Acute renal failure in zebrafish: a novel system to study a complex disease

Abstract

Acute renal failure (ARF) is characterized by a very high mortality essentially unchanged over the past 40 years. Simple vertebrate models are needed to improve our understanding of ARF and facilitate the development of novel therapies for this clinical syndrome. Here, we demonstrate that gentamicin, a commonly used nephrotoxic antibiotic, causes larval zebrafish to develop ARF characterized by histological and functional changes that mirror aminoglycoside toxicity in higher vertebrates and inability of zebrafish to maintain fluid homeostasis. We developed a novel method to quantitate renal function in larval zebrafish and demonstrate a decline in glomerular filtration rate after gentamicin exposure. The antineoplastic drug cisplatin, whose use in humans is limited by kidney toxicity, also causes typical histological changes and a decline in renal function in larval zebrafish. A specific inhibitor of Omi/HtrA2, a serine protease implicated in cisplatin-induced apoptosis, prevented renal failure and increased survival. This protective effect was confirmed in a mouse model of cisplatin-induced nephrotoxicity. Therefore, zebrafish provides a unique model system, amenable to genetic manipulation and drug screening, to explore the pathophysiology of ARF and establish novel therapies with potential use in mammals. animal model; drug development; glomerular filtration; fluorescence ACUTE RENAL FAILURE (ARF) in the intensive care unit setting is associated with mortality rates of 50 to 70%, which have not changed for several decades (10, 32), and proven therapies remain elusive. Current mammalian models of ARF have been challenged as to their applicability to the human syndrome and are often difficult to interpret (4,14). The kidneys in the commonly used mouse or rat are inaccessible, and renal tubules and vessels cannot be visualized, with the exception of a very small subpopulation near the surface of the kidney (11). Cell culture systems, on the other hand, while more simple, are inadequate because of epithelial cell dedifferentiation and absence of the three-dimensional interaction of renal tubule, blood vessel, and immune cells, which plays a central role in the pathophysiology of ARF in humans (32). Furthermore, isolated cell systems are maintained in an environment containing growth factors and cytokines that do not reflect the local environment of epithelial cells in vivo in ARF. Early larval zebrafish kidneys are notable for their anatomic simplicity, consisting of a fused midline glomerulus with one nephron on each side, adjacent to the cardinal veins, while maintaining biological complexity inherent to the kidney of higher organisms. Zebrafish are translucent, facilitating microscopic observation along the entire length of the kidney. Microinjection of mRNA, DNA, and morpholinos (modified oligonucleotides with excellent antisense properties) is well tolerated. Morphological and functional changes can be correlated with protein expression after either knockdown (morpholinos) or overexpression (mRNA) (5). Their...

Mammalian Target of Rapamycin Mediates Kidney Injury Molecule 1-Dependent Tubule Injury in a Surrogate Model

“…We previously reported a gentamicin-induced proximal tubular injury model in the zebrafish. 15 To examine the expression of Kim-1 in zebrafish after injury, adult zebrafish (.90 days postfertilization [dpf ]) were injected with gentamicin (5 mg/100 mg body wt). Quantitative RT-PCR was used to define the relative expression patterns for each kim family member at 2 days after gentamicin injection (Figure 2).…”

“…15 We injected wild-type larvae (control), and larvae expressing RFP or Kim-1-RFP with 2 ng tetramethylrhodamine-labeled 10 kDa dextran at 72 hours postfertilization (hpf) (0 hours) ( Figure 4G). Fluorescence intensities were measured over the eye at 1, 7, 24, 31, and 43 hours after injection and compared with the baseline (immediately upon equilibration after injection, 0 hours).…”

“…Wild-type zebrafish (D. rerio) were maintained on a 14/10 hour light-dark cycle at 28.5°C and fed twice daily. Fertilized eggs were raised in embryo medium at 28.5°C and staged according to the standard method as described previously 15 . All the eggs and larvae were raised in E3 medium (5 mM NaCl, 0.17 mM KCl, 0.33 mM CaCl 2 , 0.33 mM MgSO 4 , 10-5% methylene blue).…”

“…13,14 Despite the anatomic simplicity of the zebrafish pronephros (in larvae) and mesonephros (in adult fish), zebrafish models for studying AKI, polycystic kidney disease, nephronophthisis, and a range of ciliopathies have brought important insight to pathophysiologic processes in mammals. [15][16][17] Rapid breeding and development, together with ease of genetic manipulation and optical transparency, make zebrafish an ideal model organism to develop highthroughput screening for therapeutic drug discovery.…”

Nuclear Factor κB Inhibitors Alleviate and the Proteasome Inhibitor PS-341 Exacerbates Radiation Toxicity in Zebrafish Embryos

“…20 Briefly, zebrafish embryos at 24 hpf were exposed to ionizing radiation and maintained in embryo medium. At 72 hpf embryos were anesthetized using a 1:100 dilution of 4 mg/mL tricaine methanesulfonate (Sigma) and dorsally positioned on 3% methylcellulose gel.…”