Ming-Cheng Lin

Mulberry Leaf Extract Inhibits Vascular Smooth Muscle Cell Migration Involving a Block of Small GTPase and Akt/NF-κB Signals

Mulberry, the fruit of Morus alba, is commonly used in Chinese medicines because of its many pharmacologic effects. Mulberry leaves contain many phenolic antioxidants that can reduce cardiovascular disease. Atherosclerosis involves proliferation and migration of vascular smooth muscle cell (VSMC). Thus, we investigated the mechanisms by which mulberry leaf extract (MLE) might inhibit migration of VSMC. MLE was rich in polyphenols (44.82%), including gallic acid, protocatechuic acid, catechin, gallocatechin gallate, caffeic acid, epicatechin, rutin, and quercetin. MLE could inhibit the migration of A7r5 cells in a dose- and time-dependent manner. MLE also inhibited the activities of matrix metalloproteinases (MMPs) MMP-2 and MMP-9, protein expressions, and phosphorylation of FAK and Akt, and protein expressions of small guanosine triphosphatases (GTPases: c-Raf, Ras, Rac1, Cdc42, and RhoA) in a dose-dependent manner. NF-kappaB expression was also inhibited by MLE. MLE could effectively inhibit the migration of VSMC by blocking small GTPase and Akt/NF-kappaB signals.

Improvement for High Fat Diet-Induced Hepatic Injuries and Oxidative Stress by Flavonoid-Enriched Extract from Nelumbo nucifera Leaf

Nelumbo nucifera Gaertn is widespread and a popular food in central and southern Taiwan. It has also been reported to possess different therapeutic effects, but the effects of N. nucifera leaf on lipid metabolism and liver function remain unclear. In this study, a high fat diet was used to induce hyperlipidemia, hypercholesterolemia, and fatty liver in hamster. The effects of flavonoid-enriched N. nucifera leaf extract supplement and two lipid-lowing drugs, silymarin and simvastatin, on the disorders induced by high fat diet were investigated. The results showed that a 10-week application of a high fat diet to hamsters led to significant increases of body weight, plasma lipid derivatives (triglyceride, total cholesterol, and lipoproteins), lipid peroxidation, and liver damage markers (plasma aspartate aminotransferase and alanine aminotransferase). Interestingly, flavonoid-enriched N. nucifera leaf extract supplement effectively ameliorated the high fat diet-induced lipid metabolic disorders as significantly as silymarin and simvastatin did. Moreover, the flavonoid-enriched supplement alleviated the high fat diet-induced accumulation of lipids in liver, the findings showing distinguishing mechanisms from the effects of silymarin and simvastatin. These results suggested that the flavonoid-enriched N. nucifera leaf extract supplement may significantly improve the high fat diet-induced abnormal blood lipids and liver damage as significantly as the common drugs. Consequently, it is suggested that the flavonoid-enriched N. nucifera leaf extract supplement is beneficial for the improvement of lipid metabolisms and the alleviation of liver damage in high fat diet treatment.

Polyphenol-rich extract from mulberry leaf inhibits vascular smooth muscle cell proliferation involving upregulation of p53 and inhibition of cyclin-dependent kinase.

This study was carried out to investigate the impact of polyphenol-rich extract from mulberry leaf on the proliferation of vascular smooth muscle cell (VSMC) and verify its mechanism in vitro. VSMC proliferation is an important pathophysiological process in the development of atherosclerosis, which is the major cause of coronary artery disease (CAD). Polyphenol-rich foods, such as mulberry leaf, have been reported to reduce the risk of CAD. The effect of mulberry leaf extract (MLE) on cell growth was measured by a growth curve assay, on distribution of cells in the cell cycle by flow cytometry, and on cyclin-dependent kinase (CDK) activity and cell-cycle regulatory proteins by Western blot, immunoblotting, and immunoprecipitation analyses. The results showed that MLE induced phosphorylation of p53, promoted expression of p21 and p27, decreased CDK2/4 activity, inhibited phosphorylation of Rb, and thereby blocked the G1 to S transition in the cell cycle.

Mulberry Leaf Extract Inhibits the Development of Atherosclerosis in Cholesterol-Fed Rabbits and in Cultured Aortic Vascular Smooth Muscle Cells

This study used high-cholesterol-fed New Zealand white rabbits and aortic vascular smooth muscle cells (VSMCs) to investigate the impact of mulberry leaf extract (MLE) on the development of atherosclerosis. The results show that the major components of MLE are polyphenols, flavonoids, carbohydrates, proteins, and lipids, and the major contituents of mulberry leaf polyphenol extract (MLPE) are polyphenols and flavonoids. In addition to improvement of liver function, the atheroma burden and levels of serum cholesterol, triglycerides, and low-density lipoprotein (LDL) are also significantly reduced after MLE treatment. MLE and MLPE improved endothelial function, inhibited proliferation and migration of aortic VSMCs, and reduced atheromas in the vascular wall. In conclusion, this study demonstrates that, in addition to exerting hypolipidemic effects, MLE and MLPE can effectively inhibit proliferation and migration of aortic VSMCs, improve vascular endothelial function, and reduce atheroma burden, thereby preventing atherosclerosis.

Mulberry water extracts (MWEs) ameliorated carbon tetrachloride-induced liver damages in rat.

Mulberry extracts are antidiabetic and antihyperlipidemic, as well as preventive of cardiovascular disease. The current study investigates the protective mechanisms of mulberry water extracts (MWEs) in carbon tetrachloride (CCl(4))-induced hepatic injury. Oral administration of MWEs significantly reduced the lipid peroxidation triggered by CCl(4), as shown by the reduced production of thiobarituric acid-reactive substance (TBARS). The levels of serum aspartate aminotransferase (AST), alanine aminotransferase (ALT), and alkaline phosphatase (ALP) were also reduced via cotreatment with MWEs compared with CCl(4) treatment alone. Cotreatment with MWE evidently reduced CCl(4)-induced liver weight and inhibited lipid deposition and fibrogenesis. In a similar manner, cotreatment with silymarin, a well-known liver protective agent, also reversed the CCl(4)-induced effects, such as reduced TBARS formation, decreased serum AST, ALT, and ALP levels, blocked lipid accumulation, and liver fibrosis. Furthermore, MWEs attenuated the proinflammatory genes such as cyclooxygenase 2, nuclear factor kappa B, and inducible nitric oxide synthase expression. The current findings suggest that MWEs such as silymarin exhibit protective and curative effects against CCl(4)-induced liver damage and fibrosis via decreased lipid peroxidation and inhibited proinflammatory gene expression.

Gallic Acid Attenuates Oleic Acid-induced Proliferation of Vascular Smooth Muscle Cell Through Regulation of AMPK-eNOS-FAS Signaling

Vascular smooth muscle cell (VSMC) proliferation plays a central role in the pathogenesis of obesity-related atherosclerosis. The molecular mechanism of GA on oleic acid (OA)-induced proliferation of vascular smooth muscle cell is evaluated. Cells were treated with OA (150 μM), or co-treated with OA and GA (10-30 μM) for 48 h, MTT assay was performed for proliferation. Using flow cytometry analysis, the GA-treated cells caused an increase in G2/M phase. A decrease in cyclin B1 and cyclin-dependent kinase 1 (cdc2) and increase in kip/p27 and cip1/p21 were found by western blotting. Additional mechanistic studies showed that GA induced the activation of AMP-activated protein kinase (AMPK) and eNOS and the inhibition of fatty acid synthase (FAS) after stimulation with OA. Furthermore, the addition of compound C, a specific inhibitor of AMPK, reduced the activation of GA-mediated eNOS and NO production and increased the proliferation of cells. Inhibition of NOS by L-NAME had no further effect on VSMC proliferation. The present results indicate that GA was an effected and anti-atherogenic agent in VSMC. It attenuates cell cycle progression via AMPKmediated eNOS activation, which results in the production of NO and prevents atherosclerosis.

Mulberry 1-deoxynojirimycin pleiotropically inhibits glucose-stimulated vascular smooth muscle cell migration by activation of AMPK/RhoB and down-regulation of FAK.

Mulberry 1-deoxynojirimycin (DNJ), an inhibitor of α-glucosidase, has been reported to help prevent diabetes mellitus and suppress lipid accumulation. The aim of this study was to determine whether mulberry DNJ has pleiotropic effects on the development of atherosclerosis. The mechanisms by which mulberry DNJ might inhibit migration of A7r5 vascular smooth muscle cells (VSMCs) under hyperglycemic conditions mimicking diabetes were investigated. The antimigratory effects of DNJ on VSMCs were assessed by Western blot analysis of migration-related proteins and by electric cell-substrate impedance sensing (ECIS) and visualization of F-actin cytoskeleton. Two pathways of DNJ-mediated inhibition of VSMC migration were identified. The first involved AMPK activation to inhibit fatty acid synthase (FASN) and Akt activity and then RhoB activation to inhibit nuclear factor-κB (NF-κB) and matrix metalloproteinase-2 (MMP) activity. The second involved inhibition of focal adhesion kinase (FAK), Ras, and RhoA activity leading to inhibition of F-actin activity.

Mulberry water extracts inhibit rabbit atherosclerosis through stimulation of vascular smooth muscle cell apoptosis via activating p53 and regulating both intrinsic and extrinsic pathways.

Previous studies have shown that mulberry water extracts (MWEs), which contain polyphenolic compounds, have an antiatherosclerotic effect in rabbits. Apoptosis of vascular smooth muscle cells (VSMCs) is the key determinant of the number of VSMCs in remodeling. To improve the recovery from atherosclerosis pathology, it would be ideal to induce regression of atherosclerotic plaques and apoptosis of VSMCs. In this study, we treated high-cholesterol-diet-fed (HCD-fed) rabbits with MWEs, and we found that the MWEs effectively inhibited HCD-fed-induced intimal hyperplasia of vessel walls. We also found that MWEs initially activate JNK/p38 and p53, which in turn activate both Fas-ligand and mitochondria pathways, thereby causing mitochondria translocation of Bax and the reduction of Bcl-2 that trigger the cleavage of procaspases, finally resulting in apoptosis of VSMCs. In addition, 2.5-5.0 g/day of MWEs for humans may be enough to prevent atherosclerosis.

Radiolabeled Nucleosides for Predicting and Monitoring the Cancer Therapeutic Efficacy of Chemodrugs

Uncontrolled cell proliferation is an important hallmark of cancer. Cancer treatment with cytostatic chemodrugs usually results in insignificant changes in tumor size, and thus limits the applications of anatomical imaging modalities for determining the therapeutic efficacy. Positron emission tomography (PET) imaging with cell proliferation probes to assess the clinical outcome during or soon after treatment is becoming acceptable. At present, monitoring DNA synthetic pathways with radiolabeled nucleoside probes that are essential for cell proliferation has been considered a more specific approach to predict tumor response. Among the four nucleosides, thymidine analogues, such as (18)F-FLT, have undergone years of development for clinical practice, while cytidine, adenosine and guanosine analogues receive less attention. Recently, several literatures have demonstrated that PET imaging with radiolabeled cytidine and adenosine analogues may have potential to evaluate immune response after chemotherapy, and may enable the prognosis forecast. In this review, we summarize the results of recent preclinical and clinical studies regarding using radiolabeled nucleoside analogues for predicting and monitoring tumor response in cancer treatment. The preparation protocols of these nucleoside scintigraphic probes are also described.

Mulberry Water Extracts Inhibit Atherosclerosis through Suppression of the Integrin-β3/Focal Adhesion Kinase Complex and Downregulation of Nuclear Factor κB Signaling in Vivo and in Vitro

Previous studies have shown that mulberry water extracts (MWEs), which contain polyphenolic compounds, have an antiatherosclerotic effect in vivo and in vitro through stimulating apoptosis of vascular smooth muscle cells (VSMCs). Histological analysis was performed on atherosclerotic lesions from high-cholesterol diet (HCD)-fed rabbits after treatment with 0.5-1% MWEs for 10 weeks. Immunohistochemistry showed that the expressions of SMA, Ras, and matrix metalloproteinase-2 in the VSMCs were dose-dependently inhibited after MWE treatment. The antimigratory effects of MWEs on A7r5 VSMCs were assessed by western blot analysis of migration-related proteins, visualization of F-actin cytoskeleton, and reverse transcription polymerase chain reaction. The results showed that MWEs inhibited VSMC migration through reducing interactions of the integrin-β3/focal adhesion kinase complex, alterations of the cytoskeleton, and downregulation of glycogen synthase kinase 3β/nuclear factor κB signaling. Taken together, MWEs inhibited HCD-induced rabbit atherogenesis through blocking VSMC migration via reducing interactions of integrin-β3 and focal adhesion kinase and downregulating migration-related proteins.