CD247 modulates blood pressure by altering T-lymphocyte infiltration in the kidney

Abstract

The CD3 ζ chain (CD247), a gene involved in T-cell signaling, has been shown to associate with blood pressure in human genetic studies. To test the functional role of CD247 in hypertension and renal disease, zinc-finger nucleases targeting CD247 were injected into Dahl salt-sensitive (SS/JrHsdMcwi) embryos. The resulting 11-bp frameshift deletion in exon 1 of CD247 led to a predicted premature stop codon. Western blotting confirmed the absence of CD247 protein in the thymus, and flow cytometry (n=5-9 per group) demonstrated that the mutant rats (CD247(-/-)) have a >99% reduction in circulating CD3(+) T cells compared with littermate controls (CD247(+/+)). Studies were performed on age-matched, littermate male, CD247(+/+) and CD247(-/-) rats fed a 4.0% NaCl diet for 3 weeks. The infiltration of CD3(+) T cells into the kidney after high salt was significantly blunted in CD247(-/-) (1.4±0.4×10(5) cells per kidney) when compared with that in the CD247(+/+) (8.7±2.0×10(5) cells per kidney). Accompanying the reduced infiltration of T cells, mean arterial blood pressure was significantly lower in CD247(-/-) than in CD247(+/+) (134±1 versus 151±2 mm Hg). As an index of kidney disease, urinary albumin and protein excretion rates were significantly reduced in CD247(-/-) (17±1 and 62±2 mg/d, respectively) when compared with that in CD247(+/+) (49±3 and 121±5 mg/d, respectively). Glomerular and renal tubular damage were also attenuated in the CD247(-/-). These studies demonstrate that functional T cells are required for the full development of Dahl salt-sensitive hypertension and indicate that the association between CD247 and hypertension in humans may be related to altered immune cell function.

Keywords: hypertension; immune system; kidney; lymphocytes; rats.

FIGURE 1

Cytokine expression in T cells isolated from the blood and kidney of Dahl SS rats fed a high salt (4.0% NaCl) diet for three weeks. * indicates P<0.05 vs. peripheral cells.

FIGURE 2

Representative flow cytometric analysis of CD5+, CD3+ and CD45R+ cells in CD247+/+ (A,C) and CD247−/− rats (B,D). The graph illustrates mean numbers of PBMC and CD3+, CD5+ and CD45R+ lymphocytes in CD247+/+ and CD247−/− rats (E). * indicates P<0.05 vs. CD247+/+.

FIGURE 3

Changes in mean arterial pressure (A) and albumin excretion rate (B) in CD247+/+ and CD247−/− rats fed AIN-76A diet containing 0.4% NaCl followed by 21 days of 4.0% NaCl. * indicates P<0.05 vs.values obtained on the final day of 0.4% NaCl chow, † indicates P<0.05 vs. CD247+/+ on the same day.

FIGURE 4

Light microscopy of trichome stained sections of the renal outer medulla (A and B, 10× original magnification) and renal cortex (C and D, 40× original magnification) of CD247+/+ (A and C) or CD247−/− (B and D) fed 4.0% NaCl chow for three weeks. The percentage of the renal outer medulla consisting of protein casts (E), the calculated glomerular injury score (F), and the number of infiltrating B lymphocytes (CD45R+), T lymphocytes (CD3+) and macrophages/monocytes (CD11b+) in the kidney of CD247+/+ and −/− rats following three weeks of 4.0% NaCl diet are also plotted (G). * indicates P<0.05 vs. CD247−/−.