IgG Infiltration Research Articles

T follicular helper cells contribute to pathophysiology in a model of neuromyelitis optica spectrum disorders.

Neuromyelitis optica spectrum disorders (NMOSD) are inflammatory autoimmune disorders of the CNS. IgG autoantibodies targeting the aquaporin-4 water channel (AQP4-IgGs) are the pathogenic effector of NMOSD. Dysregulated T follicular helper (Tfh) cells have been implicated in loss of B cell tolerance in autoimmune diseases. The contribution of Tfh cells to disease activity and therapeutic potential of targeting these cells in NMOSD remain unclear. Here, we established an autoimmune model of NMOSD by immunizing mice against AQP4 via in vivo electroporation. After AQP4 immunization, mice displayed AQP4 autoantibodies in blood circulation, blood-brain barrier disruption, and IgG infiltration in spinal cord parenchyma. Moreover, AQP4 immunization induced motor impairments and NMOSD-like pathologies, including astrocytopathy, demyelination, axonal loss, and microglia activation. These were associated with increased splenic Tfh, Th1, and Th17 cells; memory B cells; and plasma cells. Aqp4-deficient mice did not display motor impairments and NMOSD-like pathologies after AQP4 immunization. Importantly, abrogating ICOS/ICOS-L signaling using anti-ICOS-L antibody depleted Tfh cells and suppressed the response of Th1 and Th17 cells, memory B cells, and plasma cells in AQP4-immunized mice. These findings were associated with ameliorated motor impairments and spinal cord pathologies. This study suggests a role of Tfh cells in the pathophysiology of NMOSD in a mouse model with AQP4 autoimmunity and provides an animal model for investigating the immunological mechanisms underlying AQP4 autoimmunity and developing therapeutic interventions targeting autoimmune reactions in NMOSD.

OBESITY MAY UNDERLIE SKELETAL MUSCLE AUTOIMMUNITY IN OLDER ADULTS

Abstract A mechanistic understanding of the biological basis for sarcopenia could lead to novel biomarkers and intervention targets. We recently reported a novel role of autoimmunity in sarcopenia, which is potentially mediated through cardiac troponin T (cTnT) and its autoantibody. Given the known role of obesity in both autoimmunity and sarcopenia, we further explored the association between circulating cTnT or skeletal muscle IgG and obesity in older adults. We found that, in 37 older adults (NCT01049698, BMI: 30.8 ± 2.3 kg/m²; Age: 69 ± 3.5 yrs) without any cardiac abnormality, serum cTnT was negatively correlated with skeletal muscle volume (r = -0.37, p = 0.02) and leg extension strength (r = -0.38, p = 0.02), but positively correlated with body fat/muscle volume ratio (r = 0.37, p = 0.03). In two other studies including older adults (n = 11 & 12) from a subset of subjects (NCT01049698 & NCT01298817), skeletal muscle IgG1 (GAPDH normalized level by immunoblot) was found to be positively associated with BMI or dual-energy X-ray absorptiometry derived regional fat measurements, including fat mass, abdominal subcutaneous fat, abdominal fat, or thigh fat. We also found that subjects with higher skeletal muscle IgG1 protein levels have significantly higher BMI but lower knee extensor strength. Our finding suggests that obesity may underlie increased circulating cTnT and skeletal muscle IgG infiltration. It also warrants additional investigation into its role in skeletal muscle autoimmunity in older adults.

Characteristics of B cells and immunoglobulin profile in Takayasu's arteritis.

Disorders of humoral immunity in Takayasu's arteritis (TAK) have not been well explored. This study describes the characteristics of B cells and immunoglobulin (Ig) profile in patients with TAK. Peripheral B cell populations assessed using flow cytometry and serum Ig levels assessed using a biochemical analyser in 98 newly diagnosed patients with TAK were analysed and compared with those of 31 patients with systemic lupus erythematosus (SLE) and 60 healthy controls (HCs). CD19+ B cell and IgG infiltration to the aortic tissue was evaluated by immunohistochemical staining. The proportion of peripheral CD3-CD19+ B cells and levels of serum IgG in TAK were lower than those in SLE, but higher than those in HCs. CD3-CD19+ B cell counts were higher in TAK than in HCs. Serum IgG and IgG1 levels were higher in active TAK than in non-active TAK. In TAK, positive correlations of serum IgG levels with erythrocyte sedimentation rate (ESR), C-reactive protein (CRP) level, Kerr score, and Indian Takayasu Clinical Activity Score (ITAS2010, ITAS-A) were observed before immunotherapy. After 6 months of immunotherapy, serum Ig levels significantly decreased. Positive correlations between the changes in IgG levels and values of ESR, CRP, Kerr score, and ITAS-A were detected. Immunohistochemical staining confirmed CD19+ B cell and IgG infiltration to the aortic wall in patients with TAK. Enhanced B cells might contribute to the pathogenesis of TAK, and serum IgG levels could serve as a simple, useful biomarker to assess disease activity and monitor treatment response in TAK.

Higher IgG level correlated with vitamin D receptor in the hippocampus of a pristane-induced lupus model.

Patients with systemic lupus erythematosus (SLE) may have neurological complications, characterizing neuropsychiatric lupus (NPSLE). Studies have investigated alternative therapies such as vitamin D, which has an effect on the immune system and brain, to control manifestations of SLE. Experimental lupus models may be a good alternative to best study the immunological mechanisms underlying the development of NPSLE, and the animal model of pristane-induced lupus (PIL) may mimic SLE symptoms in humans. Our objective was to evaluate central nervous system involvement and vitamin D supplementation in a PIL model. Female BALB/c mice were divided into controls (CO; n = 7), PIL (n = 9), and PIL supplemented with vitamin D (VD; n = 7). The hippocampus area was measured and immunoassays were performed for detecting vitamin D receptor (VDR) and IgG. The PIL group had a higher hippocampal IgG infiltrate when compared to the CO group. Vitamin D showed potential for reducing IgG infiltration. The hippocampus area was similar in all groups. No differences in VDR expression were observed between groups. A positive correlation was observed between the expression of VDR and IgG in the hippocampus. Our data suggest that increased IgG infiltration into the hippocampus indicated an inflammatory process that may have stimulated VDR expression. Key Points • IgG infiltrate is higher in PIL animals than controls • VDR increases along with IgG infiltrate • Hippocampal VDR expression does not increase with vitamin D supplementation.

Lack of Neuroprotective Effects of High-Density Lipoprotein Therapy in Stroke under Acute Hyperglycemic Conditions.

Introduction: The pleiotropic protective effects of high-density lipoproteins (HDLs) on cerebral ischemia have never been tested under acute hyperglycemic conditions. The aim of this study is to evaluate the potential neuroprotective effect of HDL intracarotid injection in a mouse model of middle cerebral artery occlusion (MCAO) under hyperglycemic conditions. Methods: Forty-two mice were randomized to receive either an intracarotid injection of HDLs or saline. Acute hyperglycemia was induced by an intraperitoneal injection of glucose (2.2 g/kg) 20 min before MCAO. Infarct size (2,3,5-triphenyltetrazolium chloride (TTC)-staining), blood–brain barrier leakage (IgG infiltration), and hemorrhagic changes (hemoglobin assay by ELISA and hemorrhagic transformation score) were analyzed 24 h post-stroke. Brain tissue inflammation (IL-6 by ELISA, neutrophil infiltration and myeloperoxidase by immunohisto-fluorescence) and apoptosis (caspase 3 activation) were also assessed. Results: Intraperitoneal D-glucose injection allowed HDL- and saline-treated groups to reach a blood glucose level of 300 mg/dl in the acute phase of cerebral ischemia. HDL injection did not significantly reduce mortality (19% versus 29% in the saline-injected group) or cerebral infarct size (p = 0.25). Hemorrhagic transformations and inflammation parameters were not different between the two groups. In addition, HDL did not inhibit apoptosis under acute hyperglycemic conditions. Conclusion: We observed a nonsignificant decrease in cerebral infarct size in the HDL group. The deleterious consequences of reperfusion such as hemorrhagic transformation or inflammation were not improved by HDL infusion. In acute hyperglycemia, HDLs are not potent enough to counteract the adverse effects of hyperglycemia. The addition of antioxidants to therapeutic HDLs could improve their neuroprotective capacity.

Loss of tissue-nonspecific alkaline phosphatase (TNAP) enzyme activity in cerebral microvessels is coupled to persistent neuroinflammation and behavioral deficits in late sepsis

Sepsis is a host response to systemic inflammation and infection that may lead to multi-organ dysfunction and eventual death. While acute brain dysfunction is common among all sepsis patients, chronic neurological impairment is prevalent among sepsis survivors. The brain microvasculature has emerged as a major determinant of sepsis-associated brain dysfunction, yet the mechanisms that underlie its associated neuroimmune perturbations and behavioral deficits are not well understood. An emerging body of data suggests that inhibition of tissue-nonspecific alkaline phosphatase (TNAP) enzyme activity in cerebral microvessels may be associated with changes in endothelial cell barrier integrity. The objective of this study was to elucidate the connection between alterations in cerebrovascular TNAP enzyme activity and brain microvascular dysfunction in late sepsis. We hypothesized that the disruption of TNAP enzymatic activity in cerebral microvessels would be coupled to the sustained loss of brain microvascular integrity, elevated neuroinflammatory responses, and behavioral deficits. Male mice were subjected to cecal ligation and puncture (CLP), a model of experimental sepsis, and assessed up to seven days post-sepsis. All mice were observed daily for sickness behavior and underwent behavioral testing. Our results showed a significant decrease in brain microvascular TNAP enzyme activity in the somatosensory cortex and spinal cord of septic mice but not in the CA1 and CA3 hippocampal regions. Furthermore, we showed that loss of cerebrovascular TNAP enzyme activity was coupled to a loss of claudin-5 and increased perivascular IgG infiltration in the somatosensory cortex. Analyses of whole brain myeloid and T-lymphoid cell populations also revealed a persistent elevation of infiltrating leukocytes, which included both neutrophil and monocyte myeloid derived suppressor cells (MDSCs). Regional analyses of the somatosensory cortex, hippocampus, and spinal cord revealed significant astrogliosis and microgliosis in the cortex and spinal cord of septic mice that was accompanied by significant microgliosis in the CA1 and CA3 hippocampal regions. Assessment of behavioral deficits revealed no changes in learning and memory or evoked locomotion. However, the hot plate test uncovered a novel anti-nociceptive phenotype in our septic mice, and we speculate that this phenotype may be a consequence of sustained GFAP astrogliosis and loss of TNAP activity in the somatosensory cortex and spinal cord of septic mice. Taken together, these results demonstrate that the loss of TNAP enzyme activity in cerebral microvessels during late sepsis is coupled to sustained neuroimmune dysfunction which may underlie, in part, the chronic neurological impairments observed in sepsis survivors.

The time course of blood brain barrier leakage and its implications on the progression of methamphetamine-induced seizures

The initial goals of these experiments were to determine: 1) if blood-brain barrier (BBB) breakdown was a cause or an effect of METH-induced seizures; 2) all the brain regions where BBB is disrupted as seizures progress; and 3) the correlations between body temperature and vascular leakage and neurodegeneration. A fourth objective was added after initial experimentation to determine if sub-strain differences existed in adult male C57 B6 J (Jackson laboratories, JAX) versus C57 B6N (Charles River, CR) mice involving their susceptibility to BBB breakdown and seizure severity. With the 1st “maximal” intensity myoclonic-tonic seizure (MCT) varying degrees of IgG infiltration across the BBB (≤1 mm2) were prominent in olfactory system (OS) associated regions and in thalamus, hypothalamus and neocortex. IgG infiltration areas in the OS-associated regions of the bed nucleus of the stria terminalis, septum and more medial amygdala nuclei, and the hypothalamus were increased significantly by the time continuous behavioral seizures (CBS) developed. Mice receiving METH that had body temperatures of ≥40 °C had IgG infiltration along with MCT or CBS but peak body temperatures above 40 °C did not significantly increase IgG infiltration. Neurodegeneration seen at ≥6 h was restricted to the OS in both JAX and CR mice and was most prominent in the posteromedial cortical amygdaloid nucleus. Neurodegeneration in the anterior septum (tenia tecta) was seen only in the JAX mice. We hypothesize that METH-induced hypertension and hyperthermia lead to BBB breakdown and other vascular dysfunctions in the OS brain regions resulting in OS hyperexcitation. Excitation of the OS neural network then leads to the development of seizures. These seizures in turn exacerbate the energy depletions and the reactive oxygen stress produced by hyperthermia further damaging the BBB and vascular function. These events form a recurrent cycle that results in ever increasing seizure activity and neurotoxicity.

Abstracts from The 34(th) Annual National Neurotrauma Symposium June 26-29, 2016 Lexington, Kentucky.

Abstracts from The 34(th) Annual National Neurotrauma Symposium June 26-29, 2016 Lexington, Kentucky.

Systemic inflammation affects reperfusion following transient cerebral ischaemia

Reperfusion after stroke is critical for improved patient survival and recovery and can be achieved clinically through pharmacological (recombinant tissue plasminogen activator) or physical (endovascular intervention) means. Yet these approaches remain confined to a small percentage of stroke patients, often with incomplete reperfusion, and therefore there is an urgent need to learn more about the mechanisms underlying the no-reflow phenomenon that prevents restoration of adequate microvascular perfusion. Recent evidence suggests systemic inflammation as an important contributor to no-reflow and to further investigate this here we inject interleukin 1 (IL-1) i.p. 30min prior to an ischaemic challenge using a remote filament to occlude the middle cerebral artery (MCA) in mice. Before, during and after the injection of IL-1 and occlusion we use two-dimensional optical imaging spectroscopy to record the spatial and temporal dynamics of oxyhaemoglobin concentration in the cortical areas supplied by the MCA. Our results reveal that systemic inflammation significantly reduces oxyhaemoglobin reperfusion as early as 3h after filament removal compared to vehicle injected animals. CD41 immunohistochemistry shows a significant increase of hyper-coagulated platelets within the microvessels in the stroked cortex of the IL-1 group compared to vehicle. We also observed an increase of pathophysiological biomarkers of ischaemic damage including elevated microglial activation co-localized with interleukin 1α (IL-1α), increased blood brain barrier breakdown as shown by IgG infiltration and increased pyknotic morphological changes of cresyl violet stained neurons. These data confirm systemic inflammation as an underlying cause of no-reflow in the post-ischaemic brain and that appropriate anti-inflammatory approaches could be beneficial in treating ischaemic stroke.

IgG and Complement Deposition and Neuronal Loss in Cats and Humans With Epilepsy and Voltage-Gated Potassium Channel Complex Antibodies

Voltage-gated potassium channel complex (VGKC-complex) antibody (Ab) encephalitis is a well-recognized form of limbic encephalitis in humans, usually occurring in the absence of an underlying tumor. The patients have a subacute onset of seizures, magnetic resonance imaging findings suggestive of hippocampal inflammation, and high serum titers of Abs against proteins of the VGKC-complex, particularly leucine-rich, glioma-inactivated 1 (LGI1). Most patients are diagnosed promptly and recover substantially with immunotherapies; consequently, neuropathological data are limited. We have recently shown that feline complex partial cluster seizures with orofacial involvement (FEPSO) in cats can also be associated with Abs against VGKC-complexes/LGI1. Here we examined the brains of cats with FEPSO and compared the neuropathological findings with those in a human with VGKC-complex-Ab limbic encephalitis. Similar to humans, cats with VGKC-complex-Ab and FEPSO have hippocampal lesions with only moderate T-cell infiltrates but with marked IgG infiltration and complement C9neo deposition on hippocampal neurons, associated with neuronal loss. These findings provide further evidence that FEPSO is a feline form of VGKC-complex-Ab limbic encephalitis and provide a model for increasing understanding of the human disease.

Fish oil improves motor function, limits blood–brain barrier disruption, and reduces Mmp9 gene expression in a rat model of juvenile traumatic brain injury

The effects of an oral fish oil treatment regimen on sensorimotor, blood–brain barrier, and biochemical outcomes of traumatic brain injury (TBI) were investigated in a juvenile rat model. Seventeen-day old Long-Evans rats were given a 15mL/kg fish oil (2.01g/kg EPA, 1.34g/kg DHA) or soybean oil dose via oral gavage 30min prior to being subjected to a controlled cortical impact injury or sham surgery, followed by daily doses for seven days. Fish oil treatment resulted in less severe hindlimb deficits after TBI as assessed with the beam walk test, decreased cerebral IgG infiltration, and decreased TBI-induced expression of the Mmp9 gene one day after injury. These results indicate that fish oil improved functional outcome after TBI resulting, at least in part from decreased disruption of the blood–brain barrier through a mechanism that includes attenuation of TBI-induced expression of Mmp9.

Cerebrospinal fluid/serum gradient of IgG is associated with disability at acute attacks of neuromyelitis optica

Increased blood-brain barrier (BBB) disruption can be found in patients with neuromyelitis optica (NMO); however, its clinical implication and association with disability at acute attack remains obscure. The purpose of the study was to evaluate the clinical significance of BBB disruption and the subsequent cerebrospinal fluid (CSF)/serum IgG gradient in NMO. Retrospective analysis was made of acute-stage CSF samples from NMO (n = 40) and multiple sclerosis (MS; n = 26) patients. The CSF/serum IgG gradient (QIgG), albumin ratio (Qalb), and IgG index were calculated. Multivariate regression analysis was used to identify clinical and CSF variables associated with disability at acute attacks (extended disability scale score, EDSS) in both groups. The EDSS was significantly associated with the QIgG (p < 0.001), Qalb (p = 0.012), and number of cumulative attacks (p = 0.012) in NMO but not in MS with univariate analysis. Length of spinal cord involvement was also associated with EDSS in NMO (p = 0.030). However, multivariate analysis revealed that the QIgG was only significantly associated with EDSS in NMO (0.580; 95% CI -0.257, 0.961; p = 0.002). The QIgG was also highly associated with the Qalb in NMO (p < 0.001). The QIgG may reflect systemic IgG leakage into the CNS and is strongly associated with disability at acute attacks in NMO, suggesting that BBB disruption can aggravate disease activity by facilitating systemic IgG infiltration into the CNS.

C5a alters blood‐brain barrier integrity in experimental lupus

The blood-brain barrier (BBB) is a crucial anatomic location in the brain. Its dysfunction complicates many neurodegenerative diseases, from acute conditions, such as sepsis, to chronic diseases, such as systemic lupus erythematosus (SLE). Several studies suggest an altered BBB in lupus, but the underlying mechanism remains unknown. In the current study, we observed a definite loss of BBB integrity in MRL/MpJ-Tnfrsf6(lpr) (MRL/lpr) lupus mice by IgG infiltration into brain parenchyma. In line with this result, we examined the role of complement activation, a key event in this setting, in maintenance of BBB integrity. Complement activation generates C5a, a molecule with multiple functions. Because the expression of the C5a receptor (C5aR) is significantly increased in brain endothelial cells treated with lupus serum, the study focused on the role of C5a signaling through its G-protein-coupled receptor C5aR in brain endothelial cells, in a lupus setting. Reactive oxygen species production increased significantly in endothelial cells, in both primary cells and the bEnd3 cell line treated with lupus serum from MRL/lpr mice, compared with those treated with control serum from MRL(+/+) mice. In addition, increased permeability monitored by changes in transendothelial electrical resistance, cytoskeletal remodeling caused by actin fiber rearrangement, and increased iNOS mRNA expression were observed in bEnd3 cells. These disruptive effects were alleviated by pretreating cells with a C5a receptor antagonist (C5aRant) or a C5a antibody. Furthermore, the structural integrity of the vasculature in MRL/lpr brain was maintained by C5aR inhibition. These results demonstrate the regulation of BBB integrity by the complement system in a neuroinflammatory setting. For the first time, a novel role of C5a in the maintenance of BBB integrity is identified and the potential of C5a/C5aR blockade highlighted as a promising therapeutic strategy in SLE and other neurodegenerative diseases.

Endocardial fibroelastosis associated with maternal anti-Ro and anti-La antibodies in the absence of atrioventricular block

ObjectivesThis study was designed to document the association of endocardial fibroelastosis (EFE) and maternal autoantibodies. BackgroundNeonatal lupus erythematosus is associated with the transplacental passage of maternal anti-Ro and anti-La antibodies, leading to complete atrioventricular block (CAVB). In some cases, CAVB is associated with EFE. Isolated EFE may be independently related to maternal anti-Ro and anti-La antibodies. MethodsWe identified three cases (one fetus and two infants, all female) of isolated EFE in infants born to autoantibody-positive mothers in the absence of CAVB. Demographics, echocardiograms, and pathology were reviewed. Immunohistochemical analyses for immunoglobulin (Ig)G, IgM, IgA, T-cell, B-cell, and terminal deoxynucleoleotidyl transferase-mediated dUTP-biotin nick end-labeling (TUNEL) (test for cell apoptosis) staining were performed on multiple sections of the heart in each case and compared with negative controls. ResultsTwo cases died and one received a cardiac transplant. All three cases had histologically confirmed EFE. All cases demonstrated significant diffuse IgG infiltration. To a lesser extent, myocardial tissue was also positive for IgM, CD43, and Granzyme B. None of the specimens were TUNEL positive. ConclusionsThese are the first reported cases of isolated EFE associated with maternal anti-Ro and anti-La antibodies in the absence of CAVB. The diffuse deposition of IgG and the presence of a T-cell infiltrate throughout the myocardium suggest that the transplacental passage of maternal autoantibodies induces an immune reaction within the myocardium, leading to isolated EFE. Autoantibody-mediated EFE may be an etiologic factor in cases of fetal and neonatal “idiopathic” dilated cardiomyopathy.

Appearance of immunoglobulin G and complement factor C3 in the striatum after transient focal ischemia in the rat

The pathophysiological feature of brain ischemia-infarct was investigated using immunohistochemistry and Gallyas' silver staining after transient ischemia of the middle cerebral artery (MCA) in the rat. A very strong IgG infiltration with clear-cut borders was detected at one to 3 days in ischemic core areas (lateral striatum and adjacent cortex) that fell into porencephaly later. Complement factor C3 (C3) immunoreactivity (IR) appeared similarly while albumin IR more diffusely. Microglial activation could be observed at 1 day while rounded leukocyte-like elements at 3 days after reperfusion. Data suggest that the early appearance of the IgG/C3 IR bears particular importance after transient MCA ischemia as it could predict the ensuing porencephaly in the chronic stage.