Fortified S-Allyl L-Cysteine: Animal Safety, Effect on Retinal Ischemia, and Role of Wnt in the Underlying Therapeutic Mechanism.

PURPOSE: Retinal ischemia is a medical condition associated with numerous retinal vascular disorders, such as age-related macular degeneration, glaucoma, and diabetic retinopathy. This in vitro cell and in vivo animal study investigated not only the protective effect of S-allyl L-cysteine (SAC, an active component of garlic) against retinal ischemia but also its associated protective mechanisms. METHODS: Retinal ischemia was mimicked by raising the intraocular pressure to 120 mmHg for 1 hour in one eye. The effects of pre-/postischemic administration of vehicle vs. SAC 0.18 mg vs. SAC 0.018 mg vs. SAC 0.0018 mg treatments on retina cells were evaluated through cellular viability (MTT assay), flash electroretinograms (ERGs), and fluorogold retrograde labelling (retinal ganglion cell (RGC) counting). Also, protein immunoblot was utilized to assess the role of Wnt, hypoxia inducible factor (HIF)-1α, and vascular endothelium factor (VEGF) in the proposed anti-ischemic mechanism. Lastly, the safety of drug consumption was investigated for changes in the animal's body weight, ERG waves, and blood biochemical parameters (e.g., glucose levels). RESULTS: The characteristic ischemic changes including significant reduction in ERG b-wave ratio and RGC number were significantly counteracted by pre- and postischemic low dose of SAC. Additionally, ischemia-induced overexpression of Wnt/HIF-1α/VEGF protein was ameliorated significantly by preischemic low dose of SAC. In terms of the animal safety, no significant body weight and electrophysiological differences were observed among defined different concentrations of SAC without following ischemia. In low SAC dosage and vehicle groups, various blood biochemical parameters were normal; however, high and medium concentrations of SAC significantly lowered the levels of uric acid, Hb, and MCHC. CONCLUSION: This study shows that preischemic administration of low SAC dosage has been proved to be safe and most effective against rat retinal ischemia electrophysiologically and/or histopathologically. Moreover, counteracting the ischemia-induced overexpression of Wnt/HIF-1α/VEGF might presently explain SAC's anti-ischemic mechanism.

Dry Eye: The Effect of Chi-Ju-Di-Huang-Wan Plus Si Wu Tang and the Underlying Mechanism.

Objectives: In this pilot study, the effect of 970 mg Chi-Ju-Di-Huang-Wan (CJDHW) plus 30 mg four-substance decoction (Si Wu Tang; CJDHWSWT) was evaluated, in terms of its ability to alleviate dry eye symptoms and its therapeutic mechanism. Methods: This double-masked prospective investigation has recruited dry eye patients who have been randomly selected into two groups, namely treatment (n = 15) versus nontreatment (n = 15). In the treatment group, a daily oral intake of CJDHWSWT plus eye drops systane ultra was given for 90 consecutive days. In the nontreatment group, only defined eye drops were prescribed. The examinations included Schirmer's test, fluorescein-stained superficial punctate keratitis (SPK), artificial tear consumption, tear vascular endothelium growth factor (VEGF) level, and ocular surface disease index. The drug safety tests included liver and kidney functions, and complete blood counts. The candidates were observed during the screening visit and the following three monthly follow-ups. The data were analyzed by unpaired Student's t-test. Results: Compared to no significance in the nontreatment group, CJDHWSWT significantly (p = 0.03) increased the tear secretion after 3 months of intake. Furthermore, in contrast to no significance in the treatment group, there were significant alterations, including (i) increased fluorescein-stained SPK areas (p = 0.03); (ii) increased artificial tear instillation amount (p = 0.03); (iii) elevated tear VEGF protein levels (p = 0.03) in the nontreatment group; and (iv) significant improvement in clinically relevant phenomenon (e.g., reading limit and uncomfortable feeling in windy conditions), after treatment of artificial tear plus oral intake of CJDHWSWT. As shown by the post-treatment normal defined laboratory data, there were no adverse drug effects. Conclusions: This study has supported that CJDHWSWT is safe and effective in relieving dry eye's clinically relevant symptoms/phenomena. CJDHWSWT avoided the tear VEGF upregulation probably induced by dry eye-associated hypoxia/ischemia.

Dendrobium nobile Lindley and its bibenzyl component moscatilin are able to protect retinal cells from ischemia/hypoxia by dowregulating placental growth factor and upregulating Norrie disease protein.

BACKGROUND: Presumably, progression of developmental retinal vascular disorders is mainly driven by persistent ischemia/hypoxia. An investigation into vision-threatening retinal ischemia remains important. Our aim was to evaluate, in relation to retinal ischemia, protective effects and mechanisms of Dendrobium nobile Lindley (DNL) and its bibenzyl component moscatilin. The therapeutic mechanisms included evaluations of levels of placental growth factor (PLGF) and Norrie disease protein (NDP). METHODS: An oxygen glucose deprivation (OGD) model involved cells cultured in DMEM containing 1% O2, 94% N2 and 0 g/L glucose. High intraocular pressure (HIOP)-induced retinal ischemia was created by increasing IOP to 120 mmHg for 60 min in Wistar rats. The methods included electroretinogram (ERG), histopathology, MTT assay and biochemistry. RESULTS: When compared with cells cultured in DMEM containing DMSO (DMSO+DMEM), cells subjected to OGD and pre-administrated with DMSO (DMSO+OGD) showed a significant reduction in the cell viability and NDP expression. Moreover, cells that received OGD and 1 h pre-administration of 0.1 µM moscatilin (Pre-OGD Mos 0.1 µM) showed a significant counteraction of the OGD-induced decreased cell viability. Furthermore, compared with the DMSO+OGD group (44.54 ± 3.15%), there was significant elevated NDP levels in the Pre-OGD Mos 0.1 µM group (108.38 ± 29.33%). Additionally, there were significant ischemic alterations, namely reduced ERG b-wave, less numerous retinal ganglion cells, decreased inner retinal thickness, and reduced/enhanced amacrine's ChAT/Müller's GFAP or vimentin immunolabelings. Moreover, there were significantly increased protein levels of HIF-1α, VEGF, PKM2, RBP2 and, particularly, PLGF (pg/ml; Sham vs. Vehicle: 15.11 ± 1.58 vs. 39.53 ± 5.25). These ischemic effects were significantly altered when 1.0 g/Kg/day DNL (DNL1.0 + I/R or I/R+ DNL1.0) was applied before and/or after ischemia, but not vehicle (Vehicle+I/R). Of novelty and significance, the DNL1.0 action mechanism appears to be similar to that of the anti-PLGF Eylea [PLGF (pg/ml); DNL1.0 vs. Eylea+I/R: 19.93 ± 2.24 vs. 6.44 ± 0.60]. CONCLUSIONS: DNL and moscatilin are able to protect against retinal ischemic/hypoxic changes respectively by downregulating PLGF and upregulating NDP. Progression of developmental retinal vascular disorders such as Norrie disease due to persistent ischemia/hypoxia might be thus prevented.

Xue-fu-Zhu-Yu decoction protects rats against retinal ischemia by downregulation of HIF-1α and VEGF via inhibition of RBP2 and PKM2.

BACKGROUND: Retinal ischemia-related eye diseases result in visual dysfunction. This study investigates the protective effects and mechanisms of Xue-Fu-Zhu-Yu decoction (XFZYD) with respect to retinal ischemia. METHODS: Retinal ischemia (I) was induced in Wistar rats by a high intraocular pressure (HIOP) of 120 mmHg for 1 h, which was followed by reperfusion of the ischemic eye; the fellow untreated eye acted as a control. Electroretinogram (ERG), biochemistry and histopathology investigations were performed. RESULTS: Significant ischemic changes occurred after ischemia including decreased ERG b-wave ratios, less numerous retinal ganglion cells (RGCs), reduced inner retinal thickness, fewer choline acetyltransferase (ChAT) labeled amacrine cell bodies, increased glial fibrillary acidic protein (GFAP) immunoreactivity and increased vimentin Müller immunolabeling. These were accompanied by significant increases in the mRNA/protein concentrations of vascular endothelium growth factor, hypoxia-inducible factor-1α, pyruvate kinase M2 and retinoblastoma-binding protein 2. The ischemic changes were concentration-dependently and significantly altered when XFZYD was given for seven consecutive days before or after retina ischemia, compared to vehicle. These alterations included enhanced ERG b-wave amplitudes, more numerous RGCs, enhanced inner retinal thickness, a greater number of ChAT immunolabeled amacrine cell bodies and decreased GFAP/vimentin immunoreactivity. Furthermore, decreased mRNA levels of VEGF, HIF-1α, PKM2, and RBP2 were also found. Reduced protein concentrations of VEGF, HIF-1α, PKM2, and RBP2 were also demonstrated. Furthermore, there was an inhibition of the ischemia-associated increased ratios (target protein/ß-actin) in the protein levels of VEGF, HIF-1α, PKM2, and RBP2, which were induced by Shikonin, JIB-04 or Avastin. CONCLUSION: XFZYD would seem to protect against well-known retinal ischemic changes via a synergistic inhibition of RBP2 and PKM2, as well as down-regulation of HIF-1α and a reduction in VEGF secretion.

S-allyl L-cysteine protects the retina against kainate excitotoxicity in the rat.

Excitotoxicity has been proposed to play a pivotal role in retinal ischemia. Retinal ischemia-associated ocular disorders are vision threatening. The aim was to also examine whether and how S-allyl L-cysteine (SAC) can protect the retina against kainate excitotoxicity. In vivo retinal excitotoxicity was induced by an intravitreous injection of 100 µM kainate into a Wistar rat eye for 1 day. The management and mechanisms involved in the processes were evaluated by electrophysiology, immunohistochemistry, histopathology, and various biochemical approaches. In the present study, the cultured retinal cells were shown to possess kainate receptors. The defined retinal excitotoxic changes were characterized by a decrease in electroretinogram (ERG) b-wave amplitudes, a loss of the fluorogold retrograde labeled retinal ganglion cells (RGCs), an increase in the apoptotic cells in the RGC layer, and an increase in vimentin or glial fibrillary acidic protein (GFAP) immunoreactivity, a marker for Müller cells. An up-regulation in the mRNA levels of inducible nitric oxide synthase (iNOS) and matrix metalloproteinases-9 (MMPs-9) was also detected in the retina subjected to kainate excitoxicity. Importantly, the excitotoxicity-induced alterations were significantly blunted when 100 µM SAC and/or the kainate receptor antagonist CNQX was applied. Conclusively, SAC would seem to protect the retina against kainate excitotoxicity via an inhibition of the up-regulation of iNOS and MMP-9 as well as a modulation of glial activation and apoptosis.