A novel glycoglycerolipid from Holotrichia diomphalia Bates: Structure characteristics and protective effect against DNA damage.

A lipidated polysaccharide, HDPS-2II, was isolated from the dried larva of Holotrichia diomphalia, which is used in traditional Chinese medicine. The molecular weight of HDPS-2II was 5.9 kDa, which contained a polysaccharide backbone of →4)-ß-Manp-(1 â†’ 4,6)-ß-Manp-(1 â†’ [6)-α-Glcp-(1]n â†’ 6)-α-Glcp→ with the side chain α-Glcp-(6 â†’ 1)-α-Glcp-(6 â†’ linked to the C-4 of ß-1,4,6-Manp and four types of lipid chains including 4-(4-methyl-2-(methylamino)pentanamido)pentanoic acid, 5-(3-(tert-butyl)phenoxy)hexan-2-ol, N-(3-methyl-5-oxopentan-2-yl)palmitamide, and N-(5-amino-3-methyl-5-oxopentan-2-yl)stearamide. The lipid chains were linked to C-1 of terminal α-1,6-Glcp in carbohydrate chain through diacyl-glycerol. HDPS-2II exhibited DNA protective effects and antioxidative activity on H2O2- or adriamycin (ADM)-induced Chinese hamster lung cells. Furthermore, HDPS-2II significantly ameliorated chromosome aberrations and the accumulation of reactive oxygen species (ROS), reduced γ-H2AX signaling and the expressions of NADPH oxidase (NOX)2, NOX4, P22phox, and P47phox in ADM-induced cardiomyocytes. Mechanistically, HDPS-2II suppressed ADM-induced up-regulation of NOX2 and NOX4 in cardiomyocytes, but not in NOX2 or NOX4 knocked-down cardiomyocytes, indicating that HDPS-2II could relieve intracellular DNA damage by regulating NOX2/NOX4 signaling. These findings demonstrate that HDPS-2II is a new potential DNA protective agent.

Piperlonguminine attenuates renal fibrosis by inhibiting TRPC6.

ETHNOPHARMACOLOGICAL RELEVANCE: Liuwei Dihuang (LWDH) is a classic prescription that has been used to the treatment of "Kidney-Yin" deficiency syndrome for more than 1000 years in China. Recent studies have confirmed that LWDH can prevent the progression of renal fibrosis. Numerous studies have demonstrated the critical role that TRPC6 plays in the development of renal fibrosis. Due to the complex composition of LWDH and its remarkable therapeutic effect on renal fibrosis, it is possible to discover new active ingredients targeting TRPC6 for the treatment of renal fibrosis. AIM OF STUDY: This study aimed to identify selective TRPC6 inhibitors from LWDH and evaluate their therapeutical effects on renal fibrosis. MATERIALS AND METHODS: Computer-aided drug design was used to screen the biologically active ingredients of LWDH, and their affinities to human TRPC6 protein were detected by microcalorimetry. TRPC6, TRPC3, and TRPC7 over-expressed HEK293 cells were constructed, and the selective activities of the compounds on TRPC6 were determined by measuring [Ca2+]i in these cells. To establish an in vitro model of renal fibrosis, human renal proximal tubular epithelial HK-2 cells were stimulated with TGF-ß1. The therapeutic effects of LWDH compounds on renal fibrosis were then tested by detecting the related proteins. TRPC6 was knocked-down in HK-2 cells to investigate the effects of LWDH active ingredients on TRPC6. Finally, a unilateral ureteral obstruction model of renal fibrosis was established to test the therapeutic effect. RESULTS: From hundreds of LWDH ingredients, 64 active components with oral bioavailability ≥30% and drug-likeness index ≥0.18 were acquired. A total of 10 active components were obtained by molecular docking with TRPC6 protein. Among them, 4 components had an affinity with TRPC6. Piperlonguminine (PLG) had the most potent affinity with TRPC6 and blocking effect on TRPC6-mediated Ca2+ entry. A 100 µM of PLG showed no detectable inhibition on TRPC1, TRPC3, TRPC4, TRPC5, or TRPC7-mediated Ca2+ influx into cells. In vitro results indicated that PLG concentration-dependently inhibited the abnormally high expression of α-smooth muscle actin (α-SMA), collagen I, vimentin, and TRPC6 in TGF-ß1-induced HK-2 cells. Consistently, PLG also could not further inhibit TGF-ß1-induced expressions of these protein biomarkers in TRPC6 knocked-down HK-2 cells. In vivo, PLG dose-dependently reduced urinary protein, serum creatinine, and blood urea nitrogen levels in renal fibrosis mice and markedly alleviated fibrosis and the expressions of α-SMA, collagen I, vimentin, and TRPC6 in kidney tissues. CONCLUSION: Our results showed that PLG had anti-renal fibrosis effects by selectively inhibiting TRPC6. PLG might be a promising therapeutic agent for the treatment of renal fibrosis.

The structures of two polysaccharides from Angelica sinensis and their effects on hepatic insulin resistance through blocking RAGE.

This study found two novel homogeneous polysaccharides from Angelica sinensis, APS-1I and APS-2II, binding to RAGE with a dissociation constant of 2.02 ± 0.2 and 85.92 ± 0.2 µM, respectively. APS-1I is a 17.0 kDa heteropolysaccharide, whose backbone is composed of α-1,6-Glcp, α-1,3,6-Glcp, α-1,2-Glcp, α-1,4-Galp, and α-1,3-Rhap, and whose two branches contain α-1,3,5-Araf, α-1,3-Araf, α-1,4-Galp, ß-1,3-Galp, and ß-1,4-Glcp. APS-2II is a 10.0 kDa linear glucan, that contains α-1,6-Glcp, α-1,3-Glcp, α-1,2-Glcp, and α-T-Glcp. In vitro, APS-1I demonstrated better promotion on glucose absorption and stronger repression on p-IRS-1 (Ser307), p-IRS-2 (Ser731), p-JNK, and p-P38 than APS-2II in insulin resistance (IR)-HepG2 cells. Furthermore, APS-1I treatment couldn't further decrease the inhibition on the phosphorylation of JNK and P38 produced by RAGE siRNA in IR-HepG2 cells. In vivo, APS-1I markedly improved IR and reversed the livers RAGE-JNK/p38-IRS signaling in high-fat-diet and streptozotocin-induced diabetic rats, suggesting that APS-1I could be a potential agent for improving IR in type 2 diabetes.

Structural characteristics of a hypoglycemic polysaccharide from Fructus Corni.

PFC-3 is a homogeneous polysaccharide extracted from the dried pulps of Fructus Corni with a molecular weight of 40.3 kDa. The crude polysaccharide was obtained and further purified by DEAE-Sephadex A-25 and Sephadex G-100 columns to investigate its structure and glycemic effect. The monosaccharides in the PFC-3, determined by high-performance liquid chromatography, consisted of glucose (Glc), xylose (Xyl), and galactose (Gal) with a mass molar ratio of 2.35:12.49:1.00. The methylation analysis combined with 1D (1H and 13C), and 2D NMR (1H-1H COSY, HSQC, and HMBC) further demonstrated that PFC-3 was mainly composed of 1,3-α-D-Xylp, 1,6-α-D-Galp, 1,2-α-D-Glcp, and T-α-D-Galp, and contained a backbone fragment of →6)-α-D-Galp-(1 â†’ 2)-α-D-Glcp-(1 â†’ 3)-α-D-Xylp-(1 â†’ . The hypoglycemic effect of PFC-3 in vitro was evaluated by glucose uptake and consumption assays, and the results showed that PFC-3 concentration-dependently enhanced glucose uptake and significantly improved glucose consumption in insulin-resistant HepG2 cells. Furthermore, PFC-3 significantly reduced fasting blood glucose level, glycosylated hemoglobin level, amylase activity, ameliorate lipid metabolism, and hepatic lesions in streptozotocin-induced diabetic rats. Our research provided insights into the hypoglycemic activities of PFC-3.

CAPE-pNO2 attenuates diabetic cardiomyopathy through the NOX4/NF-κB pathway in STZ-induced diabetic mice.

Diabetic cardiomyopathy (DCM) is one of the most severe cardiovascular complications in diabetes. Caffeic acid para-nitro phenethyl ester (CAPE-pNO2) could ameliorate diabetic nephropathy in the diabetic mice in our previous study. This paper was aimed to investigate the effect of CAPE-pNO2 on DCM and its potential mechanism. The DCM mice were established by intraperitoneal injection with streptozotocin (STZ, 50 mg/kg) for 5 days. When the fasting blood glucose level remains above 11.1 mmol/L, treated the mice with CAPE and CAPE-pNO2 for 8 weeks, then the mice were executed, and the samples of blood and heart tissue were collected for the subsequent experiments. The results showed that CAPE-pNO2 can alleviate CK, LDH, TC and TG levels, as well as depress the activity of ROS by down-regulating the expression of NOX4 and improving SOD activity in the serum of STZ-induced DCM mice. Meanwhile, it can also reduce the content of MDA and inhibit lipid accumulation. Besides, CAPE-pNO2 could repress the expression of TNF-α, IL-1ß and IL-6 IL - 6 via the NOX4/NF-κB pathway to improve the development of inflammation. Furthermore, it can suppress the expression of collagen and fibronectin to inhibit myocardial fibrosis through the TGF-ß1/Smad pathway, and inhibit ECM deposition by regulating TGF-ß1 directly, as shown in cardiac tissue section. Importantly, the above results showed that CAPE-pNO2 had better effects of reversing pathological changes than CAPE in a significant difference of p < 0.05. In brief, CAPE-pNO2 can prevent the heart injury of DCM mice via the NOX4/NF-κB pathway, and shows the improvement effects of anti-fibrosis, anti-oxidative and anti-inflammatory.

CAPE-pNO2 ameliorated diabetic nephropathy through regulating the Akt/NF-κB/ iNOS pathway in STZ-induced diabetic mice.

Diabetic nephropathy (DN) is one of the most severe complications of diabetes mellitus. This study aimed to determine the effects and potential mechanism of caffeic acid para-nitro phenethyl ester (CAPE-pNO2), a derivative of caffeic acid phenethyl ester (CAPE), on DN; In vivo, intraperitoneal injections of streptozotocin (STZ) were used to induce diabetes in mice; then, the mice were intraperitoneally injected daily with CAPE or CAPE-pNO2 for 8 weeks. The mice were sacrificed, and blood samples and kidney tissues were collected to measure biological indexes. The results showed that CAPE and CAPE-pNO2 could lower serum creatinine, blood urea nitrogen, 24-h albumin excretion, malondialdehyde and myeloperoxidase levels and increase superoxide dismutase activity in diabetic mice. According to HE, PAS and Masson staining, these two compounds ameliorated structural changes and fibrosis in the kidneys. In addition, the immunohistochemical and western blot results showed that CAPE and CAPE-pNO2 inhibited inflammation through the Akt/NF-κB pathway and prevented renal fibrosis through the TGF-ß/Smad pathway. In vitro, CAPE and CAPE-pNO2 inhibited glomerular mesangial cell (GMC) proliferation, arrested cell cycle progression and suppressed ROS generation. These compounds also inhibited ECM accumulation via regulating the TGF-ß1, which was a similar effect to that of the NF-κB inhibitor PDTC. More importantly, CAPE and CAPE-pNO2 could up-regulate nitric oxide synthase expression in STZ-induced diabetic mice and HG-induced GMCs. CAPE-pNO2 had stronger effects than CAPE both in vivo and in vitro. These data suggest that CAPE-pNO2 ameliorated DN by suppressing oxidative stress, inflammation, and fibrosis via the Akt/NF-κB/ iNOS pathway.