Targeting of the class II transactivator attenuates inflammation and neurodegeneration in an alpha-synuclein model of Parkinson’s disease

Abstract

BackgroundParkinson’s disease (PD) is characterized by intracellular alpha-synuclein (α-syn) inclusions, progressive death of dopaminergic neurons in the substantia nigra pars compacta (SNpc), and activation of the innate and adaptive immune systems. Disruption of immune signaling between the central nervous system (CNS) and periphery, such as through targeting the chemokine receptor type 2 (CCR2) or the major histocompatibility complex II (MHCII), is neuroprotective in rodent models of PD, suggesting a key role for innate and adaptive immunity in disease progression. The purpose of this study was to investigate whether genetic knockout or RNA silencing of the class II transactivator (CIITA), a transcriptional co-activator required for MHCII induction, is effective in reducing the neuroinflammation and neurodegeneration observed in an α-syn mouse model of PD.MethodsIn vitro, we utilized microglia cultures from WT or CIITA −/− mice treated with α-syn fibrils to investigate inflammatory iNOS expression and antigen processing via immunocytochemistry (ICC). In vivo, an adeno-associated virus (AAV) was used to overexpress α-syn in WT and CIITA −/− mice as a model for PD. Concurrently with AAV-mediated overexpression of α-syn, WT mice received CIITA-targeted shRNAs packaged in lentiviral constructs. Immunohistochemistry and flow cytometry were used to assess inflammation and peripheral cell infiltration at 4 weeks post transduction, and unbiased stereology was used 6 months post transduction to assess neurodegeneration.ResultsUsing ICC and DQ-ovalbumin, we show that CIITA −/− microglial cultures failed to upregulate iNOS and MHCII expression, and had decreased antigen processing in response to α-syn fibrils when compared to WT microglia. In vivo, global knock-out of CIITA as well as local knockdown using lentiviral shRNAs targeting CIITA attenuated MHCII expression, peripheral immune cell infiltration, and α-syn-induced neurodegeneration.ConclusionOur data provide evidence that CIITA is required for α-syn-induced MHCII induction and subsequent infiltration of peripheral immune cells in an α-syn mouse model of PD. Additionally, we demonstrate that CIITA in the CNS drives neuroinflammation and neurodegeneration. These data provide further support that the disruption or modulation of antigen processing and presentation via CIITA is a promising target for therapeutic development in preclinical animal models of PD.Electronic supplementary materialThe online version of this article (10.1186/s12974-018-1286-2) contains supplementary material, which is available to authorized users.

T cell infiltration in both human multiple system atrophy and a novel mouse model of the disease

“…All surgical procedures on male and female C57BL/6 (WT), Ccr2 reporter knock in, Tcrb −/− , or Cd4 −/− mice were performed similar to previously described work [25,26,64]. Briefly, mice were anesthetized with isoflurane and unilaterally (immunohistochemistry experiments) or bilaterally (flow cytometry experiments) injected with 2 μl of Olig001-GFP (1 × 10 13 vg/ml) or Olig001-SYN (1 × 10 13 vg/ml) into the dorsolateral striatum at a rate of 0.5 μl/min with a Hamilton syringe.…”

“…For fluorescent analyses, free-floating sections were washed with Tris-buffered saline (TBS) and labeled as previously published [25,26,64]. Briefly, sections were blocked in appropriate 5% normal serum and then labeled with anti-MHCII (clone M5/114.15.2, eBiosciences), antialpha-synuclein (phospho-Serine129 (pSer129), clone EP1536Y, Abcam), anti-tyrosine hydroxylase (TH, clone AB152, Sigma-Aldrich), anti-IBA1 (Wako), anti-TMEM119 (clone 28-3, Abcam), anti-CD3 (clone 17A2, eBioscience), anti-CD4 (clone RM4-5, BD Bioscience), anti-CD8 (clone 4SM15, eBioscience), or anti-MBP (clone SMI 99, BioLegend) antibodies diluted in 1% normal serum in TBS-Triton (TBST) overnight at 4 °C.…”

“…Images were acquired using a Nikon Eclipse Ti-E. For myelin, MHCII (I-A/I-E), and pSer129 staining quantification, slides were blinded, scanned, and quantified by the mean gray value method via imageJ (NIH) as previously described [64]. Three striatal sections per animal encompassing the dorsal striatum injection site and pSer129 expression were chosen for quantification.…”

“…Mononuclear cells were isolated 4 weeks post-transduction from striatum or lymph nodes (LNs) of mice bilaterally transduced with Olig001-SYN or Olig001-GFP control according to published methods [26,45,64]. Briefly, striatums were triturated and digested with 1 mg/mL Collagenase IV (Sigma) and 20 μg/mL DNAse I (Sigma) diluted in RPMI 1640 with 10% heat inactivated fetal bovine serum, 1% l-glutamine (Sigma), and 1% Penicillin-Streptomycin (Sigma).…”

A novel role of NLRP3-generated IL-1β in the acute-chronic transition of peripheral lipopolysaccharide-elicited neuroinflammation: implications for sepsis-associated neurodegeneration

“…In this series of studies, we assessed the consequence of lacking either NLRP3 or IL-1R1 on the induction of acute factors such as TNFα, and delayed factors including MHC-II, NOX2 (gp91), and Mac1. These delayed factors are necessary for sustained reactive microgliosis and long-lasting neuroinflammation [11,21,[58][59][60][61][62][63][64][65][66][67].…”