2, C and D). After 4 h of CY treatment, albumin extravasation was significantly increased in GSTP-null KOS 953 and WT mice over untreated controls, and albumin was slightly more in GSTP-null compared with WT mice (Fig. 2E), although this did not reach statistical significance (0.10 > p > 0.05). Fig. 2. Urinary bladder toxicity of CY in WT and GSTP-null mice. A, photomicrographs of sections from the urinary bladder of WT and GSTP-null mice treated with CY (200 mg/kg, i.p. for 24 h). Areas of lamina propria (LP), urothelium (UE), and muscularis propria … Table 1 Histopathology of CY-induced bladder injury in WT and GSTP-null mice Because histamine release from mast cells could contribute to CY-induced increase in vascular permeability and urinary bladder edema (Bjorling et al.

, 1999), the number of mast cells in bladder lamina propria was measured by acidified toluidine blue staining. There was no change in mast cell number or granulation status at 4 or 24 h after treatment in either WT or GSTP-null mice (data not shown). In contrast, a significant increase in MPO-positive (MPO+) cells was observed at 4 h post-treatment (Table 2). The number of MPO+ cells was significantly greater (by 269 �� 47%; n = 5) in CY-treated GSTP-null mice, indicating a greater level of inflammation in the GSTP-null mice, than in WT mice. Moreover, there was an ~10-times increase in the number of cells stained positive for apoptosis (TUNEL+ stain) in the lamina propria layer in WT (WT + saline, 1.3 �� 0.6; WT + CY, 13.5 �� 2.9; n = 6, 6) and GSTP-null (null + saline, 1.6 �� 0.7; null + CY, 14.6 �� 3.

6; n = 5, 5) bladders at 24 h after CY treatment. However, there was no difference in the number of or distribution of TUNEL+ cells in the urinary bladder of WT and GSTP-null mice (Fig. 3). Table 2 Neutrophil infiltration after CY treatment in WT and GSTP-null mice Fig. 3. CY-induced apoptosis in the urinary bladder. WT and GSTP-null mice were treated with CY (200 mg/kg, i.p.), and apoptosis was measured in the urinary bladder 24 h after treatment. Positive apoptosis (TUNEL) staining in the lamina propria of CY-treated … CY and Acrolein Metabolism. To assess the effect of GSTP deficiency on clearance of CY-derived acrolein, the concentration of HPMA, the primary metabolite of acrolein, was measured in the urine of mice treated with either a nontoxic dose of CY (50 mg/kg) or acrolein (2 mg/kg, i.

p.). HPMA concentration was measured by gas chromatography/mass spectrometry using 13C-HPMA as an internal standard (Fig. 4). Treatment with either CY or acrolein significantly increased urinary HPMA; however, there were no differences between WT and GSTP-null Anacetrapib mice in basal, CY-, or acrolein-induced HPMA levels (Table 3). Moreover, CY treatment had no effect on urine flow, urine albumin, urine creatinine, or urine total protein in WT and GSTP-null mice (data not shown).