A Rat Model of Rheumatoid Arthritis: TDZD-8 is Associated with the Protection Against the Induction of the Synovium Knee Joint IL-17A/GSK3β/ROS/α-SMA Axis of Fibrosis

SUMMARY: Rheumatoid arthritis (RA) that affects the synovial knee joint causes swelling of the synovial membrane and tissue damage. Interleukin-17A (IL-17A) and the enzyme glycogen synthase kinase-3β (GSK3β) are involved in the pathogenesis of RA. The link between IL-17A, GSK3β, the oxidative stress, and the profibrogenic marker alpha-smooth muscle actin (α-SMA) with and without TDZD-8, GSK3β inhibitor has not been studied before. Consequently, active immunization of rats was performed to induce RA after three weeks using collagen type II (COII) injections. The treated group received daily injection of 1 mg/kg TDZD-8 for 21 days following the immunization protocol (COII+TDZD-8). Blood and synovium tissue samples were harvested at the end of the experiment. RA development was confirmed as corroborated by a substantial increase in blood levels of the highly specific autoantibody for RA, anti-citrullinated protein antibody as well as augmentation of reactive oxidative species (ROS) levels measured as lipid peroxidation. RA induction also increased synovium tissue levels of IL-17A and the profibrogenic marker, α-SMA. All these parameters seemed to be significantly (p<0.0001) ameliorated by TDZD-8. Additionally, a significant correlation between IL-17A, ROS, and α-SMA and biomarkers of RA was observed. Thus, knee joint synovium RA induction augmented IL-17A/GSK3β/ROS/α-SMA axis mediated arthritis in a rat model of RA, which was inhibited by TDZD-8.

La artritis reumatoide (AR) que afecta la articulación sinovial de la rodilla provoca inflamación de la membrana sinovial y daño tisular. La interleucina-17A (IL-17A) y la enzima glucógeno sintasa quinasa-3β (GSK3β) están involucradas en la patogenia de la AR. No se ha estudiadol vínculo entre IL-17A, GSK3β, el estrés oxidativo y el marcador profibrogénico actina de músculo liso alfa (α-SMA) con y sin inhibidor de TDZD-8, GSK3β. En consecuencia, se realizó una inmunización activa de ratas para inducir la AR después de tres semanas usando inyecciones de colágeno tipo II (COII). El grupo tratado recibió una inyección diaria de 1 µg/ kg de TDZD-8 durante 21 días siguiendo el protocolo de inmunización (COII+TDZD-8). Se recogieron muestras de sangre y tejido sinovial al final del experimento. El desarrollo de AR se confirmó como lo corroboró el aumento sustancial en los niveles sanguíneos del autoanticuerpo altamente específico para AR, el anticuerpo antiproteína citrulinada, así como el aumento de los niveles de especies oxidativas reactivas (ROS) medidos como peroxidación lipídica. La inducción de AR también aumentó los niveles de tejido sinovial de IL-17A y el marcador profibrogénico, α-SMA. Todos estos parámetros parecían mejorar significativamente (p<0,0001) con TDZD-8. Además, se observó una correlación significativa entre IL- 17A, ROS y α-SMA y biomarcadores de AR. Por lo tanto, la inducción de AR en la sinovial de la articulación de la rodilla aumentó la artritis mediada por el eje IL-17A/GSK3β/ROS/α-SMA en un modelo de rata de AR, que fue inhibida por TDZD-8.

A rat model of rheumatoid arthritis: TDZD-8 is associated with the protection against the induction of the synovium knee joint IL-17A/GSK3beta/ROS/alpha-SMA axis of fibrosis / Un modelo de rata de artritis reumatoide: TDZD-8 está asociado con la protección contra la inducción del eje de fibrosis de la articulación sinovial de la rodilla IL-17A/GSK3ß/ROS/α-SMA

SUMMARY: Rheumatoid arthritis (RA) that affects the synovial knee joint causes swelling of the synovial membrane and tissue damage. Interleukin-17A (IL-17A) and the enzyme glycogen synthase kinase-3β (GSK3β) are involved in the pathogenesis of RA. The link between IL-17A, GSK3β, the oxidative stress, and the profibrogenic marker alpha-smooth muscle actin (α-SMA) with and without TDZD-8, GSK3β inhibitor has not been studied before. Consequently, active immunization of rats was performed to induce RA after three weeks using collagen type II (COII) injections. The treated group received daily injection of 1 mg/kg TDZD-8 for 21 days following the immunization protocol (COII+TDZD-8). Blood and synovium tissue samples were harvested at the end of the experiment. RA development was confirmed as corroborated by a substantial increase in blood levels of the highly specific autoantibody for RA, anti-citrullinated protein antibody as well as augmentation of reactive oxidative species (ROS) levels measured as lipid peroxidation. RA induction also increased synovium tissue levels of IL-17A and the profibrogenic marker, α-SMA. All these parameters seemed to be significantly (p<0.0001) ameliorated by TDZD-8. Additionally, a significant correlation between IL-17A, ROS, and α-SMA and biomarkers of RA was observed. Thus, knee joint synovium RA induction augmented IL-17A/GSK3β/ROS/α-SMA axis mediated arthritis in a rat model of RA, which was inhibited by TDZD-8.

La artritis reumatoide (AR) que afecta la articulación sinovial de la rodilla provoca inflamación de la membrana sinovial y daño tisular. La interleucina-17A (IL-17A) y la enzima glucógeno sintasa quinasa-3β (GSK3β) están involucradas en la patogenia de la AR. No se ha estudiadol vínculo entre IL-17A, GSK3β, el estrés oxidativo y el marcador profibrogénico actina de músculo liso alfa (α-SMA) con y sin inhibidor de TDZD-8, GSK3β. En consecuencia, se realizó una inmunización activa de ratas para inducir la AR después de tres semanas usando inyecciones de colágeno tipo II (COII). El grupo tratado recibió una inyección diaria de 1 µg/ kg de TDZD-8 durante 21 días siguiendo el protocolo de inmunización (COII+TDZD-8). Se recogieron muestras de sangre y tejido sinovial al final del experimento. El desarrollo de AR se confirmó como lo corroboró el aumento sustancial en los niveles sanguíneos del autoanticuerpo altamente específico para AR, el anticuerpo antiproteína citrulinada, así como el aumento de los niveles de especies oxidativas reactivas (ROS) medidos como peroxidación lipídica. La inducción de AR también aumentó los niveles de tejido sinovial de IL-17A y el marcador profibrogénico, α-SMA. Todos estos parámetros parecían mejorar significativamente (p<0,0001) con TDZD-8. Además, se observó una correlación significativa entre IL- 17A, ROS y α-SMA y biomarcadores de AR. Por lo tanto, la inducción de AR en la sinovial de la articulación de la rodilla aumentó la artritis mediada por el eje IL-17A/GSK3β/ROS/α-SMA en un modelo de rata de AR, que fue inhibida por TDZD-8.

Quercetin and resveratrol are associated with the downregulation of TNFalpha/NF-kB/inos axis-mediated acute liver injury in rats induced by paracetamol poisoning / La quercetina y el resveratrol están asociados con la regulación decreciente de la lesión hepática aguda mediada por el eje TNF-alfa/NF-kB/inos en ratas inducida por envenenamiento con paracetamol

SUMMARY: Paracetamol (known as acetaminophen, or APAP) poisoning causes acute liver damage that can lead to organ failure and death. We sought to determine that APAP overdose can augment tumor necrosis factor-alpha (TNF-α)/ nuclear factor kappa B (NF-kB)/induced nitic oxide synthase (iNOS) axis-mediated hepatotoxicity in rats, and the anti-inflammatory polyphenolic compounds, quercetin (QUR) plus resveratrol (RES) can ameliorate these parameters. Therefore, we induced acute hepatotoxicity in rats using APAP overdose (2 g/kg, orally) and the protective group of rats were treated with 50 mg/kg QUR plus 30 mg/kg RES for one week before APAP ingestion. Animals were killed at day 8. APAP poisoning caused the induction of hepatic tissue levels of TNF-α, NF-kB, and iNOS, which were significantly (p<0.05) decreased by QUR+RES. QUR+RES, also inhibited liver injury biomarkers, alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Additionally, a link between liver injury and TNF-α /NF-kB / iNOS axis mediated hepatotoxicity was observed. Thus, the presented data backing the conclusion that intoxication by paracetamol increases TNF-α / NF-kB / iNOS axis -mediated hepatotoxicity, and is protected by a combination of quercetin and resveratrol.

El envenenamiento por paracetamol (conocido como acetaminofeno o APAP) causa daño hepático agudo que puede provocar una insuficiencia orgánica y la muerte. El objetivo de este trabajo fue determinar si la sobredosis de APAP puede aumentar la hepatotoxicidad mediada por el eje del factor de necrosis tumoral alfa (TNF-α)/factor nuclear kappa B (NF-kB)/óxido nítico sintasa inducida (iNOS) en ratas, y si el polifenólico antiinflamatorio compuesto por quercetina (QUR) más resveratrol (RES) pueden mejorar estos parámetros. Por lo tanto, inducimos hepatotoxicidad aguda en ratas usando una sobredosis de APAP (2 g/kg, por vía oral). El grupo protector de ratas se trató con 50 mg/ kg de QUR más 30 mg/kg de RES durante una semana antes de la ingestión de APAP. Los animales se sacrificaron el día 8. El envenenamiento con APAP en el tejido hepático provocó la inducción de niveles de TNF-α, NF-kB e iNOS, que se redujeron significativamente (p<0,05) con QUR+RES. QUR+RES, también inhibió los biomarcadores de daño hepático, la alanina aminotransferasa (ALT) y el aspartato aminotransferasa (AST). Además, se observó una relación entre la lesión hepática y la hepatotoxicidad mediada por el eje TNF-α /NF-kB/iNOS. Por lo tanto, los datos presentados respaldan la conclusión de que la intoxicación por paracetamol aumenta la hepatotoxicidad mediada por el eje TNF-α /NF-kB / iNOS, y está protegida por una combinación de quercetina y resveratrol.

Vitamin E suppresses aortic ultrastructural alterations induced by toxic doses of monosodium glutamate / La vitamina E suprime las alteraciones ultraestructurales aórticas inducidas por dosis tóxicas de glutamato monosódico

SUMMARY: An association between certain food additives and chronic diseases is reported. Current study determined whether administering toxic doses of the food additive monosodium glutamate (MSG) into rats can induce aortopathy in association with the oxidative stress and inflammatory biomarkers upregulation and whether the effects of MSG overdose can be inhibited by vitamin E. MSG at a dose of (4 mg/kg; orally) that exceeds the average human daily consumption by 1000x was administered daily for 7 days to the rats in the model group. Whereas, rats treated with vitamin E were divided into two groups and given daily doses of MSG plus 100 mg/ kg vitamin E or MSG plus 300 mg/kg vitamin E. On the eighth day, all rats were culled. Using light and electron microscopy examinations, a profound aortic injury in the model group was observed demonstrated by damaged endothelial layer, degenerated smooth muscle cells (SMC) with vacuoles and condensed nuclei, vacuolated cytoplasm, disrupted plasma membrane, interrupted internal elastic lamina, clumped chromatin, and damaged actin and myosin filaments. Vitamin E significantly protected aorta tissue and cells as well as inhibited MSG-induced tissue malondialdehyde (MDA), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-α). The highest used vitamin E dosage was more effective. Additionally, a significant correlation was observed between the aortic injury degree and tissue MDA, TNF-α, IL-6, and superoxide dismutase (SOD) levels (p=0.001). Vitamin E effectively protects against aortopathy induced by toxic doses of MSG in rats and inhibits oxidative stress and inflammation.

RESUMEN: Se reporta una asociación entre ciertos aditivos alimentarios y enfermedades crónicas. El objetivo de este estudio fue determinar si la administración de dosis tóxicas del aditivo alimentario glutamato monosódico (MSG) en ratas puede inducir aortopatía en asociación con el estrés oxidativo y la regulación positiva de los biomarcadores inflamatorios y si el efecto de una sobredosis de MSG se puede inhibir con vitamina E. Se administró MSG diariamente durante 7 días una dosis de (4 g/kg; por vía oral) que excede el consumo diario humano promedio, en 1000x a las ratas del grupo modelo. Mientras que las ratas tratadas con vitamina E se dividieron en dos grupos y se administraron dosis diarias de MSG más 100 mg/kg de vitamina E o MSG más 300 mg/kg de vitamina E. Todas las ratas fueron sacrificadas en el octavo día. Usando exámenes de microscopía óptica y electrónica, se observó una lesión aórtica profunda en el grupo modelo demostrada por una capa endotelial dañada, células musculares lisas degeneradas (SMC) con vacuolas y núcleos condensados, citoplasma vacuolado, membrana plasmática rota, lámina elástica interna interrumpida, cromatina agrupada y filamentos de actina y miosina dañados. La vitamina E protegió significativamente el tejido y las células de la aorta, además de inhibir el malondialdehído tisular (MDA) inducido por MSG, la interleucina-6 (IL-6) y el factor de necrosis tumoral alfa (TNF-α). La dosis más alta de vitamina E utilizada fue más efectiva. Además, se observó una correlación significativa entre el grado de lesión aórtica y los niveles tisulares de MDA, TNF-α, IL-6 y superóxido dismutasa (SOD) (p=0,001). La vitamina E efectivamente protege contra la aortopatía inducida por dosis tóxicas de MSG en ratas e inhibe el estrés oxidativo y la inflamación.

Metformin Protects against Diabetic Cardiomyopathy: An Association between Desmin-Sarcomere Injury and the iNOS/mTOR/TIMP-1 Fibrosis Axis.

The intermediate filament protein desmin is essential for maintaining the structural integrity of sarcomeres, the fundamental unit of cardiac muscle. Diabetes mellitus (DM) can cause desmin to become dysregulated, following episodes of nitrosative stress, through the activation of the iNOS/mTOR/TIMP-1 pathway, thereby stimulating collagen deposition in the myocardium. In this study, type 2 diabetes mellitus (T2DM) was induced in rats. One group of animals was pre-treated with metformin (200 mg/kg) prior to diabetes induction and subsequently kept on metformin until sacrifice at week 12. Cardiac injuries developed in the diabetic rats as demonstrated by a significant (p < 0.0001) inhibition of desmin immunostaining, profound sarcomere ultrastructural alterations, substantial damage to the left ventricular tissue, collagen deposition, and abnormal ECG recordings. DM also significantly induced the cardiac expression of inducible nitric oxide synthase (iNOS), mammalian target of rapamycin (mTOR), and the profibrogenic biomarker tissue inhibitor of metalloproteinase-1 (TIMP-1). The expression of all these markers was significantly inhibited by metformin. In addition, a significant (p < 0.0001) correlation between desmin tissue levels/sarcomere damage and glycated hemoglobin, heart rate, iNOS, mTOR, and fibrosis was observed. These findings demonstrate an association between damage of the cardiac contractile unit­desmin and sarcomere­and the iNOS/mTOR/TIMP-1/collagen axis of fibrosis in T2DM-induced cardiomyopathy, with metformin exhibiting beneficial cardiovascular pleiotropic effects.

The ameliorative effect of kaempferol against CdCl2- mediated renal damage entails activation of Nrf2 and inhibition of NF-kB.

This study evaluated the nephroprotective effect of kaempferol against cadmium chloride (CdCl2) -induced nephropathy in rats. It also investigated if activation of Nrf2 is a common mechanism of action. Adult male rats ((150 ± 15 g) were divided into 4 groups (n = 8/each) as a control (1% DMSO, orally), control + kaempferol (200 mg/kg, orally), CdCl2 (50 mg/l in drinking water), and CdCl2 + kaempferol (200 mg/kg)-treated rats. All treatments were conducted for 8 weeks. Kaempferol significantly attenuated CdCl2-induced weight loss, reduction in kidney weights, and the injury in the glomeruli, proximal tubules, and distal tubules in the treated rats. It also significantly lowered serum levels of urea and creatinine, increased urine output and urinary creatinine levels and clearance but reduced urinary levels of albumin urinary albumin exertion (UAER), and urinary albumin/creatinine ratio (UACR) in these rats. In parallel, kaempferol downregulated renal levels of cleaved caspase-3 and Bax and unregulated those of Bcl2. In the kidney tissues of the control animals and CdCl2 rats, kaempferol significantly attenuated oxidative stress, inflammation and significantly boosted levels of manganese superoxide dismutase and glutathione. Also, and in both groups, kaempferol suppressed the nuclear levels of NF-κB p65, downregulated Keap1, and stimulated the nuclear activation and protein levels of Nrf2. In conclusion, kaempferol is a potential therapeutic drug to prevent CdCl2-induced nephropathy due to its anti-inflammatory and anti-oxidant effects mediated by suppressing NF- NF-κB p65 and transactivating Nrf2.

Suppression of glomerular damage and apoptosis and biomarkers of acute kidney injury induced by acetaminophen toxicity using a combination of resveratrol and quercetin.

Acute renal failure induced by a toxic dose of acetaminophen (also known as paracetamol, or APAP) is common in both humans and experimental animal models. Glomerular ultrastructural alterations induced by APAP overdose associated with the suppression of biomarkers of kidney injury have not been investigated before. Also, we investigated whether the combined polyphenolic antioxidants and anti-inflammatory compounds, resveratrol (RES) and quercetin (QUR) can protect against APAP-induced nephrotoxicity. Rats either received a single dose of APAP (2 g/kg) before being sacrificed after 24 hours or were pretreated for 7 days with combined doses of RES (30 mg/kg) and QUR (50 mg/kg) before being given a single dose of APAP and then sacrificed 24 hours post APAP ingestion. APAP significantly (p < 0.05) increased blood levels of urea, creatinine, malondialdehyde (MDA), interleukin-6 (IL-6), and tumor necrosis factor-alpha (TNF-α), which were effectively reduced by RES + QUR. In addition, APAP overdose induced the tissue expression of the apoptotic biomarker, p53, and caused profound kidney damage as demonstrated by substantial alterations to the glomerular basement membrane, podocytes, endothelial cells, widening of Bowman's space, and vacuolation of the cells lining the parietal layer, which were substantially protected by RES + QUR. Furthermore, a significant (p < 0.0001) positive correlation was observed between either glomerular basement membrane or podocyte foot processes and these parameters, urea, creatinine, MDA, and TNF-α. Thus, we conclude that APAP induces alterations to the glomerulus ultrastructure, which is protected by resveratrol plus quercetin, which also reduces blood levels of urea and creatinine, and biomarkers of oxidative stress and inflammation.

Suppression of thioacetamide-induced hepatic injury in rats treatment with resveratrol: role of mammalian target of rapamycin (mTOR) cell signaling / Supresión de la lesión hepática inducida por tioacetamida en ratas tratadas con resveratrol: rol de la vía de señalización mTOR (mammalian target of rapamycin)

Chronic hepatotoxicity is a debilitating and frequently life-threatening disease resulting in progressive liver failure. The toxic chemical, thioacetamide (TAA) is used to evaluate hepatoprotective agents, and the polyphenolic compound, resveratrol was proposed as a novel treatment for diseases with hyperactivation of the mammalian target of rapamycin (mTOR) cell signaling pathway. This analysis sought to investigate the potential protective effect of resveratrol against liver injury induced by TAA via the inhibition of hepatic mTOR. Model group rats received several injections of TAA (200 mg/kg; twice a week for 8 weeks) before being sacrificed at week 10 and the protective group was pretreated with resveratrol (20 mg/kg) daily for two weeks prior to TAA injections and continued receiving both agents until the end of the experiment. Harvested liver tissues were examined using light microscopy and liver homogenates were assayed for biomarkers of inflammation and assessed the levels of mTOR protein in all animal groups. In addition, blood samples were assayed for biomarkers of liver injury enzyme. TAA substantially damaged the hepatic tissue of the model group such as infiltration of inflammatory cells, vacuolated cytoplasm, dark pyknotic nuclei, and dilated congested blood vessel that were effectively protected by resveratrol. Resveratrol also significantly (p<0.05) inhibited TAA-induced mTOR, high sensitivity c-reactive protein (hs-CRP), tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), alanine aminotransferase (ALT) and aspartate aminotransferase (AST) in harvested liver homogenates and blood samples. Thus, we conclude that resveratrol effectively protects against TAA-induced hepatotoxicity in rats, possibly due to the inhibition of mTOR and inflammation.

La hepatotoxicidad crónica es una enfermedad debilitante y potencialmente mortal que produce insuficiencia hepática progresiva. La toxicidad del químico de la tioacetamida (TAA) se utiliza para evaluar los agentes hepatoprotectores y el compuesto polifenólico, resveratrol, se propuso como un nuevo tratamiento para enfermedades con hiperactivación de la vía de señalización celular mTOR (mammalian Target of Rapamycin). Aquí buscamos investigar el posible efecto protector del resveratrol contra la lesión hepática inducida por TAA a través de la inhibición de la vía de señalización mTOR en hepatocitos. Las ratas del grupo modelo recibieron varias inyecciones de TAA (200 mg / kg; dos veces por semana durante 8 semanas) antes de ser sacrificadas en la semana 10 y el grupo protector se trató previamente con resveratrol (20 mg / kg) diariamente durante dos semanas antes de las inyecciones de TAA y continuó recibiendo ambos agentes hasta el final del experimento. Se examinaron los tejidos hepáticos recolectados usando microscopía óptica y se analizaron los homogeneizados hepáticos para detectar biomarcadores de inflamación y se evaluaron los niveles de proteína mTOR en todos los grupos de animales. Además, se analizaron muestras de sangre para detectar biomarcadores de la enzima de lesión hepática. TAA dañó sustancialmente el tejido hepático del grupo modelo, con infiltración de células inflamatorias, citoplasma vacuolado, núcleos picnóticos oscuros y vasos sanguíneos congestionados dilatados que estaban efectivamente protegidos por el resveratrol. El resveratrol también inhibió significativamente (p <0.05) mTOR, proteína C-reactiva (hs-CRP), factor de necrosis tumoral alfa (TNF-α), interleucina-6 (IL-6), alanina aminotransferasa (ALT ) y aspartato aminotransferasa (AST) en las muestras de sangre y de hígados recolectados. En conclusión, el resveratrol protege eficazmente contra la hepatotoxicidad inducida por TAA en ratas, posiblemente debido a la inhibición de mTOR y de la inflamación.

Acetaminophen induces alterations to the renal tubular ultrastructure in a rat model of acute nephrotoxicity protected by resveratrol and quercetin / El acetaminofeno induce alteraciones en la ultraestructura tubular renal en un modelo de rata con nefrotoxicidad aguda protegida por resveratrol y quercetina

Acetaminophen (also called paracetamol, or APAP) induced nephrotoxicity is reported after accidental or intentional ingestion of an overdose of the drug. Renal tubular ultrastructural alterations induced by APAP overdose associated with the induction of biomarkers of kidney injury have not been investigated before. Also, we investigated whether the combined polyphenolic anti-inflammatory and antioxidants agents, resveratrol (RES) and quercetin (QUR) can protect against APAP-induced acute kidney injury. The model group of rats received a single dose of APAP (2 g/kg), whereas the protective group of rats was pre-treated for 7 days with combined doses of RES (30 mg/kg) and QUR (50 mg/kg) before being given a single dose of APAP. All rats were then sacrificed one day post APAP ingestion. Harvested kidney tissues were prepared for transmission electron microscopy (TEM) staining and blood samples were assayed for urea, creatinine, and biomarkers of inflammation and oxidative stress. TEM images and blood chemistry analysis showed that APAP overdose induced kidney damage as demonstrated by substantial alterations to the proximal convoluted tubule ultrastructure, and a significant (p<0.05) increase in urea, creatinine, tumor necrosis factor-alpha (TNF-a), and malondialdehyde (MDA) blood levels, which were protected by RES+QUR. These findings indicate that APAP induces alterations to the renal tubular ultrastructure, which is inhibited by resveratrol plus quercetin, which also decreases blood levels of kidney injury biomarkers.

El objetivo de este trabajo fue estudiar la nefrotoxicidad inducida por acetaminofeno (también llamado paracetamol o APAP) después de la ingestión accidental o intencional de una sobredosis de la droga. Las alteraciones ultraestructurales tubulares renales inducidas por sobredosis de APAP asociadas con la inducción de biomarcadores de daño renal no se han investigado. Además, estudiamos si los agentes combinados antiinflamatorios y antioxidantes polifenólicos, el resveratrol (RES) y la quercetina (QUR) pueden proteger contra la lesión renal aguda inducida por APAP. El grupo modelo de ratas recibió una dosis única de APAP (2 g / kg), mientras que el grupo protector de ratas se trató previamente durante 7 días con dosis combinadas de RES (30 mg / kg) y QUR (50 mg / kg) antes de recibir una dosis única de APAP. Todas las ratas se sacrificaron un día después de la ingestión de APAP. Los tejidos renales fueron preparados para el análisis a través de la microscopía electrónica de transmisión (MET). En las muestras de sangre se determinaron la urea, creatinina y los biomarcadores de inflamación y estrés oxidativo. Las imágenes MET y el análisis químico de la sangre mostraron que la sobredosis de APAP inducía daño renal, como lo demuestran las alteraciones sustanciales en la ultraestructura del túbulo contorneado proximal, y además, de un aumento significativo (p <0,05) de la urea, creatinina, factor de necrosis tumoral alfa y niveles sanguíneos de malondialdehído, protegidos por RES + QUR. Estos hallazgos indican que APAP induce alteraciones en la ultraestructura tubular renal, inhibida por el resveratrol más quercetina, que también disminuye los niveles sanguíneos de biomarcadores de daño renal.

Suppression of hepatic apoptosis induced by acetaminophen using a combination of resveratrol and quercetin: an association of oxidative stress and interleukin-11 / Suspensión de la apoptosis hepática inducida por acetaminofén mediante una combinación de resveratrol y quercetina: una asociación de estrés oxidativo e interleucina-1

We sought to determine whether the combined polyphenolic compounds, resveratrol and quercetin can substantially protect against modulation of hepatic biomarkers of apoptosis and survival, p53-Bax axis and B-cell lymphoma 2 (Bcl-2) in an animal model of acetaminophen-induced acute liver injury via the association of oxidative stress and interleukin-11 (IL-11). The model group of rats received a single dose of acetaminophen (2 g/kg), whereas the protective group of rats was pre-treated for 7 days with combined doses of resveratrol (30 mg/kg) and quercetin (50 mg/kg) before being given a single dose of acetaminophen. All rats were then sacrificed 24 hours post acetaminophen ingestion. Acetaminophen overdose induced acute liver injury as demonstrated by profound liver parenchymal damage and increased levels of the liver injury enzymes, alanine aminotransferase (ALT) and aspartate aminotransferase (AST). Acetaminophen significantly (p<0.05) modulated malondialdehyde (MDA), p53, apoptosis regulator Bax, Bcl-2, IL-11, interleukin-6 (IL-6), ALT, AST, superoxide dismutase (SOD), and glutathione peroxidase (GPx), which were significantly protected by resveratrol plus quercetin. We further demonstrated a significant (p<0.01) correlation between IL-11 scoring and the levels of p53, Bax, Bcl-2, and MDA. Thus, resveratrol plus quercetin effectively protect against acetaminophen-induced apoptosis, which is associated with the inhibition of oxidative stress and IL-11.

En el estudio se intentó determinar si los compuestos polifenólicos combinados, el resveratrol y la quercetina pueden proteger sustancialmente contra la modulación de los biomarcadores hepáticos de apoptosis y supervivencia, el eje p53-Bax y el linfoma de células B 2 (Bcl-2) en un modelo animal de lesión hepática aguda inducida por acetaminofén, a través de la asociación del estrés oxidativo y la interleucina-11 (IL-11). El grupo modelo de ratas recibió una dosis única de acetaminofén (2 g / kg), mientras que el grupo protector de ratas fue tratado durante 7 días con dosis combinadas de resveratrol (30 mg / kg) y quercetina (50 mg / kg) antes de recibir una dosis única de acetaminofén. Todas los animales fueron sacrificados 24 horas después de la ingestión de acetaminofén. La sobredosis de acetaminofén indujo una lesión hepática aguda, como se observó en el daño profundo del parénquima hepático y el aumento de los niveles de las enzimas en la lesión hepática, alanina aminotransferasa (ALT) y aspartato aminotransferasa (AST). Acetaminofén moduló significativamente (p <0.05) malondialdehído (MDA), p53, regulador de apoptosis Bax, Bcl2, IL-11, interleucina-6 (IL-6), ALT, AST, superóxido dismutasa (SOD) y glutatión peroxidasa ( GPx), los que se encontraron significativamente protegidos por el resveratrol y quercetina. Además se determinó una correlación significativa (p <0.01) entre la puntuación de IL-11 y los niveles de p53, Bax, Bcl-2 y MDA. En conclusión, el resveratrol más la quercetina protegen de manera efectiva contra la apoptosis inducida por acetaminofén, asociada con la inhibición del estrés oxidativo y la IL-11.

Exercise augments the modulatory effects of vitamin E on pre-diabetes-induced aortopathy: a potential role of adiponectin.

Background: We tested the hypothesis that vitamin E may protect against pre-diabetes-induced aortic injury (aortopathy), and exercise can augment the action of vitamin E.Material and methods: Rats were either fed with a high fat and fructose diet (HFD) (model group) or a standard laboratory chow (control group) for 15 weeks before being sacrificed. The three protective groups were treated with vitamin E (HFD + Vit E), swimming exercises (HFD + Ex), and vitamin E plus swimming exercises (HFD + VitE + Ex), respectively.Results: Aortopathy was developed in the model group as demonstrated by substantial tissue ultrastructural alterations, which were partially protected by vitamin E and effectively protected with vitamin E plus swim exercise. Also, swimming exercises significantly (p < .05) increased the modulatory effects of vitamin E on dyslipidemia, insulin resistance, blood pressure, oxidative stress, inflammation, leptin, and adiponectin, except coagulation and thrombosis.Conclusions: Swim exercise augments the protective effects of vitamin E in a pre-diabetic animal model.

Suppression of monosodium glutamate-induced acute kidney injury and renal ultrastructural damage in rats by vitamin E / Supresión de lesiones renales agudas inducidas por glutamato monosódico y daño ultraestructural renal en ratas por vitamina E

Food additives and flavour enhancers used in the food industry are potential health risks. We tested the hypothesis that the food additive and flavour enhancer, monosodium glutamate (MSG), which is the sodium salt of glutamic acid can induce ultrastructural alterations to the kidney, and the antioxidant vitamin E can protect against acute kidney injuries induced by a toxic dose of MSG in a rat model of the disease. The model group of rats received a daily dose of MSG (4 gm/kg) for 7 days, whereas the protective groups were either received a 100 mg/kg vitamin E plus MSG or 300 mg/kg vitamin E plus MSG for 7 days. Rats were then sacrificed on day 8. Transmission and light microscopy images revealed substantial kidney damage induced by MSG in the model group as demonstrated by degenerated epithelial cells with Pyknotic nuclei, swollen mitochondria, damaged brush margins, dilated tubules, and widening of Bowman's space with shrinkage and deformity of some glomeruli. Treatment of the model group with vitamin E showed a substantial protection of kidney tissue and renal ultrastructure by 300 mg/kg vitamin E compared to a partial protection by 100 mg/kg vitamin E. In addition, MSG significantly (p<0.05) increased serum levels of urea and creatinine, which were significantly (p<0.05) decreased with vitamin E. However, for serum creatinine, high doses of vitamin E (300 mg/kg) were more effective than lower doses (100 mg/kg) of vitamin E. These results indicate that vitamin E at 300 mg/kg effectively protects against MSG-induced acute kidney injury in rats.

Los aditivos alimentarios y los potenciadores del sabor utilizados en la industria alimentaria son riesgos potenciales para la salud. Probamos la hipótesis de que el aditivo alimentario y el potenciador del sabor, glutamato monosódico (MSG), la sal sódica del ácido glutámico, puede inducir alteraciones ultraestructurales del riñón, y que las propiedades antioxidantes de la vitamina E, pueden proteger contra las lesiones renales inducidas por una dosis tóxica de MSG en un modelo de rata. El grupo modelo de ratas recibió una dosis diaria de MSG (4 g / kg) durante 7 días, mientras que los grupos protectores recibieron una dosis de 100 mg / kg de vitamina E más MSG o 300 mg / kg de vitamina E más MSG durante 7 días. Las ratas se sacrificaron el día 8. Las imágenes de microscopía óptica y de transmisión revelaron un daño renal sustancial inducido por el MSG en el grupo modelo, como lo demuestran las células epiteliales degeneradas con núcleos picnóticos, mitocondrias hinchadas, bordes dañados, túbulos dilatados y ensanchamiento del espacio de Bowman, además de la deformidad de algunos glomérulos. El tratamiento del grupo modelo con vitamina E mostró una protección sustancial del tejido renal y la ultraestructura renal de 300 mg / kg de vitamina E en comparación con una protección parcial de 100 mg / kg de vitamina E. Además, el MSG aumentó significativamente (p <0,05) en el suero los niveles de urea y creatinina, disminuyeron significativamente (p <0,05) con la vitamina E. Sin embargo, para la creatinina sérica, las dosis altas de vitamina E (300 mg / kg) fueron más efectivas que las dosis más bajas (100 mg / kg) de vitamina E. Estos resultados indican que la vitamina E a 300 mg / kg protege eficazmente contra la lesión renal aguda inducida por MSG en ratas.

Metformin suppresses aortic ultrastrucural damage and hypertension induced by diabetes: a potential role of advanced glycation end products.

Cardiovascular disease secondary to diabetes represents a significant challenge to the health community. The advanced glycation end products (AGEs) play an important role in diabetes-mediated vascular injury. We tested whether metformin can suppress aortic AGEs production and protect against aortic injuries (aortopathy) and hypertension in streptozotocin-induced type 2 diabetes mellitus (T2DM) animal model. T2DM was induced in rats two weeks after being fed on a high carbohydrate and fat diet (HCFD), and continued on a HCFD until being sacrificed at week 12 (model group). The protective group was put on metformin two weeks before diabetic induction and continued on metformin and HCFD until the end of the experiment, at week 12. Using electron microscopy examinations, we observed in the model group substantial damage to the ultrastructure of aortic endothelial and vascular smooth muscle layers as demonstrated by markedly distorted vacuolated endothelial and vascular smooth muscle cells with pyknotic nuclei detached from the underlying basement membrane, which were protected by metformin. Also, metformin significantly (p < .05) decreased both systolic and diastolic blood pressure, aortic levels of AGEs, and blood levels of oxidative stress and inflammatory biomarkers. We conclude that metformin protects against T2DM-induced aortopathy and hypertension, possibly via the inhibition of AGEs, inflammation, and oxidative stress.

Vitamin E ameliorates alterations to the articular cartilage of knee joints induced by monoiodoacetate and diabetes mellitus in rats.

We recently reported an animal model of osteoarthritis (OA) induced by a combination of the chondrocyte glycolysis inhibitor, monoiodoacetate (MIA) and the agent that induces diabetes mellitus, streptozotocin (STZ). Here we investigated the potential protective effect of the antioxidant and anti-inflammatory agent, vitamin E against MIA+STZ-induced OA. Therefore, rats were either injected once with MIA (2 mg/50 µL) + 65 mg/kg STZ before being sacrificed after 8 weeks (model group) or were treated immediately after MIA+STZ injections with vitamin E (600 mg/kg; thrice a week) before being sacrificed after 8 weeks (treatment group). Using scanning and transmission electron microscopy examinations, we observed in the model group a substantial damage to the articular cartilage of the knee joint as demonstrated by the destruction of the chondrocytes, territorial matrix, disrupted lacunae, collagen fibers, and profound chondrocyte ultrastructural alterations such as degenerated chondrocyte, irregular cytoplasmic membrane, damaged mitochondria and rough endoplasmic reticulum, vacuolated cytoplasm, presence of lipid droplets and different sizes of lysosomes, which were substantially but not completely protected by vitamin E. H&E stained sections of knee joint articular cartilage showed that MIA+STZ induced damage to the chondrocyte and territorial matrix. Vitamin E also significantly (p < .05) inhibited MIA+STZ-induced blood levels of the inflammatory biomarkers, tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) that are known to be modulated in OA and diabetes. We conclude that vitamin E protects against MIA+STZ-induced knee joints injuries in rats, which is associated with the inhibition of biomarkers of inflammation.

Melatonin Improves Memory Deficits in Rats with Cerebral Hypoperfusion, Possibly, Through Decreasing the Expression of Small-Conductance Ca2+-Activated K+ Channels.

This study investigated the expression pattern, regulation of expression, and the role of hippocampal small-conductance Ca2+-activated K+ (SK) channels in memory deficits after cerebral hypoperfusion (CHP) with or without melatonin treatment, in rats. Adults male Wistar rats (n = 20/group) were divided into (1) a sham (2) a sham + melatonin (3) a two-vessel occlusion (2-VO) model, and (4) a 2-VO + melatonin. Melatonin was administered (i.p.) to all rats at a daily dose of 10 mg kg-1 for 7 days starting at the time of 2-VO-induction. In contrast to 2-VO rats, melatonin increased the latency of the passive avoidance learning test and decreased time to find the hidden platform in Water Morris Test in all tested rats. In addition, it concomitantly downregulated SK1, SK2, and SK3 channels, downregulated mRNA levels of TNFα and IL-1ß, enhanced BDNF levels and activity of PKA levels, and restored the levels of cholinergic markers in the hippocampi of the treated-rats. Mechanistically, melatonin significantly prevented CHP-induced activation of ERK1/2, JNK, and P38 MAPK at least by inhibiting ROS generation and enhancing the total antioxidant potential. In cultured hypoxic hippocampal neurons, individual blockage of MAPK signaling by the MEK1/2 inhibitor (U0126), but not by the P38 inhibitor (SB203580) or JNK inhibitor (SP600125), completely prevented the upregulation of all three kinds of SK channels. These data clearly confirm that upregulation of SK channels plays a role in CHP-induced memory loss and indicate that melatonin reverses memory deficits after CHP in rats, at least by, downregulation of SK1, SK2, and SK3 channels in their hippocampi.

Insulin Suppresses Type 1 Diabetes Mellitus-Induced Ventricular Cardiomyocyte Damage Associated with the Inhibition of Biomarkers of Inflammation and Oxidative Stress in Rats.

AIMS: We sought to determine whether insulin can protect against type 1 diabetes mellitus (T1DM)-induced cardiac ultrastructural alterations in an animal model of the disease. This has not been investigated before. METHODS: Rats were either injected once with 65 mg/kg streptozotocin (STZ) before being sacrificed after 8 weeks or were treated with a daily injection of insulin 2 days by STZ and continued until being sacrificed. RESULTS: Harvested tissues obtained from left ventricles in the untreated T1DM rats showed substantial damage to the cardiomyocyte ultrastructure as demonstrated by disintegrated myofibrils and their sarcomeres, damaged mitochondria and lipid droplets, which was substantially protected by insulin. Insulin also significantly inhibited T1DM-induced hyperglycemia (p < 0.001), dyslipidemia (p < 0.0001), malondialdehyde (MDA; p < 0.0001), tumor necrosis factor-alpha (TNF-α; p < 0.001) and interleukin-6 (p < 0.001). We further demonstrated a significant (p ≤ 0.001) correlation between either sarcomere or mitochondrial injury scoring and the serum levels of glucose, dyslipidemia, and biomarkers of oxidative stress (OxS) and inflammation. CONCLUSIONS: These results indicate that insulin effectively suppresses left ventricular cardiomyocyte ultrastructural damage, which substantially slows down the progression of diabetic cardiomyopathy for 8 weeks in a rat model of T1DM, possibly due to the glycemic control and inhibition of dyslipidemia, OxS and inflammation.

Pre-diabetes induces ultrastructural alterations in the large blood vessel aorta in rats / La prediabetes induce alteraciones ultraestructurales en la aorta de los grandes vasos sanguíneos en ratas

Excessive consumption of carbohydrate and fat increases the risk of cardiovascular disease. We sought to determine the potential ultrastructural alterations in large blood vessels induced by a high fat and fructose diet (HFD) in a rat model of prediabetes. Rats were either fed with HFD (model group) or a standard laboratory chow (control group) for 15 weeks before being sacrificed. The harvested thoracic aorta tissues were examined using transmission electron microscopy (TEM), and blood samples were assayed for biomarkers of pre-diabetes.TEM images showed that HFD induced profound pathological changes to the aortic wall layers, tunica intima and tunica media ultrastructures in the pre-diabetic rats as shown by apoptotic endothelial cells with pyknotic nuclei, damaged basal lamina, deteriorated smooth muscle cells that have irregular plasma membranes, shrunken nucleus with clumped nuclear chromatin, damaged mitochondria and few cytoplasmic lipid droplets and vacuoles. In addition, HFD significantly (p<0.05) decreased adiponectin and increased biomarkers of lipidemia, glycaemia, inflammation, oxidative stress, vascular injury such as soluble intercellular adhesion molecule-1 (sICAM-1), soluble vascular cell adhesion protein 1 (sVCAM-1), endothelin-1 (ET-1), and coagulation and thrombosis such as Von Willebrand factor (vWF), and plasminogen activator inhibitor-1 (PAI-1), compared to normal levels of these parameters in the control group. Thus, we demonstrated that feeding rats with a HFDisable to develop a pre-diabetic animal model that is useful to study the aortic ultrastructural alterations.

El consumo excesivo de carbohidratos y grasas aumenta el riesgo de enfermedades cardiovasculares. Intentamos determinar las posibles alteraciones ultraestructurales en los grandes vasos sanguíneos, inducidas por una dieta alta en grasas y fructosa (HFD) en un modelo de rata de prediabetes. Las ratas se alimentaron con HFD (grupo modelo) o una comida de laboratorio estándar (grupo de control) durante 15 semanas antes de ser sacrificadas. Los tejidos de la aorta torácica recolectados se examinaron mediante microscopía electrónica de transmisión (TEM) y las muestras de sangre se analizaron para detectar biomarcadores de prediabetes. Las imágenes TEM mostraron que HFD indujo cambios patológicos profundos en las capas de la pared aórtica, túnica íntima y túnica media en la ratas pre-diabéticas como lo muestran las células endoteliales apoptóticas con núcleos picnóticos, lámina basal dañada, células musculares lisas deterioradas que tienen membranas plasmáticas irregulares, núcleo encogido con cromatina nuclear aglomerada, mitocondrias dañadas y pocas gotitas lipídicas citoplásmicas y vacuolas. Además, HFD presentó disminución significativa de adiponectina (p <0,05), y aumento de biomarcadores de lipidemia, glucemia, inflamación, estrés oxidativo, lesión vascular como la molécula de adhesión intercelular soluble 1 (sICAM-1), proteína de adhesión de células vasculares soluble 1 (sVCAM-1), endotelina 1 (ET-1), y la coagulación y la trombosis, como el factor de Von Willebrand (vWF), y el inhibidor del activador del plasminógeno-1 (PAI -1), en comparación con los niveles normales de estos parámetros en el grupo de control. Por tanto, la alimentación de ratas con HFD es capaz de desarrollar un modelo animal prediabético que es útil para estudiar las alteraciones ultraestructurales aórticas.

Swim exercise inhibits hemostatic abnormalities in a rat model of obesity and insulin resistance.

BACKGROUND: We sought to determine whether swim exercise can inhibit high carbohydrate and fat diet (HCFD)-induced biomarkers of coagulation and thrombosis. MATERIAL AND METHODS: Rats were either fed with HCFD (model group) or a standard laboratory chow (control group) for 15 weeks. Swim exercise-'treated' rats started swim exercise training from the 11th week until being sacrificed, on Week 15. RESULTS: HCFD caused a significant increase in blood glucose, insulin resistance (HOMA-IR), lipidemia, and inflammatory biomarkers. In addition, HCFD significantly modulated coagulation and thrombosis biomarkers; fibrinogen, plasminogen activator inhibitor-1, von Willebrand factor, prothrombin time, activated partial thromboplastin time, blood clotting and bleeding time, and ADP-induced platelet aggregation that was effectively inhibited by swimming exercises. CONCLUSIONS: We demonstrate that in an animal model of obesity and insulin resistance, there is a significant change in hemostasis, which is ameliorated by swim exercise.

Unacylated ghrelin stimulates steroidogenesis in lean rats and reverses reproductive dysfunction in high fat diet-fed rats.

This study investigated the effect of sub-chronic administration of unacylated ghrelin (UAG) on steroidogenesis, sperm parameter, and reproductive function in lean and high fat diet (HFD)-induced obese male rats. Rats were divided into 4 groups (n = 12 each) as 1) Control-fed standard diets (STD): (10 kcal%), 2) STD + UAG (200 ng/kg, i.p.), 3) HFD obese: fed HFD (45 kcal%), and 4) HFD + UAG. Diet regimen was continued for 16 weeks after which normal saline as a vehicle or UAG was administered to desired groups for the next 4 weeks. In vitro, testicular slices were incubated with increasing concentrations of UAG (10-8-10-6 M) in the presence or absence of GSH-R1a antagonist, [D-Lys-3]-GHRP-6 (10-6 M). UAG significantly increased the circulatory levels of FSH, LH and testosterone, increased testicular testosterone levels and sperm count and motility in lean and obese rats and reduced sperm morphological abnormalities and increased pregnancy rate and number of pups at birth in HFD-obese rats. Associated with the reduction in the final body and fat masses weights and independent of food intake, UAG post-therapy to both lean and HFD-fed rats significantly lowered fasting blood glucose and insulin levels, lowered HOMA-IR value, enhanced OGTT and ITT, lowered circulatory leptin levels, downregulated aromatase expression in adipose and testicular tissue and inhibited HFD-induced testicular oxidative stress and activation of cleaved caspase-3. Dysregulation of testicular levels of StAR, SF-1, CYP11A1 in the testis of both groups as well as in the in vitro preparation, in a dose-dependent manner, independent of GSH-R1a and not associated with activation of STAT3, a mediator of leptin signaling was apparent. In conclusion, administration of UAG can enhance reproductive function in lean rats and reverses HFD-induced reproductive dysfunction in obese rats. Abbreviations: AG: acylated ghrelin; BMI: body mass index; CHOL: cholesterol;FSH: follicular stimulating hormone; GHS: growth hormone secretagogues; GSH: reduced glutathione; HFD: high fat diet; HOMA-IR: homeostasis model assessment of insulin resistance: IR: insulin resistance; OGTT: oral glucose tolerance test; ITT: insulin tolerance test; LH: luteinizing hormone; MDA: malondialdehyde; STAT3: signal transducer and activator of transcription; SOD: superoxide dismutase; STD: standard diet; SF-1: steroidogenic factor-1; StAR: steroidogenic acute regulatory protein; CYP11A1: cholesterol side-chain cleavage enzyme (or P450scc); TGs: triglycerides; UAG: unacylated ghrelin.

Insulin protects against hepatocyte ultrastructural damage induced by type 1 diabetes mellitus in rats.

Diabetic complications that affect vital organs such as the heart and liver represent a major public health concern. The potential protective effects of the hormone insulin against hepatocyte ultrastructural alterations induced secondary to type 1 diabetes mellitus (T1DM) in a rat model of the disease have not been investigated before. Therefore, rats were injected once with 65 mg/kg streptozotocin (T1DM group) and the protection group (T1DM+Ins) received a daily injection of insulin 48 h post diabetic induction by streptozotocin and continued until being sacrificed at week 8. The harvested liver tissues were examined using transmission electron microscopy (TEM) and blood samples were assayed for biomarkers of liver injury enzyme, glycemia, lipidemia, inflammation, and oxidative stress. TEM images showed that T1DM induced profound hepatocyte ultrastructural alterations as demonstrated by pyknotic nucleus, condensation of chromatin, irregular nuclear membrane, swollen mitochondria, dilated rough endoplasmic reticulum, damaged intercellular space, and accumulation of few lipid droplets inside the hepatocyte cytoplasm, which were substantially protected with insulin. In addition, the blood chemistry profile complements the TEM data as demonstrated by an increase in serum levels of alanine aminotransferase (ALT), dyslipidemia, C-reactive protein (CRP), tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and malondialdehyde (MDA) by T1DM that were significantly (p < 0.05) reduced with insulin injections. Thus, we conclude that insulin effectively protects against T1DM-induced liver injury in rats for a period of 8 weeks, possibly due to the inhibition of inflammation, oxidative stress, and dyslipidemia.