Yung Hsin Cheng

The generation of induced pluripotent stem cells for macular degeneration as a drug screening platform: identification of curcumin as a protective agent for retinal pigment epithelial cells against oxidative stress

Age-related macular degeneration (AMD) is one retinal aging process that may lead to irreversible vision loss in the elderly. Its pathogenesis remains unclear, but oxidative stress inducing retinal pigment epithelial (RPE) cells damage is perhaps responsible for the aging sequence of retina and may play an important role in macular degeneration. In this study, we have reprogrammed T cells from patients with dry type AMD into induced pluripotent stem cells (iPSCs) via integration-free episomal vectors and differentiated them into RPE cells that were used as an expandable platform for investigating pathogenesis of the AMD and in-vitro drug screening. These patient-derived RPEs with the AMD-associated background (AMD-RPEs) exhibited reduced antioxidant ability, compared with normal RPE cells. Among several screened candidate drugs, curcumin caused most significant reduction of ROS in AMD-RPEs. Pre-treatment of curcumin protected these AMD-RPEs from H2O2-induced cell death and also increased the cytoprotective effect against the oxidative stress of H2O2 through the reduction of ROS levels. In addition, curcumin with its versatile activities modulated the expression of many oxidative stress-regulating genes such as PDGF, VEGF, IGFBP-2, HO1, SOD2, and GPX1. Our findings indicated that the RPE cells derived from AMD patients have decreased antioxidative defense, making RPE cells more susceptible to oxidative damage and thereby leading to AMD formation. Curcumin represented an ideal drug that can effectively restore the neuronal functions in AMD patient-derived RPE cells, rendering this drug an effective option for macular degeneration therapy and an agent against aging-associated oxidative stress.

Sustained delivery of latanoprost by thermosensitive chitosan-gelatin-based hydrogel for controlling ocular hypertension

Glaucoma is an irreversible ocular disease that may lead to progressive visual field loss and eventually to blindness with inadequately controlled intraocular pressure (IOP). Latanoprost is one of the most potent ocular hypotensive compounds, the current first-line therapy in glaucoma. However, the daily instillation required for efficacy and undesirable side-effects are major causes of treatment adherence failure and persistence in glaucoma therapy. In the present study, we developed an injectable thermosensitive chitosan/gelatin/glycerol phosphate (C/G/GP) hydrogel as a sustained-release system of latanoprost for glaucoma treatment. The latanoprost-loaded C/G/GP hydrogel can gel within 1min at 37°C. The results show a sustained release of latanoprost from C/G/GP hydrogel in vitro and in vivo. The latanoprost-loaded C/G/GP hydrogel showed a good in vitro and in vivo biocompatibility. A rabbit model of glaucoma was established by intravitreal injection of triamcinolone acetonide. After a single subconjunctival injection of latanoprost-loaded C/G/GP hydrogel, IOP was significantly decreased within 8days and then remained at a normal level. The results of the study suggest that latanoprost-loaded C/G/GP hydrogel may have a potential application in glaucoma therapy.

Thermosensitive chitosan-based hydrogel as a topical ocular drug delivery system of latanoprost for glaucoma treatment.

Ocular hypertension is a major risk factor for the development and progression of glaucoma. Frequent and long-term application of latanoprost often causes undesirable local side effects, which are a major cause of therapeutic failure due to loss of persistence in using this glaucoma medical therapy. In the present study, we developed a thermosensitive chitosan-based hydrogel as a topical eye drop formulation for the sustained release of latanoprost to control ocular hypertension. The developed formulation without preservatives may improve compliance and possibly even efficacy. The results of this study support its biocompatibility and sustained-release profile both in vitro and in vivo. After topical application of latanoprost-loaded hydrogel, triamcinolone acetonide-induced elevated intraocular pressure was significantly decreased within 7 days and remained at a normal level for the following 21 days in rabbit eyes. This newly developed chitosan-based hydrogel may provide a non-invasive alternative to traditional anti-glaucoma eye drops for glaucoma treatment.

Aspirin attenuates vinorelbine-induced endothelial inflammation via modulating SIRT1/AMPK axis

Vinorelbine (VNR), a semisynthetic vinca alkaloid acquired from vinblastine, is frequently used as the candidate for intervention of solid tumors. Nevertheless, VNR-caused endothelial injuries may lead a mitigative effect of clinical treatment efficiency. A growing body of evidence reveals that aspirin is a potent antioxidant and anti-inflammation drug. We investigated whether aspirin attenuate VNR-induced endothelial dysfunction. Human endothelial cells (EA.hy 926) were treated with VNR to cause endothelial inflammation. Western blotting, ROS assay, ELISA were used to confirm the anti-inflammatory effect of aspirin. We confirmed that VNR suppresses SIRT1 expression, reduced LKB1 and AMPK phosphorylation as well as enriched PKC activation in treated endothelial cells. Furthermore, the membrane translocation assay displayed that the levels of NADPH oxidase subunits p47phox and Rac-1 in membrane fractions of endothelial cells were higher in cells that had been treated with VNR for than in untreated cells. We corroborated that treatment of Aspirin significantly diminishes VNR-repressed SIRT1, LKB1 and AMPK phosphorylation and VNR-promoted NADPH oxidase activation, however, those findings were vanished by SIRT1 and AMPK siRNAs. Our data also shown that Aspirin represses VNR-activated TGF-beta-activated kinase-1 (TAK1) activation, inhibited the interaction of TAK1/TAK-binding protein1 (TAB1), suppressed NF-kappa B activation and pro-inflammatory cytokine secretion. We demonstrated a novel connection between VNR-caused oxidative damages and endothelial dysfunction, and provide further insight into the protective effects of aspirin in VNR-caused endothelial dysfunction.

Ginkgo biloba extract inhibits oxidized low-density lipoprotein (oxLDL)-induced matrix metalloproteinase activation by the modulation of the lectin-like oxLDL receptor 1-regulated signaling pathway in human umbilical vein endothelial cells

BACKGROUNDnThe overexpression of matrix metalloproteinases (MMPs) induced by oxidized low-density lipoprotein (oxLDL) has been found in atherosclerotic lesions. Previous reports have identified that oxLDL, via the upregulation of lectin-like ox-LDL receptor 1 (LOX-1), modulates the expression of MMPs in endothelial cells. Ginkgo biloba extract (GbE), from Ginkgo biloba leaves, has often been considered as a therapeutic compound for cardiovascular and neurologic diseases. However, further investigation is needed to ascertain the probable molecular mechanisms underlying the antiatherogenic effects of GbE. The aim of this study was to investigate the effects of GbE on oxLDL-activated MMPs of human endothelial cells and to test the involvement of LOX-1 and protein kinase C (PKC)-α, extracellular signal-regulated kinase (ERK), and peroxisome proliferator-activated receptor-γ (PPAR-γ).nnnMETHODSnHuman umbilical vein endothelial cells were stimulated with oxLDL, with or without GbE treatment. LOX-1 signaling and MMPs expression were tested by Western blotting or activity assay. Further, protein expression levels of PKC-α, ERK, nuclear factor-κB, and PPAR-γ were investigated by Western blotting.nnnRESULTSnGbE inhibited the oxLDL-caused upregulation of MMP-1, MMP-2, and MMP-3. Pretreating with GbE reduced oxLDL-activated LOX-1 expression. Furthermore, pharmacologic inhibitors of free radicals, Ca(++), PKC, and GbE, inhibited the oxLDL-induced ERK and nuclear factor-κB activation. Lastly, GbE ameliorated the oxLDL-inhibited PPAR-γ function.nnnCONCLUSIONSnData obtained in this study indicate that GbE actives its protective effects by regulating the LOX-1-mediated PKC-α/ERK/PPAR-γ/MMP pathway, resulting in the suppression of reactive oxygen species formation and, ultimately, the reduction of MMPs expression in endothelial cells treated with oxLDL.

Baicalein protects against oxLDL-caused oxidative stress and inflammation by modulation of AMPK- alpha

Atherosclerosis is considered to be a form of chronic inflammation and a disorder of lipid metabolism. Oxidative transformations in the lipid and apolipoprotein B (Apo B) constituent of low density lipoprotein drive the initial step in atherogenesis due to macrophage scavenger receptors identify oxidized LDL (oxLDL) but non-oxidized LDL. The human vascular endothelial cells fact a critical role in vasodilation, provides a nonadhesive surface for circulation, reduces vascular smooth muscle proliferation, inflammation, thrombus formation and platelet aggregation. Assembly of oxLDL contribute to stimulation of endothelial cells with up-regulation of adhesion molecules, increase oxidative stress to the vascular endothelium and inhibition of NO-mediated vasodilation. When adhesion molecules are over-expressed on the surface of endothelial cells under oxLDL stimulation, it will recruit monocytes to the arterial wall. Then adherent monocytes will migrate into the subendothelial space and subsequently differentiate into macrophages. In the subendothelial space, oxLDL will be taken up by macrophages, thereby causing the substantial cholesterol accumulation and the foam cells production.

SIRT1 inhibition causes oxidative stress and inflammation in patients with coronary artery disease

Coronary artery disease (CAD) is the primary critical cardiovascular event. Endothelial cell and monocyte dysfunction with subsequent extravagant inflammation are the main causes of vessel damage in CAD. Thus, strategies that repress cell death and manage unsuitable pro-inflammatory responses in CAD are potential therapeutic strategies for improving the clinical prognosis of patients with CAD. SIRT1 (Sirtuin 1) plays an important role in regulating cellular physiological processes. SIRT1 is also thought to protect the cardiovascular system by means of its antioxidant, anti-inflammation and anti-apoptosis activities. In the present study, we found that the SIRT1 expression levels were repressed and the acetylated p53 expression levels were enhanced in the monocytes of patients with CAD. LOX-1/oxidative stress was also up-regulated in the monocytes of patients with CAD, thereby increasing pro-apoptotic events and pro-inflammatory responses. We also demonstrated that monocytes from CAD patients caused endothelial adhesion molecule activation and the adherence of monocytes and endothelial cells. Our findings may explain why CAD patients remain at an increased risk of long-term recurrent ischemic events and provide new knowledge regarding the management of clinical CAD patients.

Effects of thermosensitive chitosan-gelatin based hydrogel containing glutathione on Cisd2-deficient chondrocytes under oxidative stress

Aging is considered as a primary risk factor in the development of osteoarthritis (OA) which associated with mitochondrial dysfunction and oxidative stress. CDGSH iron sulfur domain 2 (Cisd2) deficiency causes mitochondrial dysfunction and drive premature aging. In the present study, thermosensitive chitosan-gelatin based hydrogel containing glutathione was developed as injectable drug delivery system for administration by minimal invasive surgery for the treatment of OA. Cisd2 deficiency (Cisd2-/-) mouse induced pluripotent stem cells-derived chondrocytes were established and characterized. The results suggested that 100μM of glutathione may be an optimal concentration to treat Cisd2-/- chondrocytes without cytotoxicity. The developed hydrogel showed sustained release profile of the glutathione and could decrease the reactive oxygen species level. Post-treatment of glutathione-loaded hydrogel could rescue Cisd2-/- chondrocytes from oxidative damage via increasing catalase activity, down-regulation of inflammation, and decreasing apoptosis. These results suggest that thermosensitive glutathione-loaded hydrogel may be a potential antioxidant therapeutic strategy for treating Cisd2-/- chondrocytes in the near future.

Chicoric acid is a potent anti-atherosclerotic ingredient by anti-oxidant action and anti-inflammation capacity

Atherosclerotic cardiovascular disease is linked to both oxidative stress and endothelial cell dysfunction. Chicoric acid has antioxidant and anti-inflammatory properties. In the present investigation, we demonstrated that chicoric acid inhibits oxidized low-density lipoprotein (oxLDL)-facilitated dysfunction in human umbilical vein endothelial cells (HUVECs). Oxidative injuries were tested by investigating the formation of intracellular reactive oxygen species (ROS) and by examining the activity of antioxidant enzymes and the function of endothelial nitric oxide synthase (eNOS). We also confirmed that chicoric acid mitigates apoptotic features caused by oxLDL, such as the subsequent break down of mitochondrial transmembrane potential and the activation of Bax, which promote DNA strand breaks and activate caspase-3. Moreover, our data revealed that chicoric acid attenuated the oxLDL activation of NF-?B, the attachment of THP-1 cells and the overexpression of adhesion molecules in human endothelial cells. The results of this study suggest a potential molecular mechanism through which chicoric acid inhibits oxLDL-induced human endothelial dysfunction.

Baicalein is an available anti-atherosclerotic compound through modulation of nitric oxide-related mechanism under oxLDL exposure

OxLDL facilitate reactive oxygen species (ROS) formation and up-regulation of the executioner caspase-3 via the mitochondrial apoptotic pathway involves several critical steps in human endothelial cells. Previous studies reported that oxLDL-facilitated endothelial oxidative stress is associated with impairment of eNOS and up-regulation of inducible nitric oxide synthase (iNOS). Baicalein is the most abundant component that has anti-HIV, anti-tumor, anti-oxidant and free radical scavenging functions. In this present study, we shown that baicalein hinibits oxLDL-caused endothelial dysfunction through suppression of endothelial inflammation and oxidative stress that causes to cellular apoptosis. Specifically, baicalein reduces the elevation of ROS concentration, which subsequently inhibits the oxLDL-decreased expression of anti-oxidant enzymes, enriches the bioavailability of NO, stabilizes the mitochondrial membrane, thereby inhibiting the discharge of cytochrome c from mitochondria, a molecule required for the activation of the pro-apoptotic protein caspase 3. However, inhibition of eNOS impairs the anti-apoptotic and anti-inflammatory effects of baicalein. These results provide new insight into the possible molecular mechanisms by which baicalein protects against atherogenesis by NO-related pathways.