Purpose: In our previous study, exacerbation of albuminuria was observed in A1 adenosine receptor knockout (A1AR^−/-) mice with diabetic nephropathy (DN), but the mechanism was unclear. Here, we investigated the relationship of megalin loss and albuminuria, to identify the protective effect of A1AR in megalin loss associated albuminuria by inhibiting pyroptosis-related caspase-1/IL-18 signaling of DN.
Methods: We successfully collected DN patients’ samples and built diabetes mice models induced by streptozotocin. Megalin, cubilin, and A1AR expression were detected in kidney tissue samples from DN patients and mice through immunohistochemical and immunofluorescent staining. A1AR, caspase-1, interleukin-18 (IL-18) expression were analyzed using Western blotting in wild-type and A1AR ^−/-mice. Human renal proximal tubular epithelial cells (PTC) were cultured with high glucose to observe the effect of A1AR agonist and antagonist on caspase-1/IL-18 and megalin injury.
Results: The loss of megalin, co-localized with A1AR at PTC, was associated with the level of albuminuria in diabetic patients and mice. The injury of megalin-cubilin was accompanied with the A1AR upregulation (1.30±0.1 vs 0.98±0.2, P =0.042), the caspase-1 (1.33±0.1 vs 1.0±0.2, P =0.036), and IL-18 (1.26±0.2 vs 0.96±0.2, P =0.026) signaling activation in mice with DN. More severe pathological injury, 24 hrs urine albumin excretion (170.8±4.1 μg/d vs 132.0±2.9 μg/d vs 17.9±2.8 μg/d, P <0.001) and megalin-cubilin loss were observed in A1AR ^−/-DN mice with more pronounced caspase-1 (1.52±0.03 vs 1.20±0.01, P =0.017) and IL-18 (1.42±0.02 vs 1.21±0.02, P =0.018) secretion. High glucose could stimulate the secretion of caspase-1 (1.72 times, P ≤0.01) and IL-18 (1.64 times, P ≤0.01), which was abolished by A1AR agonist and aggravated by A1AR antagonist.
Conclusion: A1AR played a protective role in proximal tubular megalin loss associated albuminuria by inhibiting the pyroptosis-related caspase-1/IL-18 signaling in DN.
Keywords: A1 adenosine receptor, diabetic nephropathy, caspase-1, megalin