T-Helper Cells & Cytokines In MS

Pro- and anti-inflammatory Cytokines are thought to participate in the development and regulation of AutoImmunity in Multiple Sclerosis (MS), a DeMyelinating Disease of the Central Nervous System (CNS).

We analyzed the percentage of Interferon-gamma (IFN-) and InterLeukin-4 (IL-4) producing cells in the peripheral blood of both active and stable MS patients, and of healthy controls.

After short-term stimulation, Cytokine-producing cells were IntraCellularly stained and sorted.

Significantly lower percentages of IFN- and IL-4-producing T-Cells were found in stable MS patients than in controls, and in active than in stable patients.

The diminution affected CD4⁺ (Th1, Th2) and CD8⁺ (Tc1) phenotypes.

Tc2 cells were not detected. The Th1/Th2 ratio did not differ in active and stable MS, nor in controls.

The fact that Th2 and Tc1 cell percentages were higher in stable than in active MS possibly indicates that these cells play a down-modulating role in the Immune Response.

In contrast, a role in exacerbating the Immune Response is not attributable to Th1 cells, given their reduction in acute MS.

Our data do not support the hypothesis that MS is a Th1/Th2 paradigmatic disease;

Rather, they suggest that sequestration in the CNS, or activation-induced Apoptosis (whether in vivo or in vitro) may explain reduced levels of IFN- and IL-4 -producing subsets in the peripheral blood of clinically acute patients.

PhosphoDiEsterase-4 (PDE4) Inhibitors have the potential to modulate Immune Responses from the Th1 toward the Th2 phenotype and are considered candidate therapies for Th1-mediated AutoImmune Disorders.

However, depending on the model and cell types employed, studies of atopic individuals have come to the opposite conclusion, i.e., that PDE Inhibitors may be beneficial in Asthma.

Using in vitro ImmunoPharmacologic techniques we analyzed the effects of PDE4 and PDE3 inhibitors on human Immune Cells to address these discrepancies and broaden our understanding of their mechanism of action.

Our results indicate that PDE Inhibitors have complex inhibitory effects within in vivo achievable concentration ranges on Th1-mediated Immunity, whereas Th2-mediated responses are mostly unaffected or enhanced.

The Th2 skewing of the developing Immune Response is explained by the effects of PDE Inhibitors on several factors contributing to T-Cell priming: the Cytokine milieu; the type of costimulatory signal, i.e., up-regulation of CD86 and down-regulation of CD80; and the Ag avidity.

The combination of PDE4 and PDE3 Inhibitors expresses synergistic effects and may broaden the therapeutic window.

Finally, we observed a differential sensitivity to PDE Inhibition in AutoReactive vs foreign Ag-specific T-Cells and cells derived from Multiple Sclerosis patients vs those derived from healthy donors.

This suggests that PDE Inhibition weakens the strength of the T-Cell stimulus and corrects the underlying disease-associated Cytokine skew in T-Cell-mediated AutoImmune Disorders.

These new findings broaden the understanding of the ImmunoModulatory actions of PDE Inhibitors and underscore their promising drug profile for the treatment of AutoImmune Disorders.

IL-18 is a Cytokine which plays an important role in Th1 response through its ability to induce IFN- production in T-Cells and NK Cells.

The purpose of the study was to measure IL-18 levels in Serum and CSF of 21 patients with the Relapsing/Remitting form of MS.

9 with active Gadolinium enhancing lesions in MRI and 12 Without Enhancing lesions, and to compare results with control group consisting of 11 patients with diagnosis of NeurAsthenia and Tension Headache.

IL-18 concentration in the CSF and Sera was measured by ELISA. We found a highly significant increase of both IL-18 CSF and Serum levels in MS patients in comparison with the control group.

In patients with active MRI lesions the levels of IL-18 in CSF and Serum were significantly higher in comparison with the levels found in patients without enhancing lesions.

The results suggest involvement of IL-18 in ImmunoPathogenesis of MS especially in the active stages of the disease.

Serum but not CSF concentrations of the Interferon--inducing Cytokine InterLeukin-18 (IL-18) were significantly augmented in patients with MS.

As compared to both healthy controls and patients with Other Neurologic Diseases.

Patients with MS with Chronic Secondary/Progressive disease had significantly higher Serum levels than those with Relapsing/Remitting MS.

In the latter group, IL-18 levels were higher in patients with acute exacerbation as compared to those with stable disease.

Myelin-specific EnCephalitoGenic T-Cells, when passively transferred into rats or mice, cause an Experimental AutoImmune Disease. Previous studies by our group have shown that:

1. The same cells also significantly reduce post-traumatic degeneration in these animals after injury to the Central Nervous System
2. This beneficial AutoImmunity is a Physiological response
3. Animals differ in their ability to resist injurious conditions
4. The ability to resist post-traumatic degeneration correlates with resistance to the development of an AutoImmune Disease

Here we show that Optic Nerve Neurons in both resistant and susceptible rat strains can be protected from secondary degeneration after crush injury by immunization with Myelin Basic Protein emulsified in complete or incomplete Freund's Adjuvant.

We provide evidence that potentially destructive AutoImmunity (causing AutoImmune Disease) and beneficial AutoImmunity (causing improved Neuronal survival) both result from activity of the same Myelin-specific, ProInflammatory Th1 cells.

We further show that following passive transfer of such Th1 Cells, the expression of their beneficial potential depends on the activity of an additional T-Cell (CD4⁺) population.

By identifying the additional cellular component of AutoImmune NeuroProtection, we may be able to take meaningful steps toward achieving NeuroProtection without risk of accompanying AutoImmune Disease.

CD4⁺ CD25(high) regulatory T-Cells (Tregs) of patients with Relapsing/Remitting (RR) Multiple Sclerosis (MS), in contrast to those of patients with Secondary/Progressive (SP) MS, show a reduced suppressive function.

In this study, we analysed forkhead box P3 (FOXP3) at the single-cell level in MS patients and controls (healthy individuals and patients with Other Neurological Diseases) by means of intracellular flow cytometry.

Our data revealed a reduced number of peripheral blood CD4⁺ CD25(high) FOXP3⁺ T-Cells and lower FOXP3 protein expression per cell in RR-MS patients than in SP-MS patients and control individuals, which was correlated with the suppressive capacity of Tregs in these patients.

Interestingly, Interferon-beta (IFN-ß)-treated RR-MS patients showed restored numbers of FOXP3⁺ Tregs.

Furthermore, a higher percentage of CD4⁺ CD25(high) FOXP3⁺ Tregs in RR-MS patients, as compared with controls and SP-MS patients, expressed CD103 and CD49d, Adhesion Molecules involved in T-Cell recruitment towards inflamed tissues.

This was consistent with a significantly increased number of CD27⁺ CD25(high) CD4⁺ T-Cells in the CerebroSpinal Fluid (CSF), as compared with peripheral blood, in RR-MS patients.

Taken together, these data show aberrant FOXP3 expression at the single-cell level correlated with Treg dysfunction in RR-MS patients.

Our results also suggest that Tregs accumulate in the CSF of RR-MS patients, in an attempt to down-regulate local inflammation in the Central Nervous System.

Summary Multiple Sclerosis (MS) is a Chronic Inflammatory DeMyelinating Disease of the Central Nervous System and a defect in the regulatory T-Cell subset seems to be involved in the pathogenesis of the disease.

Foxp3 is a transcription factor that is selectively expressed in CD4⁺ CD25⁺ regulatory T-Cells and is required for their development and function.

T-bet is a key transcription factor for the development of T-Helper 1 (Th1) Cells.

We found that both the percentage of circulating CD4⁺ CD25⁺ Foxp3⁺ cells and Foxp3 expression were lower in Relapsing/Remitting (RR) MS patients during relapses than during remission.

Otherwise, the percentage of CD4⁺ T-bet⁺ T-Cells and T-bet expression in CD4⁺ T-Cells were higher in Relapsing than in Remitting RRMS patients.

CD4⁺ CD25⁺ T-Cells both from Relapsing and from Remitting RRMS patients showed significantly less capacity than corresponding cells from healthy subjects to suppress autologous CD4⁺ CD25^- T-Cell proliferation, despite a similar Foxp3 expression level.

CD4⁺ CD25⁺ T-Cells from healthy subjects and patients in remission clearly reduced T-bet mean fluorescence intensity (MFI) in CD4⁺ CD25^- T-Cells up to a ratio of 1:10.

Whereas CD4⁺ CD25⁺ T-Cells from patients in relapse were able to reduce T-bet expression only at a high ratio.

Our data indicate that the increased number of regulatory T (T-reg) Cells and the increased Foxp3 expression in circulating CD4⁺ CD25⁺ T-Cells may contribute to the maintenance of tolerance in the remission phase of MS.

Moreover, the inhibitory capacity of CD4⁺ CD25⁺ T-Cells seems to be impaired in relapsing patients under inflammatory conditions, as shown by the high levels of T-bet expression in CD4⁺T-Cells.