Neuronal CCL2 is upregulated during hepatic encephalopathy and contributes to microglia activation and neurological decline

Abstract

BackgroundAcute liver failure leads to systemic complications with one of the most dangerous being a decline in neurological function, termed hepatic encephalopathy. Neurological dysfunction is exacerbated by an increase of toxic metabolites in the brain that lead to neuroinflammation. Following various liver diseases, hepatic and circulating chemokines, such as chemokine ligand 2 (CCL2), are elevated, though their effects on the brain following acute liver injury and subsequent hepatic encephalopathy are unknown. CCL2 is known to activate microglia in other neuropathies, leading to a proinflammatory response. However, the effects of CCL2 on microglia activation and the pathogenesis of hepatic encephalopathy following acute liver injury remain to be determined.MethodsHepatic encephalopathy was induced in mice via injection of azoxymethane (AOM) in the presence or absence of INCB 3284 dimesylate (INCB), a chemokine receptor 2 inhibitor, or C 021 dihydrochloride (C021), a chemokine receptor 4 inhibitor. Mice were monitored for neurological decline and time to coma (loss of all reflexes) was recorded. Tissue was collected at coma and used for real-time PCR, immunoblots, ELISA, or immunostaining analyses to assess the activation of microglia and consequences on pro-inflammatory cytokine expression.ResultsFollowing AOM administration, microglia activation was significantly increased in AOM-treated mice compared to controls. Concentrations of CCL2 in the liver, serum, and cortex were significantly elevated in AOM-treated mice compared to controls. Systemic administration of INCB or C021 reduced liver damage as assessed by serum liver enzyme biochemistry. Administration of INCB or C021 significantly improved the neurological outcomes of AOM-treated mice, reduced microglia activation, reduced phosphorylation of ERK1/2, and alleviated AOM-induced cytokine upregulation.ConclusionsThese findings suggest that CCL2 is elevated systemically following acute liver injury and that CCL2 is involved in both the microglia activation and neurological decline associated with hepatic encephalopathy. Methods used to modulate CCL2 levels and/or reduce CCR2/CCR4 activity may be potential therapeutic targets for the management of hepatic encephalopathy due to acute liver injury.

Bile Acid Signaling Is Involved in the Neurological Decline in a Murine Model of Acute Liver Failure

“…17,18 Cerebral edema was assessed in all mice using the wet/dry weight method described by Baskaya et al 20 Water content was expressed as a percentage of brain weight, calculated as follows: ([wet weight À dry weight]/wet weight) Â 100%. Increased brain water content of 1% to 2% in mice is indicative of cerebral edema and is characterized by increased intracranial pressure.…”

“…40 Increased expression of the inflammatory chemokine ligand 2 has been attributed to AOM-induced HE progression and knockout of the IL-1 receptor, or the tumor necrosis factor receptor confers neuroprotection after AOM-induced HE. 18,41 Data from the AOM model of acute liver failure indicate that ammonia concentrations are elevated only in the later stages of HE, well beyond the onset of overt neurological impairment, 42 indicating that other factors must be playing a role in the early stages of encephalopathy onset. The data presented herein suggest that serum bile acids may also be playing a role in the pathogenesis of HE.…”

Neuroinflammatory Signals during Acute and Chronic Liver Diseases

“…Both microglia activation (as demonstrated by an ameboidal phenotype) and motility (as demonstrated by analysis of the turnover rate) were shown to be altered in the cerebral cortex at late stages of HE when severe neurological symptoms were evident, coinciding with the appearance of brain edema [4]. Furthermore, increased number of microglia [5][6][7] and increased reactive phenotype [6] has been demonstrated in the cerebral cortex of AOMinjected mice. This HE-associated microgliosis could be atenuated with anti-inlammatory treatment modalities [5][6][7], which also atenuated or delayed various neurocognitive deicits [8].…”

“…Furthermore, increased number of microglia [5][6][7] and increased reactive phenotype [6] has been demonstrated in the cerebral cortex of AOMinjected mice. This HE-associated microgliosis could be atenuated with anti-inlammatory treatment modalities [5][6][7], which also atenuated or delayed various neurocognitive deicits [8]. While mice injected with TAA displayed a signiicant reduction in locomotor activity, which was accompanied by increased expression of certain proinlammatory cytokines and chemokines, no microglia activation was observed [8].…”

“…Mechanisms of Neuroinflammationdysregulated leading to an increased production of CCL2 [5] and a downregulation of fractalkine [6] and subsequent microglia activation [5,6]. This imbalance may, at least in part, be atributable to aberrant bile acid signaling in the frontal cortex during acute liver failure [29].…”