Knockdown of ApoL1 in Zebrafish Larvae Affects the Glomerular Filtration Barrier and the Expression of Nephrin

Abstract

APOL1, a secreted high-density lipoprotein, is expressed in different human tissues. Genetic variants of APOL1 are described to be associated with the development of end stage renal diseases in African Americans. In human kidney, APOL1 is mainly expressed in podocytes that are responsible for proper blood filtration. Since mice do not express ApoL1, the zebrafish is an ideal model to study the role of ApoL1. Injection of morpholinos against zApoL1 into zebrafish eggs and larvae, respectively, induces severe edema indicating a leakage of the filtration barrier. This was demonstrated in zApoL1 knockdown larvae by intravascular injection of fluorescently-labeled 10- and 500-kDa dextrans and by clearance of the vitamin D-binding protein from the circulation. Immunohistochemistry and RT-PCR revealed the reduction of nephrin, a podocyte-specific protein essential for blood filtration. Coinjection of human nephrin mRNA rescued the zApoL1 knockdown induced phenotype. Reduced APOL1 and nephrin levels were also found in biopsies of patients suffering from end stage renal diseases. Our results demonstrate that zApoL1 is essential for proper blood filtration in the zebrafish glomerulus and that zApoL1 affects the expression of nephrin.

The Cell Biology of APOL1

“…In normal human kidney the APOL1 protein localizes to the podocyte, the proximal tubule and the endothelium of extraglomerular arterioles and small arteries 27 , a finding subsequently replicated by others 28, 29 . Interestingly, the amount of APOL1 protein in the podocyte and proximal tubules was decreased in FSGS or HIV associated nephropathy (HIVAN), two kidney diseases associated with APOL1 risk variants 27, 29 .…”

“…Although inflammatory mediators consistently increase APOL1 expression in vitro , immunohistochemical studies of human kidney sections of individuals with kidney diseases demonstrate decreased abundance of APOL1 protein in the podocyte and proximal tubule 27, 29 . Subjects with kidney diseases and the APOL1 risk genotype [two risk alleles] have similar APOL1 abundance by immunohistology compared to subjects with kidney disease and the low risk genotype [one on no risk alleles].…”

“…Interestingly, the amount of APOL1 protein in the podocyte and proximal tubules was decreased in FSGS or HIV associated nephropathy (HIVAN), two kidney diseases associated with APOL1 risk variants 27, 29 . Although, the APOL1 signal in the vascular endothelium appeared similar in disease and non-disease kidney sections, the vascular media of renal arterioles and small arteries stained positive only in the setting of kidney disease 27 .…”

APOL1 variants change C-terminal conformational dynamics and binding to SNARE protein VAMP8

“…We and others have previously shown that the overall fluorescence intensity of APOL1 signal in the glomeruli of focal segmental glomerulosclerosis (FSGS) and HIV-associated nephropathy (HIVAN) samples was diminished relative to glomeruli from samples without renal pathology (12,14). In this study, we further characterized APOL1 localization and distribution in podocytes to determine if these phenotypes were dependent upon kidney disease diagnosis or APOL1 genotype.…”

“…APOL1-coding polymorphisms, G1 and G2, encode APOL1 proteins that circumvent SRA-dependent resistance and restore parasite killing, but, if present on both alleles, they markedly increase CKD odds in African American patients (2,3). We and others have shown that APOL1 is localized in human kidney glomeruli (12)(13)(14) and that circulating APOL1 levels do not associate with categorical or quantitative CKD phenotypes (15,16). Others have shown that poor kidney allograft outcomes are associated with the APOL1 genotype of the transplanted kidney but not the recipient (17,18).…”