HECTD2 knockdown ameliorates Pseudomonas-induced lung injury in vivo

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HECTD2 knockdown ameliorates Pseudomonas-induced lung injury in vivo

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Fig. 6. HECTD2 knockdown ameliorates Pseudomonas-induced lung injury in vivo. HECTD2 knockdown ameliorates Pseudomonas-induced lung injury in vivo. C57BL/6J mice were infected intratracheally with Lenti-control shRNA or Lenti-HECTD2 shRNA (107 PFU per mouse) for 144 hours, and mice were inoculated with PA103 [104 colony-forming units (CFU) per mouse] for 18 hours. Mice were euthanized, and lungs were lavaged with saline, harvested, and then homogenized. (A to C) Lavage protein, cell count, and bacteria count were measured. (D to F) Lavage cytokine secretion was measured. (G) Survival studies of mice that were administered PA103 (intratracheally, 105 PFU per mouse, n = 8 mice per group) were determined (time, hours). Mice were carefully monitored over time; moribund, preterminal animals were immediately euthanized and recorded as deceased. Kaplan-Meier survival curves were generated using SPSS software (P = 0.001). (H) H&E staining was performed on lung samples from (A). (I) PIAS1, HECTD2, and actin immunoblots from homogenized lung samples. Scale bar, 100 μm; original magnification, ×10. Data are average of two experiments (Student’s t test on means ± SEM). (A to G) n = 6 to 9 mice per group. NS, not significant. Source data in fig. S13 and table S6. Tiffany A. Coon et al., Sci Transl Med 2015;7:295ra109 Copyright © 2015, American Association for the Advancement of Science