Upregulation of Megalin, Cubilin, NGAL mRNA expression in kidney may represent tubular injury and apoptosis in chronic condition of rat diabetic model

@#Introduction: Diabetes mellitus (DM) leads to microvascular injury development and produces diabetes nephropathy (DN) with proteinuria, tubular injury, apoptosis and autophagy with upregulation of Bax, BASP and mTORC-1. Megalin, Cubilin and Neutrophil Gelatinase Associated Lipocalin (NGAL) play role in acute pathological condition of kidney injury, however its expression in chronic and slowly progressive kidney injury such as DN has not been elucidated yet. This study focuses upregulation of Megalin, Cubilin and NGAL in association with tubular injury and apoptosis in DN condition. Materials and methods: Diabetic condition was performed with intraperitoneal injection of Streptozotocin 60 mg/kg body weight (BW) in Sprague Dawley rats (2 months old, n=24), and were kept for 1, 2, and 4 months (DM1, DM2, and DM4, respectively). Control group was injected with NaCl 0.9%. Serum glucose level and proteinuria score were assessed, furthermore tubular injury score was quantified based on Periodic-Acid Schiff (PAS) staining. Reverse Transcriptase-PCR (RT-PCR) was carried out for NGAL, Megalin, Cubilin, m-TOR, Bax, and BASP-1 mRNA expression. Data were analyzed using SPSS 22 software. Results: DM led to kidney injury in this model with significant higher glucose level, proteinuria and tubular injury, especially in DM4 group which represented chronic phase of DN and CKD. These findings associated with upregulation of Megalin,Cubilin and NGAL mRNA expression in DM groups, especially in DM4 group. DM4 group also revealed higher expression of Bax, BASP and mTOR mRNA expression which demonstrated apoptosis. Conclusion: Megalin, Cubilin and NGAL upregulation may represent tubular injury and apoptosis as progression of DN.

El síndrome urémico hemolítico y manejo renal de proteínas / Hemolytic uremic syndrome and renal handling of proteins

El síndrome urémico hemolítico (SUH) está definido por la tríada de anemia hemolítica microangiopática, trombocitopenia e insuficiencia renal aguda. En Argentina constituye la primera causa de insuficiencia renal aguda en pediatría. Aproximadamente, del 2% al 4% de los pacientes mueren durante la fase aguda de la enfermedad, y solo un tercio del 96% restante que sobrevive lo hace con secuelas renales, como la persistencia de la proteinuria. Un individuo adulto sano filtra alrededor de 5000 mg/día de proteínas, si bien la excreción en orina es escasa (150 mg/día). La escasa cantidad de proteínas excretadas indica la presencia de un mecanismo de reabsorción a nivel del túbulo proximal. Por lo tanto, la reabsorción tubular renal desempeña un papel muy importante ya que, ante una función glomerular normal, es el principal mecanismo encargado de evitar la depleción proteica corporal. Desde hace aproximadamente 30 años se sabe que la albúmina es reabsorbida en el túbulo proximal. La reabsorción proteica se produce por un mecanismo de endocitosis mediada por el receptor dependiente de clatrina y por endocitosis de fase líquida. Clásicamente se ha descrito que el mecanismo básico del daño renal en el SUH típico y en el atípico es una microangiopatía trombótica, pero de diferentes causas. Sin embargo, debe tenerse en cuenta que la fisiopatología de esta enfermedad es más compleja de lo que se creía, ya que la alteración tubular que surge va a evolucionar en fallas en el mecanismo de endocitosis de proteínas que se suman a las eliminadas por las alteraciones a nivel de la barrera de filtración glomerular.

Hemolytic uremic syndrome (HUS) is defined by the triad of hemolytic anemia microangiopathic, thrombocytopenia and acute renal failure. In Argentina it constitutes the first cause of acute renal failure in Pediatrics. Approximately 2-4% of patients die during the acute phase of the disease, and only a third of the remaining 96% survive with renal sequelae, such as the persistence of proteinuria. A healthy adult filters around 5000 mg/day of proteins, with an excretion in urine of 150 mg/day. The little quantity of proteins excreted indicates the presence of a reabsorption mechanism at the level of the proximal tubule. Therefore, the tubular reabsorption plays a very important role since it is the main mechanism responsible for preventing the depletion of protein. For approximately 30 years, it has been known that albumin is reabsorbed in the proximal tubule. Protein reabsorption occurs by a clathrin-dependent receptor mediated endocytosis mechanism and by fluid phase endocytosis. The basic mechanism of renal damage in typical and atypical HUS has been described as a thrombotic microangiopathy, but of different causes. However, the pathophysiology of this disease is more complex than what was believed since the emerging tubular alteration will ewvolve into failures of the protein endocytosis mechanism that are added to the alterations at the level of the glomerular filtration barrier.

Progress of the renal chloride channel CLC-5 and Dent disease / 国际儿科学杂志

Chloride channel CLC-5 is a voltage-dependent gated channel.The voltage-gated characteristic of CLC-5 is not only regulated by glutamate E211 and E268,but also by lysine K210.In proximal renal tubule,CLC-5 can interact with megalin protein,and adjust the reabsorption of albumin together.CLC-5 can also affect the activities of the sodium hydrogen exchanger isoform 3 in proximal renal tubule.CLCN5 gene mutations can lead to Dent disease,and recent studies have found some new pathogenic mutants of CLC-5,VS05G,L266V and G446A,and so on.CLCN5 can also mutate together with ORCL1,and then result in Dent disease.However,the regulatory mechanism of the voltage-gated channel,physiological functions and molecular mechanism,and Dent disease are still not entirely clear.In this paper,we will review these problems of CLC-5.

Bone marrow cells contribute to tubular epithelium regeneration following acute kidney injury induced by mercuric chloride.

Background & objectives: Acute tubular necrosis (ATN) caused by renal ischaemia, renal hypo-perfusion, or nephrotoxic substances is the most common form of acute kidney injury (AKI). There are a few treatment options for this life-threatening disease and the mortality rate exceeds 50 per cent. In critical cases of AKI the only option is renal transplantation. In the present study we evaluated whether bone marrow cells (BMCs) are involved in regeneration of kidney tubules following acute tubular necrosis in the mouse. Methods: Six to eight week old C57BL6/J and congenic enhanced green fluorescence protein (eGFP) mice were used. The relative contributions of eGFP-expressing BMCs were compared in two different approaches to kidney regeneration in the mercuric chloride (HgCl2)-induced mouse model of AKI: induced engraftment and forced engraftment. In vitro differentiation of lineage-depleted (Lin-) BMCs into renal epithelial cells was also studied. Results: In the forced engraftment approach, BMCs were found to play a role in the regeneration of tubules of renal cortex and outer medulla regions. About 70 per cent of donor-derived cells expressed megalin. In vitro culture revealed that Lin- BMCs differentiated into megalin, E-cadherin and cytokeratin-19 (CK-19) expressing renal epithelial cells. Interpretation & conclusions: The present results demonstrate that Lin- BMCs may contribute in the regeneration of renal tubular epithelium of HgCl2-induced AKI. This study may also suggest a potential role of BMCs in treating AKI.

New Insights into the Roles of Megalin/LRP2 and the Regulation of its Functional Expression

Since the discovery of the low-density lipoprotein receptor (LDLR) and its association with familial hypercholesterolemia in the early 1980s, a family of structurally related proteins has been discovered that has apolipoprotein E as a common ligand, and the broad functions of its members have been described. LRP2, or megalin, is a member of the LDLR family and was initially called gp330. Megalin is an endocytic receptor expressed on the apical surface of several epithelial cells that internalizes a variety of ligands including nutrients, hormones and their carrier proteins, signaling molecules, morphogens, and extracellular matrix proteins. Once internalized, these ligands are directed to the lysosomal degradation pathway or transported by transcytosis from one side of the cell to the opposite membrane. The availability of megalin at the cell surface is controlled by several regulatory mechanisms, including the phosphorylation of its cytoplasmic domain by GSK3, the proteolysis of the extracellular domain at the cell surface (shedding), the subsequent intramembrane proteolysis of the transmembrane domain by the gamma-secretase complex, and exosome secretion. Based on the important roles of its ligands and its tissue expression pattern, megalin has been recognized as an important component of many pathological conditions, including diabetic nephropathy, Lowe syndrome, Dent disease, Alzheimer's disease (AD) and gallstone disease. In addition, the expression of megalin and some of its ligands in the central and peripheral nervous system suggests a role for this receptor in neural regeneration processes. Despite its obvious importance, the regulation of megalin expression is poorly understood. In this review, we describe the functions of megalin and its association with certain pathological conditions as well as the current understanding of the mechanisms that underlie the control of megalin expression.

Expression of endocytic receptor megalin/cubilin in embryonic mouse kidneys / 解剖学报

Objective To investigate the expression of endocytic receptor megalin and cubilinin in the developing mouse kidneys, and the correlation between the expression and the development of the renal tubules. Methods Expression of megalin and cubilin in developing mouse kidneys was examined at different embryonic days (E) using immunohistochemistry. Meanwhile, the ultrastructure of developing proximal tubules related to endocytosis was observed at transimission electron microscope level. Results At E9.5, megalin and cubilin were co-expressed in apical plasma membrane of the mesonephric ducts and mesonephric tubules. From E11 to E18, in the metanephros, the expression of both receptors were seen at the free surface and apical plasma of uretic bud, but weakly in all renal tubules of S shaped body at early differentiatial stage. With the mature of proximal tubule development, they were both confined to the brush border and the apical plasma of the proximal tubules in juxtamedullary cortex.Conclusion The endocytic receptors, megalin and cubilin are expressed in apical part of nearly all renal tubule epithelia in early development, and confined to free surface of mature proximal tubules, suggesting that with the mature of proximal tubules, the two receptors are generally involved in collaborating to facilitate, the reabsorption of ultrafiltration.