Minocycline attenuates experimental autoimmune encephalomyelitis in rats by reducing T cell infiltration into the spinal cord

doi:10.1016/j.jneuroim.2009.11.009

Journal of Neuroimmunology

Volume 219, Issues 1–2, 26 February 2010, Pages 33-37

https://doi.org/10.1016/j.jneuroim.2009.11.009 Get rights and content

Abstract

We investigated the anti-inflammatory effects of minocycline in EAE, an animal model of MS. Minocycline, administered for two weeks after the clinical onset, significantly decreased the cumulative and mean clinical scores of EAE. This was associated with the reduction of both CD4⁺ and CD8⁺ T cell numbers in the spinal cord and the downregulation of LFA-1 on T cells without affecting the cytokine production profile. The predominant cytokine produced by T cells in the spleen was IFN-γ whereas in the CNS it was IL-17. Our results indicate that minocycline regulates T cell infiltration into the CNS without modifying the dominant cytokine production.

Introduction

Multiple sclerosis (MS) is the most common demyelinating disease of young adults in which the early changes of inflammation and myelin loss result in chronic demyelination with axon loss. These chronic changes lead to patients entering the secondary progressive stage of the disease from which there is no improvement. Although there has been a considerable advance in treatment of MS in the last two decades, the available therapies have limited effectiveness and there is still a great need for development of new therapies.

In recent years minocycline has emerged as a potential drug to treat MS (Brundula et al., 2002, Popovic et al., 2002). Minocycline is a semi-synthetic antibiotic from the tetracycline family which, in addition to its anti-microbial properties, exerts pleiotropic effects on the immune system. It is widely used as an anti-inflammatory drug to treat skin disorders or arthritis. Within the tetracycline family, minocycline has the highest permeability across the blood brain barrier which makes it suitable for the treatment of CNS disorders. Minocycline has been used experimentally in several animal models of demyelinating and neurodegenerative disorders including MS (Nikodemova et al., 2007), Parkinson's disease (Du et al., 2001), ALS (Zhu et al., 2002) and ischemia (Yrjanheikki et al., 1998). Some studies also suggest that in addition to its anti-inflammatory effects minocycline also exerts neuroprotective properties (Wilkins et al., 2004) which make this drug even more suitable for treatment of MS. In clinical trials, minocycline decreased the number of relapses and reduced gadolinium-enhancing lesions in patients with the relapsing–remitting form of MS (Metz et al., 2004, Zhang et al., 2008). Despite these successes, the underlying mechanisms of action of this drug in EAE or MS are not fully understood.

We and others have previously shown that minocycline significantly attenuates the clinical severity of experimental allergic encephalomyelitis (EAE), an animal model of MS (Brundula et al., 2002, Nikodemova et al., 2007, Popovic et al., 2002). We have demonstrated that minocycline decreased the size and number of inflammatory lesions and myelin loss and inhibited microglial activation in the spinal cord (Nikodemova et al., 2007, Popovic et al., 2002). However, the mechanisms by which minocycline suppresses inflammatory responses in the CNS have not been fully delineated. Previously, we showed that minocycline inhibits microglial MHC II expression which is necessary for reactivation of T cells that had infiltrated into the CNS (Nikodemova et al., 2007). In addition, it appears that the suppressive effects of minocycline on EAE were associated with immune deviation in the periphery (Popovic et al., 2002). Minocycline has also been shown to inhibit activities of matrix metalloproteinases, enzymes that are important in facilitating the passage of lymphocytes across the blood brain barrier (Brundula et al., 2002, Paemen et al., 1996). These data suggest that several mechanisms may contribute to the overall anti-inflammatory effects of minocycline.

In this study we investigated the effects of minocycline on the infiltration of T cells into the spinal cord in an EAE model. We found that minocycline treatment, when started after the disease onset, suppressed the progression of clinical symptoms of EAE that was primarily associated with reduced number of CD4⁺ and CD8⁺ T cells present in the spinal cord. However, minocycline treatment did not alter Th1, Th2 and Th17 cytokine responses in the spinal cord or spleen suggesting that regulating the number of infiltrating T cells without modifying their cytokine production profile is sufficient for eliciting anti-inflammatory effects of minocycline.

Section snippets

Animals

Female Dark Agouti (DA) rats (155–165 g) were purchased from Harlan Sprague Dawley (Madison, WI). Animals were housed at standard condition of 12 h light/dark cycle and fed ad lib. All experiments were conducted with approval of the Research Animal Resources Center of the University of Wisconsin, Madison.

Induction of EAE and minocycline treatment

EAE was induced by immunization with 10 μg MOG in complete Freund's adjuvant as we described in detail previously (Nikodemova et al., 2007). Minocycline treatment was started at the clinical onset

Minocycline reduces clinical severity of EAE

As we have previously described (Nikodemova et al., 2007), the onset of the disease occurs 10–14 day post-immunization followed by a biphasic course of the disease. In the present study, minocycline treatment was started at the first signs of clinical disease and continued for two weeks. The effects of minocycline on disease course are summarized in Table 1. The mean clinical score was 2.16 in untreated animals, while it was significantly decreased to 1.08 in minocycline-treated animals.

Discussion

Minocycline has been shown to alleviate several CNS disorder in animal models presumably due to its anti-inflammatory and neuroprotective properties, however, the exact mechanisms of minocycline actions are still unclear. We have reported previously that minocycline attenuated clinical symptoms of EAE and delayed the onset of the disease when administered at the time of immunization (Popovic et al., 2002). These effects were accompanied by the decreased number and size of inflammatory lesions

Acknowledgement

This work was supported by a grant from the National Multiple Sclerosis Society (TR3761).

References (14)

K.J. Dugger et al.
Effector and suppressor roles for LFA-1 during the development of experimental autoimmune encephalomyelitis
J. Neuroimmunol.
(2009)
M. Nikodemova et al.
Minocycline down-regulates MHC II expression in microglia and macrophages through inhibition of IRF-1 and protein kinase C (PKC)alpha/betaII
J. Biol. Chem.
(2007)
L. Paemen et al.
The gelatinase inhibitory activity of tetracyclines and chemically modified tetracycline analogues as measured by a novel microtiter assay for inhibitors
Biochem. Pharmacol.
(1996)
V. Brundula et al.
Targeting leukocyte MMPs and transmigration: minocycline as a potential therapy for multiple sclerosis
Brain
(2002)
Y. Du et al.
Minocycline prevents nigrostriatal dopaminergic neurodegeneration in the MPTP model of Parkinson's disease
Proc. Natl. Acad. Sci. U S A
(2001)
Y. Komiyama et al.
IL-17 plays an important role in the development of experimental autoimmune encephalomyelitis
J. Immunol.
(2006)
J. Lee et al.
Mycobacterium bovis bacille Calmette–Guerin infection in the CNS suppresses experimental autoimmune encephalomyelitis and Th17 responses in an IFN-gamma-independent manner
J. Immunol.
(2008)

There are more references available in the full text version of this article.

Cited by (34)

Distinct origins, gene expression and function of microglia and monocyte-derived macrophages in CNS myelin injury and regeneration
2018, Clinical Immunology
Citation Excerpt :
Accumulation of macrophages within MS lesions, derived from both microglia and monocytes, is positively correlated with active demyelination [42–43]. Indeed, macrophages contribute to white matter pathology, as reducing macrophage activation using minocycline attenuates clinical presentation in EAE [44–46]. The deleterious functions of CNS macrophages include antigen presentation to cytotoxic T cells, stripping away myelin from axons, synaptic degradation and secretion of molecules toxic to neural cells such as pro-inflammatory cytokines, glutamate and reactive oxygen/nitrogen species [39,47–49].
Central nervous system (CNS) injury incurs a rapid innate immune response, including that from macrophages derived from endogenous microglia and circulating monocytes infiltrating the lesion site. One example of such injury is the demyelination observed in the autoimmune disease multiple sclerosis (MS), where macrophages are implicated in both myelin injury and regeneration. Although initially microglia and monocyte-derived macrophages were considered to have identical origins, gene expression, and function, recent advances have revealed important distinctions in all three categories and have caused a paradigm shift in view of their unique identity and roles. This has important consequences for understanding their individual contribution to neurological function and therapeutic targeting of these populations in diseases like MS. Here, we address the differences between CNS endogenous and exogenously-derived macrophages with a particular focus on myelin damage and regeneration.
Antibiotics acting as neuroprotectants via mechanisms independent of their anti-infective activities
2013, Neuropharmacology
Citation Excerpt :
The aforementioned neuroprotective mechanisms of minocycline have been assessed in a number of animal models of neurologic diseases. Minocycline was consistently beneficial in animal models of Alzheimer's disease (Fan et al., 2007; Seabrook et al., 2006), amyotrophic lateral sclerosis (Zhu et al., 2002), and multiple sclerosis [i.e., autoimmune encephalomyelitis (Maier et al., 2007; Nikodemova et al., 2010; Zemke and Majid, 2004)]. Inconsistent results were observed in studies involving animals models of Huntington's disease (Chen et al., 2000; Diguet et al., 2004; Ryu et al., 2006; Smith et al., 2003; Stack et al., 2006), ischemic injury (Carty et al., 2008; Matsukawa et al., 2009; Sakata et al., 2012; Tsuji et al., 2004), Parkinson's disease (Diguet et al., 2004; Du et al., 2001; Peng et al., 2006; Radad et al., 2010; Yang et al., 2003), and spinal cord injuries (Kim and Suh, 2009).
This review considers available evidence that some antibiotics have ancillary neuroprotective effects. Notably, β-lactam antibiotics are believed to increase the expression of glutamate transporter GLT1, potentially relieving the neurological excitotoxicity that characterizes disorders like amyotrophic lateral sclerosis. Minocycline has shown promise in reducing the severity of a number of neurological diseases, including multiple sclerosis, most likely by reducing apoptosis and the expression of inflammatory mediators in the brain. Rapamycin inhibits the activity of a serine/threonine protein kinase that has a role in the pathogenesis of numerous neurologic diseases. Herein we examine the unique neuroprotective aspects of these drugs originally developed as anti-infective agents.
Is minocycline useful for therapy of acute viral encephalitis?
2012, Antiviral Research
Minocyline is a tetracycline derivative with anti-inflammatory, anti-apoptotic, and anti-oxidant properties. Therapy has proved useful in some experimental models of both noninfectious and infectious neurological diseases and also in clinical trials in humans, including acute traumatic cervical spinal cord injury. In models of viral encephalitis, treatment has shown both beneficial and deleterious effects. In reovirus infection in mice, minocycline delayed the disease, but did not improve either the morbidity or mortality of the disease. In neuroadapted Sindbis virus infection of mice, minocycline prevented disease, but therapy needed to be given before clinical signs were present in most of the animals. In experimental rabies in neonatal mice minocycline aggravated the disease, likely related to anti-inflammatory effects. Minocycline has also been shown to aggravate disease in a mouse model of Huntington disease, in a monkey model of Parkinson disease, and in a mouse model of hypoxic-ischemic brain injury. Hence, there is experimental evidence of benefit of minocycline in both infectious and noninfectious neurological diseases, but there is a lack of benefit and harmful effects in other diseases. This may reflect multiple mechanisms of actions that cannot be predicted in a new disease or in an infection caused by a specific viral agent. Minocycline therapy is a double-edged sword and this drug should not be given empirically to patients with acute viral encephalitis for anticipated neuroprotective effects. Much more work needs to be done in experimental models in animals as well as in clinical trials. Because patient enrollment in clinical trials on acute viral encephalitis has proven to be difficult, funding will be a challenge.
The prospects of minocycline in multiple sclerosis
2011, Journal of Neuroimmunology
Citation Excerpt :
Administration of minocycline for two weeks could inhibit microglia expression of MHC II and the reactivation of T cells and thus attenuate the clinical severity of EAE (Nikodemova et al., 2007). It was recently found that regulating the activation and the number of infiltrating T cells without modifying their cytokine production profile was sufficient for minocycline to elicit anti-inflammatory effects and suppressed the progression of clinical symptoms of EAE (Nikodemova et al., 2010). Microglia, the resident antigen-presenting cell of the CNS, plays a critical role in MHC II-dependent antigen presentation and thus the reactivation of CNS-infiltrated encephalitogenic T cells (Nikodemova et al., 2007).
Multiple sclerosis (MS) is the most common inflammatory demyelinating disease of the central nervous system (CNS). Although there are several approved drugs for MS, not all patients respond optimally to these drugs. More effective, well-tolerated therapeutic strategies for MS are necessary, either through the development of new medication or combination of existing ones. Minocycline is a traditional antibiotic with profound anti-inflammatory and neuropropective effects and good tolerance for long-term use. The encouraging results from the animal model and clinical experiments on minocycline make it a promising candidate for MS treatment whether used alone or combined with other drugs. In this review, we summarized the pharmacological actions of minocycline and focused on its therapeutic effects and safety in experimental autoimmune encephalomyelitis (EAE) and MS. The data obtained here showed that minocycline would be an effective and safe therapy for MS.
Minocycline reduces ethanol drinking
2011, Brain, Behavior, and Immunity
Citation Excerpt :
Minocycline is a second generation tetracycline antibiotic that has immune modulatory actions independent of it bactericidal actions (e.g., Kielian et al., 2007; Kloppenburg et al., 1994; Sewell et al., 1996). As an immune modulator, minocycline can down-regulate expression of pro-inflammatory genes and mediators in both the central and peripheral immune systems (Henry et al., 2008; Homsi et al., 2009; Huang et al., 2009; Kloppenburg et al., 1996; Nikodemova et al., 2010). To test the effect of minocycline on alcohol intake, drinking in both male and female C57Bl/6J mice was measured using a free choice voluntary drinking model.
Alcoholism is a disease characterized by continued alcohol consumption despite recurring negative consequences. Thus, medications that reduce the drive to consume alcohol can be beneficial in treating alcoholism. The neurobiological systems that regulate alcohol consumption are complex and not fully understood. Currently, medications are available to treat alcoholism that act either by causing accumulation of a toxic metabolite of ethanol, or by targeting specific transmitter receptors. The purpose of our study was to investigate a new potential therapeutic pathway, neuroimmune interactions, for effects on ethanol consumption. We hypothesized that neuroimmune activity of brain glia may have a role in drinking. We utilized minocycline, a second generation tetracycline antibiotic that has immune modulatory actions, to test our hypothesis because it is known to suppress microglia, and to a lesser extent astroglia, activity following many types of insults to the brain. Treatment with 50 mg/kg minocycline significantly reduced ethanol intake in male and female C57Bl/6J mice using a free choice voluntary drinking model. Saline injections did not alter ethanol intake. Minocycline had little effect on water intake or body weight change. The underlying mechanism whereby minocycline reduced ethanol intake requires further study. The results suggest that drugs that alter neuroimmune pathways may represent a new approach to developing additional therapies to treat alcoholism.
Minocycline suppresses activation of nuclear factor of activated T cells 1 (NFAT1) in human CD4<sup>+</sup> T cells
2011, Journal of Biological Chemistry
Citation Excerpt :
Minocycline has also shown therapeutic effects on T cells in experimental autoimmune encephalomyelitis, an animal model of multiple sclerosis. Minocycline treatment of rodents with experimental autoimmune encephalomyelitis attenuated the severity of encephalitis, decreased T cell transmigration in vitro, and reduced CD4+ and CD8+ T cell infiltration into the spinal cord in vivo (1, 11). In the context of HIV infection, T cell activation is closely correlated with disease progression and remains elevated even after long term administration of highly active antiretroviral therapy (12–14).
Minocycline is a tetracycline family antibiotic that has anti-inflammatory and immunomodulatory properties. These properties have shown promise in the treatment of conditions such as rheumatoid arthritis, Huntington disease, and multiple sclerosis. As lymphocyte activation is involved in the pathogenesis of many of these diseases, T cells are postulated to be a primary target in minocycline therapy. Previous studies have demonstrated attenuation of CD4⁺ T cell activation by minocycline, but a specific mechanism has not been elucidated. In this study, we investigated the effect of minocycline on the activity of three key transcription factors regulating CD4⁺ T cell activation: NF-κB, AP-1 (activator protein 1), and NFAT (nuclear factor of activated T) cells. Our data demonstrate that minocycline selectively impairs NFAT-mediated transcriptional activation, a result of increased phosphorylation and reduced nuclear translocation of the isoform NFAT1. Minocycline increased the activity of the NFAT kinase GSK3 and decreased intracellular Ca²⁺ flux, both of which facilitate NFAT1 phosphorylation. These findings provide a novel mechanism for minocycline induced suppression of CD4⁺ T cell activation and may better inform the application of minocycline as an immunomodulatory agent.

View all citing articles on Scopus

¹: These authors contributed equally to this work.

View full text

Article preview

Journal of Neuroimmunology

Abstract

Introduction

Section snippets

Animals

Induction of EAE and minocycline treatment

Minocycline reduces clinical severity of EAE

Discussion

Acknowledgement

References (14)

Effector and suppressor roles for LFA-1 during the development of experimental autoimmune encephalomyelitis

J. Neuroimmunol.

Minocycline down-regulates MHC II expression in microglia and macrophages through inhibition of IRF-1 and protein kinase C (PKC)alpha/betaII

J. Biol. Chem.

The gelatinase inhibitory activity of tetracyclines and chemically modified tetracycline analogues as measured by a novel microtiter assay for inhibitors

Biochem. Pharmacol.

Targeting leukocyte MMPs and transmigration: minocycline as a potential therapy for multiple sclerosis

Brain

Minocycline prevents nigrostriatal dopaminergic neurodegeneration in the MPTP model of Parkinson's disease

Proc. Natl. Acad. Sci. U S A

IL-17 plays an important role in the development of experimental autoimmune encephalomyelitis

J. Immunol.

Mycobacterium bovis bacille Calmette–Guerin infection in the CNS suppresses experimental autoimmune encephalomyelitis and Th17 responses in an IFN-gamma-independent manner

J. Immunol.

Cited by (34)

Distinct origins, gene expression and function of microglia and monocyte-derived macrophages in CNS myelin injury and regeneration

Antibiotics acting as neuroprotectants via mechanisms independent of their anti-infective activities

Is minocycline useful for therapy of acute viral encephalitis?

The prospects of minocycline in multiple sclerosis

Minocycline reduces ethanol drinking

Minocycline suppresses activation of nuclear factor of activated T cells 1 (NFAT1) in human CD4<sup>+</sup> T cells