Adriana Izquierdo

Role of the Renin-Angiotensin System on the Parathyroid Hormone–Related Protein Overexpression Induced by Nephrotoxic Acute Renal Failure in the Rat

Parathyroid hormone-related protein (PTHrP), a mitogenic factor for renal cells, is overexpressed in acute renal failure (ARF). Recent data support an association between PTHrP and the renin-angiotensin system in the damaged kidney. The effects of angiotensin II (Ang II) inhibitors (quinapril, enalapril, and/or losartan) on PTHrP and the PTH1 receptor (PTH1R) expression in rats with either folic acid (FA)- or gentamicin-induced ARF were analyzed. The decreased renal function and the PTHrP upregulation and PTH1R downregulation induced by the nephrotoxins were inhibited by the Ang II blockers. In tubuloepithelial cells NRK-52E, the rapid (10 min) increase in PTHrP mRNA by FA, associated with a perinuclear relocalization of Ang II/AT1 receptor, was inhibited by losartan but not candesartan, which traps Ang II receptors at the cell surface. Maximal PTHrP protein overexpression by FA (at 24 to 72 h)-or by exogenous Ang II-was abolished by both Ang II antagonists. PTHrP upregulation by FA was preceded by increased extracellular signal-regulated kinase (ERK) phosphorylation and inhibited by the ERK inhibitor PD098059. FA also activated cAMP response element-binding (CREB) protein, and this was prevented by losartan in these cells. Moreover, PTHrP mRNA overexpression by either FA or Ang II occurred in NRK 52E that were transfected with a CREB construct but not the dominant-negative CREB133 construct. These findings demonstrate that the decreased renal function and PTHrP overexpression in nephrotoxin-damaged kidney depends on renin-angiotensin system. In this setting, intracellular Ang II/AT1 receptor recycling seems to be related to PTHrP induction through ERK and CREB activation in tubuloepithelial cells.

Accelerated renal disease is associated with the development of metabolic syndrome in a glucolipotoxic mouse model

SUMMARY Individuals with metabolic syndrome are at high risk of developing chronic kidney disease (CKD) through unclear pathogenic mechanisms. Obesity and diabetes are known to induce glucolipotoxic effects in metabolically relevant organs. However, the pathogenic role of glucolipotoxicity in the aetiology of diabetic nephropathy is debated. We generated a murine model, the POKO mouse, obtained by crossing the peroxisome proliferator-activated receptor gamma 2 (PPARγ2) knockout (KO) mouse into a genetically obese ob/ob background. We have previously shown that the POKO mice showed: hyperphagia, insulin resistance, hyperglycaemia and dyslipidaemia as early as 4 weeks of age, and developed a complete loss of normal β-cell function by 16 weeks of age. Metabolic phenotyping of the POKO model has led to investigation of the structural and functional changes in the kidney and changes in blood pressure in these mice. Here we demonstrate that the POKO mouse is a model of renal disease that is accelerated by high levels of glucose and lipid accumulation. Similar to ob/ob mice, at 4 weeks of age these animals exhibited an increased urinary albumin:creatinine ratio and significantly increased blood pressure, but in contrast showed a significant increase in the renal hypertrophy index and an associated increase in p27Kip1 expression compared with their obese littermates. Moreover, at 4 weeks of age POKO mice showed insulin resistance, an alteration of lipid metabolism and glomeruli damage associated with increased transforming growth factor beta (TGFβ) and parathyroid hormone-related protein (PTHrP) expression. At this age, levels of proinflammatory molecules, such as monocyte chemoattractant protein-1 (MCP-1), and fibrotic factors were also increased at the glomerular level compared with levels in ob/ob mice. At 12 weeks of age, renal damage was fully established. These data suggest an accelerated lesion through glucolipotoxic effects in the renal pathogenesis in POKO mice.

Parathyroid hormone-related protein induces hypertrophy in podocytes via TGF-β1 and p27Kip1: implications for diabetic nephropathy

BACKGROUND Hypertrophy of podocytes is characteristic in diabetic nephropathy (DN). Previously, we observed the upregulation of parathyroid hormone-related protein (PTHrP) and its receptor PTH1R, in experimental DN, associated with renal hypertrophy. Herein, we test the hypothesis that PTHrP participates in the mechanism of high glucose (HG)-induced podocyte hypertrophy. METHODS On mouse podocytes, hypertrophy was assessed by protein content/cell and [H(3)]leucine incorporation. Podocytes were stimulated with HG (25 mM), PTHrP(1-36) (100 nM), angiotensin II (AngII) (100 nM) or TGF-beta(1) (5 ng/mL) in the presence or absence of PTHrP-neutralizing antibodies (alpha-PTHrP), the PTH1R antagonist JB4250 (10 microM), PTHrP silencer RNA (siRNA) or TGF-beta(1) siRNA. Protein expression was analysed by western blot and immunohistochemistry. RESULTS HG-induced hypertrophy was abolished in the presence of either alpha-PTHrP or PTHrP siRNA. This effect was associated with an inhibition of the upregulation of TGF-beta(1) and p27(Kip1). JB4250 also inhibited HG-induced p27(Kip1) upregulation. Interestingly, whilst HG and AngII were unable to stimulate the expression of p27(Kip1) on PTHrP siRNA-transfected podocytes, TGF-beta(1) was still able to upregulate p27(Kip1) in these cells. Moreover, HG and PTHrP-induced hypertrophy as well as p27(Kip1) upregulation were abolished on TGF-beta(1) siRNA-transfected podocytes. Furthermore, the glomeruli of transgenic PTHrP-overexpressing mice showed a constitutive overexpression of TGF-beta(1) and p27(Kip1) to a degree similar to that of diabetic animals. CONCLUSIONS PTHrP seems to participate in the hypertrophic signalling triggered by HG. In this condition, AngII induces the upregulation of PTHrP, which might induce the expression of TGF-beta(1) and p27(Kip1). These findings provide new insights into the protective effects of AngII antagonists in DN, opening new paths for intervention.

Parathyroid hormone-related protein is a hypertrophy factor for human mesangial cells: Implications for diabetic nephropathy.

Hypertrophy of human mesangial cells (HMC) is among the earliest characteristics in patients with diabetic nephropathy (DN). Recently, we observed the upregulation of parathyroid hormone (PTH)‐related protein (PTHrP) in experimental DN, associated with renal hypertrophy. Herein, we first examined whether PTHrP was overexpressed in human DN, and next assessed the putative role of this protein on high glucose (HG)‐induced HMC hypertrophy. As previously found in mice, kidneys from diabetic patients showed an increased tubular and glomerular immunostaining for PTHrP. In HMC, HG medium increased PTHrP protein expression associated with the development of hypertrophy as assessed by cell protein content. This effect was also induced by PTHrP(1–36). HG and PTHrP(1–36)‐induced hypertrophy were associated with an increase in cyclin D1 and p27Kip1 protein expression, a decreased cyclin E expression, and the prevention of cyclin E/cdk2 complex activation. Both PTHrP neutralizing antiserum (α‐PTHrP) and the PTH/PTHrP receptor antagonist (JB4250) were able to abolish HG induction of hypertrophy, the aforementioned changes in cell cycle proteins, and also TGF‐β1 up‐regulation. Moreover, the capability of both HG and PTHrP(1–36) to induce HMC hypertrophy was abolished by α‐TGFβ1. These data show for the first time that PTHrP is upregulated in the kidney of patients with DN. Our findings also demonstrate that PTHrP acts as an important mediator of HG‐induced HMC hypertrophy by modulating cell cycle regulatory proteins and TGF‐β1. J. Cell. Physiol. 227: 1980–1987, 2012.

A transgenic mouse model for studying the role of the parathyroid hormone-related protein system in renal injury

Parathyroid hormone- (PTH-) related protein (PTHrP) and its receptor, the PTH1 receptor (PTH1R), are widely expressed in the kidney, where PTHrP exerts a modulatory action on renal function. PTHrP is known to be upregulated in several experimental nephropathies such as acute renal failure (ARF), obstructive nephropathy (ON) as well as diabetic nephropathy (DN). In this paper, we will discuss the functional consequences of chronic PTHrP overexpression in the damaged kidney using a transgenic mouse strain overexpressing PTHrP in the renal proximal tubule. In both ARF and ON, PTHrP displays proinflammatory and profibrogenic actions including the induction of epithelia to mesenquima transition. Moreover, PTHrP participates in the mechanisms of renal hypertrophy as well as proteinuria in experimental DN. Angiotensin II (Ang II), a critical factor in the progression of renal injury, appears to be, at least in part, responsible for endogenous PTHrP upregulation in these pathophysiological settings. These findings provide novel insights into the well-known protective effects of Ang II antagonists in renal diseases, paving the way for new therapeutic approaches.

Early peroxisome proliferator-activated receptor gamma regulated genes involved in expansion of pancreatic beta cell mass

BackgroundThe progression towards type 2 diabetes depends on the allostatic response of pancreatic beta cells to synthesise and secrete enough insulin to compensate for insulin resistance. The endocrine pancreas is a plastic tissue able to expand or regress in response to the requirements imposed by physiological and pathophysiological states associated to insulin resistance such as pregnancy, obesity or ageing, but the mechanisms mediating beta cell mass expansion in these scenarios are not well defined. We have recently shown that ob/ob mice with genetic ablation of PPARγ2, a mouse model known as the POKO mouse failed to expand its beta cell mass. This phenotype contrasted with the appropriate expansion of the beta cell mass observed in their obese littermate ob/ob mice. Thus, comparison of these models islets particularly at early ages could provide some new insights on early PPARγ dependent transcriptional responses involved in the process of beta cell mass expansionResultsHere we have investigated PPARγ dependent transcriptional responses occurring during the early stages of beta cell adaptation to insulin resistance in wild type, ob/ob, PPARγ2 KO and POKO mice. We have identified genes known to regulate both the rate of proliferation and the survival signals of beta cells. Moreover we have also identified new pathways induced in ob/ob islets that remained unchanged in POKO islets, suggesting an important role for PPARγ in maintenance/activation of mechanisms essential for the continued function of the beta cell.ConclusionsOur data suggest that the expansion of beta cell mass observed in ob/ob islets is associated with the activation of an immune response that fails to occur in POKO islets. We have also indentified other PPARγ dependent differentially regulated pathways including cholesterol biosynthesis, apoptosis through TGF-β signaling and decreased oxidative phosphorylation.

Novel Role of Parathyroid Hormone-Related Protein in the Pathophysiology of the Diabetic Kidney: Evidence from Experimental and Human Diabetic Nephropathy

Parathyroid hormone-related protein (PTHrP) and its receptor type 1 (PTH1R) are extensively expressed in the kidney, where they are able to modulate renal function. Renal PTHrP is known to be overexpressed in acute renal injury. Recently, we hypothesized that PTHrP involvement in the mechanisms of renal injury might not be limited to conditions with predominant damage of the renal tubulointerstitium and might be extended to glomerular diseases, such as diabetic nephropathy (DN). In experimental DN, the overexpression of both PTHrP and the PTH1R contributes to the development of renal hypertrophy as well as proteinuria. More recent data have shown, for the first time, that PTHrP is upregulated in the kidney from patients with DN. Collectively, animal and human studies have shown that PTHrP acts as an important mediator of diabetic renal cell hypertrophy by a mechanism which involves the modulation of cell cycle regulatory proteins and TGF-β1. Furthermore, angiotensin II (Ang II), a critical factor in the progression of renal injury, appears to be responsible for PTHrP upregulation in these conditions. These findings provide novel insights into the well-known protective effects of Ang II antagonists in renal diseases, paving the way for new therapeutic approaches.

Parathyroid Hormone-Related Protein as a Mediator of Renal Damage: New Evidence from Experimental as well as Human Nephropathies

End-stage renal disease (ESRD) is a public health problem worldwide, with an increasing incidence and prevalence, poor prognosis and high cost (Ministerio de Salud, 2005). In 2005 the prevalence of ESRD in the USA was 1,131 patients per million inhabitants, with an incidence of 296 new patients per year per million inhabitants. In 2008, the adjusted incident rate for patients age 45–64 fell to the same level seen in 1998 -605 per million population. The rate for those aged 75 and older declined 0.9 percent between 2007 and 2008. However, racial and ethnic discrepancies persist with 2008 incident rates in the African American and Native American populations 3.6 and 1.8 times greater, respectively, than the rate among whites, and the rate in the Hispanic population 1.5 times higher than that of non-Hispanics (U. S. Renal Data System, 2010). Diabetic nephropathy (DN) is one of the major causes of ESRD in the Western world. The incidence of nephropathy owing to type 1 diabetes is declining; however, diabetes mellitus type 2 is now the most common single cause of renal insufficiency in the USA, Japan and Europe (Tisher & Hostetter, 1994). It has been estimated that 4% of the world’s current population is diabetic, and that in Europe there will be more than 32 million patients with diabetes in 2010, causing an increase in related complications, such as DN. This pathology has been recognized as a worldwide medical catastrophe (Ziyadeh & Sharma, 2003). DN appears clinically in 10-50% of diabetic patients after 10-20 years of evolution of the disease. These patients show a progressive increase in urinary excretion of protein, starting from an initial microalbuminuria to clear nephrotic syndrome with progressive decline of renal function (Cooper et al., 1999; Katoh et al., 2000; Tisher & Hostetter, 1994; Yotsumoto et al., 1997).

Entraining synthetic genetic oscillators

We propose a new approach for synchronizing a population of synthetic genetic oscillators, which consists in the entrainment of a colony of repressilators by external modulation. We present a model where the repressilator dynamics is affected by periodic changes in temperature. We introduce an additional plasmid in the bacteria in order to correlate the temperature variations with the enhancement of the transcription rate of a certain gene. This can be done by introducing a promoter that is related to the heat shock response. This way, the expression of that gene results in a protein that enhances the overall oscillations. Numerical results show coherent oscillations of the population for a certain range of the external frequency, which is in turn related to the natural oscillation frequency of the modified repressilator. Finally we study the transient times related with the loss of synchronization and we discuss possible applications in biotechnology of large-scale production coupled to synchronization events induced by heat shock.

La proteína relacionada con la paratirina: un nuevo mediador en la nefropatía diabética

Resumen La proteina relacionada con la paratirina (PTHrP) y su receptor tipo 1 (PTH1R) se expresan ampliamente en el rinon, tanto a nivel glomerular como tubular, donde es capaz de modular la funcion renal. Recientemente hemos analizado el posible papel de esta proteina en un modelo experimental de nefropatia diabetica (ND). Utilizando ratones CD1 controles y ratones transgenicos, con sobreexpresion renal de PTHrP, hemos observado que con ND hay un incremento significativo en la expresion del sistema renal de PTHrP/PTH1R tanto a nivel glomerular como tubular. Esta activacion se asocio con el desarrollo de hipertrofia y proteinuria. Asimismo, el analisis de regresion logistica puso de manifiesto que en los animales con mayor expresion renal de PTHrP y PTH1R hay un riesgo mayor de desarrollar proteinuria. En estudios in vitro, observamos que una elevada concentracion de glucosa incrementa la expresion del sistema PTHrP/PTH1R en celulas glomerulares (podocitos y celulas mesangiales) y en tubuloepiteliales mediante un mecanismo que implica a la angiotensina II. La PTHrP, factor capaz de modular la hemodinamica glomerular, podria ademas participar en la fisiopatologia asociada a la hiperfiltracion glomerular. Datos preliminares de nuestro grupo senalan que la PTHrP, a traves p27 Kip1 (regulador negativo del ciclo celular), forma parte de los mecanismos moleculares implicados en la hipertrofia del rinon diabetico. Por otra parte, hemos observado un aumento de PTHrP en el glomerulo de pacientes diabeticos, proporcionando las primeras evidencias de la implicacion de la PTHrP en la patologia renal humana. Finalmente, la infiltracion de macrofagos en el rinon diabetico ha sido motivo de renovado interes y recientemente se ha demostrado acciones proinflamatorias/profibrogenicas de la PTHrP, efectos que favorecerian el proceso inflamatorio asociado a la ND. Estos hallazgos indican a la PTHrP como un nuevo agente involucrado en los cambios renales asociados a la diabetes.