LIPOPOLYSACCHARIDE　PRECONDITIONING　AMELIORATES　HEAT-INDUCED　ACUTE　KIDNEY　INJURY

Heat-related illnesses (HRIs) cause systemic inflammation and multiple organ dysfunctions including acute kidney injury (AKI) which can progress to chronic kidney disease (CKD). Therefore, we need to develop safe and effective medical countermeasures for HRIs and HRI-induced AKI. It is known that a preceding weak/faint stimulus to the immune system increases the ability of the immune system to adapt to subsequent strong stimuli; this is referred to as “trained immunity”. We previously reported that repeated low-dose lipopolysaccharide (LPS) injections (LPS preconditioning) significantly increase resistance to bacterial infection in mice and attenuate the associated inflammatory response. We think that LPS preconditioning, therefore, may also be effective in preventing HRI-induced AKI.

We used adult male C57/BL6J mice (8-12 weeks). The mice were randomly divided into LPS preconditioning and saline groups (n = 14 each). The mice in the LPS preconditioning group were intraperitoneally injected with low-dose LPS (5 µg/kg) dissolved in 500 µL saline for 3 consecutive days. The saline group received an intraperitoneal injection of 500 μL saline alone for three consecutive days. Twenty-four hours later, the mice were then heat stressed in an environmental chamber at 40°C temperature/60% humidity for 30 min without food and water. Mice were sacrificed immediately prior to or 24 hours, 96 hours after heat stress to obtain blood, urine, and kidney samples. Additionally, kidney macrophages were analyzed using flow cytometry to evaluate phenotype changes associated with inflammation.

There was no significant difference in body weight or rectal temperature between the two groups at immediately and 1 hour after heat stress. Although the survival rate was not statistically different between the LPS preconditioning and saline groups, serum creatinine (Cr) levels were significantly lower in the LPS preconditioning group four days after heat stress, while the serum blood urea nitrogen (BUN) levels of the two groups did not differ to a statistically significant extent. We also measured the urinary kidney injury molecule-1(KIM-1) level, a biomarker of AKI biomarkers that is elevated when proximal tubules are injured. The saline group showed a marked elevation in urinary KIM-1 one day after heat stress, while the LPS preconditioning group did not show this elevation. Pathologically, tubular injuries, such as cast formation and loss of brush border, were observed after heat stress in the saline group, whereas these kidney lesions were not observed in the LPS preconditioning group. The tubular injury score was significantly lower in the LPS preconditioning group than in the saline group. The expression of heat shock protein (Hsp) 70 in tubular cells increased in LPS preconditioned mice before heat stress. Additionally, kidney resident macrophages in LPS preconditioned mice decreased expression of MHC class II and CD80 while increasing expression of CD206 at 96 hours after heat stress, suggesting a polarization to M2 phenotype. 

We have demonstrated that LPS preconditioning has the potential to prevent heat-induced AKI. The enhancement of the Hsp70 expression in tubular cells and the suppression of the MHC II expression in kidney macrophages may be part of the underlying mechanism. Further clinical studies are needed.