Biogem

Nephrology Area

NEPHROLOGY AREA

( prof. Francesco Trepiccione)

The aim of the Translational Nephrology laboratory is to advance the knowledge of the molecular mechanisms underlying kidney diseases in order to identify new therapeutic targets or new prevention and diagnosis factors. The nephrology laboratory is divided into three thematic areas:

- Rare diseases (Anna Iervolino, PhD)

Starting from the generation of mouse and zebrafish models of kidney diseases, we study the causal mechanisms of specific rare nephrological diseases, such as Fanconi syndrome, Bartter syndrome, nephrogenic diabetes insipidus, Bardet-Biedl syndrome and focal segmental glomerulosclerosis. We recently started also a preclinical study for gene therapy of specific tubulopathies.

- Renal Molecular and Cellular Physiology (Yoko Suzumoto, PhD)

It studies the renal function at the cellular and molecular level in different physiopathological conditions in order to identify the mechanisms underlying complex diseases such as arterial hypertension. Also in this case, using a mouse model of renal-dependent hypertension, it is investigated at the molecular level the possible interaction between gut microbiome and arterial hypertension. Other projects concern the clinical use of paraoxonase-2 and the implementation of anti-aging strategies of the peritoneal membrane during peritoneal dialysis.

- Kidney-Brain (Prof. Giovambattista Capasso)

Kidney failure, especially the advanced one, is often complicated by a significant reduction in cognitive abilities. In the laboratory, using experimental models of chronic kidney failure and the technologies necessary for the characterization of cognitive functions in rodents, we study the factors responsible for this correlation, also using new imaging technologies, in order to identify the relevant molecular processes and possibly identify potential new therapies.

In addition to the common techniques of molecular biology and histology, our laboratory has the expertise and the technology necessary for the study of the nephron’s individual tubular portions through:

Kidney micropuncture: a technique that makes possible the collection and/or injection of compounds directly into the tubular or vascular lumen of the individual kidney’s tubular segments.

Live imaging through two-photon microscopy: a technique that allows the exploration of physiological processes such as blood perfusion, glomerular filtration, reabsorption of substances directly in the anesthetized living animal. It can be applied to different organs; it is currently used in our laboratory for the kidney and peritoneal membrane.

Physiological characterization of renal functions: this uses a number of protocols adapted to rodents for the study of glomerular filtration in vivo, of the major stimuli for the study of the transport of sodium, chlorine, magnesium, bicarbonate and potassium.

Biomedical Research

Internal seminar

YOKO SUZUMOTO PhD, 'Characterization of renal Paraoxonase 2 (PON2)' Monday 20th September

Paraoxonase 2 (PON2) is a membrane-associated enzyme that possesses high hydrolytic activity against bacterial signaling molecule 3-oxo-C12 homoserine lactone, thus exhibits the defense against pathogenic bacteria such as Pseudomonas aeruginosa. In addition, PON2 shows redox function leading to the prevention of atherosclerosis. Furthermore, accumulating data suggest the involvement of PON2 in several human diseases such as Diabetes Mellitus, Alzheimer’s disease and pancreatic cancer. In particular, recent report demonstrated the potential involvement of PON2 in the pathogenesis of venous thromboembolism in COVID-19 patients. In the present study, regulatory mechanisms and potential roles of renal PON2 were investigated utilizing both in vivo and in vitro systems. In Milan normotensive rats strain, under high aldosterone condition induced by low sodium diet, PON2 was up-regulated in parallel with proteolytic activation of alpha-ENaC in the renal cortex. These phenomena were through the activation of mineralocorticoid receptor, since aldosterone antagonist spironolactone blocked both ENaC activation and PON2 up-regulation. Furthermore, spironolactone and canrenone were shown to inhibit PON2 lactonase activity in vitro. Finally, aldosterone induced-upregulation of PON2 was accompanied by decreased reactive oxygen species (ROS) levels in the renal cortex, in particular superoxide levels. These results indicate hormone aldosterone as a novel regulator of PON2 in the kidney, and suggest that PON2 could preserve the kidney from aldosterone-induced ROS-related damages, such as fibrosis, through the modulation of ROS levels.

VINCENZO COSTANZO PhD, In vivo evaluation of renal function using intravital multiphoton microscopyMonday 27th September

2-photon microscopy (2PM) represents the “gold standard” technique to carry out in vivo renal pathophysiology studies. Indeed, compared to previous methods, 2PM offers greater laser penetration into tissues thanks to the reduced light scattering, low photo-toxicity that allows to monitor the samples over the time, and lowered fluorescence loss. Biogem has been equipped with a 2PM laboratory, thanks to the generous donation of the “Terzo Pilastro Internazionale” foundation, led by Prof. Emmanuele F.M. Emanuele. In this study we focused on 3 main research projects: the study of glomerular function by assessing the single nephron glomerular filtration rate (SNGFR), the study of tubular function by evaluating the reabsorption of glucose and proteins in the proximal tubules, and the detection and quantification of renal fibrosis in vivo and ex vivo. In the Translational Nephrology laboratory, an innovative approach to measure in vivo the SNGFR has been developed, since it represents one of the most important renal parameters capable of providing precise and real-time information about renal function. This approach has been validated in experimental models of acute kidney injury and it guarantees great precision and reliability in the measurements, offering results comparable to previous methods. For studies of tubular function, in particular the reabsorption of glucose in the proximal renal tubules, a GLUT2 cko mouse model mimicking the Fanconi Bickel syndrome has been used. It is a genetic disease characterized by hepato-renal glucose accumulation, impaired glucose and galactose metabolism, and proximal tubule dysfunction. By 2PM we demonstrated in vivo that GLUT2 cko mice have an impaired mechanism of glucose reabsorption compared to the control group. Finally, an approach to detect and quantify renal fibrosis both in animal models and tissue sections has been developed. This is possible by exploiting the optical phenomenon of the "second harmonic generation" (SHG), which is an intrinsic property of 2PM allowing the detection of specific tissue molecules, such as collagen fibers and myosin, without any exogenous labeling. This method guarantees high specificity reducing the variability due to the classic histological staining techniques. SHG coupled with machine learning software is able to perform image classification and represents a valid tool for quantifying renal fibrosis, not only in animal models but also for routinely clinical practice. The microscopy approaches developed in this project will increase the knowledge of many pathological conditions, such as acute renal failure and renal fibrosis, and may be used to develop new drugs able to restore physiological conditions.

Biogem

NEPHROLOGY AREA

Biomedical Research

Internal seminar

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