*2.4. SH045 Treatment Leads to Less Renal Immune Cell Infiltration*

To evaluate inflammatory cell infiltration in UUO kidneys, we examined macrophages and T cell presence using immunofluorescence. Kidney cross sections were immunolabelled with the macrophage marker F4/80 and T cell marker CD4 as described previously [17]. As shown in Figure 4A,B, excessive CD4-positive cells infiltration was observed in renal interstitium of UUO kidneys in comparison to control kidneys (Figure 4A,B). Similarly, the number of F4/80-positive cells in UUO kidneys was also markedly increased compared to control kidneys (Figure 4C,D). In accordance with ameliorated inflammatory cytokine and chemokine expression, SH045 treatment decreased UUO-induced macrophage and T cell infiltration (Figure 4A–D). Thus, these data suggest that TRPC6 inhibition reduces renal inflammation in the UUO model of NZO mice.

**Figure 4.** SH045 impact on renal inflammatory cell accumulation after UUO. (**A**) Representative images of control and UUO-injured kidneys stained with CD4+ T cells (magnification: 400×). Rectangles represent single-cell magnifications. Scale bars are 50 μm. (**B**) Quantification in renal infiltration of CD4+ T cells (control *n* = 6, UUO *n* = 8). (**C**) Representative images of control and UUO-injured kidneys stained with F4/80+ macrophages (magnification: 400×). Rectangles represent single-cell magnifications. Scale bars are 50 μm. (**D**) Quantification in renal infiltration of F4/80+ macrophages (control *n* = 6, UUO *n* = 8). Data expressed as means ± SD. Two-way ANOVA followed by Sidak's multiple comparisons post hoc test. \* *p* < 0.05, \*\*\* *p* < 0.001, and \*\*\*\* *p* < 0.0001 defined as significant. ns, not statistically significant. AU, arbitrary units.
