The Role of BMAL1 in LPS-Induced Depressive Behavior and Related “Inflammatory Storm”

Introduction

According to the 2019 Global Burden of Disease Study, mental disorders are listed as one of the top 10 causes of global burden, with depression being a major contributor. Over 350 million people worldwide suffer from depression, making it the most common cause of disability globally. Despite the availability of numerous antidepressants in clinical practice, more than 30% of patients are resistant to first-line treatment with selective serotonin reuptake inhibitors (SSRIs) due to their limited effectiveness in initial treatment. Although the N-methyl-D-aspartate receptor (NMDAR) antagonist ketamine has attracted great interest due to its powerful and rapid antidepressant properties and has been reported as a new direction for treating depression, its exact mechanism of action remains unclear. Therefore, in-depth research on the pathogenesis of depression and the development of new effective drug treatments is an urgent task.

In addition to the known pathophysiological effects of stress and gut-brain axis dysfunction, increasing evidence suggests that neuroinflammation or neuroimmune mechanisms play an important role in the progression of depression. Infected patients are more likely to develop depression, and depressed patients also excessively release inflammatory factors known as “inflammatory storm” or “cytokine storm”, such as C-reactive protein (CRP), interleukin-6 (IL-6), and tumor necrosis factor-α (TNF-α). Moreover, recent meta-analyses show that patients unresponsive to antidepressants have abnormal inflammatory processes, including high baseline levels of CRP. Our previous studies found that rats induced with depression by chronic stress or lipopolysaccharide (LPS) exhibited depressive behavior, accompanied by increased plasma concentrations of CRP and IL-6, and decreased brain-derived neurotrophic factor (BDNF). These findings support the pathobiological overlap between inflammation and depression.

Microglia are key regulators of inflammatory responses in the central nervous system (CNS) and are the resident macrophages of the CNS. The activation state of microglia determines whether they are pro-inflammatory or neuroprotective. Activated microglia promote the release of inflammatory factors, leading to neuronal damage. Our previous research results found that microglia accumulate in the cerebellum and hippocampus of Alzheimer’s disease (AD) model rats.

Multiple studies have shown that inflammation alters circadian rhythms by disrupting biological clock mechanisms, and therapies targeting IL-6 can improve sleep disorders. As important regulators of microglia, imbalances in the initiator receptor family may affect internal activities and produce various results in different diseases. Furthermore, circadian rhythm disorders are common in mental disorders, and depression has a bidirectional relationship with circadian rhythm disorders. Circadian disruption may promote the occurrence and severity of depression, and conversely, depression can further disrupt circadian rhythms. Synchronizing the brain clock with the external environment can alleviate depressive symptoms.

Based on the above background, this study aims to explore the role of circadian clock genes, especially brain and muscle ARNT-like protein 1 (BMAL1), in the relationship between inflammation and depression, using LPS-challenged rats and BV2 cells as research models.

Research Source

This study was conducted by scholars Xu Dan-Dan, Hou Zhi-Qi, Xu Ya-Yun, et al., from institutions including Anhui Medical University and the General Hospital of the People’s Liberation Army. The paper was accepted for publication in January 2024 by the Journal of Neuroimmune Pharmacology.

Experimental Design and Procedures

Experimental Animals and Model Establishment

Two-month-old male SD rats were used for the experiment. After one week of adaptation, they were randomly divided into control and LPS model groups. The model group rats received intraperitoneal injections of 0.5mg/kg LPS (derived from E. coli, serotype 055:B5) every other day, while the control group received an equal volume of sterile saline. Forced swim tests (FST) were conducted 24 hours after administration, and a series of behavioral tests were performed after four injections were completed.

Behavioral Tests

All behavioral tests were conducted in a quiet, soundproof room, including open field test (OFT), elevated plus maze test (EPM), Y-maze test (Y-Maze), and sucrose preference test (SPT). The experimental results showed that rats in the model group exhibited depressive-like behavior after four LPS injections, confirming the successful induction of the depression model.

Inflammatory Storm Detection

LPS injection resulted in significantly increased serum concentrations of IL-6, TNF-α, and CRP in the model group rats, as well as increased protein expression levels of IL-6 and TNF-α in the hippocampus, confirming that LPS induced an “inflammatory storm”.

HPA Axis Activity and Adrenal Cortex Morphology Observation

LPS treatment caused a significant increase in cortisol (CORT) concentration and hypothalamic CRH mRNA levels in the model group rats, with mild to moderate morphological changes in the adrenal cortex, indicating HPA axis hyperactivity.

BDNF and Synaptic Plasticity-Related Protein Expression Detection

As the LPS challenge time increased, the expression of BDNF and synaptic-associated proteins (such as Syt-1) gradually decreased in the hippocampus and hypothalamus, suggesting the crucial role of BDNF and synaptic plasticity in depression.

Circadian Rhythm-Related Indicator Detection

After LPS treatment, significant changes were observed in the model group rats’ anal temperature fluctuations, serum melatonin, and corticosterone concentrations. Hippocampal BMAL1 protein expression increased, while PER2, CRY2, and CLOCK proteins significantly decreased. The opposite was observed for BMAL1 expression in the hypothalamus, suggesting circadian rhythm disruption.

Cell Experiments

Using BV2 microglial cells as an in vitro model, it was found that LPS treatment led to decreased cell viability, increased phagocytic ability, increased TNF-α and IL-6 protein expression, and increased BMAL1 protein expression. After downregulating BMAL1 expression through small interfering RNA (siRNA) technology, the pathological changes caused by LPS were reversed, indicating the important role of BMAL1 in inflammatory responses.

Research Results and Significance

Main Results

1. LPS induced depressive-like behavior in rats, manifested as increased immobility time in FST and decreased sucrose preference index in SPT.
2. LPS caused HPA axis hyperactivity and “inflammatory storm”, with significant increases in serum and brain TNF-α and IL-6.
3. LPS led to decreased expression of BDNF and synaptic-related proteins in the rat hippocampus and hypothalamus, with increased BMAL1 in the hippocampus but decreased in the hypothalamus.
4. In BV2 cell experiments, LPS treatment increased BMAL1, TNF-α, and IL-6 protein expression, and inhibiting BMAL1 could reverse these changes.

Conclusions and Highlights

This study reveals the potential bridging role of BMAL1 between inflammation and depression and points out that BMAL1 imbalance may lead to inflammatory storms and circadian rhythm disorders, thereby triggering pathological changes in depression. This finding helps explore new antidepressant drug targets and provides new ideas for disease prevention and treatment.

Research Significance and Value

This study not only provides a new mechanism for BMAL1 in inflammation-related depression but also links circadian genes with depression and neuroinflammation, expanding the understanding of the pathogenesis of depression. At the same time, the research results show that regulating BMAL1 and its related pathways may become a new method for depression treatment, providing valuable information for the development of future treatment strategies.