Heterotrimeric G-proteins play a crucial role in the control of renal epithelial cell function during homeostasis and in response to injury. In this report, G-protein bg subunit (Gbg) dimer activity was evaluated during the process of tubular repair after renal ischemia-reperfusion injury (IRI) in male Sprague Dawley rats. Rats were treated with a small molecule inhibitor of Gbg activity, gallein (30 or 100 mg/kg), 1 hour after reperfusion and every 24 hours for 3 additional days. After IRI, renal dysfunction was prolonged after the high-dose gallein treatment in comparison with vehicle treatment during the 7-day recovery period. Renal tubular repair in the outer medulla 7 days after IRI was significantly (P , 0.001) attenuated after treatment with high-dose gallein (100 mg/kg) in comparison with low-dose gallein (30 mg/kg), or the vehicle and fluorescein control groups. Gallein treatment significantly reduced (P , 0.05) the number of proliferating cell nuclear antigen-positive tubular epithelial cells at 24 hours after the ischemia-reperfusion phase in vivo. In vitro application of gallein on normal rat kidney (NRK-52E) proximal tubule cells significantly reduced (P , 0.05) S-phase cell cycle entry compared with vehicletreated cells as determined by 59-bromo-29-deoxyuridine incorporation. Taken together, these data suggest that Gbg signaling contributes to the maintenance and repair of renal tubular epithelium and may be a novel therapeutic target for the development of drugs to treat acute kidney injury.

Herein we describe lung vascular injury and repair using a rodent model of Pseudomonas aeruginosa pneumonia-induced acute respiratory distress syndrome (ARDS) during: 1) the exudative phase (48-hour survivors) and 2) the reparative/fibro-proliferative phase (1-week survivors). Pneumonia was induced by intratracheal instillation of P. aeruginosa strain PA103, and lung morphology and pulmonary vascular function were determined subsequently. Pulmonary vascular function was assessed in mechanically ventilated animals in vivo (air dead space, P a O 2 , and lung mechanics) and lung permeability was determined in isolated perfused lungs ex vivo (vascular filtration coefficient and extravascular lung water). At 48 hours post infection, histological analyses demonstrated capillary endothelial disruption, diffuse alveolar damage, perivascular cuffs, and neutrophil influx into lung parenchyma. Infected animals displayed clinical hallmarks of ARDS, including increased vascular permeability, increased dead space, impaired gas exchange, and decreased lung compliance. Overall, the animal infection model recapitulated the morphological and functional changes typically observed in lungs from patients during the exudative phase of ARDS. At 1 week post infection, there was lung histological and pulmonary vascular functional evidence of repair when compared with 48 hours post infection; however, some parameters were still impaired when compared with uninfected controls. Importantly, lungs displayed increased fibrosis and cellular hyperplasia reminiscent of lungs from patients during the fibro-proliferative phase of ARDS. Control, sham inoculated animals showed normal lung histology and function. These data represent the first comprehensive assessment of lung pathophysiology during the exudative and reparative/fibro-proliferative phases of P. aeruginosa pneumonia-induced ARDS, and position this pre-clinical model for use in interventional studies aimed at advancing clinical care.

We hypothesized that increased 3‐phosphoinositide‐dependent kinase 1 (PDK1) activation contributes to arterial wall thickening following vascular injury. Vascular smooth muscle cells (VSMCs) were pretreated with BX‐912, a small molecule inhibitor of PDK1, prior to stimulation with PDGF. Western analysis confirmed inhibition of PDK1 activity in the presence of BX‐912 by decreased phosphorylation of PDK1 (ser241) by ~60% and Akt (thr308) by ~ 70%. Conversely, Akt phosphorylation at ser473 was only mildly affected. To examine VSMC migration, both scratch wound assays and modified Boyden chamber studies were completed. In both instances, wound‐ and PDGF‐induced chemotaxis was attenuated following BX‐912 treatment. Following wire injury of the carotid artery in C57BL6/J mice, in vivo PDK1 phosphorylation (ser 241) was assessed by immunoblotting in both injured and contralateral control carotids at 3, 7, and 14 days post injury. PDK1 phosphorylation was markedly increased at day 3 (~5 fold vs. non‐injured), remained slightly elevated at day 7 (~2 fold) and returned close to baseline by later timepoints following injury. Our findings indicate that BX‐912 is effective in attenuating PDK1 activity and chemotaxis in VSMCs and that increased PDK1 activity occurs following vascular injury. Thus, PDK1 inhibition, via BX‐912 treatment, could potentially be utilized to attenuate wall remodeling in response to injury. Support: NIH HL084159

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