The complex effects of the slow-releasing hydrogen sulfide donor GYY4137 in a model of acute joint inflammation and in human cartilage cells

The role of hydrogen sulfide (H2 S) in inflammation remains unclear with both pro- and anti-inflammatory actions of this gas described. We have now assessed the effect of GYY4137 (a slow-releasing H2 S donor) on lipopolysaccharide (LPS)-evoked release of inflammatory mediators from human synoviocytes (HFLS) and articular chondrocytes (HAC) in vitro. We have also examined the effect of GYY4137 in a complete Freund's adjuvant (CFA) model of acute joint inflammation in the mouse. GYY4137 (0.1-0.5 mM) decreased LPS-induced production of nitrite (NO2 (-) ), PGE2 , TNF-a and IL-6 from HFLS and HAC, reduced the levels and catalytic activity of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and reduced LPS-induced NF-?B activation in vitro. Using recombinant human enzymes, GYY4137 inhibited the activity of COX-2, iNOS and TNF-a converting enzyme (TACE). In the CFA-treated mouse, GYY4137 (50 mg/kg, i.p.) injected 1 hr prior to CFA increased knee joint swelling while an anti-inflammatory effect, as demonstrated by reduced synovial fluid myeloperoxidase (MPO) and N-acetyl-ß-D-glucosaminidase (NAG) activity and decreased TNF-a, IL-1ß, IL-6 and IL-8 concentration, was apparent when GYY4137 was injected 6 hrs after CFA. GYY4137 was also anti-inflammatory when given 18 hrs after CFA. Thus, although GYY4137 consistently reduced the generation of pro-inflammatory mediators from human joint cells in vitro, its effect on acute joint inflammation in vivo depended on the timing of administration.

Tyrosine nitration of prostacyclin synthase is associated with enhanced retinal cell apoptosis in diabetes

The risk of diabetic retinopathy is associated with the presence of both oxidative stress and toxic eicosanoids. Whether oxidative stress actually causes diabetic retinopathy via the generation of toxic eicosanoids, however, remains unknown. The aim of the present study was to determine whether tyrosine nitration of prostacyclin synthase (PGIS) contributes to retinal cell death in vitro and in vivo. Exposure of human retinal pericytes to heavily oxidized and glycated LDL (HOG-LDL), but not native forms of LDL (N-LDL), for 24 hours significantly increased pericyte apoptosis, accompanied by increased tyrosine nitration of PGIS and decreased PGIS activity. Inhibition of the thromboxane receptor or cyclooxygenase-2 dramatically attenuated HOG-LDL-induced apoptosis without restoring PGIS activity. Administration of superoxide dismutase (to scavenge superoxide anions) or L-N(G)-nitroarginine methyl ester (L-NAME, a nonselective nitric oxide synthase inhibitor) restored PGIS activity and attenuated pericyte apoptosis. In Akita mouse retinas, diabetes increased intraretinal levels of oxidized LDL and glycated LDL, induced PGIS nitration, enhanced apoptotic cell death, and impaired blood-retinal barrier function. Chronic administration of tempol, a superoxide scavenger, reduced intraretinal oxidized LDL and glycated LDL levels, PGIS nitration, and retina cell apoptosis, thereby preserving the integrity of blood-retinal barriers. In conclusion, oxidized LDL-mediated PGIS nitration and associated thromboxane receptor stimulation might be important in the initiation and progression of diabetic retinopathy.

Berries:emerging impact on cardiovascular health

Berries are a good source of polyphenols, especially anthocyanins, micronutrients, and fiber. In epidemiological and clinical studies, these constituents have been associated with improved cardiovascular risk profiles. Human intervention studies using chokeberries, cranberries, blueberries, and strawberries (either fresh, or as juice, or freeze-dried), or purified anthocyanin extracts have demonstrated significant improvements in LDL oxidation, lipid peroxidation, total plasma antioxidant capacity, dyslipidemia, and glucose metabolism. Benefits were seen in healthy subjects and in those with existing metabolic risk factors. Underlying mechanisms for these beneficial effects are believed to include upregulation of endothelial nitric oxide synthase, decreased activities of carbohydrate digestive enzymes, decreased oxidative stress, and inhibition of inflammatory gene expression and foam cell formation. Though limited, these data support the recommendation of berries as an essential fruit group in a heart-healthy diet.

Pigment epithelium-derived factor mitigates inflammation and oxidative stress in retinal pericytes exposed to oxidized low-density lipoprotein

Oxidized and/or glycated low-density lipoprotein (LDL) may mediate capillary injury in diabetic retinopathy. The mechanisms may involve pro-inflammatory and pro-oxidant effects on retinal capillary pericytes. In this study, these effects, and the protective effects of pigment epithelium-derived factor (PEDF), were defined in a primary human pericyte model. Human retinal pericytes were exposed to 100 microg/ml native LDL (N-LDL) or heavily oxidized glycated LDL (HOG-LDL) with or without PEDF at 10-160 nM for 24 h. To assess pro-inflammatory effects, monocyte chemoattractant protein-1 (MCP-1) secretion was measured by ELISA, and nuclear factor-kappaB (NF-kappaB) activation was detected by immunocytochemistry. Oxidative stress was determined by measuring intracellular reactive oxygen species (ROS), peroxynitrite (ONOO(-)) formation, inducible nitric oxide synthase (iNOS) expression, and nitric oxide (NO) production. The results showed that MCP-1 was significantly increased by HOG-LDL, and the effect was attenuated by PEDF in a dose-dependent manner. PEDF also attenuated the HOG-LDL-induced NF-kappaB activation, suggesting that the inhibitory effect of PEDF on MCP-1 was at least partially through the blockade of NF-kappaB activation. Further studies demonstrated that HOG-LDL, but not N-LDL, significantly increased ONOO(-) formation, NO production, and iNOS expression. These changes were also alleviated by PEDF. Moreover, PEDF significantly ameliorated HOG-LDL-induced ROS generation through up-regulation of superoxide dismutase 1 expression. Taken together, these results demonstrate pro-inflammatory and pro-oxidant effects of HOG-LDL on retinal pericytes, which were effectively ameliorated by PEDF. Suppressing MCP-1 production and thus inhibiting macrophage recruitment may represent a new mechanism for the salutary effect of PEDF in diabetic retinopathy and warrants more studies in future.

Free radical-mediated lipid peroxidation and systemic nitric oxide bioavailability:implications for postexercise hemodynamics

The metabolic vasodilator mediating postexercise hypotension (PEH) is poorly understood. Recent evidence suggests an exercise-induced reliance on pro-oxidant-stimulated vasodilation in normotensive young human subjects, but the role in the prehypertensive state is not known.

Susceptibility of streptozotocin-induced diabetic rat retinal function and ocular blood flow to acute intraocular pressure challenge.

PURPOSE: To consider whether STZ-induced hyperglycemia renders rat retinal function and ocular blood flow more susceptible to acute intraocular pressure (IOP) challenge.

METHODS: Retinal function (electroretinogram, ERG) was measured during acute IOP challenge (10-100 mmHg, 5 mmHg increments, 3 min/step, vitreal cannulation) in adult Long-Evans rats (6-week old, citrate: n=6, STZ: n=10) 4 weeks after citrate buffer or streptozotocin (STZ, 65 mg/kg, blood glucose > 15 mmol/l) injection. At each IOP, dim and bright flash (-4.56, -1.72 log cd.s.m^-2) ERG responses were recorded to measure inner retinal and ON-bipolar cell function, respectively. Ocular blood flow (laser Doppler flowmetry, citrate; n=6, STZ; n=10) was also measured during acute IOP challenge. Retinae were isolated for qPCR analysis of nitric oxide synthase mRNA expression endothelial, eNos; inducible, iNos; neuronal, nNos).

RESULTS: STZ-induced diabetes increased the susceptibility of inner retinal (IOP at 50% response, 60.1, CI: 57.0-62.0 mmHg vs. citrate: 67.5, CI: 62.1-72.4 mmHg) and ON-bipolar cell function (STZ: 60.3, CI: 58.0-62.8 mmHg vs. citrate: 65.1, CI: 58.0-62.78 mmHg) and ocular blood flow (43.9, CI: 40.8-46.8 vs. citrate: 53.4, CI: 50.7-56.1 mmHg) to IOP challenge. Citrate eyes showed elevated eNos mRNA (+49.7%) after IOP stress, an effect not found in STZ-diabetic eyes (-5.7%, P<0.03). No difference was observed for iNos or nNos (P>0.05) following IOP elevation.

CONCLUSIONS: STZ-induced diabetes increased functional susceptibility during acute IOP challenge. This functional vulnerability is associated with a reduced capacity for diabetic eyes to upregulate eNOS expression and to autoregulate blood flow in response to stress.

Current therapeutic strategies to mitigate the eNOS dysfunction in ischaemic stroke

Impairment of endothelial nitric oxide synthase (eNOS) activity is implicated in the pathogenesis of endothelial dysfunction in many diseases including ischaemic stroke. The modulation of eNOS during and/or following ischaemic injury often represents a futile compensatory mechanism due to a significant decrease in nitric oxide (NO) bioavailability coupled with dramatic increases in the levels of reactive oxygen species that further neutralise NO. However, applications of a number of therapeutic agents alone or in combination have been shown to augment eNOS activity under a variety of pathological conditions by potentiating the expression and/or activity of Akt/eNOS/NO pathway components. The list of these therapeutic agents include NO donors, statins, angiotensin-converting enzyme inhibitors, calcium channel blockers, phosphodiesterase-3 inhibitors, aspirin, dipyridamole and ellagic acid. While most of these compounds exhibit anti-platelet properties and are able to up-regulate eNOS expression in endothelial cells and platelets, others suppress eNOS uncoupling and tetrahydrobiopterin (an eNOS stabiliser) oxidation. As the number of therapeutic molecules that modulate the expression and activity of eNOS increases, further detailed research is required to reveal their mode of action in preventing and/or reversing the endothelial dysfunction.