GSTM1 Deletion Exaggerates Kidney Injury in Experimental Mouse Models and Confers the Protective Effect of Cruciferous Vegetables in Mice and Humans

Gstm1 KO mice have higher BP and renal oxidative stress at baseline. (A) SBP measured by radiotelemetry was significantly higher in Gstm1 KO mice: WT 129.0±4.9 versus KO 135.6±2.7 mm Hg; P=0.02; n=6 each. (B) Urinary ACR was not different between groups: WT 27.3±7.2 versus KO 29.1±10.1 µg/mg; P=0.73; n=6 each. (C) Twenty four hour excretion of urinary 8-isoprostane was elevated in Gstm1 KO mice: WT 6.6±1.2 versus KO 14.2±3.1 ng/100 mg body wt/24 h; P=0.001; n=5/6. All comparisons by t test. *P<0.05; **P<0.01.

Deletion of Gstm1 in the Nx CDK model results in poor survival and exaggerated HTN and kidney injury. (A) Survival curve showing significant mortality in Gstm1 KO mice after Nx; P=0.01 versus WT; n=8 each. Time of censoring for mice that were used for terminal experiments are marked by ×. (B) SBP measured by radiotelemetry was significantly higher in Gstm1 KO mice: WT 135.0±8.4 versus KO 150.6±13.7 mm Hg; P=0.04; n=4/7. (C) Renal superoxide levels measured by lucigenin luminescence and normalized to dry tissue weight were higher in Gstm1 KO mice: WT 74.5±36.1 versus KO 250.4±153.6; P=0.04; n=6 each. (D) Urinary ACR was higher in Gstm1 KO mice: WT 235.1±60.3 versus KO 748.4±460.4 µg/mg; P=0.04; n=6 each. (E) Kidney pathology scores were greater in Gstm1 KO mice for all compartments (total: WT 8.3±2.3 versus KO 18.7±3.6; P<0.001) and in most individual compartments (glomeruli: WT 6.5±1.2 versus KO 14.5±2.7; P<0.001; tubules: WT 1.0±0.0 versus KO 1.5±0.8; P=0.20; interstitium: WT 0.8±1.2 versus KO 2.7±0.8; P=0.01), n=6 each. (F) Area fractions were greater in Gstm1 KO mice for glomerular to total noninfarcted kidney (G/T, left: WT 2.3±0.7 versus KO 3.3±0.7; P=0.03) and mesangial to glomerular (M/G, right: WT 23.7±6.1 versus KO 35.7±5.5; P=0.005), n=6 each. (G) Percent of scratched area that remained uncovered after 14 hours was reduced in Gstm1 KO mice: WT 69.3±11.5 versus KO 54.2±8.6; P=0.007; n=9 each. All comparisons by t test. *P<0.05; **P<0.01; ***P<0.001.

Deletion of Gstm1 in the AngII HTN model results in exaggerated oxidative stress and kidney injury that is ameliorated by Tempol. (A) SBP measured by radiotelemetry was significantly higher in Gstm1 KO mice treated with Tempol (KO+T): WT 157.8±6.7 versus KO 163.7±14.1 versus KO+T 195.6±13.2 mm Hg; P=0.001; n=5/6/4. (B) Urinary ACR was not significantly different: WT 5108.2±1302.9 versus KO 4657.9±1262.2 µg/mg; P=0.62; n=5/4. (C) Renal superoxide levels measured by lucigenin luminescence and normalized to dry tissue weight were higher in Gstm1 KO mice: WT 41.6±20.8 versus KO 128.25±30.5 versus KO+T 53.8±18.4; P<0.001; n=6/6/4. (D) Kidney pathology scores were greater in Gstm1 KO mice for all compartments (total: WT 8.8±6.1 versus KO 19.8±4.1 versus KO+T 7.3±3.8; P=0.002) and in individual compartments (glomeruli: WT 7.7±4.2 versus KO 15.2±2.7 versus KO+T 7.0±3.6; P=0.003; tubules: WT 0.8±1.3 versus KO 2.3±0.8 versus KO+T 0.3±0.5; P=0.01; interstitium: WT 0.3±0.8 versus KO 2.3±1.5 versus KO+T 0.0±0.0; P=0.006), n=6/6/4. (E) Renal mRNA levels of genes involved in inflammation normalized to Hprt in WT mice at baseline (WTBL) and with HTN (WT), n=6/5 and KO mice at baseline (KOBL), with HTN (KO) and with HTN treated with Tempol (KO+T), n=6/6/4. Expression levels were elevated in the KO mice for MCP-1 (KOBL 0.0054±0.0050 versus KO 0.048±0.016 versus KO+T 0.019±0.0068; P<0.001) and CXCL-1 (KOBL 0.0071±0.0027 versus KO 0.068±0.028 versus KO+T 0.012±0.0051; P<0.001). WT comparisons in (E) by t test. Comparisons in other panels and KO comparisons in (E) by one-way ANOVA. *P<0.05; **P<0.01; ***P<0.001; ****P<0.001.

Gstm1 KO mice in the AngII HTN model have increased renal inflammation associated with the deletion of Gstm1 in the parenchyma. (A) Flow cytometry data of renal leukocytes normalized to WT counts showed Gstm1 KO mice have increased populations (P values: CD4+T=0.04; CD8+T=0.05; B=0.16; PMN=0.007; F4/80_High=0.004), n=3/4. Comparisons by one-way ANOVA. *P<0.05; **P<0.01. (B) Flow cytometry data of renal leukocytes for BM chimeras generated from BM crosstransplantation (genotypes listed as donor-recipient) normalized to WT-WT showed recipient genotype was the statistically significant factor (donor P values: CD4+T=0.82; CD8+T=0.95; B=0.87; PMN=0.70; F4/80_High=0.31; recipient P values: CD4+T=5.9×10⁻⁵; CD8+T=0.30; B=1.1×10⁻⁵; PMN=1.5×10⁻⁶; F4/80_High=5.0×10⁻⁷), n=4/5/5/4. Comparisons by two-way ANOVA.

Treatment with SRBP ameliorated kidney injury only in Gstm1 KO mice. In all panels, WT and KO mice are the same as those in Figure 3, with the data reproduced here for ease of comparison. (A) SBP measured by radiotelemetry was elevated in WT mice fed SRBP (WT+SRBP 176.9±5.9 mm Hg; P=0.001; n=6) but not in Gstm1 KO mice fed SRBP (KO+SRBP 168.9±3.5 mm Hg; P=0.24; n=6). (B) Renal superoxide levels measured by lucigenin luminescence and normalized to dry tissue weight were not affected by SRBP in WT mice (WT+SRBP 57.7±31.5 counts/min per milligram dry tissue; P=0.02; n=6) but were reduced in Gstm1 KO mice (KO+SRBP 50.4±33.9 counts/min per milligram dry tissue; post hoc versus KO P=0.004; n=5). (C) Urinary ACR was reduced by SRBP in Gstm1 KO mice (KO+SRBP 1671.5±683.6 µg/mg; post hoc versus KO P=0.05; n=5), but not WT mice (WT+SRBP 4314.4±2155.3 µg/mg; P=0.47; n=6). (D) Kidney pathology scores were unaffected by SRBP in WT mice (total: 5.4±1.5; P=0.24; glomeruli: 4.4±0.5; P=0.11; tubules: 1.0±1.0; P=0.82; interstitium: 0.0±0.0; P=0.36; n=5) but were reduced in Gstm1 KO mice (total: 4.6±1.7; post hoc versus KO P=2.0×10⁻⁶; glomeruli: 3.6±0.9; post hoc versus KO P=2.4×10⁻⁷; tubules: 1.0±1.0; post hoc versus KO P=0.10; interstitium: 0.0±0.0; P=0.004; n=5). Comparisons between WT and WT+SRBP by t test. Comparisons between WT+SRBP, KO, and KO+SRBP by one-way ANOVA. *P<0.05; **P<0.01; ***P<0.001; ****P<0.001.