Podocyte Developmental Defects Caused by Adriamycin in Zebrafish Embryos and Larvae: A Novel Model of Glomerular Damage

Abstract

The zebrafish pronephros is gaining popularity in the nephrology community, because embryos are easy to cultivate in multiwell plates, allowing large number of experiments to be conducted in an in vivo model. In a few days, glomeruli reach complete development, with a structure that is similar to that of the mammalian counterpart, showing a fenestrated endothelium and a basement membrane covered by the multiple ramifications of mature podocytes. As a further advantage, zebrafish embryos are permeable to low molecular compounds, and this explains their extensive use in drug efficacy and toxicity experiments. Here we show that low concentrations of adriamycin (i.e. 10 and 20 µM), when dissolved in the medium of zebrafish embryos at 9 hours post-fertilization and removed after 48 hours (57 hpf), alter the development of podocytes with subsequent functional impairment, demonstrated by onset of pericardial edema and reduction of expression of the podocyte proteins nephrin and wt1. Podocyte damage is morphologically confirmed by electron microscopy and functionally supported by increased clearance of microinjected 70 kDa fluorescent dextran. Importantly, besides pericardial edema and glomerular damage, which persist and worsen after adriamycin removal from the medium, larvae exposed to adriamycin 10 and 20 µM do not show any myocardiocyte alterations nor vascular changes. The only extra-renal effect is a transient delay of cartilage formation that rapidly recovers once adriamycin is removed. In summary, this low dose adriamycin model can be applied to analyze podocyte developmental defects, such as those observed in congenital nephrotic syndrome, and can be taken in consideration for pharmacological studies of severe early podocyte injury.

Human Pluripotent Stem Cell–Derived Kidney Model for Nephrotoxicity Studies

“…Puromycin, which is a primary glomerular toxicant and a secondary a tubular toxicant, exhibited statistically significant increased levels of both NPHS1/WT1 and HO-1, with the increase in NPHS1/WT1 being slightly more than the tubular marker. Doxorubicin treatment showed increased NPHS1/WT1 levels at 30 mM; this is consistent with studies showing that zebrafish embryos treated with 10-20 mM doxorubicin exhibited altered podocyte development with functional impairment and reduction of NPHS1 and WT1 (Zennaro et al, 2014). Although changes in NPHS1 and WT1 expression level are seen when subjected to glomerular toxicants, additional studies need to be done to confirm the sensitivity of these markers for toxicity.…”

Isoniazid causes heart looping disorder in zebrafish embryos by the induction of oxidative stress

“…4 The expression of four cardiac-specific transcript factors in zebrafish embryos after 48 hpf of isoniazid exposure. * P < 0.05, ** P < 0.1, *** P < 0.001, ns means no significantly different many reports that environmental poisons and drugs such as doxorubicin [41], chlorpyrifos [42], and dinitrobenzene cause abnormal development of various organs by the induction of ROS accumulation. In our study, we observed similar consequences, that ROS was highly concentrated in the heart and cardiac looping was impaired in the treatment group, which suggested that isoniazid caused cardiac dysplasia by the upregulation of ROS.…”

Isoniazid causes heart looping disorder in zebrafish embryos by the induction of oxidative stress

“…Recently, an article reported that high levels of ROS lead to heart looping disorder during heart development [41,42]. There are also many reports that environmental poisons and drugs such as doxorubicin [43], chlorpyrifos [44], and dinitrobenzene cause abnormal development of various organs by the induction of ROS accumulation. In our study, we observed similar consequences, that ROS was highly concentrated in the heart and cardiac looping was impaired in the treatment group, which suggested that isoniazid caused cardiac dysplasia by the upregulation of ROS.…”