8 research outputs found

    Effects of maslimic acid and related triterpene derivatives on kidney function of male Sprague-Dawley rats.

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    Ph. D. University of KwaZulu-Natal, Durban 2014.Reports indicate that hyperglycaemia leads to development of kidney complications which result in sodium retention, decrease in glomerular filtration rate (GFR) and high blood pressure. Various biochemical processes such as polyol pathway, AGEs formation are thought to gives rise to a development and progression of these complications. Clinical trials show that there is currently no commercially available compound that lowers blood glucose concentration while alleviating diabetic nephropathy (DN). Current methods involve the use of ACE blockers which are associated with side effects. Previous reports in our laboratories indicate that triterpene constituents of Syzygium spp. such as oleanolic acid (OA) possess hypoglycaemic and renoprotetive effects in STZ-induced diabetic rats. The important question is whether MA, a related triterpene also possesses the same properties. MA is a hydrophobic triterpene and we therefore synthesised derivatives to improve solubility, bioavailability and efficacy. Accordingly this study was designed to investigate the effects of MA and related triterpene derivatives on renal function of STZ-induced diabetic rats

    Mechanisms of Regulation of Proximal Tubule Sodium Transporters in Obesity-Induced Hypertension

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    Hypertension is one of the common complications of obesity. Using a rat model of diet induced obesity and hypertension we investigated some of the mechanisms that are involved in regulation of blood pressure in obesity. The first aim of this study was to determine the role of proximal tubule transporters on the renal sodium handling in obese hypertensive (OP) and lean normotensive (OR) rats. An acute increase in renal perfusion pressure resulted in a blunted natriuretic response in OP vs. OR rats and indicated that increased sodium reabsorption in the proximal tubule is casual, at least in part, for hypertension in OP rats. Subsequently, protein expression and activity of Na,K-ATPase and NHE3 were increased in obese rats compared to lean rats. Moreover, in OP rats more NHE3 was associated with its active pool located in the microvillus region. Together, these results suggest that hypertension in obese rats is characterized by the impaired pressure-natriuresis and diuresis that can be explained by the increased activity of proximal tubule sodium transporters. Previous results from our lab determined that peroxisome proliferator activated receptor Îł (PPARÎł) has reduced expression and activity in the kidney of OP vs. OR rats. Therefore, in the second aim we investigated the effect of PPARÎł ligand activation on expression and activity of proximal tubule Na+ transporters in OP and OR rats. In addition, by employing in vitro studies using proximal tubule epithelial cells, we determined whether pioglitazone exerts its effect via direct PPARÎł activation. Pioglitazone reduced systolic blood pressure in obese rats while having no effect in lean rats. However, it increased sodium retention in the lean group. Pioglitazone increased Na,K-ATPase activity in OP rats, while its protein expression was increased in both groups. In contrast, NHE3 activity was reduced in obese rats treated with pioglitazone and protein expression was decreased in both groups. Pioglitazone did not have an effect on NHE3 localization in obese rats, but in lean rats, it had tendency to redistribute NHE3 towards the more active membrane pool. In cells transiently transfected to overexpress or silence PPARÎł, we demonstrated that pioglitazone reduced Na,K-ATPase and NHE3 abundance via PPARÎł activation. Collectively, the results indicated that pioglitazone reduced blood pressure in the obese group most likely by decreasing activity of NHE3. However, other factors besides trafficking are involved in the transporter regulation. Pioglitazone did not reduce blood pressure in lean rats, suggesting that the metabolic milieu is an important determinant of the pioglitazone differential effect on the blood pressure and on the proximal tubule transporters. Nitric oxide (NO) plays an important role in regulating pressure natriuresis and diuresis and its availability seems to be altered in obese animals and humans. The third aim was designed to examine the role of NO on blood pressure, pressure natriuresis and expression of sodium transporters NHE3 and Na,K-ATPase in OP and OR rats. To determine the role of NO, we performed in vivo study using L-NAME for chronic NO inhibition. The NO inhibition did not change glomerular filtration rate in either of the groups. Natriuresis and diuresis was significantly decreased only in treated OR rats. Also, NHE3 protein expression and activity were significantly elevated in treated vs. non-treated OR rats, with no significant changes in OP rats. Moreover, L-NAME caused a shift of NHE3 to the active pool located in microvillus region in OR group only. In conclusion, normotensive OR rats are more susceptible to NO deficiency and the mechanism involves an increase in activity of NHE3 with the transporter redistribution playing a significant role. In addition, we investigated in cell culture whether hormones elevated in obesity can modulate Na,K-ATPase and NHE3 via cGMP production. In vitro experiments provided some evidence that angiotensin II and insulin interact with the NO signaling pathway at the level of cGMP. cGMP could affect transporter activity by phosphorylation which could account for the effects determined in vivo

    Rôle du système rénine-angiotensine intrarénal dans l’hypertension et les dommages rénaux chez les souris transgéniques diabétiques

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    Plusieurs expériences et études cliniques ont démontré que l’activation du système rénine-angiotensine (RAS) peut induire l’hypertension, un facteur de risque majeur pour les maladies cardiovasculaires et rénales. L’angiotensinogène (Agt) est l’unique substrat du RAS. Cependant, il n’a pas encore été démontré si l’activation du RAS intrarénal peut à elle seule induire des dommages rénaux, indépendamment de l’hypertension systémique, et ainsi jouer un rôle prépondérant dans la progression de la néphropathie diabétique. Afin d’explorer le rôle du RAS intrarénal dans les dommages rénaux, un diabète a été induit par l’injection de streptozotocin chez des souris transgéniques (Tg) surexprimant l’Agt de rat dans les cellules des tubules proximaux du rein (RPTC). Les souris Tg diabétiques ont été traitées soit avec des inhibiteurs du RAS (perindopril et losartan), de l’insuline ou une combinaison des deux pour 4 semaines avant d’être euthanasiées. Pour une autre étude, des souris Tg non-diabétiques ont été traitées soit avec des inhibiteurs du RAS, l’hydralazine (vasodilatateur) ou l’apocynine (inhibiteur de la NADPH oxydase) pour une période de 8 semaines avant l’euthanasie. Des souris non-Tg ont été utilisées comme contrôles. Des cellules immortalisées de tubule proximal de rat (IRPTC) transfectées de manière stable avec un plasmide contenant l’Agt ou un plasmide contrôle ont été employées comme modèle in vitro. Nos résultats ont démontré que les souris Tg présentaient une augmentation significative de la pression systolique, l’albuminurie, l’apoptose des RPTC et l’expression de gènes pro-apoptotiques par rapport aux souris non-Tg. Les mêmes changements ont été observés chez les souris Tg diabétiques par rapport aux souris non-Tg diabétiques. L’insuline et/ou les inhibiteurs du RAS ont permis d’atténuer ces changements, sauf l’hypertension qui n’était réduite que par les inhibiteurs du RAS. Chez les IRPTC transfectées avec l’Agt in vitro, les hautes concentrations de glucose augmentent l’apoptose et l’activité de la caspase-3 par rapport aux cellules contrôles et l’insuline et/ou les inhibiteurs du RAS empêchent ces augmentations. En plus des changements physiologiques, les RPTC des souris Tg présentent aussi une augmentation significative de la production des espèces réactive de l’oxygène (ROS) et de l’activité de la NADPH oxydase, ainsi qu’une augmentation de l’expression du facteur de croissance transformant-beta 1 (TGF-β1), de l’inhibiteur activateur du plasminogène de type 1 (PAI-1), des protéines de la matrice extracellulaire, du collagène de type IV et de la sousunité p47 de la NADPH oxydase. Le traitement des souris Tg avec l’apocynine et le perindopril a permis d’améliorer tous ces changements, sauf l’hypertension qui n’était pas corrigée par l’apocynine. D’autre part, l’hydralazine a prévenu l’hypertension, sans modifier l’albuminurie, l’apoptose des RPTC ou l’expression des gènes pro-apoptotiques. Ces résultats montrent bien que l’activation du RAS intrarénal et l’hyperglycémie agissent de concert pour induire l’albuminurie et l’apoptose des RPTC, indépendamment de l’hypertension systémique. La génération des ROS via l’activation de la NADPH oxydase induit en partie l’action du RAS intrarénal sur l’apoptose des RPTC, la fibrose tubulo-interstitielle et l’albuminurie chez les souris Tg. D’autre part, une expérience en cours a tenté d’encore mieux délimiter les effets de l’activation du RAS intrarénal, tout en éliminant la néphrotoxicité du STZ. Pour cette étude, les souris Tg surexprimant l’Agt de rat dans leurs RPTC ont été croisées aux souris Ins2Akita, un modèle spontané de diabète de type I, afin de générer des souris Akita-rAgt-Tg. Les résultats préliminaires indiquent que le RAS intrarénal est activé dans les souris Akita et que la combinaison avec l’hyperglycémie induit du stress du réticulum endoplasmique (ER) dans les RPTC in vivo. Le stress du ER contribue à l’apoptose des RPTC observée dans le diabète, à tout le moins dans le modèle Akita. Le traitement avec des inhibiteurs du RAS permet d’atténuer certains des dommanges rénaux observés dans les souris Akita-rAgt-Tg.Experimental and clinical studies have shown that renin-angiotensin system (RAS)activation may lead to hypertension, a major cardiovascular and renal risk factor. Angiotensinogen (Agt) is the sole substrate of the RAS. However, it is unclear whether intrarenal RAS activation alone could induce kidney injury independently of systemic hypertension and play an important role in the progression of diabetic nephropathy (DN). To explore the role of intrarenal RAS in kidney injury, transgenic (Tg) mice overexpressing rat Agt in their renal proximal tubular cells (RPTCs) were rendered diabetic by streptozotocin (STZ). Diabetic Tg mice were treated with RAS blockers (perindopril and losartan), insulin or a combination of both and then euthanized after 4 weeks of treatment. In a separate study, non-diabetic Tg mice were treated with RAS blockers or hydralazine (a vasodilator) or apocynin (an NADPH oxidase inhibitor) and then euthanized after 8 weeks of treatment. Non-Tg littermates served as controls in both studies. Immortalized rat proximal tubule cells (IRPTCs) stably transfected with Agt cDNA or control plasmid were used in the experiments as an in vitro model. Our results showed that non-diabetic Tg mice displayed a significant increase in systolic blood pressure (SBP), albuminuria, RPTC apoptosis, and proapoptotic gene expression. Diabetic Tg mice had a further increase of albuminuria, RPTC apoptosis, and proapoptotic gene expression, though the SBP of the diabetic Tg mice was similar to that of non-diabetic Tg mice. RAS blockers and/or insulin treatments markedly attenuated these changes, except that insulin had no impact on hypertension. In vitro, high-glucose melieu significantly increased apoptosis and caspase-3 activity in Agt stable transfectants compared to control cells, and these changes were attenuated by insulin and/or RAS blockers. Furthermore, non-diabetic Tg mice showed significantly elevated reactive oxygen species (ROS) production and NADPH oxidase activity, as well as enhanced expression of transforming growth factor-beta 1 (TGF-β1), plasminogen activator inhibitor-1 (PAI-1), extracellular matrix proteins, collagen type IV, and NADPH oxidase subunit p47 in their RPTC. Treatment with apocynin and perindopril ameliorated these changes, but apocynin had no effect on SBP. In contrast, hydralazine prevented hypertension but not albuminuria, RPTC apoptosis, or proapoptotic gene expression. These data indicate that intrarenal RAS activation and hyperglycemia act in concert to induce albuminuria and RPTC apoptosis independent of systemic hypertension. ROS generation via NADPH oxidase activation mediates, at least in part, intrarenal RAS action on RPTC apoptosis, tubulointerstitial fibrosis and albuminuria in Tg mice. On the other hand, in an on-going experiment, to avoid the nephro-toxic effects of STZ and further delineate the effects of intrarenal RAS activation, Tg mice overexpressing rat Agt in their RPTCs were crossbred with Ins2Akita mice, a spontaneous type I diabetes model, to generate Akita-rAgt-Tg mice. Preliminary data indicated that hyperglycaemia and intrarenal RAS activation induced endoplasmic reticulum (ER) stress in RPTC in vivo, and the ER stress pathway contributed to RPTC apoptosis in diabetes, at least in the Akita model. RAS blockade was effective in attenuating some parameters of renal injury in AkitarAgt-Tg mice

    Ubiquitination in Health and Diseases

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    Ubiquitination is a biological process mediated by ubiquitin itself, the E1 ubiquitin-activating enzyme, E2 ubiquitin-conjugating enzyme, E3 ubiquitin ligase, and deubiquitinating enzyme, respectively. Currently, these multiple biological steps are revealed to participate in various life phenomena, such as cell proliferation, regulation of cell surface proteins expression, and mitochondrial function, which are profoundly related to human health and diseases. Although clinical applications targeting ubiquitination are still limited compared to those directed toward kinase systems such as tyrosine kinases, multiple enzymatic consequences should be future therapeutic implications. This Special Issue of IJMS entitled “Ubiquitination in Health and Disease” successfully published15 distinguished manuscripts, with a total of 66 international authors and. This book provides the latest and most useful information for researchers and scientists in this field
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