Hypertensive Kidney Disease and The Role of Toll-Like Receptor 4

Abstract

Introduction: Hypertension is one of the leading causes of kidney disease. Persistently high, uncontrolled blood pressure (BP) can constrict the blood vessels within the kidneys, impairing their ability to regulate fluids and eliminate waste. This deterioration of renal function can result in serious health complications. Moreover, hypertension can initiate a self-perpetuating cycle of innate immune activation, which contributes to structural changes and worsens kidney dysfunction. Research from our laboratory and others has identified toll-like receptors (TLRs) as key mediators in this immune response associated with hypertensive kidney injury. Of the thirteen known TLRs in mammals, TLR4 plays a particularly significant role: its activation during hypertension triggers a range of cellular responses, some of which are pathological and contribute to kidney damage. Conversely, studies have shown that genetic mutation (beneficial) or pharmacological inhibition of TLR4 can limit organ injury, including kidney damage, in hypertensive models. Despite these findings, the precise mechanisms by which TLR4 contributes to hypertensive kidney damage and dysfunction remain unclear.

Objectives: The main objective of this dissertation was to investigate whether hypertension, via the stress response, activates TLR4, subsequently triggering inflammatory pathways, tissue remodeling, and kidney dysfunction. A further objective was to determine if mutation or inhibition of TLR4 can mitigate pathological kidney remodeling and dysfunction associated with hypertension.

Methods: In vivo experiments were conducted using both female and male C3H/HeOuJ and C3H/HeJ mice, aged 10–12 weeks. The C3H/HeOuJ mice possess normal TLR4 function (designated TLR4N in this study), whereas C3H/HeJ mice carry a spontaneous mutation in the TLR4 gene (designated TLR4M). Hypertension and related pathologies were induced in these mice by infusing them with angiotensin II (Ang-II, 1000 ng/kg/min) using Alzet mini-osmotic pumps for four weeks. Physiological and pathological parameters, including systolic and mean arterial pressure (MAP) and kidney function as indicated by glomerular filtration rate (GFR), were monitored throughout and at the conclusion of the 4-week period. At the end of the experiments, kidneys were collected for biochemical, histological, protein, and mRNA analyses. For the in vitro model, mouse kidney mesangial cells were cultured in a humidified incubator at 37°C with 5% CO2 until they reached 60% to 70% confluence, displaying a uniform density. Cells were seeded in DMEM/F-12 (50/50) medium supplemented with 5% fetal bovine serum, antibiotics, and L-glutamine. In a 6-well plate, the cells were pre-treated with 200 nM TAK-242 (a TLR4 inhibitor) for one hour, followed by exposure to 200 nM Angiotensin-II for 24 hours. Subsequently, the cells were collected for further laboratory analysis. Appropriate controls were used in all experiments.

Results: Results showed that Ang II significantly elevated blood pressure in TLR4N mice, whereas TLR4M mice were resistant to this increase under the same Ang II treatment. In TLR4N mice, hypertension also led to a significant upregulation of TLR4 expression in the kidney, increased levels and activity of matrix metalloproteinases (MMPs), and reduced levels of their natural inhibitors, tissue inhibitors of matrix metalloproteinases (TIMPs). Similar results were observed in in vitro experiments with mesangial cells. Additionally, TLR4N hypertensive mice exhibited elevated inflammatory markers, increased apoptosis, and pyroptosis in the kidneys, all of which were mediated by activation of the NF-κB signaling pathway. Moreover, TLR4N mice displayed increased collagen genes expression, glomerular hypercellularity, reduced renal blood flow, higher resistive index, and impaired renal function following Ang II-induced hypertension. Notably, all these adverse changes were mitigated in TLR4M mice subjected to the same hypertensive regimen.

Summary and Conclusion: Hypertension activates the TLR4 receptor, triggering a cascade of immune responses that disrupts the balance of MMPs and TIMPs in the renal vasculature. This activation also drives pyroptosis and apoptosis via the NF-kB signaling pathway, ultimately leading to impaired kidney function. Conversely, TLR4 mutation or inhibition offers targeted strategies to mitigate vascular damage and enhance renal function in hypertension. Together, these findings indicate that hypertension promotes kidney remodeling and dysfunction through TLR4 activation, while TLR4 mutation (TLR4M) or inhibition confers protection against renal remodeling and improves kidney function in hypertensive conditions. Further research is required to validate these model-based findings in preclinical and clinical studies.