The decreases of nephrin and nuclear WT1 in podocytes may cause albuminuria during the experimental sepsis in mice

Abstract

Sepsis is induced by infectious challenges, and septic organ failure often occurs under local and systemic inflammation. Albuminuria is also evident during sepsis, but little is known about the molecular basis of septic albuminuria. Using lipopolysaccharide (LPS)-treated mice as a sepsis model, we found that the loss of nephrin, a key component for maintaining podocyte slit diaphragm, became evident in accordance with the onset of albuminuria, especially 36 h post-LPS challenge (i.e., albumiuric stage). Likewise, nephrin mRNA levels were decreased to 13% of saline-treated mice. Such a transcriptional suppression of nephrin was associated with the loss of nucleus-localized Wilms tumor-1 (WT1), a transcriptional factor for up-regulating nephrin gene. Thereafter, urinary albumin levels were decreased in mice between 72 and 96 h post-LPS challenge (i.e., recovery-stage). Notably, nuclear localization of WT1 seemed to be normalized, and nephrin mRNA and protein levels returned near the basal level 72 h post-LPS challenge. During LPS-mediated sepsis, there was a transient increase in blood interleukin-1β, a suppressor of nephrin production in podocytes. Therefore, down-regulation of nephrin by the loss in nuclear WT1, along with hyper-cytokinemia, may underlie the mechanisms by which albuminuria is induced by infectious stresses.Septic syndrome is elicited by infectious agents of bacteria, virus, and fungus. The infectious components (including bacterial cell walls) function as foreign ligands of toll-like receptors, and systemic inflammation elicits thrombosis and eventually multiple organ failure (4, 26). Furthermore, albuminuria is also a common event during septic diseases (6,25,28), along with inflammatory responses. For example, lipopolysaccharide (LPS) of Escherichia coli, or histone-like protein of Streptococcal pyogenesis, often triggers albuminuria and glomerular nephritis (1,32,33), and this is associated with over-production of pro-inflammatory cytokines by activated macrophages (10,22). However, it is still unclear how sepsis-induced albuminuria is linked with an increase in pro-inflammatory cytokines (i.e., cytokine storm). Recent merging evidences provide a central dogma that glomerular podocytes have a key function to prevent the onset of albuminuria (2, 16, 31). Podocytes form foot processes, highly dynamic cellular extensions that are connected by slit diaphragm. As an important protein for podocytes, nephrin (encoded by NPHS1 gene) is well known (16,31). Nephrin is a transmembrane protein of the

Decreases in Podocin, CD2-Associated Protein (CD2AP) and Tensin2 May Be Involved in Albuminuria during Septic Acute Renal Failure

“…In this regard, we reported that the loss of nephrin, another key molecule of SD [9], becomes evident during sepsis through a loss in nuclear localization of WT1, a key transcriptional factor of nephrin [ 16]. In this study, we focused on podocin, because it is a key anchorage that strengthens FP-FP contact via a bridge between nephrin and CD2AP [21].…”

“…The albuminuria was evaluated using SDS-PAGE, where its concentration (mg/ml) was determined, based on the band size of bovine serum albumin under densitometry [16].…”

“…Morphometry: We quantified the glomerular expressions of podocin, CD2AP and tensin2 according to a staining score [16]. Briefly, >20 glomeruli were randomly chosen, and expression levels in each glomerulus were scored based on the fluorescence extent, as followed: 0; none, 1; faint, 2; mild, 3; moderate, and 4; strong.…”

“…Briefly, >20 glomeruli were randomly chosen, and expression levels in each glomerulus were scored based on the fluorescence extent, as followed: 0; none, 1; faint, 2; mild, 3; moderate, and 4; strong. The extent of molecular expressions was expressed as the individual index (i.e., total score/glomerular number examined) [16].…”

“…Immunoprecipitation and Western blotting: Kidney tissue homogenates were prepared from mice in lysis buffer, as reported [16,25]. The renal extraction was incubated overnight at 4°C with goat anti-podocin antibody.…”