PB0423 - Protein phosphatase 1 alpha from platelets dampen endotoxin -induced proinflammatory signaling and kidney tubular apoptosis and injury

Background: Tubular epithelial cell dysfunction due to apoptosis is observed in endotoxin-associated AKI. Although thrombocytopenia is associated with higher mortality in AKI patients, the role of platelets in kidney injury remains unclear.

Aims: Since platelets participate in wound-healing, we hypothesized that platelets have a protective role during the early phase of endotoxemia–induced kidney injury.

Methods: Blood was collected pre and 70 hours post E.coli infection in a swine model of sepsis and immunoblotted with anti-PP1ca (catalytic subunit of PP1a). Human (HK2) and mouse (BUMPT) proximal tubular cells were challenged with 10 mcg/ml of LPS in the presence or absence of human or mouse platelets. Cells were assayed for apoptosis and phosphorylation status of P38 MAPK. Wild type (WT) mice and platelet specific deletion of PP1ca (PP1ca-/-) mice was challenged with 20 mg/kg LPS and renal injury and apoptosis studied by qPCR, TUNEL.

Results: Compared to pre-infection platelets, E. coli-infected swine platelets showed a 2.8 fold increase in PP1ca. Compared to the WT mice, LPS-induced early kidney injury markers Kim1 and NGAL was increased by 2 and 1 fold in PP1ca-/- mice. TUNEL staining was increased 2 fold in PP1ca-/- mice. LPS-induced apoptotic injury was reduced (40%) in HK2 cells and (30%) in BUMPT cells incubated with human or WT mouse platelets. PP1ca-/- platelets reduced only 10% of LPS-induced injury. Lastly LPS-induced P38 MAPK phosphorylation in BUMPT cells was markedly decreased upon incubation with WT but not PP1ca-/- platelets.

Conclusion(s): Our studies show that PP1ca is upregulated in platelets during E.coli infection in swine. Platelet specific deletion of PP1ca in mice exacerbates endotoxin-induced renal injury and apoptosis. LPS-induced cell death and increased phosphorylation of P38 MAPK in proximal tubular cells was mitigated by platelet PP1ca. These studies suggest that the loss of a protective role played by platelet PP1ca may contribute to thrombocytopenia- associated kidney injury.