Intrawound Vancomycin Powder Reduces Bacterial Load in Contaminated Open Fracture Model.

OBJECTIVES: To compare the effectiveness of both vancomycin powder and antibiotic bead placement to irrigation and debridement alone in prevention of infection in a contaminated open fracture model in rats. METHODS: In a previously described model of contaminated open fractures, 45 rats had simulated open fractures created, stabilized, and contaminated with Staphylococcus aureus. They were then treated 6 hours later with 3 interventions: irrigation and debridement alone (control group) or in combination with placement of polymethyl methacrylate beads containing vancomycin and tobramycin powders (antibiotic bead group) or placement of 10 mg of intrawound vancomycin powder (powder group). Rats were allowed to recover and then killed 14 days later for harvest of femurs and plates. Femurs and plates were both incubated overnight, and bacterial colonies were counted in each group for comparison. RESULTS: Quantitative counts of bacteria in bone showed significantly reduced growth in both bead and powder groups when compared with control group (P < 0.0001). Quantitative counts of bacteria in plates showed significantly reduced growth in both bead and powder groups when compared with control group (P < 0.0003; 0.029). No significant differences were seen in bacterial growth between bead and powder groups for either bones (P = 0.13) or plates (P = 0.065). CONCLUSIONS: When compared with irrigation and debridement alone, placement of intrawound vancomycin powder significantly decreased bacterial load in a contaminated open fracture model in rats similar to placing antibiotic beads. This may provide an additional adjuvant treatment that does not require a secondary surgery for bead removal.

Protonation of key acidic residues is critical for the K⁺-selectivity of the Na/K pump.

The sodium-potassium (Na/K) pump is a P-type ATPase that generates Na(+) and K(+) concentration gradients across the cell membrane. For each hydrolyzed ATP molecule, the pump extrudes three Na(+) and imports two K(+) by alternating between outward- and inward-facing conformations that preferentially bind K(+) or Na(+), respectively. Remarkably, the selective K(+) and Na(+) binding sites share several residues, and how the pump is able to achieve the selectivity required for the functional cycle is unclear. Here, free energy-perturbation molecular dynamics (FEP/MD) simulations based on the crystal structures of the Na/K pump in a K(+)-loaded state (E2·P(i)) reveal that protonation of the high-field acidic side chains involved in the binding sites is crucial to achieving the proper K(+) selectivity. This prediction is tested with electrophysiological experiments showing that the selectivity of the E2P state for K(+) over Na(+) is affected by extracellular pH.

Selectivity of externally facing ion-binding sites in the Na/K pump to alkali metals and organic cations.

The Na/K pump is a P-type ATPase that exchanges three intracellular Na(+) ions for two extracellular K(+) ions through the plasmalemma of nearly all animal cells. The mechanisms involved in cation selection by the pump's ion-binding sites (site I and site II bind either Na(+) or K(+); site III binds only Na(+)) are poorly understood. We studied cation selectivity by outward-facing sites (high K(+) affinity) of Na/K pumps expressed in Xenopus oocytes, under voltage clamp. Guanidinium(+), methylguanidinium(+), and aminoguanidinium(+) produced two phenomena possibly reflecting actions at site III: (i) voltage-dependent inhibition (VDI) of outwardly directed pump current at saturating K(+), and (ii) induction of pump-mediated, guanidinium-derivative-carried inward current at negative potentials without Na(+) and K(+). In contrast, formamidinium(+) and acetamidinium(+) induced K(+)-like outward currents. Measurement of ouabain-sensitive ATPase activity and radiolabeled cation uptake confirmed that these cations are external K(+) congeners. Molecular dynamics simulations indicate that bound organic cations induce minor distortion of the binding sites. Among tested metals, only Li(+) induced Na(+)-like VDI, whereas all metals tested except Na(+) induced K(+)-like outward currents. Pump-mediated K(+)-like organic cation transport challenges the concept of rigid structural models in which ion specificity at site I and site II arises from a precise and unique arrangement of coordinating ligands. Furthermore, actions by guanidinium(+) derivatives suggest that Na(+) binds to site III in a hydrated form and that the inward current observed without external Na(+) and K(+) represents cation transport when normal occlusion at sites I and II is impaired. These results provide insights on external ion selectivity at the three binding sites.