medicarpin was found to be highly distributed in body tissues and minimally excreted via urine or feces.

Metabolites of Medicarpin and Their Distributions in Rats.

Medicarpin is a bioactive pterocarpan that has been attracting increasing attention in recent years. However, its metabolic fate in vivo is still unknown. To clarify its metabolism and the distribution of its metabolites in rats after oral administration, the HPLC-ESI-IT-TOF-MSn technique was used. A total of 165 new metabolites (13 phase I and 152 phase II metabolites) were tentatively identified, and 104, 29, 38, 41, 74, 28, 24, 15, 42, 8, 10, 3, and 17 metabolites were identified in urine, feces, plasma, the colon, intestine, stomach, liver, spleen, kidney, lung, heart, brain, and thymus, respectively. Metabolic reactions included demethylation, hydrogenation, hydroxylation, glucuronidation, sulfation, methylation, glycosylation, and vitamin C conjugation. M1 (medicarpin glucuronide), M5 (vestitol-1'-O-glucuronide) were distributed to 10 organs, and M1 was the most abundant metabolite in seven organs. Moreover, we found that isomerization of medicarpin must occur in vivo. At least 93 metabolites were regarded as potential new compounds by retrieving information from the Scifinder database. This is the first detailed report on the metabolism of ptercarpans in animals, which will help to deepen the understanding of the metabolism characteristics of medicarpin in vivo and provide a solid basis for further studies on the metabolism of other pterocarpans in animals.

Glyceollins Modulate Tumor Development and Growth in a Mouse Xenograft Model of Human Colon Cancer in a p53-Dependent Manner.

Glyceollins are soybean-derived phytoalexins that induce the nuclear factor (erythroid-derived 2)-like 2 (Nrf2) signaling pathway, which is involved in the detoxification of carcinogens and the removal of reactive oxygen species (ROS). Recent studies, however, have indicated that Nrf2 induction stimulates the development of pre-existing tumors and confers resistance to chemotherapy by elevating drug metabolism and by efficient scavenging of ROS produced by the Warburg effect, which is regulated, in turn, by the p53 tumor suppressor. This study, therefore, aimed at examining whether glyceollins could accelerate tumor growth in the presence of active p53, using a xenograft BALB/c nude mouse model transplanted subcutaneously with p53 wild-type and p53 null HCT116 human colon cancer cells. Glyceollins were orally administered at a dose of either 1 or 4 mg/kg body weight after xenografting HCT116 cells, and tumor growth and volume were monitored for 2 weeks. A high dose of glyceollins resulted in a significant increase in the average volume of p53 wild-type HCT116 xenografts, but not of p53 null HCT116 xenografts. However, a low dose of glyceollins had no effect on the tumor growth regardless of p53 presence. Interestingly, antioxidant enzymes, including heme oxygenase-1 (HO-1) and NAD(P)H: quinone oxidoreductase, were prominently induced by glyceollins in p53 wild-type xenografts, compared with p53 null xenografts. These results suggest that a high dose of glyceollins possibly promotes the growth of p53 wild-type colon cancer through activation of the Nrf2-mediated signaling pathway and, in particular, strong induction of HO-1 expression. Therefore, the consumption of Nrf2 activators, including glyceollins, should be carefully monitored for patients suffering from certain types of cancer and/or receiving chemotherapy.

Protective Effect of Glyceollins in a Mouse Model of Dextran Sulfate Sodium-Induced Colitis.

Glyceollins, which are derived from daidzein in soybean in response to various stimuli or stresses, have been reported to activate antioxidant/detoxifying enzymes in a nuclear factor (erythroid-derived 2)-like 2 (Nrf2)-dependent manner, in addition to exerting anti-inflammatory effects in murine macrophages. As the Nrf2 signaling pathway is known to antagonize nuclear factor (NF)-κB signaling, glyceollins likely have the potential to prevent or treat inflammatory bowel disease. Thus, this study was conducted to examine whether glyceollins could inhibit dextran sulfate sodium (DSS)-induced colitis in a mouse model. Ulcerative colitis (UC) was induced in male BALB/c mice by administering drinking water with 4% DSS for 5 days. Glyceollins (4 or 10 mg/kg of body weight) were orally administered 48 h before and after DSS treatment. We found that glyceollins alleviated histological colon damage and inflammation induced by DSS treatment. More specifically, glyceollins reduced plasma levels of inflammatory cytokines, such as tumor necrosis factor-α and interleukin-6, which were otherwise markedly increased by DSS treatment. Markers of tissue damage, including malondialdehyde and 8-hydroxy-2-guanosine, were significantly increased by DSS treatment; however, this effect was mitigated through concomitant treatment with glyceollins. Furthermore, nuclear accumulation of NF-κB p65 and the expression of inducible nitric oxide synthase were upregulated by glyceollins, consistent with the observed modulation of inflammatory markers. In conclusion, glyceollins have therapeutic potential for UC and merit further clinical study.

The pterocarpanquinone LQB-118 induces apoptosis in acute myeloid leukemia cells of distinct molecular subtypes and targets FoxO3a and FoxM1 transcription factors.

Acute myeloid leukemia (AML) patients' outcome is usually poor, mainly because of drug resistance phenotype. The identification of new drugs able to overcome mechanisms of chemoresistance is essential. The pterocarpanquinone LQB-118 compound has been shown to have a potent cytotoxic activity in myeloid leukemia cell lines and patient cells. Our aim was to investigate if LQB-118 is able to target FoxO3a and FoxM1 signaling pathways while sensitizing AML cell lines. LQB-118 induced apoptosis in both AML cell lines HL60 (M3 FAB subtype) and U937 (M4/M5 FAB subtype). Cell death occurred independently of alterations in cell cycle distribution. In vivo administration revealed that LQB-118 was not cytotoxic to normal bone marrow-derived cells isolated from mice. LQB-118 induced FoxO3a nuclear translocation and upregulation of its direct transcriptional target Bim, in HL60 cells. However, LQB-118 induced FoxO3a nuclear exclusion, followed by Bim downregulation, in U937 cells. Concomitantly, LQB-118 exposure reduced FoxM1 and Survivin expression in U937 cells, but this effect was more subtle in HL60 cells. Taken together, our data suggest that LQB-118 has a selective and potent antitumor activity against AML cells with distinct molecular subtypes, and it involves differential modulation of the signaling pathways associated with FoxO3a and FoxM1 transcription factors.

[Study on protective mechanism of kushenin injection on colonic mucosa of experimental colitis rats].

OBJECTIVE: To study the effect of NOD2 on colitis pathogenesis in experimental rats, and discuss therapeutical effect and mechanism of kushenin injection (OMT) on colitis in experimental rats. METHOD: Fourty Sprague-Dawley (SD) rats were randomly divided into four groups: the normal control group, the model group, the SASP group, and the OMT group, with 10 rats in each group. Except the normal control group, models were established in the remaining three groups with TNBS. The OMT group was injected with kushenin injection, the SASP group was orally administered with mesalazine suspension, the model group and the normal group were orally administered with distilled water for 15 days. Colon lesion score and histological score of experimental rats were observed. Expression of NOD2, NF-kappaB p65 protein in rats colonic mucous was detected by immunohistochemistry. Expression of IL-6 in rat colon mucous was detected by ELISA. RESULT: Compared with normal control group, the expression of NOD2, NF-kappaB p65 and IL-6 in colonic mucosa of the model group were significantly increased (P < 0.01). The SASP group and the OMT group showed lower expressions of NOD2, NF-kappaB p65 and IL-6 in colonic mucosa than the model group (P < 0.01, P < 0.05). CONCLUSION: The over expression of colonic mucosa proteins NOD2 and NF-kappaB p65 and increasing secretion of IL-6 take part in the appearance and development of ulcerative colitis. OMT can attenuate ulcerative colitis and protect colonic mucosa by inhibiting expression of NOD2, NF-kappaB p65 and decreasing IL-6.

Glyceollins, soy isoflavone phytoalexins, improve oral glucose disposal by stimulating glucose uptake.

Soy glyceollins, induced during stress, have been shown to inhibit cancer cell growth in vitro and in vivo. In the present study, we used prediabetic rats to examine the glyceollins effect on blood glucose. During an oral glucose tolerance test (OGTT), the blood glucose excursion was significantly decreased in the rats treated with oral administration of either 30 or 90 mg/kg glyceollins. Plasma analysis demonstrated that glyceollins are absorbed after oral administration, and duration of exposure extends from 20 min to at least 4 h postadministration. Exposure of 3T3-L1 adipocytes to glyceollins significantly increased both insulin-stimulated and basal glucose uptake. Basal glucose uptake was increased 1.5-fold by exposure to 5 µM glyceollin in a dose-response manner. Coincubation with insulin significantly stimulated maximal glucose uptake above basal uptake levels and tended to increase glucose uptake beyond the levels of either stimulus alone. On a molecular level, polymerase chain reaction showed significantly increased levels of glucose transporter GLUT4 mRNA in 3T3-L1 adipocytes, especially when the cells were exposed to 5 µM glyceollins for 3 h in vitro. It correlated with elevated protein levels of GLUT4 detected in the 5 µM glyceollin-treated cells. Thus, the simulative effect of the glyceollins on adipocyte glucose uptake was attributed to up-regulation of glucose transporters. These findings indicate potential benefits of the glyceollins as an intervention in prediabetic conditions as well as a treatment for type 1 and type 2 diabetes by increasing both the insulin-mediated and the basal, insulin-independent, glucose uptake by adipocytes.

Glyceollin-elicited soy protein consumption induces distinct transcriptional effects as compared to standard soy protein.

Glyceollins are stress-induced compounds in soybeans with bioactive properties distinct from parent soy isoflavones. The goals of this study were to evaluate the effects of dietary glyceollin-enriched and standard soy protein isolates and identify candidate target pathways of glyceollins on transcriptional profiles within mammary gland tissue. Thirty female postmenopausal cynomolgus monkeys were randomized to diets containing one of three protein sources for 3 weeks: (1) control casein/lactalbumin (C/L), (2) standard soy protein containing 194 mg/day isoflavones (SOY), and (3) glyceollin-enriched soy protein containing 189 mg/day isoflavones + 134 mg/day glyceollins (GLY). All diets contained a physiologic dose of estradiol (E2) (1 mg/day). All doses are expressed in human equivalents scaled by caloric intake. Relative to the control C/L diet, the GLY diet resulted in greater numbers of differentially regulated genes, which showed minimal overlap with those of SOY. Effects of GLY related primarily to pathways involved in lipid and carbohydrate metabolism, including peroxisome proliferator-activated receptor (PPAR)-γ and AMP-activated protein kinase (AMPK) signaling, adipocytokine expression, triglyceride synthesis, and lipase activity. Notable genes upregulated by the GLY diet included PPAR-γ, adiponectin, leptin, lipin 1, and lipoprotein lipase. The GLY diet also resulted in lower serum total cholesterol, specifically nonhigh-density lipoprotein cholesterol, and increased serum triglycerides as compared to the C/L diet. No effects of GLY or SOY were seen on serum insulin, adipocytokines, or vascular and bone turnover markers. These preliminary findings suggest that glyceollin-enriched soy protein has divergent effects from standard soy with some specificity for adipocyte activity and nutrient metabolism.

Effectiveness of the local or oral delivery of the novel naphthopterocarpanquinone LQB-118 against cutaneous leishmaniasis.

OBJECTIVES: This paper describes the antileishmanial properties of LQB-118, a new compound designed by molecular hybridization, orally active in Leishmania amazonensis-infected BALB/c mice. METHODS: In vitro antileishmanial activity was determined in L. amazonensis-infected macrophages. For in vivo studies, LQB-118 was administered intralesionally (15 µg/kg/day, five times a week), intraperitoneally (4.5 mg/kg/day, five times a week) or orally (4.5 mg/kg/day, five times a week) to L. amazonensis-infected BALB/c mice throughout experiments lasting 85 or 105 days. At the end of the experiments, serum levels of alanine aminotransferase, aspartate aminotransferase and creatinine were measured as toxicological parameters. RESULTS: LQB-118 was active against intracellular amastigotes of L. amazonensis [50% inhibitory concentration (IC(50)) 1.4 µM] and significantly less so against macrophages (IC(50) 18.5 µM). LQB-118 administered intralesionally, intraperitoneally or orally was found to control both lesion and parasite growth in L. amazonensis-infected BALB/c mice, without altering serological markers of toxicity. CONCLUSIONS: These results demonstrate that the molecular hybridization of a naphthoquinone core to pterocarpan yielded a novel antileishmanial compound that was locally and orally active in an experimental cutaneous leishmaniasis model.

Application of kushenin on patients with chronic hepatitis C after renal transplantation.

OBJECTIVE: To evaluate the efficacy of kushenin in treating patients with chronic hepatitis C after renal transplantation. METHODS: Fifty-five patients were randomly assigned by lottery to the treatment group (29 cases) and control group (26 cases). The same immunosuppression therapy was given to all patients in both groups. Patients in the treatment group were treated with kushenin 0.6 g once a day, while those in the control group were treated with conventional liver protective agents such as vitamins. The treatment duration of both groups was 3 months. The incidences of serious hepatitis and acute rejection reaction, serum biochemistry parameters including indicators of liver and kidney functions, hepatic fibrosis index, and serum HCV-RNA were compared between the two groups. RESULTS: (1) The incidence of serious hepatitis in the treatment group and the control group was 3.45% (1/29 cases) and 11.54% (3/26 cases), respectively, which was insignificantly different between the two groups (P=0.335). (2) The incidence of acute rejection in the treatment group was 6.90% (2/29 cases) and that in the control group was 7.69% (2/26 cases), showing insignificant difference (P=0.335). (3) The differences in serum alanine aminotransferase (ALT), direct bilirubin (DBIL), hyaluronic acid (HA), propeptide collagen type III (PC III), laminin (LN), collagen type IV (Col IV) levels between the two groups were insignificant before transplantation (P>0.05), while the above-mentioned parameters in the treatment group were significantly lower than those in the control group after transplantation (P<0.05). The difference in serum creatinine (SCr) and endogenous creatinine clearance rate (CCr) between the two groups was insignificant before and after transplantation (P>0.05). (4) The negative conversion rate of HCV-RNA in the treatment group was 31.03% (9/29 cases), significantly higher than the value of 11.54% (3/26 cases) in the control group after transplantation (P<0.05). (5) The levels of serum ALT and DBIL in patients with HCV-RNA converted to negative were significantly lower than those with still-positive HCV-RNA (P<0.05). CONCLUSIONS: Kushenin has a certain effect on inhibiting the proliferation of HCV, protecting liver cells, and anti-liver fibrosis. On the other hand, it has no obvious influence on renal allograft function. Thus, the drug is clinically safe and effective for use in treating patients with chronic hepatitis C after renal transplantation.

New isoflavones and pterocarpane with hepatoprotective activity from the stems of Erycibe expansa.

The methanolic extract from the stems of Erycibe expansa was found to show a hepatoprotective effect on D-galactosamine-induced cytotoxicity in primary cultured mouse hepatocytes. By bioassay-guided separation, two new prenylisoflavones and a pterocarpane, erycibenins A (1), B (2), and C (3), were isolated from the active fraction (the EtOAc-soluble fraction) together with ten isoflavones (4-13) and seven pterocarpanes (14-20). The stereostructures of the new compounds were determined on the basis of chemical and physicochemical evidence including modified Mosher's method. In addition, the isolated constituents, erycibenin A (1, IC50 = 79 microM), genistein (6, 29 microM), orobol (7, 36 microM), and 5,7,4'-trihydroxy-3'-methoxyisoflavone (8, 55 microM) exhibited inhibitory activity on D-galactosamine-induced cytotoxicity in primary cultured mouse hepatocytes.