Divergent Sex-Specific Effects of Toll-like Receptor 4 and Cluster of Differentiation 14 in Dahl Salt-Sensitive Hypertension

Abstract

Renal inflammation accompanies hypertension and renal damage in Dahl Salt-Sensitive rats fed a high salt diet (HS, 4.0% NaCl). Via a transcriptomics approach, we previously found cluster of differentiation 14 (CD14) and toll-like receptor 4 (TLR4), partner molecules involved in innate immune system activation, to be upregulated in renal medullary tissue of Dahl SS rats after a HS challenge. A subsequent study demonstrated that both CD14 and TLR4 are upregulated in macrophages infiltrating the kidneys of Dahl SS fed HS. Interestingly, CD14 deletion on the Dahl SS background confers an amplification of hypertension and renal damage in females but has no effect in males. Given this finding, we examined sex-specific effects of TLR4 deletion in the pathogenesis of Dahl SS hypertension. The current study tested the hypothesis that TLR4 knockout differentially modulates salt-sensitivity in males versus females. A mutation was generated in the TLR4 gene on the Dahl SS background using CRISPR-Cas9 technology to create a 1-basepair frameshift mutation resulting in a premature stop codon. SS TLR4+/+ and SS TLR4-/- males (n=10) and females (n=6), maintained on a low salt chow (LS, 0.4% NaCl), underwent telemeter implantation at approximately 7 weeks of age for continuous monitoring of blood pressure. Following recovery and baseline blood pressure recordings, animals were switched to a HS chow for 3 weeks. Urine samples were collected once during the LS period and weekly throughout the HS challenge. Kidneys were flushed and harvested for analysis. Mean arterial pressure (MAP) was not different between the groups during the baseline period (117.3 ± 1, 116.9 ± 1, 117.3 ± 1, 116.9 ± 2 mmHg). Throughout the HS challenge, MAP was significantly reduced in the male SS TLR4-/- compared to male SS TLR4+/+ (day 19: 150.8 ± 4 vs. 162.5 ± 5 mmHg, p=0.010), but MAP was unchanged in female SS TLR4-/- compared to SSTLR4+/+ (151.6 ± 4 vs 155.3 ± 4 mmHg, p=0.554). Similarly, urinary albumin excretion was reduced in the male SS TLR4-/- versus SS TLR4+/+ (170.3 ± 27 vs 321.3 ± 63 mg/day, p=0.023), indicative of reduced renal injury in these animals. Albuminuria was not observed to be different between female SS TLR4-/- and SS TLR4+/+ (149.7 ± 24 vs 145.9 ± 14, p=0.448). At the end of the HS challenge, immune cells were isolated from the kidneys of these animals and characterized by flow cytometric analysis. We observed no difference in either the circulating or infiltrating immune cell profile between wildtypes and knockouts. These results indicate a male-specific effect of TLR4 deletion and suggest that TLR4 promotes salt-sensitive hypertension and renal damage in males. This finding contrasts with the effect of TLR4’s partner molecule, CD14, which attenuates SS hypertension in females. Further studies are needed to elucidate the molecular mechanisms of TLR4/CD14 activation and signaling during salt-sensitive hypertension. HL161231 This is the full abstract presented at the American Physiology Summit 2023 meeting and is only available in HTML format. There are no additional versions or additional content available for this abstract. Physiology was not involved in the peer review process.