Systemic Inflammation Disrupts Circadian Rhythms and Diurnal Neuroimmune Dynamics

Abstract

Circadian rhythms regulate physiological processes in approximately 24 h cycles, and their disruption is associated with various diseases. Inflammation may perturb circadian rhythms, though these interactions remain unclear. This study examined whether systemic inflammation induced by an intraperitoneal injection of lipopolysaccharide (LPS) could alter central and peripheral circadian rhythms and diurnal neuroimmune dynamics. Mice were randomly assigned to two groups: the saline control group and the LPS group. The diurnal expression of circadian clock genes and inflammatory cytokines were measured in the hypothalamus, hippocampus, and liver. Diurnal dynamic behaviors of microglia were also assessed. Our results revealed that the LPS perturbed circadian gene oscillations in the hypothalamus, hippocampus, and liver. Furthermore, systemic inflammation induced by the LPS could trigger neuroinflammation and perturb the diurnal dynamic behavior of microglia in the hippocampus. These findings shed light on the intricate link between inflammation and circadian disruption, underscoring their significance in relation to neurodegenerative diseases.

Keywords: circadian disruption; clock gene expression; microglia; peripheral circadian clocks.

Figure 1

The intraperitoneal injection of an LPS induced systemic inflammation and perturbed the diurnal expression of proinflammatory cytokines in the liver. The mRNA expressions of (A) IL-1β, (B) IL-6, (C) MCP-1, and (D) TNF-α in the liver at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21) were measured by RT-qPCR and relative to the endogenous gene GAPDH. Data are expressed as mean ± SEM. n = 3–5 mice per group for each time point. * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001 by unpaired two-tailed Student’s t-test.

Figure 2

The intraperitoneal injection of an LPS induced systemic inflammation and perturbed the diurnal expression of proinflammatory cytokines in the hypothalamus. The mRNA expressions of (A) IL-1β, (B) IL-6, (C) MCP-1, and (D) TNF-α in the hypothalamus at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21) were measured by RT-qPCR and relative to the endogenous gene GAPDH. Data are expressed as mean ± SEM. n = 3–5 mice per group for each time point. * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001 by unpaired two-tailed Student’s t-test.

Figure 3

The intraperitoneal injection of the LPS induced systemic inflammation and altered the diurnal expression of proinflammatory cytokines in the hippocampus. The mRNA expressions of (A) IL-1β, (B) IL-6, (C) MCP-1, and (D) TNF-α in the hippocampus at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21) were measured by RT-qPCR and relative to the endogenous gene GAPDH. Data are expressed as mean ± SEM. n = 3–5 mice per group for each time point. * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001 by unpaired two-tailed Student’s t-test.

Figure 4

The intraperitoneal injection of the LPS induced the activation of microglia and perturbed the diurnal dynamic behavior of microglia in the CA1 of the hippocampus. (A) Representative confocal images of the immunohistochemical staining of DAPI and Iba1 in the CA1 of the hippocampus sections of the control and LPS groups at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21) (Scale bar = 20 μm; 10 μm for inset). The (B) number of Iba1 positive cells, (C) the relative fluorescence intensity of Iba1 positive cells, the (D) summed process length per Iba1 positive cell, and the (E) summed endpoints per Iba1 positive cell in the CA1 of the hippocampus sections were quantified at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21). Data are expressed as mean ± SEM. n = 3–5 mice per group for each time point. * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001 by unpaired two-tailed Student’s t-test.

Figure 5

The intraperitoneal injection of the LPS induced the activation of microglia and perturbed the diurnal dynamic behavior of microglia in the CA3 of the hippocampus. (A) Representative confocal images of the immunohistochemical staining of DAPI and Iba1 in the CA3 of the hippocampus sections of the control and LPS groups at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21) (Scale bar = 20 μm; 10 μm for inset). The (B) number of Iba1 positive cells, (C) the relative fluorescence intensity of Iba1 positive cells, the (D) summed process length per Iba1 positive cell, and the (E) summed endpoints per Iba1 positive cell in the CA3 of the hippocampus sections were quantified at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21). Data are expressed as mean ± SEM. n = 3–5 mice per group for each time point. * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001 by unpaired two-tailed Student’s t-test.

Figure 6

The intraperitoneal injection of the LPS induced the activation of microglia and perturbed the diurnal dynamic behavior of microglia in the DG of the hippocampus. (A) Representative confocal images of the immunohistochemical staining of DAPI and Iba1 in the DG of the hippocampus sections of the control and LPS groups at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21) (Scale bar = 20 μm; 10 μm for inset). The (B) number of Iba1 positive cells, (C) the relative fluorescence intensity of Iba1 positive cells, the (D) summed process length per Iba1 positive cell, and the (E) summed endpoints per Iba1 positive cell in the DG of the hippocampus sections were quantified at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21). Data are expressed as mean ± SEM. n = 3–5 mice per group for each time point. * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001 by unpaired two-tailed Student’s t-test.

Figure 7

The intraperitoneal injection of the LPS disrupted the diurnal oscillations of the circadian genes in the liver. The mRNA expression of (A) Bmal1, (B) Clock, (C) Cry1, (D) Cry2, (E) Per1, (F) Per2, and (G) Per3 in the liver at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21) were measured by RT-qPCR and relative to the endogenous gene GAPDH. Data are expressed as mean ± SEM. n = 3–5 mice per group for each time point. * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001 by unpaired two-tailed Student’s t-test.

Figure 8

The intraperitoneal injection of an LPS disrupted the diurnal oscillations of the circadian genes in the hypothalamus. The mRNA expression of (A) Bmal1, (B) Clock, (C) Cry1, (D) Cry2, (E) Per1, (F) Per2, and (G) Per3 in the hypothalamus at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21) were measured by RT-qPCR and relative to the endogenous gene GAPDH. Data are expressed as mean ± SEM. n = 3–5 mice per group for each time point. * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001 by unpaired two-tailed Student’s t-test.

Figure 9

The intraperitoneal injection of the LPS disrupted the diurnal oscillations of the circadian genes in the hippocampus. The mRNA expressions of (A) Bmal1, (B) Clock, (C) Cry1, (D) Cry2, (E) Per1, (F) Per2, and (G) Per3 in the hippocampus at 4 different time points (ZT 3, ZT 9, ZT 15, and ZT 21) were measured by RT-qPCR and relative to the endogenous gene GAPDH. Data are expressed as mean ± SEM. n = 3–5 mice per group for each time point. * p < 0.05; ** p < 0.01; *** p < 0.001; and **** p < 0.0001 by unpaired two-tailed Student’s t-test.