Carla Iarlori

Peripheral cytokines profile in Parkinson's disease

Higher levels of proinflammatory cytokines are found in Parkinsons disease (PD) patients brains and inflammation is thought to be a major contributor to the neurodegeneration. During the inflammatory process, microglial release of proinflammatory cytokines act on the endothelium of blood-brain barrier (BBB) cells to stimulate upregulation of adhesion molecules. Consequently, this upregulation leads to the recruitment of passing T cells and monocytes, which express the counter receptors, that then go on to release more cytokines [Whitton, P.S., 2007. Inflammation as a causative factor in the aetiology of Parkinsons disease, Br. J. Pharmacol. 50, 963-976; Kortekaas, R., Leenders, K.L., Van Oostrom, J.C., Vaalburg, W., Bart, J., Willemsen, A.T., Hendrikse, N.H., 2005. Blood-brain barrier dysfunction in parkinsonian midbrain in vivo, Ann. Neurol. 57, 176-179]. In addition, a systemic inflammatory response results in the production of cytokines which circulate in the blood and communicate with neurons within the brain. Thus, a central inflammatory reaction interacts with peripheral blood mononuclear cells (PBMCs) modulating immune activity. The present study investigates levels of production and expression of cyto/chemokines by PBMCs in PD patients. Basal and LPS-induced levels of MCP-1, RANTES, MIP-1alpha, IL-8, IFNgamma, IL-1beta and TNFalpha were significantly higher in PD patients than in HC subjects (p<0.001), as determined by RT-PCR and Elisa methods. Cyto/chemokine levels were significantly correlated with UPDRS III and H/Y stage (p<0.001). The Pearsons correlation coefficient (R) was also used to assess the strength of the relationship between NF-kappaBp65 levels and all studied cyto/chemokines and between NF-kappaBp65, UPDRS III and H/Y score in PD patients. The overall results strengthen and extend the knowledge of the peripheral dysregulation in the cytokine network associated with PD.

Treatment with an acetylcholinesterase inhibitor in Alzheimer patients modulates the expression and production of the pro-inflammatory and anti-inflammatory cytokines.

Elevated levels of cytokines have been detected in brains of Alzheimers disease (AD) patients, and altered peripheral levels of IL-1beta, TNFalpha and IL-6 have been reported in these patients. We studied the ability of PBMC from patients with AD, matched with a control group, to release pro- and anti-inflammatory cytokines, and the effect of AChEI treatment on cytokine release. Our data indicates that AChEI treatment down-regulates IL-1, IL-6 and TNF, and up-regulates the expression and production of IL-4 in PBMC in AD patients, and that AChEI leads to the remodelling of the cytokine network, probably acting on the lymphocytic cholinergic system.

Peripheral Chemokine Receptors, Their Ligands, Cytokines and Alzheimer's Disease

Cerebral inflammation as well as systemic immunological alterations has been reported in Alzheimers disease (AD). We aimed to determine whether spontaneous and mitogen stimulated production of peripheral blood mononuclear cell (PBMC) cytokines, chemokines and chemokine receptors in clinically diagnosed patients with AD were unregulated. PBMC were purified from AD patients and from healthy controls. Supernatants were analyzed for cytokine levels by ELISA methods. mRNA expression was determined by RT-PCR. Expression of chemokine receptors CCR2 and CCR5 was determined by cytofluorimetric analysis. Both CCR5 and CCR2 expression were increased in AD patients respect to control subjects and the expression of CCR2 and CCR5 was more frequent on CD4+ and less frequent on CD8+ cells. Levels of Th1-type cytokine IFNgamma and chemokine RANTES were increased and levels of Th2-type cytokine IL-4 and chemokine MCP-1 were reduced in AD patients compared with those of control subjects. Acetylcholinesterase inhibitor pyridostigmine bromide (AChEI)-therapy reduced CCR2, CCR5, RANTES and IFNgamma expression and production in AD patients. CCR5, CCL5/RANTES, CCL2/MCP-1 and IFNgamma expression and production were increased in PBMC treated with amyloid-beta1-42. Addition of AChEI to PBMC suppresses CCL5/RANTES and IFNgamma. The observed patterns of cyto-chemokine involvement strengthen the questions regarding the inflammatory theory in AD, and raise a pathophysiologic role for selective alteration of cyto-chemokine network.

Acetylcholinesterase inhibitors effects on oncostatin-M, interleukin-1β and interleukin-6 release from lymphocytes of Alzheimer's disease patients

Many factors are involved in the pathogenesis of Alzheimers disease (AD), and inflammatory-immunologic activation seems to play a major role. One strategy for treatment of AD has been to use acetylcholinesterase (AChE) inhibitors to increase the levels of acetylcholine and enhancing cholinergic activity in the affected regions of the brain. Cholinergic compounds modulate the immune system, therefore secretion, by peripheral blood mononuclear cells (PBMC), of cytokines was investigated in age-matched controls and in AD patients. Cytokines released by PBMC from AD patients enrolled as pre-treatment patients (T0) and as post-treatment with AchEI (T1), were detected by ELISA assay. The result showed an increase in oncostatin M, interleukin-1 beta (IL-1 beta) and interleukin-6 (IL-6) secretion in AD patients compared to healthy controls, and a decrease of cytokine levels in each AD patients treated for 1 month with an acetylcholinesterase inhibitor (AchEI). In conclusion, the results of this study show that the complex pathology in AD may be reflected in a pattern of altered cytokine secretion from PBMC.

RANTES production and expression is reduced in relapsing-remitting multiple sclerosis patients treated with interferon-β-1b

RANTES (regulated upon activation, normal T-cell expressed and secreted), a CC chemokine, appears to play a role in the pathogenesis of relapsing-remitting multiple sclerosis (RR-MS), enhancing the inflammatory response within the nervous system. We have demonstrated that RANTES production is increased in RR-MS compared to controls. Interferon-beta-1b (IFN-beta-1b) treatment reduces RANTES production in sera and peripheral blood adherent mononuclear cell (PBAM) supernatants both in relapse and remission. IFN-beta-1b also reduces RANTES expression in PBAM. Our results suggest that RANTES modulation might represent one of the mechanisms of action of IFN-beta-1b in RR-MS.

Alzheimer patients treated with an AchE inhibitor show higher IL-4 and lower IL-1 beta levels and expression in peripheral blood mononuclear cells.

The study evaluates the expression and production of cytokines in peripheral blood mononuclear cells of patients with Alzheimer disease treated or not treated with acetylcholinesterase inhibitor, which enhances neuronal transmission. Cytokines associated with brain inflammation such as interleukin (IL)-1β, IL-6, and tumor necrosis factor-α have been implicated in the regulation of amyloid peptide protein synthesis. The anti-inflammatory cytokine, IL-4, may suppress the activity of IL-1β. Patients were assessed for clinical and immunologic features at baseline and after 1 month of treatment with Donepezil, an acetylcholinesterase inhibitor. Peripheral blood mononuclear cells were cultured with and without phytohemagglutinin stimulation. IL-1β and IL-4 levels were measured by enzyme-linked immunosorbent assay. Reverse transcriptase-polymerase chain reaction was used to determine the expression of cytokines in peripheral mononuclear cells. Compared with untreated patients and healthy control subjects, IL-1β levels and expression decreased in Alzheimer disease patients treated with Donepezil (P < 0.001). In contrast, IL-4 levels and expression were significantly higher in Alzheimer patients treated with the acetylcholinesterase inhibitor. This increment was observed in both unstimulated and phytohemagglutinin-stimulated peripheral blood mononuclear cells.

Expression and production of two selected beta-chemokines in peripheral blood mononuclear cells from patients with Alzheimer's disease.

MCP-1 and RANTES are molecules that regulate monocyte and T-lymphocyte recruitment towards sites of inflammation. We sought to evaluate the role of these chemokines in Alzheimers disease (AD), and the effect of acetylcholinesterase inhibitor (AchEI) therapy on their release from peripheral blood mononuclear cells (PBMC). MCP-1 and RANTES mRNA expressions were determined by RT-PCR and the amount of secreted chemokines was assayed using specific ELISA methods from purified PBMC from each AD patients (n = 40) at the time of enrolment (T0) and after 1 month of treatment with AchEI (T1) and from 20 healthy age and sex-matched subjects (HC). We found that expression and production of MCP-1 in AD patients was significantly lower than in HC subjects. After 1 month of therapy with AchEI (Donepezil), MCP-1 levels increased in each patient. However, higher levels were detected for RANTES in AD patients compared to control subjects and in AD patients treated with Donepezil. MCP-1 and RANTES have a compensatory role in balancing the impaired mechanisms involved in immune response during ageing. Our present findings suggest that these two chemokines are both involved in AD pathogenesis and might reflect different states of activation and/or responsiveness of PBMC from AD patients, contributing to the impaired of the peripheral immune system in these patients.

The acetylcholinesterase inhibitor, Donepezil, regulates a Th2 bias in Alzheimer's disease patients

The increased pro-inflammatory cytokine production was previously observed in Alzheimers disease (AD). We sought to explore whether acetylcholinesterase inhibitor (AChEI) therapy ameliorates clinical symptoms in AD through down-regulation of inflammation. Expression and release of monocyte chemotactic protein-1 (MCP-1), a positive regulator of Th2 differentiation, and interleukin (IL)-4, an anti-inflammatory cytokine from peripheral blood mononuclear cells (PBMC) in AD patients, were investigated. PBMC were purified from AD patients at time of enrollment (T0) and after 1 month of treatment with AChEI (T1) and from healthy controls (HC). Supernatants were analyzed for cytokine levels by ELISA methods. mRNA expression were determined by RT-PCR. Expression and production of MCP-1 and IL-4 were significantly increased in AD subjects under therapy with the AChEI Donepezil, compared to the same AD patients at time of enrollment (P < 0.001). Our data suggest another possible explanation for the ability of Donepezil [diethyl(3,5-di-ter-butyl-4-hydroxybenzyl)phosphonate] to delay the progression of AD; in fact, Donepezil may modulate MCP-1 and IL-4 production, which may reflect a general shift towards type Th0/Th2 cytokines which could be protective in AD disease. The different amounts of MCP-1 and IL-4 observed might reflect the different states of activation and/or responsiveness of PBMC, that in AD patients could be kept in an activated state by pro-inflammatory cytokines.

Interferon β-1b modulates MCP-1 expression and production in relapsing–remitting multiple sclerosis

Monocyte chemoattractant protein-1 (MCP-1) seems to be involved in the pathogenesis of multiple sclerosis (MS). We found that in unstimulated (PHA ) and PHA-stimulated (PHA + ) peripheral blood mononuclear cells (PBMC), MCP-1 and TNFa levels are higher in stable untreated MS patients. Interferon gamma (IFNg) is higher in relapsing patients in PHA cultures and in stable patients in PHA + cultures. Chronic IFNb-1b treatment down-regulates TNFa, IFNg and MCP-1 production except for TNFa in relapsing patients. IFNb-1b, in vitro, increases MCP-1, TNFa and IFNg spontaneous production in all patients. Multivariate analysis suggests that MCP-1 production is dependent from clinical status and not from TNFa and IFNg production. Logistic regression analysis shows that MCP-1 production is significantly modified by treatment. Further studies are needed to clarify the role of MCP-1 in MS. D 2002 Elsevier Science B.V. All rights reserved.

Low-dose oral methotrexate treatment in chronic progressive multiple sclerosis.

Abstract. We aimed to further assess the safety and efficacy of low-dose oral methotrexate (LDOM) treatment for chronic progressive MS (CPMS). We studied 20 CPMS patients, including 16 with secondary progressive MS who had shown disease progression in the previous year. The mean follow-up was 23 months. The mean EDSS score was 6.3 ± 1.1 before treatment and 6.4 ± 1.1 after one year of treatment. At one year, 15 of 20 patients were still being treated, and 10 were stable. Twelve patients have completed 18 months of treatment, and eight are stable. Two patients stopped treatment because of side effects, two more because they did not perceive benefit, and one was lost to follow-up. Six patients had mild, transient increases in liver enzymes not requiring treatment interruption, and two had localized herpes zoster. Magnetic resonance imaging (MRI) performed before treatment and at one year remained unchanged in responders. We confirm that LDOM is safe in carefully selected and monitored CPMS patients. MTX is inexpensive and, given its anti-inflammatory and immunomodulatory properties, may be used as add-on therapy in non-responders to interferon beta, although hepatic toxicity may be a problem in long-term treatment.