Fingolimod Modulates Peripheral Effector and Regulatory T Cells in MS Patients

Multiple sclerosis (MS) is a complex neurological disease where, in genetically predisposed individuals, the unbalanced interplay between pathogenic and regulatory T cells will result in the progression of the autoimmune assault to neural antigens. Fingolimod (FTY720), an oral sphingosine 1-phosphate modulator recently approved for the treatment of MS, inhibits the egress of T cells from lymph nodes acting specifically on naïve and memory T cells and sparing effector T cells. Here we characterized IL-17 and IFNγ producing effector CD4 and CD8 positive T cells as well as CD4 positive CD25^highCD127^low regulatory T cells in MS patients before and 1 month after treatment was started. We observed that fingolimod did not significantly affect the percentage of CCR6 and CD161 positive T cells in both CD4 and CD8 compartments. In contrast, it significantly reduced the levels of both CD4+ CCR6+ CD161+ and CD8+ CCR6+ CD161+ producing IFNγ alone or in combination with IL-17. The percentage of IL-17 secreting cells in both subsets was affected by the treatment to a lesser extent. Finally, we observed that CD4+ CD25^highCD127^low regulatory T cells were decreased in MS patients compared to healthy controls and fingolimod significantly increased their frequencies. All together these findings demonstrate that fingolimod functionally modulates the ability of potentially pathogenic effector cells to produce relevant pro-inflammatory cytokines and increases the number of circulating regulatory T cells possibly contributing in restoring a balance between these populations.     

Interferon‐γ–dependent inhibition of B cell activation by bone marrow–derived mesenchymal stem cells in a murine model of systemic lupus erythematosus

Objective

Bone marrow–derived mesenchymal stem cells (BM‐MSCs) are multipotent cells characterized by immunomodulatory properties and are therefore considered a promising tool for the treatment of immune‐mediated diseases. This study was undertaken to assess the influence of murine BM‐MSCs on the activation of B cells in (NZB × NZW)F₁ mice as an animal model of systemic lupus erythematosus (SLE).

Methods

We evaluated the in vitro effects of BM‐MSCs on the proliferation and differentiation to plasma cells of splenic mature B cell subsets, namely follicular and marginal zone B cells isolated from (NZB × NZW)F₁ mice. Lupus mice were also treated with BM‐MSCs, and serum autoantibodies, proteinuria, histologic changes in the kidney, and survival rates were monitored.

Results

BM‐MSCs inhibited antigen‐dependent proliferation and differentiation to plasma cells of follicular and marginal zone B cells in vitro. This inhibitory effect was dependent on interferon‐γ (IFNγ) and was mediated by cell‐to‐cell contact, involving the programmed death 1 (PD‐1)/PD ligand pathway. In vivo treatment with BM‐MSCs did not affect the levels of anti–double‐stranded DNA antibodies or proteinuria. However, a reduction in glomerular immune complex deposition, lymphocytic infiltration, and glomerular proliferation was observed.

Conclusion

Our findings indicate that BM‐MSCs affect B cell receptor–dependent activation of both follicular and marginal zone B cells from lupus mice. This inhibitory effect is IFNγ‐dependent and cell contact–dependent. MSCs in vivo do not affect the production of autoantibodies, the level of proteinuria, or the mortality rates. Nonetheless, the significant improvement in histologic findings in the kidney supports the potential role of MSCs in the prevention of glomerular damage.

     

Human mesenchymal stem cells modulate B-cell functions

Human mesenchymal stem cells (hMSCs) suppress T-cell and dendritic-cell function and represent a promising strategy for cell therapy of autoimmune diseases. Nevertheless, no information is currently available on the effects of hMSCs on B cells, which may have a large impact on the clinical use of these cells. hMSCs isolated from the bone marrow and B cells purified from the peripheral blood of healthy donors were cocultured with different B-cell tropic stimuli. B-cell proliferation was inhibited by hMSCs through an arrest in the G0/G1 phase of the cell cycle and not through the induction of apoptosis. A major mechanism of B-cell suppression was hMSC production of soluble factors, as indicated by transwell experiments. hMSCs inhibited B-cell differentiation because IgM, IgG, and IgA production was significantly impaired. CXCR4, CXCR5, and CCR7 B-cell expression, as well as chemotaxis to CXCL12, the CXCR4 ligand, and CXCL13, the CXCR5 ligand, were significantly down-regulated by hMSCs, suggesting that these cells affect chemotactic properties of B cells. B-cell costimulatory molecule expression and cytokine production were unaffected by hMSCs. These results further support the potential therapeutic use of hMSCs in immune-mediated disorders, including those in which B cells play a major role.     

Whole-body biodistribution and radiation dosimetry of the new cardiac tracer 99mTc-N-DBODC

Our purpose was to evaluate the safety profile and biodistribution behavior in healthy human volunteers of the new myocardial perfusion tracer bis[(dimethoxypropylphosphanyl)ethyl]ethoxyethylamine N,N'-bis(ethoxyethyl)dithiocarbamato nitrido technetium(V) (99mTc-N-DBODC). METHODS: Ten healthy male volunteers were injected with 99mTc-N-DBODC under both stress and rest conditions. Anterior and posterior planar gamma-camera images were collected at 5, 30, 60, 240, and 1,440 min after injection, with organ uptake quantified by region-of-interest analysis. Tracer kinetics in body fluids were determined by collecting blood and urine samples at different time points. RESULTS: After injection, 99mTc-N-DBODC showed significant accumulation in the myocardium and prolonged retention. Under rest conditions, uptake in the heart, lungs, and liver at 5 min after injection was 1.67% +/- 0.13%, 1.16% +/- 0.07%, and 10.85% +/- 1.72%, respectively, of administered activity. Under stress conditions, heart uptake was significantly higher (2.07% +/- 0.22%). Radioactivity in the liver decreased to 3.64% +/- 0.98% and 2.37% +/- 0.48% at 60 and 240 min, respectively, after injection. This rapid liver clearance led to favorable heart-to-liver ratios, reaching values of 0.74 +/- 0.13 at rest and 1.26 +/- 0.28 during exercise 60 min after tracer administration. Radiation dose estimates were comparable to those obtained with other myocardial perfusion cationic compounds. CONCLUSION: The high uptake in the myocardium and the fast liver washout of 99mTc-N-DBODC will allow SPECT images of the left ventricle to be acquired early and with excellent quality.     

The prevalence of multiple sclerosis in the north–west Italian province of Genoa

The objective of this study was to assess the prevalence of multiple sclerosis (MS), calculated as point prevalence on 31 December 1997, in the province of Genoa, North–western Italy. Methods The province of Genoa is located in North–western Italy, an area of 1835 km². On the point prevalence day the population consisted of 913,218 inhabitants. MS cases were identified by analysing archives of the hospitals with neurological or rehabilitation wards, neurologists serving the community, files of local chapters of the Italian MS society, all requests for oligoclonal bands analysis on CSF in the studied area. Patients included in the study were MS cases diagnosed before 31 December 1997 according to the Poser criteria resident in the province under study. Results A total of 857 subjects were alive and residing in the province of Genoa on the prevalence day. The overall crude prevalence rate was 94 per 100,000 (95% CI 88–100); 291 were males (34%) with a crude prevalence of 67 per 100,000 (95 % CI 60–76) and 566 were females (66%) with a prevalence of 118 per 100,000 (95% CI 108–128). The female/male ratio was 1.9. When age and sex were adjusted to the Italian standard population of 1991 prevalence was 85 per 100,000. Five hundred and thirty two out of the 857 patients agreed to be interviewed. The interviewed sample was representative of the prevalence sample: sex and gender distributions were identical in the two samples. The overall mean age was 48 (± 13) years (48 ± 12 years in males; 48 ± 14 years in females). Mean disease duration was 15 (± 10) years for males and 16 (± 11) years for females. Two hundred and ninety one (55 %) subjects had a relapsing remitting (RR) clinical course, 150 (28%) were secondary progressive (SP) and 91 (17%) were primary progressive (PP). Mean EDSS score was 5 (± 2; median 5). The mean age at time of onset was 33 (±10) years for males and 32 (± 11) years for females. The disease onset was monosymptomatic in 76% (n = 407) patients and polysymptomatic in 24% (n = 125). The mean length of time between clinical onset and diagnosis was 5 (± 6) years. Conclusion We confirmed that the province of Genoa is a very high risk area for MS. We found a high rate of patients with a PP course; also the proportion of patients with high disability scores is greater compared to previous studies.     

Predictions of mortality from pleural mesothelioma in Italy: a model based on asbestos consumption figures supports results from age-period-cohort models

Italy was the second main asbestos producer in Europe, after the Soviet Union, until the end of the 1980s, and raw asbestos was imported on a large scale until 1992. The Italian pattern of asbestos consumption lags on average about 10 years behind the United States, Australia, the United Kingdom and the Nordic countries. Measures to reduce exposure were introduced in the mid-1970s in some workplaces. In 1986, limitations were imposed on the use of crocidolite and in 1992 asbestos was definitively banned. We have used primary pleural cancer mortality figures (1970-1999) to predict mortality from mesothelioma among Italian men in the next 30 years by age-cohort-period models and by a model based on asbestos consumption figures. The pleural cancer/mesothelioma ratio and mesothelioma misdiagnosis in the past were taken into account in the analysis. Estimated risks of birth cohorts born after 1945 decrease less quickly in Italy than in other Western countries. The findings predict a peak with about 800 mesothelioma annual deaths in the period 2012-2024. Results estimated using age-period-cohort models were similar to those obtained from the asbestos consumption model.     

Health status description of populations living in three areas of Tuscany (Livorno,Orbetello and Piombino) through causes of death distribution

By assessing mortality causes, the authors have described the health status of people living in 3 areas of Tuscany around Livorno, Orbetello and Piombino municipalities. Direct standard mortality and local standard mortality ratios were assessed as regards the years 1988?1997. Total mortality turned out to be equal or lower than the Italian average, with the exception of the women from Livorno municipality. Cancer mortality is higher in Livorno area, among females in Orbetello area and males in both Orbetello and Piombino municipalities. As regards regional mortality, the situation in Livorno seems to be the worst, followed by Orbetello and Piombino.     

An alternative approach to the preparation of (188)Re radiopharmaceuticals from generator-produced [(188)ReO(4)](-): efficient synthesis of (188)Re(V)-meso-2,3-dimercaptosuccinic acid

A new efficient approach for the preparation of (188)Re radiopharmaceuticals starting from [(188)ReO(4)](-), produced at a carrier-free level through the (188)W/(188)Re generator system, is described. The reaction procedure was based on the combined action of different reagents and has been applied in detail to the preparation of the therapeutic agent (188)Re(V)-DMSA (H(2)DMSA [meso-2,3-dimercaptosuccinic acid]). The most efficient combination required the use of SnCl(2), oxalate ions, and gamma-cyclodextrin. These were reacted with [(188)ReO(4)](-) and H(2)DMSA to afford the final radiopharmaceutical in high radiochemical purity, at room temperature, and in weakly acidic solution. The role played by the various reagents in the reaction was investigated. It was found that SnCl(2) behaved as the actual reducing agent, whereas oxalate and gamma-cyclodextrin greatly enhanced the ease of reduction of [(188)ReO(4)](-) through the action of two hypothetical mechanisms. In the first step of the reaction, oxalate ions gave rise to the formation of Re(VII) complexes with the concomitant expansion of the coordination sphere of the metal. This process strongly favored the electron transfer between Sn(2+) and Re(+7) centers, giving rise to intermediate reduced rhenium complexes. These species were further stabilized by the formation of transient host-guest aggregates with gamma-cyclodextrin and finally converted into (188)Re(V)-DMSA through simple replacement of the coordinated ligands by H(2)DMSA.