Early prognosticators of later TSPO-PET-measurable microglial activation in multiple sclerosis.

BACKGROUND: Factors driving increased innate immune cell activation in multiple sclerosis (MS) brain are not well understood. As higher prevalence of microglial/macrophage activation in association with chronic lesions and diffusely in the normal appearing white matter predict more rapid accumulation of clinical disability, it is of high importance to understand processes behind this. Objective of the study was to explore demographic, clinical and paraclinical variables associating with later positron emission tomography (PET)-measurable innate immune cell activation. METHODS: PET-imaging using a TSPO-binding [11C]PK11195 was performed to evaluate microglial activation in patients with relapsing-remitting MS aged 40-55 years with a minimum disease duration of five years (n = 37). Medical records and diagnostic MR images were reviewed for relevant early MS disease-related clinical and paraclinical parameters. RESULTS: More prominent microglial activation was associated with higher number of T2 lesions in the diagnostic MRI, a higher immunoglobulin G (IgG) index in the diagnostic CSF and Expanded Disability Status Scale (EDSS) ≥ 2.0 five years after diagnosis. CONCLUSION: The number of T2 lesions in MRI, and CSF immunoglobulin content measured by IgG index at the time of MS diagnosis associated with later TSPO-PET-measurable innate immune cell activation. This suggests that both focal and diffuse early inflammatory phenomena impact the development of later progression-related pathology.

Dimethyl fumarate decreases short-term but not long-term inflammation in a focal EAE model of neuroinflammation.

BACKGROUND: Dimethyl fumarate (DMF) is an oral immunomodulatory drug used in the treatment of autoimmune diseases. Here, we sought to study whether the effect of DMF can be detected using positron emission tomography (PET) targeting the 18-kDa translocator protein (TSPO) in the focal delayed-type hypersensitivity rat model of multiple sclerosis (fDTH-EAE). The rats were treated orally twice daily from lesion activation (day 0) with either vehicle (tap water with 0.08% Methocel, 200 µL; control group n = 4 (3 after week four)) or 15 mg/kg DMF (n = 4) in 0.08% aqueous Methocel (200 µL) for 8 weeks. The animals were imaged by PET using the TSPO tracer [18F]GE-180 in weeks 0, 1, 2, 4, 8, and 18 following lesion activation, and the non-displaceable binding potential (BPND) was calculated. Immunohistochemical staining for Iba1, CD4, and CD8 was performed in week 18, and in separate cohorts of animals, following 2 or 4 weeks of treatment. RESULTS: Using the fDTH-EAE model, DMF reduced the [18F]GE-180 BPND in the DMF-treated animals compared to control animals after 1 week of treatment (two-tailed unpaired t test, p = 0.031), but not in weeks 2, 4, 8, or 18 when imaged in vivo by PET. Immunostaining for Iba1 showed that DMF had no effect on the perilesional volume or the core lesion volume after 2 or 4 weeks of treatment, or at 18 weeks. However, the optical density (OD) measurements of CD4+ staining showed reduced OD in the lesions of the treated rats. CONCLUSIONS: DMF reduced the microglial activation in the fDTH-EAE model after 1 week of treatment, as detected by PET imaging of the TSPO ligand [18F]GE-180. However, over an extended time course, reduced microglial activation was not observed using [18F]GE-180 or by immunohistochemistry for Iba1+ microglia/macrophages. Additionally, DMF did affect the infiltration of CD4+ and CD8+ T-lymphocytes at the fDTH-EAE lesion.

Cessation of anti-VLA-4 therapy in a focal rat model of multiple sclerosis causes an increase in neuroinflammation.

BACKGROUND: Positron emission tomography (PET) can be used for in vivo evaluation of the pathology associated with multiple sclerosis. We investigated the use of longitudinal PET imaging and the 18-kDa translocator protein (TSPO) binding radioligand [18F]GE-180 to detect changes in a chronic multiple sclerosis-like focal delayed-type hypersensitivity experimental autoimmune encephalomyelitis (fDTH-EAE) rat model during and after anti-VLA-4 monoclonal antibody (mAb) treatment. Thirty days after lesion activation, fDTH-EAE rats were treated with the anti-VLA-4 mAb (n = 4) or a control mAb (n = 4; 5 mg/kg, every third day, subcutaneously) for 31 days. Animals were imaged with [18F]GE-180 on days 30, 44, 65, 86 and 142. Another group of animals (n = 4) was used for visualisation the microglia with Iba-1 at day 44 after a 2-week treatment period. RESULTS: After a 2-week treatment period on day 44, there was a declining trend (p = 0.067) in [18F]GE-180-binding in the anti-VLA-4 mAb-treated animals versus controls. However, cessation of treatment for 4 days after a 31-day treatment period increased [18F]GE-180 binding in animals treated with anti-VLA-4 mAb compared to the control group (p = 0.0003). There was no difference between the groups in TSPO binding by day 142. CONCLUSIONS: These results demonstrated that cessation of anti-VLA-4 mAb treatment for 4 days caused a transient rebound increase in neuroinflammation. This highlights the usefulness of serial TSPO imaging in the fDTH-EAE model to better understand the rebound phenomenon.

Humoral response to John Cunningham virus during pregnancy in multiple sclerosis.

BACKGROUND: Pregnancy induces an immunosuppressive state in the mother to ensure immunological acceptance of the foetus. Impairment of cell-mediated immune responses may render the mother susceptible to intracellular pathogens. It is not presently known whether pregnancy alters the immunosurveillance for John Cunningham virus (JCV), an opportunistic pathogen associated with natalizumab treatment for multiple sclerosis (MS). OBJECTIVE: To evaluate whether the humoral immune response to JCV is altered during pregnancy among MS patients and healthy controls to get insight to potential pregnancy-induced alterations related to immune response to JCV during pregnancy. METHODS: Serum anti-JCV-antibody-indices (JCV-Ab-index) were determined by a two-step second-generation enzyme-linked immunosorbent assay in 49 MS patients during and after pregnancy and in 49 healthy controls during pregnancy. For comparison, total IgG levels and antibodies against Epstein-Barr, cytomegalo and measles viruses were similarly measured. RESULTS: The JCV-Ab-indices of MS patients were not altered during the pregnancy (1st vs. 3rd trimester, 0.62 vs. 0.77, p = 0.99). Contrary to this, in the healthy controls JCV-Ab-indices (p = 0.005), antibody levels to the other viruses, and total IgG levels (p < 0.0001) decreased significantly during pregnancy. CONCLUSIONS: JCV-Ab levels remain unaltered during MS pregnancy, while the total IgG concentration is reduced/diluted due to increasing plasma volumes during the course of pregnancy. This may imply a biologically significant alteration in the immune response to JCV during MS pregnancy.

Anti-JC virus seroprevalence in a Finnish MS cohort.

BACKGROUND: The risk of progressive multifocal leukoencephalopathy (PML) caused by the JC virus (JCV) is increased in patients with multiple sclerosis receiving biological therapies. OBJECTIVES: To determine the seroprevalence of anti-JCV antibodies in Finnish patients with multiple sclerosis (MS) and clinically isolated syndrome and to assess the clinical risk factors for JCV seropositivity. METHODS: The JCV seroprevalence was analyzed in 503 patients using a second-generation two-step ELISA. Sixty-seven patients underwent longitudinal serological evaluation over 4.5 years. RESULTS: The overall seroprevalence of JCV was 57.4%. The seropositivity was higher in men than in women, tended to increase with age, and was not affected by different immunomodulatory therapies. However, in patients with ongoing natalizumab treatment (n = 72), the anti-JCV antibody screening index was lower than in patients without such therapy [median 0.3 (range 0.1-3.1) vs 0.6 (0.1-3.1), respectively, P = 0.01]. Over 4.5 years, 4/19 (21%) initially seronegative patients converted to seropositivity, whereas 4/48 (8.3%) initially seropositive patients reverted to seronegativity. Fluctuations in serostatus were observed in 3/67 patients. CONCLUSION: The study confirmed a high anti-JCV antibody prevalence in patients with MS and its association with age and male gender but not with disease-modifying therapies. Our data suggest that therapy with natalizumab may cause a decrease in anti-JCV antibody levels, suggesting an immunosuppressive effect of natalizumab without an impact on JCV seroprevalence. The results of studies performed until now confirm the predictive value of anti-JCV antibody measurement in the assessment of PML risk; however, changes in serostatus need to be considered.

Hormonal and gender-related immune changes in multiple sclerosis.

Similarly to many other autoimmune diseases, multiple sclerosis (MS) is more common among women than men, and its incidence among women is rising. There are also qualitative differences in the disease course between men and women, with male patients experiencing increased disease progression, brain atrophy, and cognitive impairment. During pregnancy, women with MS typically have a greatly reduced relapse rate, whereas very soon after the delivery, the disease activity returns, often even at a higher level than seen in the prepregnancy year. The reasons for the increased postpartum activity are not entirely clear, but factors such as the abrupt decrease in estrogen levels immediately after the delivery and the loss of the immunosuppressive state of pregnancy are likely of importance. There is compelling evidence that estrogen, progesterone, and testosterone control MS pathology by influencing immune responses and by contributing to repair mechanisms in the nervous system. Hormones may thus offer important insights into MS disease prevention and treatment. In this review, the possible reasons for the sex bias in autoimmune diseases will be discussed. The pregnancy-related alterations in MS, including the effect of pregnancy on disease activity, long-term disability accumulation, and prevalence will be reviewed, as well as the hormonal and immunological mechanisms potentially underlying these changes. Finally, the present thinking on the effect of hormones on the changing incidence of MS will be elucidated.

Multiple sclerosis and vitamin D during pregnancy and lactation.

BACKGROUND: Both pregnancy and high vitamin D concentration seem to generate a protective environment against multiple sclerosis (MS) relapses. Longitudinal case-control analysis of vitamin D concentrations during pregnancy and lactation of MS mothers is lacking. AIMS OF THE STUDY: To examine serum 25-hydroxyvitamin-D3 levels of MS patients during and after pregnancy and compare these to the levels measured in healthy controls. METHODS: Fifteen relapsing-remitting MS mothers underwent repeated testing for 25-hydroxyvitamin-D3 at 10-12, 26-28 and 35-37 gestational weeks and 1, 3 and 6 months post-partum. An identical series of samples was collected from six control mothers. RESULTS: The prevalence of vitamin D deficiency (<50 nmol/l) during pregnancy was high (73%) among MS patients. Vitamin D levels were significantly higher during pregnancy when compared to early post-partum values among MS patients. At the end of the follow-up period, the vitamin D levels returned to levels observed in early pregnancy. In healthy controls, the alterations during and after pregnancy were similar in nature, but the vitamin D concentrations were higher at all time points when compared to MS patients (P = 0.037). CONCLUSIONS: Vitamin D deficiency during the pregnancy and lactation seems to be common in mothers with MS and needs to be treated adequately.

Increased thyroid autoimmunity among women with multiple sclerosis in the postpartum setting.

BACKGROUND: Multiple sclerosis (MS) patients are predisposed to thyroid abnormalities, but the risk for pregnancy-related thyroid pathology among MS patients has not been evaluated. OBJECTIVES: The objectives of this research are to prospectively evaluate the prevalence of thyroid autoimmunity among MS patients in relation to pregnancy, and to investigate its impact on pregnancy outcome, postpartum depression and fatigue. METHODS: Forty-six pregnant MS patients underwent repeat testing for serum thyroid antibodies (Abs), clinical evaluation and thyroid hormone measurement. Results were compared to 35 age-matched healthy mothers. RESULTS: At six months postpartum 35.3% of MS patients presented elevated levels of thyroid Abs compared to 5.7% of controls, p = 0.01. Mean thyroid Ab concentrations among MS patients were significantly reduced during pregnancy and returned to maximal levels at six months postpartum. The proportion of individuals with postpartum thyroid dysfunction did not differ significantly between MS patients and healthy controls (3.4% vs 2.9%, p = 1.00). Elevated thyroid Ab levels did not increase the risk for adverse pregnancy outcome, fatigue or postpartum depression. CONCLUSIONS: Considering the tendency of MS mothers to develop thyroid autoimmunity postpartum and in association to treatments, we recommend screening MS patients for thyroid dysfunction (TSH) during early pregnancy and after delivery.

Is the modulatory effect of pregnancy in multiple sclerosis associated with changes in blood apoptotic molecules?

OBJECTIVE: We examined whether the modulatory effect of pregnancy on multiple sclerosis (MS) is associated with changes in the apoptotic molecules in sera. SUBJECTS AND METHODS: The serum levels of tumor necrosis factor-related apoptosis-inducing ligand (sTRAIL), sFas, Fas ligand (sFasL) and macrophage migration inhibitory factor were analyzed from 19 MS patients and 14 controls during late pregnancy and post-partum. The obtained results were related to disease activity and the progression of MS. RESULTS: Disease activity decreased during pregnancy. The levels of sTRAIL and sFasL increased from late pregnancy to post-partum situation in both MS patients and controls, but in MS patients the changes in the levels of sTRAIL from late pregnancy to post-partum were smaller than in controls. CONCLUSIONS: Post-partum upregulation of TRAIL and FasL seems to be caused by physiologic reactivation of the mother's immune system after pregnancy. An increased risk of relapses in MS post-partum may be associated with changes in the immunomodulatory potential of these apoptotic molecules.

Pregnancy outcome in women with multiple sclerosis: results from a prospective nationwide study in Finland.

The majority of individuals obtaining the diagnosis of multiple sclerosis are women of childbearing age. They are naturally concerned as to how multiple sclerosis affects the course of pregnancy and the developing foetus. The objective of this study was to prospectively evaluate the incidence of pregnancy complications and delivery risks, and to follow the natural course of multiple sclerosis during and after pregnancy in a cohort of Finnish patients with multiple sclerosis. Sixty-one patients with multiple sclerosis who became pregnant during the years 2003-2005 were prospectively followed-up from early pregnancy until 6 months postpartum. Multiple sclerosis relapses, Expanded Disability Status Scale rates and obstetric details were recorded. The results were compared with the statistics obtained from Finnish Medical Birth Register from the year 2004. We found that patients with multiple sclerosis were no more likely to experience pregnancy complications than Finnish pregnant women generally, but they had a greater likelihood for a need of artificial insemination (4.9% vs. 0.9%; p = 0.0009). Subjects with multiple sclerosis were more likely to undergo assisted vaginal delivery than the at-large cohort (16.4% vs. 6.5%; p = 0.0017). We conclude that pregnancy does not seem to pose a woman with multiple sclerosis to a greater risk for pregnancy complications when compared with women in general. The potential need for instrumental delivery should, however, be taken into account when planning the delivery of a mother with multiple sclerosis.

Apparent Diffusion Coefficient Histograms in the Follow-up of Relapsing-Remitting Multiple Sclerosis.

Diffusion-weighted (DW) magnetic resonance (MR) imaging in addition to conventional magnetic resonance MR images provide valuable information in multiple sclerosis (MS). Increased diffusivity measured with diffusion-weighted imaging (DWI) has been demonstrated in normal appearing brain tissue in MS. So far, longitudinal changes taking place in whole-brain histograms in patients with active relapsing-remitting MS have not been evaluated. The aim of this study was to evaluate how apparent diffusion coefficient (ADC) histograms are altered during the follow-up of active relapsing-remitting MS patients. Nine patients were studied twice by MRI with a three-month interval. All patients had active newly diagnosed MS with two to three relapses during the year preceding the first MRI, and interferon-beta treatment was initiated after obtaining the first image. ADC histograms were produced after removing extracranial tissues and cerebrospinal fluid from the images. Additionally, brain volume index (BVI) and lesion volume on FLAIR images were measured. Five patients had signs of disease activity in the follow-up MRI. In the four patients without signs of disease activity the change in ADC histogram parameters was less than 2%. In patients with disease activity both increase (one case) and decrease (four cases) in histogram parameters were detected. Changes in BVI or lesion volume did not significantly correlate with histogram changes. The number of new T2-lesions showed a positive correlation with mean (r=0.79, P=0.014) and upper quartile (r=0.77, P=0.021) value change. Alterations in disease activity lead to histogram changes; both shifts to lower values and shifts to higher values are possible. The histogram changes are mostly related to subtle inflammatory changes in normal appearing brain tissue during inflammatory activity and their resolution during less active inflammatory conditions.

Immunoregulatory factors in multiple sclerosis patients during and after pregnancy: relevance of natural killer cells.

Multiple sclerosis (MS) ameliorates typically during pregnancy but after the delivery the relapse rate often increases. Our study was conducted to understand the immunoregulatory mechanisms accompanying this phenomenon. MS patients were followed-up prospectively during pregnancy and 6 months postpartum, with immunological characterization of the peripheral blood. Groups of age- and parity-matched healthy pregnant women, and age- and sex-matched non-pregnant women and non-pregnant MS patients were studied as controls. In our patient cohort, the annualized relapse rate was 1.0 +/- 1.0 relapses/woman/year (mean +/- standard deviation) during the year before pregnancy, but dropped to 0.2 +/- 0.9 during the third trimester (P = 0.02). After the delivery the relapse rate increased again to 1.4 +/- 1.9 (1-3 months postpartum versus third trimester P = 0.003). While percentages of peripheral blood CD3, CD4, CD8 and CD19 immune cell subsets were unchanged over the observation period, reduced disease activity during the last trimester was associated with a significant increase in the percentage of circulating CD56(bright) natural killer (NK) cells. Simultaneously, the proportion of circulating CD56(dim) NK cells was clearly reduced. No alteration was noted in CD4+ CD25(high) forkhead box P3+ regulatory T cells. Production of interferon-gamma by peripheral blood lymphocytes was down-regulated significantly during pregnancy in comparison to the postpartum period, resulting in an increased T helper type 2 (Th2) : Th1 ratio during pregnancy. In conclusion, pregnant state in MS patients is characterized by an increase in the percentage of CD56(bright) NK cells and by enhanced Th2 type cytokine secretion. Our findings suggest a potential role for CD56(bright) regulatory NK cells in the control of autoimmune inflammation during pregnancy in MS.

Vascular adhesion protein-1 in human ischaemic stroke.

AIMS: Leukocyte extravasation exacerbates tissue injury after ischaemic stroke. Vascular adhesion protein-1 (VAP-1) is an endothelial adhesion molecule with the potential capacity to guide transmigration of inflammatory cells into ischaemic brain. Moreover, VAP-1 could worsen ischaemic brain injury due to its function as a semicarbazide-sensitive amine oxidase (SSAO) producing toxic metabolites from primary amines. The purpose of this study was to elucidate these aspects of VAP-1-function in the pathogenesis of human ischaemic stroke. METHODS: We studied VAP-1 expression in infarcted and control brains post mortem using immunohistochemistry. Levels of soluble VAP-1 (sVAP-1) in the serum of patients with acute stroke and in control sera were determined using enzyme-linked immunosorbent assay. RESULTS: In the acute phase of ischaemic stroke, the frequency of VAP-1-stained vessels was strongly diminished in the ipsilateral hemisphere but in the contralateral hemisphere it was comparable with the expression in the control brains. In the serum of acute stroke patients with a symptom duration <6 h the level of sVAP-1 was significantly increased (652 +/- 224 ng/ml; mean +/- SD) when compared with an age- and sex-matched control group (542 +/- 104 ng/ml; P < 0.05). CONCLUSIONS: As both cell surface and sVAP-1 possess vasculopathy-promoting SSAO enzymatic activity, our results imply that by inducing SSAO-derived toxic metabolites, VAP-1 might aggravate ischaemic vascular changes. The subsequent release of sVAP-1 into circulation could be further examined as a potential marker of early ischaemic vasculopathy.

Magnetic resonance imaging of the brain used to detect early post-partum activation of multiple sclerosis.

Post-partum relapses are a frequent phenomenon in multiple sclerosis (MS). The purpose of this study was to evaluate the timing and extent of new or growing T2-lesions after delivery in a cohort of Finnish MS patients. In addition to serial magnetic resonance imaging (MRI), the patients were followed up clinically with determination of relapse rate and expanded disability status scale. The annualized relapse rate was decreased during the last trimester of pregnancy [mean 0.14, standard deviation (SD) 0.14] when compared with the time before pregnancy (mean 0.64, SD 0.14; P = 0.04) and to time post-partum (mean 1.50, SD 0.45; P = 0.0002). New or enlarging lesions were detected in the post-partum images in 14 of 28 patients. Gadolinium-enhancing lesions in post-partum MRI were present in eight of 13 patients. There was a significant increase in the number of T2-lesions (P = 0.0009), in the total volume of MS-lesions measured from fluid-attenuated inversion recovery images (P = 0.0126) and in the number of diffusion weighted imaging hyperintense lesions (P = 0.0098) in the post-partum images. The clinical results support the earlier findings of decreased disease activity in late pregnancy. The clinical and MRI findings indicate that post-partum activation is an early and common phenomenon amongst mothers with MS.

Expansion of CD56Bright natural killer cells in the peripheral blood of multiple sclerosis patients treated with interferon-beta.

We studied how interferon-beta (IFN-beta) treatment of relapsing-remitting multiple sclerosis (MS) affects subgroups of natural killer cells (NK cells). Following IFN-beta treatment, there was an expansion of CD56(Bright) NK-cells in the peripheral blood of MS patients, while at the same time the proportion of CD56(Dim) cells was diminished. In a control group, the proportion of CD56(Bright) NK-cells was significantly higher in secondary lymphoid tissues compared to the peripheral blood of the same individual. Our findings confirm that CD56(Bright) NK-cells preferably locate within the secondary lymphoid tissues, where they may interact with T cells and thereby contribute to the control of the disease activity in MS.

25-Hydroxyvitamin D levels in serum at the onset of multiple sclerosis.

Past sun exposure and vitamin D supplementation have been associated with a reduction in the risk of MS. We measured the serum concentration of 25-hydroxyvitamin D (25[OH]D) at the time of MS diagnosis in 40 MS patients and 40 controls. We found no difference in the serum levels of 25(OH)D between MS patients and controls when all samples or samples obtained during winter months were compared, but MS patients had significantly lower serum 25(OH)D concentrations in June to September than the controls. The vitamin D stores were adequate for bone metabolism (> 37 nmol/L) in 70% of MS patients throughout the year and within the hypovitaminosis level (< 37 nmol/L) in 30% of MS patients at some time of the year. During MS-relapses, 25(OH)D levels were lower than in remission, but mostly within the reference range observed in relation with normal bone metabolism. We conclude that the vitamin D stores in most MS patients are adequate for their normal bone metabolism. However, lower vitamin D levels during MS relapses than in remission suggest that vitamin D could be involved in the regulation of the clinical disease activity of MS. The optimal serum levels of vitamin D for the regulation of immune responses remain to be determined.