O-Glycosylation Regulates Ubiquitination and Degradation of the Anti-Inflammatory Protein A20 to Accelerate Atherosclerosis in Diabetic ApoE-Null Mice

Background: Accelerated atherosclerosis is the leading cause of morbidity and mortality in diabetic patients. Hyperglycemia is a recognized independent risk factor for heightened atherogenesis in diabetes mellitus (DM). However, our understanding of the mechanisms underlying glucose damage to the vasculature remains incomplete. Methodology/Principal Findings: High glucose and hyperglycemia reduced upregulation of the NF-κB inhibitory and atheroprotective protein A20 in human coronary endothelial (EC) and smooth muscle cell (SMC) cultures challenged with Tumor Necrosis Factor alpha (TNF), aortae of diabetic mice following Lipopolysaccharide (LPS) injection used as an inflammatory insult and in failed vein-grafts of diabetic patients. Decreased vascular expression of A20 did not relate to defective transcription, as A20 mRNA levels were similar or even higher in EC/SMC cultured in high glucose, in vessels of diabetic C57BL/6 and FBV/N mice, and in failed vein grafts of diabetic patients, when compared to controls. Rather, decreased A20 expression correlated with post-translational O-Glucosamine-N-Acetylation (O-GlcNAcylation) and ubiquitination of A20, targeting it for proteasomal degradation. Restoring A20 levels by inhibiting O-GlcNAcylation, blocking proteasome activity, or overexpressing A20, blocked upregulation of the receptor for advanced glycation end-products (RAGE) and phosphorylation of PKCβII, two prime atherogenic signals triggered by high glucose in EC/SMC. A20 gene transfer to the aortic arch of diabetic ApoE null mice that develop accelerated atherosclerosis, attenuated vascular expression of RAGE and phospho-PKCβII, significantly reducing atherosclerosis. Conclusions: High glucose/hyperglycemia regulate vascular A20 expression via O-GlcNAcylation-dependent ubiquitination and proteasomal degradation. This could be key to the pathogenesis of accelerated atherosclerosis in diabetes

Eficiência do laser diodo 980 nm em comparação à da glicose a 75% na oclusão de veias da orelha de coelhos The efficiency of the diode laser 980 nm compared to glucose 75% in occlusion of the veins in rabbit ears

CONTEXTO: Por ser o laser um método novo no tratamento das varizes, há muitos mitos e dúvidas com relação à sua eficácia; assim, surgiu a necessidade de compará-lo a substância esclerosante mais utilizada em nosso meio (glicose hipertônica). OBJETIVO: Comparar a eficiência do laser diodo 980 nm à glicose 75% na oclusão de veias em orelha de coelho. MÉTODOS: Ensaio aleatório em animais de laboratório por 21 dias. A amostra consistiu de orelhas de coelhos machos adultos. Grupo L (laser): 15 orelhas tratadas com laser; grupo G (glicose 75%): 15 orelhas tratadas com glicose a 75%. Variáveis primárias: veias esclerosadas e/ou ocluídas. Variáveis complementares: volume da substância administrada, complicações e peso. O tamanho da amostra foi estimado em 30 orelhas. Foi realizado o teste exato de Fisher associado ao Risco Relativo (RR), calculando-se o intervalo de confiança (IC) de 95% para as variáveis acima. RESULTADOS: A incidência de esclerose ou oclusão venosa no grupo G foi de 53% (8/15; IC95%: 27-79) e no grupo L, 20% (3/15; IC95%: 4-49). O p bicaudal foi de 0,1281, o RR usando a aproximação de Katz foi de 2,66; IC95%: 0,87-8,15. CONCLUSÃO: A eficiência do laser diodo 980 nm em comparação à da glicose 75% na oclusão de veias para o modelo experimental estudado foi equivalente.<br>BACKGROUND: The laser is a new treatment to varicose veins and there is several myths and doubts in relation to its efficacy; then, there is the need to compare it with the most commonly sclerosing solution (hypertonic glucose) used in our specialty. OBJECTIVE: To compare the efficiency of the diode laser 980nm to the glucose 75% in the occlusion of veins from the ear of rabbits. METHODS: Aleatory trial in laboratory animals during 21 days. The sample consisted of ears from male adult rabbits. Group L (laser): 15 ears treated with laser; group G (glucose 75%): 15 ears treated with glucose 75%. Primary variables: sclerotic and/or occluded veins. Complementary variables: volume of the managed substance, complications and weight of rabbit. The sample size was estimated in 30 ears. The statistical analysis was carried out by Fisher''s exact test associated to the Relative Risk (RR), calculating the confidence interval of 95% for the mentioned variables. RESULTS: The incidence of sclerosis or venous occlusion in group G was 53% (8/15; 95%CI: 27-79) and in group L was 20% (3/15; 95% CI: 4-49). Two-tailed p was 0.1281, RR using the approximation of Katz was 2.66; 95%CI: 0.87-8.15. CONCLUSION: The efficiency of the diode laser 980 nm in comparison to glucose 75% in occlusion of veins in this experimental model was equivalent

4-aminopyridyl-based lead compounds targeting CYP51 prevent spontaneous parasite relapse in a chronic model and improve cardiac pathology in an acute model of Trypanosoma cruzi infection

Chagas disease, caused by the protozoan Trypanosoma cruzi, is the leading cause of heart failure in Latin America. The clinical treatment of Chagas disease is limited to two 60 year-old drugs, nifurtimox and benznidazole, that have variable efficacy against different strains of the parasite and may lead to severe side effects. CYP51 is an enzyme in the sterol biosynthesis pathway that has been exploited for the development of therapeutics for fungal and parasitic infections. In a target-based drug discovery program guided by x-ray crystallography, we identified the 4-aminopyridyl-based series of CYP51 inhibitors as being efficacious versus T.cruzi in vitro; two of the most potent leads, 9 and 12, have now been evaluated for toxicity and efficacy in mice.Both acute and chronic animal models infected with wild type or transgenic T. cruzi strains were evaluated. There was no evidence of toxicity in the 28-day dosing study of uninfected animals, as judged by the monitoring of multiple serum and histological parameters. In two acute models of Chagas disease, 9 and 12 drastically reduced parasitemia, increased survival of mice, and prevented liver and heart injury. None of the compounds produced long term sterile cure. In the less severe acute model using the transgenic CL-Brenner strain of T.cruzi, parasitemia relapsed upon drug withdrawal. In the chronic model, parasitemia fell to a background level and, as evidenced by the bioluminescence detection of T. cruzi expressing the red-shifted luciferase marker, mice remained negative for 4 weeks after drug withdrawal. Two immunosuppression cycles with cyclophosphamide were required to re-activate the parasites. Although no sterile cure was achieved, the suppression of parasitemia in acutely infected mice resulted in drastically reduced inflammation in the heart.The positive outcomes achieved in the absence of sterile cure suggest that the target product profile in anti-Chagasic drug discovery should be revised in favor of safe re-administration of the medication during the lifespan of a Chagas disease patient. A medication that reduces parasite burden may halt or slow progression of cardiomyopathy and therefore improve both life expectancy and quality of life