C-reactive protein concentration has limited value in the diagnosis of meningoencephalitis of unknown origin in dogs.

OBJECTIVES: To evaluate blood and cerebrospinal fluid (CSF) concentrations of C-reactive protein (CRP) in dogs with meningoencephalitis of unknown origin (MUO); to evaluate whether blood CRP concentration is associated with epidemiological, clinicopathologic, and MRI findings; and to investigate blood CRP predictive power in survival. ANIMALS: 30 client-owned dogs with MUO, 15 client-owned dogs with steroid-responsive meningitis arteritis (SRMA; positive control group), and 15 healthy dogs (negative control group). METHODS: Blood CRP concentration was measured in each group, while it was performed in CSF only in the MUO and SRMA groups. The analysis of epidemiological data included breed, age, sex, duration of clinical signs, and history of seizures. Blinded analysis of MRI was performed based on a classification grid, and traditional CSF analysis parameters were assessed. The predictive power of blood CRP concentration regarding survival at 6 months was investigated. RESULTS: Of the 30 dogs with MUO, 9 (30%) had an increased CRP concentration in blood, and 3 (10%) showed a measurable CRP in CSF. Median blood CRP concentration in dogs with MUO was 0.1 mg/L (range, 0.1 to 102 mg/L), which was not statistically different from the healthy dog group but significantly lower than the SRMA control group. Only the duration of clinical signs was positively associated with an increased blood CRP level. Blood CRP concentration was not associated with survival at 6 months. CLINICAL RELEVANCE: Blood CRP concentration is of limited value for the diagnosis and prognosis of dogs with MUO. Chronicity of the disease may be associated with an increased concentration of blood CRP.

Amylin, Aß42, and Amyloid in Varicella Zoster Virus Vasculopathy Cerebrospinal Fluid and Infected Vascular Cells.

BACKGROUND: Varicella zoster virus (VZV) vasculopathy is characterized by persistent arterial inflammation leading to stroke. Studies show that VZV induces amyloid formation that may aggravate vasculitis. Thus, we determined if VZV central nervous system infection produces amyloid. METHODS: Aß peptides, amylin, and amyloid were measured in cerebrospinal fluid (CSF) from 16 VZV vasculopathy subjects and 36 stroke controls. To determine if infection induced amyloid deposition, mock- and VZV-infected quiescent primary human perineurial cells (qHPNCs), present in vasculature, were analyzed for intracellular amyloidogenic transcripts/proteins and amyloid. Supernatants were assayed for amyloidogenic peptides and ability to induce amyloid formation. To determine amylin's function during infection, amylin was knocked down with small interfering RNA and viral complementary DNA (cDNA) was quantitated. RESULTS: Compared to controls, VZV vasculopathy CSF had increased amyloid that positively correlated with amylin and anti-VZV antibody levels; Aß40 was reduced and Aß42 unchanged. Intracellular amylin, Aß42, and amyloid were seen only in VZV-infected qHPNCs. VZV-infected supernatant formed amyloid fibrils following addition of amyloidogenic peptides. Amylin knockdown decreased viral cDNA. CONCLUSIONS: VZV infection increased levels of amyloidogenic peptides and amyloid in CSF and qHPNCs, indicating that VZV-induced amyloid deposition may contribute to persistent arterial inflammation in VZV vasculopathy. In addition, we identified a novel proviral function of amylin.

Clinical characteristics, breed differences, and quality of life in North American dogs with acute steroid-responsive meningitis-arteritis.

BACKGROUND: Steroid-responsive meningitis-arteritis (SRMA) is a common inflammatory neurologic disorder of dogs for which certain breeds are predisposed. OBJECTIVES: To determine whether breed differences exist in clinical features, treatment response, and relapse in a population of North American dogs with SRMA, and to evaluate the effect of disease on dogs' quality of life (QoL). ANIMALS: Sixty-one client-owned dogs with SRMA: 29 dogs identified through an American Kennel Club-Canine Health Foundation survey and 32 dogs from North Carolina (NC) State Veterinary Hospital. METHODS: Retrospective case series. Caregivers completed an online survey to assess QoL. RESULTS: Breeds represented most often included the Golden Retriever (n = 12), Bernese Mountain Dog (10), Wirehaired Pointing Griffon (9), Boxer (9), and Beagle (6). No breed differences were identified with respect to clinical severity, diagnostic findings, or outcome. Twenty-nine dogs (48%) had ≥1 disease relapse. There was a significant effect of cerebrospinal fluid nucleated cell count on the frequency of disease relapse (P = .003), but no relationship was identified between treatment protocol and relapse. Dogs' QoL was associated with the severity of corticosteroid-related adverse effects (P = .03), which were dose-related (r = .24, P = .02) and more prevalent in Wirehaired Pointing Griffons than in other breeds (P = .04). CONCLUSION AND CLINICAL IMPORTANCE: Golden Retrievers and Wirehaired Pointing Griffons should be considered among the breeds recognized to develop SRMA. Treatment with higher corticosteroid dosages is correlated with more severe adverse effects and worse QoL, but it may not improve clinical outcome.

The endocannabinoid system in canine Steroid-Responsive Meningitis-Arteritis and Intraspinal Spirocercosis.

Endocannabinoids (ECs) are involved in immunomodulation, neuroprotection and control of inflammation in the central nervous system (CNS). Activation of cannabinoid type 2 receptors (CB2) is known to diminish the release of pro-inflammatory factors and enhance the secretion of anti-inflammatory cytokines. Furthermore, the endocannabinoid 2-arachidonoyl glycerol (2-AG) has been proved to induce the migration of eosinophils in a CB2 receptor-dependent manner in peripheral blood and activate neutrophils independent of CB activation in humans. The aim of the current study was to investigate the influence of the endocannabinoid system in two different CNS inflammatory diseases of the dog, i.e. Steroid-Responsive Meningitis-Arteritis (SRMA) and Intraspinal Spirocercosis (IS). The two main endocannabinoids, anandamide (AEA) and 2-AG, were quantified by mass spectrometry in CSF and serum samples of dogs affected with Steroid- Responsive Meningitis-Arteritis in the acute phase (SRMA A), SRMA under treatment with prednisolone (SRMA Tr), intraspinal Spirocercosis and healthy dogs. Moreover, expression of the CB2 receptor was evaluated in inflammatory lesions of SRMA and IS and compared to healthy controls using immunohistochemistry (IHC). Dogs with SRMA A showed significantly higher concentrations of total AG and AEA in serum in comparison to healthy controls and in CSF compared to SRMA Tr (p<0.05). Furthermore, dogs with IS displayed the highest ECs concentrations in CSF, being significantly higher than in CSF samples of dogs with SRMA A (p<0.05). CSF samples that demonstrated an eosinophilic pleocytosis had the highest levels of ECs, exceeding those with neutrophilic pleocytosis, suggesting that ECs have a major effect on migration of eosinophils in the CSF. Furthermore, CB2 receptor expression was found in glial cells in the spinal cord of healthy dogs, whereas in dogs with SRMA and IS, CB2 was strongly expressed not only in glial cells but also on the cellular surface of infiltrating leukocytes (i.e. neutrophils, eosinophils, lymphocytes, plasma cells, and macrophages) at lesion sites. The present study revealed an upregulated endocannabinoid system in dogs with inflammatory CNS diseases, highlighting the endocannabinoid system as a potential target for treatment of inflammatory CNS diseases.

Th17-skewed immune response and cluster of differentiation 40 ligand expression in canine steroid-responsive meningitis-arteritis, a large animal model for neutrophilic meningitis.

BACKGROUND: Steroid-responsive meningitis-arteritis (SRMA) is an immune-mediated disorder characterized by neutrophilic pleocytosis and an arteritis particularly in the cervical leptomeninges. Previous studies of the disease have shown increased levels of IL-6 and TGF-ß1 in cerebrospinal fluid (CSF). In the presence of these cytokines, naive CD4+ cells differentiate into Th17 lymphocytes which synthesize interleukin 17 (IL-17). It has been shown that IL-17 plays an active role in autoimmune diseases, it induces and mediates inflammatory responses and has an important role in recruitment of neutrophils. The hypothesis of a Th17-skewed immune response in SRMA should be supported by evaluating IL-17 and CD40L, inducing the vasculitis. METHODS: An enzyme-linked immunosorbent assay (ELISA) was performed to measure IL-17 and CD40L in serum and CSF from a total of 79 dogs. Measurements of patients suffering from SRMA in the acute state (SRMA A) were compared with levels of patients under treatment with steroids (SRMA T), recurrence of the disease (SRMA R), other neurological disorders, and healthy dogs, using the two-part test. Additionally, secretion of IL-17 and interferon gamma (IFN-γ) from the peripheral blood mononuclear cells (PBMCs) was confirmed by an enzyme-linked immunospot (ELISpot) assay. RESULTS: Significant higher levels of IL-17 were found in CSF of dogs with SRMA A compared with SRMA T, other neurological disorders and healthy dogs (p < 0.0001). In addition, levels of CD40L in CSF in dogs with SRMA A and SRMA R were significantly higher than in those with SRMA T (p = 0.0004) and healthy controls (p = 0.014). Furthermore, CSF concentrations of IL-17 and CD40L showed a strong positive correlation among each other (rSpear = 0.6601; p < 0.0001) and with the degree of pleocytosis (rSpear = 0.8842; p < 0.0001 and rSpear = 0.6649; p < 0.0001, respectively). IL-17 synthesis from PBMCs in SRMA patients was confirmed; however, IL-17 is mainly intrathecally produced. CONCLUSIONS: These results imply that Th17 cells are inducing the autoimmune response in SRMA and are involved in the severe neutrophilic pleocytosis and disruption of the blood-brain barrier (BBB). CD-40L intrathecal synthesis might be involved in the striking vasculitis. The investigation of the role of IL-17 in SRMA might elucidate important pathomechanism and open new therapeutic strategies.

Relapses in dogs with steroid-responsive meningitis-arteritis.

OBJECTIVES: To describe relapse rates in steroid-responsive meningitis-arteritis and to describe clinical and laboratory parameters in dogs with and without relapses. METHODS: Seventy-four dogs with steroid-responsive meningitis-arteritis were retrospectively identified and assigned to one of three groups: (1) without relapse; (2) at least one relapse and (3) unknown relapse status. The following parameters are reported for the first two groups: sex, age, breed, body weight, nucleated cell count, total protein concentration and percentage of neutrophils on initial cerebrospinal fluid analysis, immunoglobulin A in serum and initial cerebrospinal fluid analysis, nucleated cell count on cerebrospinal fluid analysis at 3-month re-evaluation, C-reactive protein in serum and initial cerebrospinal fluid analysis and at 3-month re-evaluation. RESULTS: Relapses occurred in 32 · 4% of dogs (one relapse: 62 · 5%; two relapses: 25 · 0%; three relapses: 8 · 3%; four relapses: 4 · 2%), 55 · 4% were relapse-free and in 12 · 2% the relapse status was unknown. C-reactive protein in serum and cerebrospinal fluid on 3-month re-evaluation was normal in 80% and 75% of dogs with relapses, respectively. In dogs without relapse, C-reactive protein in serum and cerebrospinal fluid on 3-month re-evaluation was normal in 100% and 90% of dogs, respectively. CLINICAL SIGNIFICANCE: Relapses are frequent but no reliable predictive indicator has emerged in this study. Nevertheless, elevated C-reactive protein in serum warrants continuing therapy; normal C-reactive protein in serum does not exclude future relapse.

Interleukin-6, vascular endothelial growth factor and transforming growth factor beta 1 in canine steroid responsive meningitis-arteritis.

BACKGROUND: Steroid Responsive Meningitis-Arteritis (SRMA) is a common cause of inflammation of the canine central nervous system (CNS). To investigate if transforming growth factor beta 1 (TGF-ß1), interleukin-6 (IL-6) and vascular endothelial growth factor (VEGF) are involved in the production of excessive immunoglobulin A (IgA), the induction of acute phase proteins and in the development of a systemic necrotizing vasculitis, characteristic of SRMA, these three signalling proteins were evaluated. RESULTS: Cerebrospinal fluid (CSF) and serum samples of dogs during the acute phase of SRMA (SRMA) were tested for IL-6, VEGF and TGF- ß1. Results were compared to those of dogs affected with SRMA during treatment (SRMA Th) and during relapse (SRMA R), to dogs with other meningoencephalomyelitides (ME), with miscellaneous non-inflammatory diseases of the CNS (CNS-Mix), with idiopathic epilepsy (IE), with systemic inflammatory diseases (Syst. Infl.) and with healthy dogs (Healthy). Concentrations of IL-6 and VEGF in CSF were significantly elevated in the SRMA group compared to the other disease categories (p<0.05). The CSF concentrations of TGF-ß1 were increased in SRMA group, but statistically significant differences were found only in comparison with Healthy and CNS-Mix groups. No differences were detected in the serum concentrations of TGF-ß1 between the different groups. In untreated SRMA patients, a positive correlation (rSpear = 0.3549; P=0.0337) between concentrations of TGF-ß1 and IgA concentration was found in CSF, while concentrations of IL-6 and VEGF in CSF positively correlated with the degree of pleocytosis (rSpear=0.8323; P<0.0001 and rSpear=0.5711; P=0.0166, respectively). CONCLUSIONS: Our results suggest that these three signalling proteins are biomarkers of disease activity in SRMA. VEGF might play an important role in the development of a systemic arteritis. TGF-ß1 is considered to be involved in the excessive IgA production, while IL-6 in the pleocytosis. The combined intrathecal increase of TGF-ß1 and IL-6 detected in SRMA could possibly force CD4 progenitors to differentiate towards the newly described Th17 lymphocyte subset and enhance the autoimmune response.

Evaluation of a microsphere-based immunofluorescence assay for the determination of Immunoglobulin A concentrations in cerebrospinal fluid of dogs.

The simultaneous increase of immunoglobulin A (IgA) in serum and cerebrospinal fluid (CSF) is a characteristic finding in dogs suffering from canine steroid-responsive meningitis-arteritis (SRMA). The study aimed at developing and evaluating a microsphere-based immunofluorescence assay (MIA) for the measurement of IgA, trying to fulfill the need of a quicker method using only small volumes of CSF. Microsphere beads were coated with goat-anti-dog IgA antibodies and bound IgA was detected by a mouse-anti-dog IgA antibody in combination with a PE-labeled goat-anti-mouse IgG. CSF from 44 dogs were tested for IgA and compared with an in-house utilized ELISA. Using clinical relevant reference ranges, the new method showed a good agreement (84.17%) with the ELISA. A method comparison revealed a moderate agreement only. These findings indicate that the MIA will not replace the ELISA, but it opens the possibility for further research with microsphere-based assays.

[Evaluation of decreased glucose levels in the cerebrospinal fluid of dogs]. / Untersuchungen zu erniedrigten Glukosewertenim Liquor cerebrospinalis des Hundes.

OBJECTIVE: The aim of the study was to evaluate the glucose ratio (glucose level in the cerebrospinal fluid [CSF]/blood glucose level) as a quickly available marker for detecting bacterial meningoencephalomyelitis (BM). MATERIAL AND METHODS: Blood and CSF samples of 328 dogs were reviewed and evaluated retrospectively. Following the neurological diagnosis, the dogs were assigned to seven different groups: steroid-responsive meningitis-arteritis (SRMA), intervertebral disc disease (IVDD), neoplasia of the central nervous system (N), idiopathic epilepsy (IE), bacterial meningoencephalomyelitis (BM), meningoencephalomyelitis of other origin (ME) and healthy dogs. RESULTS: The median of the CSF-glucose level (mmol/l) and the median of the glucose ratio in the SRMA group displayed the lowest values and differed significantly from the CSF-glucose levels of dogs in the groups IVDD, N, IE and healthy dogs (CSF-glucose level: p<0.01; glucose ratio: p<0.05). In the BM group, both parameters did not differ significant- ly from other groups, but displayed similar low levels as in the SRMA group. There was a negative correlation between the CSF cell count and the CSF-glucose ratio (Spearman correlation coefficient -0.322, p=0.01, R²=0.108). CONCLUSION: The CSF-glucose concentration cannot be used as a distinct marker to differentiate BM from other inflammatory CNS-diseases, especially from SRMA usually accompanied by severe pleocytosis. Low CSF-glucose levels appear to be caused by elevated CSF cell counts rather than by bacterial metabolism. CLINICAL RELEVANCE: For a definitive diagnosis of bacterial meningoencephalomyelitis in dogs, the detection of microorganisms remains necessary.

Extracellular hsp70 release in canine Steroid Responsive Meningitis-Arteritis.

The role of extracellular 70 kDa heat shock protein 70 (ehsp70) in central nervous system inflammation is vastly understudied, despite evidence supporting the ability to drive a pro-inflammatory state. We investigated the presence of ehsp70 in cerebrospinal fluid (CSF) and serum of dogs with Steroid Responsive Meningitis-Arteritis (SRMA), with the hypothesis that an ehsp70 response would occur, and might play a role in the pathogenesis of this disease. Samples from 30 dogs acutely affected with SRMA, and 30 dogs treated with corticosteroids and currently in clinical remission from SRMA were compared with normal dogs. Serum and CSF concentrations of ehsp70 were quantified using an enzyme-linked immunosorbent assay. An ehsp70 response occurred in the CSF of dogs with SRMA and this response was attenuated by corticosteroid treatment. There was no correlation between serum and CSF concentrations of ehsp70, supporting local production and release of ehsp70 and not simply leakage from serum. Dogs with SRMA thus represent a powerful spontaneous model by which to study the role of ehsp70 in CNS inflammation.

Pathogenetic factors for excessive IgA production: Th2-dominated immune response in canine steroid-responsive meningitis-arteritis.

Canine steroid-responsive meningitis-arteritis (SRMA) is a systemic inflammatory disease with a predominant manifestation within the cervical meninges, increased immunoglobulin A (IgA) levels in serum and cerebrospinal fluid (CSF), and a shift of the B:T cell ratio towards a higher percentage of B cells. A Th2-dominated immune response associated with SRMA was therefore hypothesised. Pellets of peripheral blood mononuclear cells (PBMNCs) and CSF white blood cells (CSF WBCs) from dogs in the acute phase of SRMA (n=16) and under glucocorticoid treatment for SRMA (n=16) were investigated for interleukin (IL)-2, interferon (IFN)-γ, IL-4, IL-5 and IL-10 mRNA expression by means of reverse-transcriptase real-time polymerase chain reaction. Results were compared with those of dogs with other inflammatory (n=9) and neoplastic disorders (n=10) of the central nervous system. A tendency towards low levels of Th1 response related cytokines (IL-2, IFN-γ) and high IL-4 expression was observed indicating a Th2-skewed immune response. The pronounced IL-4 production may be an important pathogenetic factor for excessive IgA production in the acute phase of SRMA and for those cases under glucocorticoid treatment.

Steroid responsive meningitis-arteritis: a prospective study of potential disease markers, prednisolone treatment, and long-term outcome in 20 dogs (2006-2008).

BACKGROUND: Previous multidrug studies have identified the value of prednisolone in treating steroid responsive meningitis-arteritis (SRMA) and the potential value of acute phase proteins (APPs) and immunoglobulin A (IgA) in diagnosis and monitoring. HYPOTHESIS: (1) Prednisolone monotherapy is a successful immunosuppressive modality in the treatment of SRMA; (2) protein markers are useful in identifying the potential for relapse. ANIMALS: Twenty client-owned dogs with SRMA presented to the University of Glasgow Small Animal Hospital between May 2006 and May 2008. METHODS: A prospective, observational study: CBC, biochemistry, and cerebrospinal fluid (CSF) analyses were performed. C-reactive protein (CRP), serum amyloid-A, alpha-1-acid glycoprotein, and haptoglobin (Hp) were assessed in the serum. IgA concentrations were determined in the serum and CSF. RESULTS: Clinical resolution of SRMA was achieved in all 20 dogs. Serum CRP concentration remained increased at remission in 16/20 dogs whereas CSF cytology was within normal limits in 20/20 dogs. Serum APPs decreased significantly on treatment (P<.05) except Hp, which remained unaltered. Serum and CSF IgA concentrations remained increased for the duration of treatment. CONCLUSIONS AND CLINICAL IMPORTANCE: The prednisolone regimen presented was successful in treating SRMA without the need for additional drugs. Serum APPs are of use in the diagnosis and management of SRMA, particularly in relation to identifying relapse. Serum and CSF IgA concentrations remain increased throughout disease, aiding in diagnosis but not contributing to the management of SRMA.

Concentrations of acute-phase proteins in dogs with steroid responsive meningitis-arteritis.

BACKGROUND: Measurement of concentrations of acute-phase proteins (APPs) is used as an aid in the diagnosis of a variety of diseases in animals. OBJECTIVE: To determine the concentration of APPs in dogs with steroid responsive meningitis-arteritis (SRMA) and other neurologic diseases. ANIMALS: One hundred and thirty-three dogs with neurologic diseases, 6 dogs with sepsis, and 8 healthy dogs were included in the study. Thirty-six dogs had SRMA (31 of which had monitoring), 14 dogs had other meningoencephalitides (ME), 32 had disk disease (IVDD/DLSS), 26 had tumors affecting the central nervous system (TCNS), and 25 had idiopathic epilepsy (IE). METHODS: Prospective, observational study: C-reactive protein (CRP), alpha(2)-macroglobulin (AMG), and albumin concentrations were determined in the serum or plasma. CRP was also measured in the cerebrospinal fluid. RESULTS: Serum CRP was significantly higher in dogs with SRMA (x=142 microg/mL+/-75) and sepsis (x=114 microg/mL+/-67) in comparison with dogs with other neurologic diseases (x=2.3-21 microg/mL; P< .001). There was no significant difference detected in AMG between groups. Serum albumin concentration was significantly lower (P< .01) in dogs with SRMA (x=3.2 g/dL+/-0.41) than in other groups (x=3.6-3.9 g/dL). Serum CRP concentration of SRMA dogs correlated with alkaline phosphatase levels (r=0.515, P= .003). CONCLUSIONS AND CLINICAL IMPORTANCE: CRP concentrations in serum are useful in diagnosis of dogs with SRMA. Serum CRP could be used as a monitoring parameter in treatment management of these dogs.

Disproportionally strong increase of B cells in inflammatory cerebrospinal fluid of dogs with Steroid-responsive Meningitis-Arteritis.

Steroid-responsive Meningitis-Arteritis (SRMA) is a systemic inflammatory disease of juvenile to young adult dogs with a relapsing course and most prominent manifestation in the cervical meninges. Immunophenotyping and flow cytometric measurement of lymphocytes in peripheral blood (PB) and CSF was performed in the acute phase of SRMA (n=12) and during glucocorticosteroid treatment (n=10). Values were compared to those from dogs with other neurologic diseases (n=63) and healthy individuals (n=7). Dogs with SRMA had high CD4:CD8alpha ratios in PB and low T:B cell ratios in PB and CSF suggesting that a T(H)2-mediated immune response occurs. The T:B cell ratio in CSF was markedly lower than that in PB indicating that either a selective recruitment of B cells or, alternatively, their strong intrathecal proliferation takes place. SRMA appears to be a valuable animal model for the investigation of compartmentalization of immune responses and for studies on differences in local central nervous system and systemic immune responses.

Chemotactic activity and IL-8 levels in the cerebrospinal fluid in canine steroid responsive meningitis-arteriitis.

Steroid responsive meningitis-arteriitis (SRMA) is a systemic immune disorder, characterized by inflammatory-stenosing lesions of the meningeal arteries and meningitis. The predilection of the disease for the central nervous system (CNS) remains unexplained. In this study, chemotactic activity and chemotactic factors were measured in the cerebrospinal fluid (CSF) of dogs with SRMA. CSF of dogs with SRMA exerted a marked chemotactic activity for leukocytes. Neutrophils were attracted to a similar degree as by CSF from animals with bacterial encephalitis. Chemotactic activity was also noted for mononuclear cells, however, by far weaker than in CSF from animals with viral encephalitis. While the inflammatory process could be suppressed with glucocorticoid treatment, the chemotactic activity of CSF persisted. We could identify IL-8-like activities using a desensitization assay in the CSF of animals with SRMA and also found increased IgA levels. Increased chemotactic activity for polymorphonuclear leukocytes correlated positively with the levels of IL-8-like activity in CSF. Our observations clearly suggest that in SRMA chemotactic factors are generated in the CNS. These include IL-8, but probably also others. The intensity of this production appears to correlate with IgA levels in the CSF suggesting either a causal link or reflecting the severity of the inflammation.