Blood samples were collected from each dog before (time  = 0), and 2, 4, 24, 48, and 96 h after either hydrodynamic injection or slow injection (1,000 ml) in 1 or 2 h through the peripheral vein as controls. The serum biochemical analysis and hemogram were performed by the Cleveland Office of Marshfield Labs and BML Inc. (Shibuya-ku, Tokyo, Japan). The cytokines were analyzed by the GeneticLab. Co. Ltd. (Sapporo, Hokkaido, Japan) using the Luminex 200 System with the MILLIPLEXMAP Canine Cytokine/Chemokine Kit (MILLIPORE, CCYTO-90K) [18] (link), [19] (link). Tissue samples for hematoxylin and eosin (H&E) staining were collected before, immediately following, and 3 h after the hydrodynamic injection to rats, and before, immediate following, and 24 h after the injection to dogs. Three different liver sections from each of the 3 rats and 2 dogs were stained, images were captured, and a quantitative analysis of the sinusoidal area was performed using ImageJ software (version 1.6.0_20, National Institutes of Health, USA) as previously reported [20] (link).
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