Search

Chronic inflammatory polyneuropathies (CIP) may be associated with monoclonal gammopathies (MGUS) directed or not against neuropathy-related antigens. We analysed peripheral T and B cell subsets and cytokine network in 13 patients with CIP without MGUS (CIP), 10 with CIP with MGUS anti-MAG (CIP-MAG) and 5 with CIP with MGUS without anti-MAG activity (CIP-MGUS). We evaluated, by flow cytometry, CD3+, CD3 + DRII+, CD4+, CD8+, CD16+ and CD19+ cells subset distribution, and intracellular IL2, INFg, IL1, IL6, IL10 and TNFa production by CD4+, CD8+ and CD14+ cells. Results were compared with age-matched healthy controls (HC); statistical analysis was performed. Compared to HC, CIP patients had a significant decrease of CD3+ and B cells, CIP-MAG patients had an increase of CD3+ and NK and a decrease of CD3 + DRII+ and B cells. Comparing the different subgroups, we found a decrease of CD3 + DRII+ and B cells in CIP-MAG versus CIP, and an increase of CD4+ in CIP-MGUS versus CIP. Compared to HC, CIP showed an increase of INFg in CD4+ and of IL6 in CD14+. CIP-MAG showed an increase of IL10 in CD4+, IL1 and IL10 in CD8+, and IL1 in CD14+, versus HC and other subgroups. Our results suggest that in CIP-MAG patients it is possible to identify a distinct immunological profle. [ABSTRACT FROM AUTHOR]

Chronic sensory ataxic neuropathies (CSAN) are characterised by a predominant involvement of sensory large fibers. We previously demonstrated the presence of serum antibodies against neuropathy-related antigens in about 50% of these patients. The aim of our study was to analyse peripheral T and B cell subsets and cytokine network in CSAN patients in order to evaluate their immunological profile. We studied 10 CSAN patients. ELISA test detected the presence of neuropathy-related autoantibodies in 5 out of 10 patients. We evaluated, by flow cytometry, CD3+, CD3 + DRII+, CD4+, CD8+, CD16+ and CD19+ cells subset distribution, and intracellular IL2, INFg, IL1, IL6, IL10 and TNFa production by CD4+, CD8+ and CD14+ cells. Results were compared with healthy controls (HC); statistical analysis was performed. Compared to HC, CSAN patients showed no significant difference in lymphocyte subset distribution. We found a significant increase of IL6 in CD4+, CD8+ and CD14+ cells. INFg and TNFa were increased in CD4+ cells. TNFa was decreased in CD14+. Our results show an increase of pro-inflammatory cytokines not associated with imbalance of T and B cells subsets. These data support an activation of the immune system in CSAN, as previously suggested by the detection of neuropathy-related autoantibodies. [ABSTRACT FROM AUTHOR]

To investigate the signs of inflammatory processes in Alzheimer's disease (AD), we examined peripheral blood mononuclear cells (PBMC) from 51 AD patients (29 with mild and 22 with moderately severe dementia) and 51 age-matched healthy controls (HC), using flow cytometry to analyse the absolute number and the percentage of T, B and NK cells. We also studied the surface expression of CD25, CD28, CD57, CD71, CD45RA and CD45RO markers on cells CD4+ and CD8+. In 30 AD patients and 20 HC the production of IL-2, IFN-gamma, IL-10 and TNF-alpha by PBMC after stimulation with [25-35], [1-40] and [1-16] beta-amyloid (betaA) fragments was also evaluated. A significant decrease in circulating B and CD8+CD28- cells, as well as an increase in CD8+ cells expressing CD71+ and CD28+, was observed in AD patients. A significant decrease in IL-10 production was also found after stimulation of PMBC with betaA [1-40]. The decreased IL-10 production was not related to disease severity. The observed imbalance of immune peripheral cell subpopulations and decreased IL-10 production point to a reduction of suppressor cell function in AD patients.

Endothelin (ET)-1 is an angiogenic factor that, among others, is secreted by endothelial cells during development of several neoplasias. In particular, Kaposi sarcoma (KS) skin lesions show overexpression of the ET-1 system. Spindle cells, which characterize tumor lesions, are of endothelial origin and during disease are infected by human herpesvirus 8 (HHV-8). The majority of these cells are latently infected, suggesting that latent genes are sufficient for maintenance of viral infection and development of KS. The establishment of a reliable infection system is required to better understand the role of viral and cellular angiogenic factors involved in KS progression. For this purpose, we used human microvascular endothelial cells (HMEC-1) to establish an ET-1-producing model of infection with HHV-8. Viral particles purified from BCBL-1 cells were used to infect HMEC-1 monolayer, and infection was assessed by polymerase chain reaction, reverse transcription polymerase chain reaction, and confocal microscopy. Mitochondrial activity and cell viability, measured at 24, 48, and 72 hours after infection by 3,(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide assay, was reduced in HHV-8-infected cells compared with control. In contrast, 1 week after infection, HHV-8-positive cells showed higher mitochondrial functionality. Endothelin production was measured in culture media collected at 24, 48, and 72 hours after infection. The levels of endothelin precursor big endothelin-1 was increased 3 days after infection, although big ET-1 and ET-1 production did not differ significantly between infected and uninfected cells. These results indicate this model as a useful tool to further characterize the effects of HHV-8 in the early and late phases of infection, and to determine its ability to interfere with the endothelin system.

We investigated the apoptosis of myelin basic protein (MBP)-specific T lymphocytes in multiple sclerosis (MS) patients with acute (AMS) or stable (SMS) MS by evaluating the expression of apoptosis markers on peripheral cells. Cells of healthy controls (HC) were evaluated as well. Results showed that mitogen-stimulated apoptosis was comparable among patients and controls, whereas MBP-stimulated CD4+ and CD8+ 7-AAD+ and 7-AAD+ Fas+ cell (apoptotic cells) were significantly reduced in AMS patients. A reduction of the apoptotic rate of myelin-specific CD4+ and CD8+ T lymphocytes could be involved in the immune-mediated destruction of the myelin sheath seen in AMS patients.

Aim: Plasmodium falciparum (P. falciparum) malaria is the most important parasitic infection of humans, responsible for about 2,000,000 deaths every year. Cytoadherence of P. falciparum parasitized erythrocytes (pRBC) to vascular endothelium contributes to the pathogenesis of severe malaria causing microcirculatory obstruction and subsequent tissue hypoxia. Several cytokines and vasoactive mediators are involved in this process. The aim of this paper was to investigate the production of endothelin-1 (ET-1), a potent vasoconstrictor agent, by endothelial cells from large vessels (human umbilical vein endothelial cells, HUVEC) or the microvasculature (human microvascular endothelial cells, HMEC-1), co-cultured with different strains of P. falciparum pRBC under normoxic or hypoxic conditions., Methods: HMEC-1, immortalized by SV 40 large Tontigen, were maintained in MCDB 131 medium supplement ed with 10% fetal calf serum, 10 ng/ml of epidermal growth factor, 1 microg/ml of hydrocortisone, 2 mM glutamine, 100 U/ml of penicillin, 100 microg/ml of streptomycin and 20 mM Hepes buffer. The levels of ET-1 in the supernatants were measured by immunoenzymatic assay., Results: The results indicated that IL1-beta and hypoxia were able to induce ET-1 production by both HUVEC and HMEC-1. However, the co-incubation of HUVEC or HMEC-1 with pRBC induced a dose-dependent decrease of both constitutive and IL1- or hypoxia-induced ET-1 production. The inhibition was independent from the parasite strain used and from the origin of endothelial cells., Conclusion: These results show that pRBC by modulating both constitutive and stimulated ET-1 release from endothelial cells can induce modifications of the vascular tone in different anatomical districts. This could be of relevance in the pathogenesis of severe malaria.

The modulation of CD44, VCAM-1 and CD71 expression was analysed by flow cytometry in the 1321N1 astrocytoma cell line in the presence of interleukin-1beta (IL1beta), tumour necrosis factor-alpha (TNFalpha) and 1-40 or 25-35 beta-amyloid (Abeta) fragments. The percentage of 1321N1 astrocytoma cell line expressing these markers increased significantly after treatment with TNFalpha or IL1beta. The presence of Abeta 1-40 fragment, alone or in combination with IL1beta, induced an increase in the percentage of cells expressing CD44, but not VCAM-1. However, the concomitant presence of Abeta 1-40 fragment and of IL1beta or TNFalpha caused an increase in the percentage of CD71 positive cells. In contrast, the shorter Abeta 25-35 fragment was always inactive. These results indicates that Abeta 1-40 fragment, in association with cytokines, can activate this astrocyte-derived cell line and add further elements in favour of the hypothesis that beta-amyloid can act as immunological mediator., (Copyright 2003 Wiley-Liss, Inc.)

In this study, we investigated the production of endothelin 1 (ET-1) by a human microvascular endothelial cell line, HMEC-1, co-cultured with Plasmodium falciparum-parasitized red blood cells (pRBCs). The results indicate that hypoxia increased the basal level of ET-1 production by HMEC-1 cells after 24 or 48 h of treatment. However, the co-incubation of HMEC-1 cells with pRBCs, but not with uninfected RBCs, induced a dose-dependent decrease of both constitutive and hypoxia-induced ET-1 production. The inhibition was not due to a decrease in cell viability, as lactate dehydrogenase release remained constant. These results indicate that pRBCs are able to interfere with both the constitutive and stimulated ET-1 release from the microvascular endothelium, thus inducing local modifications of the vascular tone and of the inflammatory response. This could be of relevance in the pathogenesis of the most severe forms of P. falciparum infections, such as cerebral malaria or malaria during pregnancy.

Peripheral blood mononuclear cells of multiple sclerosis (MS) patients were stimulated with myelin basic protein (MBP) together with anti-CD28 monoclonal antibody and staphylococcal enterotoxin B to optimize cytokine production by antigen-specific cells. Type 1 (IL-2, IL-12, IFNgamma) and pro-inflammatory (TNFalpha, IL-1beta, IL-6) cytokines were augmented in CD4+, CD8+, and CD14+ cells of acute MS patients and of patients undergoing disease reactivation. These cytokines were reduced in IFNbeta-treated and in stable MS patients; type 2 cytokines (IL-4, IL-10) were increased in these patients. Similar immune profiles are seen in MS patients in whom remission is naturally or pharmacologically (IFNbeta) achieved. Cytokine alterations are particularly evident in CD14+ cells, underlying their critical role in the modulation of the immune response.

Immune parameters were analyzed in peripheral blood mononuclear cells (PBMC) and cervical mucosa biopsy specimens of human immunodeficiency virus (HIV)-seronegative women sexually exposed to HIV (exposed seronegative [ESN]), HIV-infected women, and healthy women without HIV exposure. HIV was not detected in PBMC or cervical mucosa biopsy specimens of ESN women. However, interleukin (IL)-6, IL-10, IL-12, interferon (IFN)-gamma, and tumor necrosis factor (TNF)-alpha and -beta mRNA were elevated in PBMC and cervical mucosa biopsy specimens of ESN and HIV-infected women; CCR5 and CXCR4 mRNA were augmented in cervical mucosa biopsy specimens, but not in PBMC, of ESN and HIV-infected women; HIV-specific IFN-gamma-secreting cells were detected in vaginal washes of ESN and HIV-infected women; and phenotypic alterations were present in PBMC of ESN women. These results suggest that active HIV infection is not required for T cell activation; immune alterations occur in women in whom HIV infection cannot be detected virologically or clinically.

In order to investigate the potential role of endothelins (ETs) and nitric oxide (NO) in the pathogenesis of multiple sclerosis (MS) we evaluated the levels of these vasoactive mediators in cerebrospinal fluid (CSF) of relapsing remitting MS patients and in a group of subjects with other neurological diseases (OND) and in a control group of subjects without neurological disease. Eighty patients affected from clinically diagnosed MS were selected, 44 of them were studied during an acute clinical attack and 36 in a stable phase. The OND group included 21 subjects affected by degenerative non inflammatory (n=9) and inflammatory (n=12) neurological disease while the control group included 22 subjects with cancer of the prostate (n=11) and with bladder disease (n=11). ET levels were significantly increased in CSF of relapsing remitting MS patients with an acute clinical attack in comparison with those in a stable phase, the OND group and the control group. Moreover significant differences were observed among the four groups with regard to the NO levels: MS patients in a stable and acute phase like OND group have high levels of NO compared to the control group. Since the blood-brain barrier index values did not differ significantly between the three groups, the data of this study suggest an important role for NO and ET in cerebral microcirculation in MS patients.

Human myelin basic protein (hMBP) gene is one of the candidate genes in the complex mosaic of multiple sclerosis (MS) susceptibility. In this study we verified the distribution of the polymorphism of the region 5' flanking the first exon of the hMBP gene, in 97 relapsing remitting, 74 primary progressive Italian MS patients, and in 236 healthy controls, using polymerase chain reaction (PCR) and gel electrophoresis analysis in this region from 1116 - 1540 nt. Three different band patterns were observed: one homozygote with a 354 bp long fragment, one homozygote with 424 bp long fragment and one heterozygote with both bands present. The short fragment was statistically more frequent in RRMS patients than in HC (P<0.05). The long fragment was more present in HC. Similarly the short homozygous pattern (354 bp/354 bp) was significantly higher in the RRMS patients versus the healthy controls (P<0.01). The sequence analysis of the hMBP alleles showed that while the long fragments matched the prototype sequence completely, all the short fragments showed a deletion of 70 bp from nt 1177 to nt 1247, which explains the short 354 bp allele detected by PCR. Moreover two single mismatches in positions 1386 (T-->C) and 1431 (G-->A), were present only in the short hMBP fragment.

To verify the possible effect of IFN-beta treatment on auto antibodies development in multiple sclerosis (MS) we studied 69 MS patients before and during the treatment with IFN-beta 1b (n=35) and IFN-beta 1a (n=20) for 27 and 12 months respectively, and, as controls, 14 untreated MS patients. The serum, collected every 3 months from all the patients, was investigated for the presence of antinuclear (ANA), anti-smooth muscle (ASMA), anti-mitochondrial (AMA), anti-native DNA (nDNA) anti-cardiolipin (aCL), anti-parietal cells (APCA), anti-microsomal (AMC) and anti-tireoglobulin (ATG) antibodies. Among the IFN-beta 1b-treated MS patients an increase of the frequency and of the level of ANA, AMC and ATG was observed. ASMA and ANA antibodies were already present in about 45% of the MS patients before the treatment and fluctuated over the time. In one patient the treatment was interrupted after 6 months because of the occurrence of high ASMA level and of an autoimmune hepatitis. The data obtained in the smaller number of MS patients treated with IFN-beta 1a were very similar. No increase in aCL level was observed during both the IFN treatments. Our results indicate that the treatment with IFN-beta induces an increase of AMC and ATG antibodies in MS patients and confirm that, although rare, autoimmune diseases could be observed. The possible effect of these auto antibodies on the treatment efficacy and on MS clinical course need to be further investigated.

To verify the possible role of human herpesviruses as triggering or aggravating factors in relapsing-remitting multiple sclerosis (RRMS) clinical acute attack, we studied the prevalence of some herpesviruses in the peripheral blood mononuclear cells (PBMCs) collected from 22 MS patients during an MS relapse and in a stable phase and from 18 healthy controls (HC). DNA belonging to Herpes simplex virus type 1 (HSV-1) and type 2 (HSV-2), Human cytomegalovirus (HCMV), Epstein-Barr virus (EBV) and Human Herpes virus 6 (HHV-6) has been searched by specific nested polymerase chain reaction (n-PCR). EBV and HHV6 DNA has been detected with high frequency in acute and stable MS and in healthy controls without significant differences. HCMV DNA was observed both in acute and stable MS but not in HC, and, more interestingly, HSV-1 DNA was only found in 13% of acute MS, while both stable MS and healthy controls were negative. On the basis of these results we focused on HSV-1, and to confirm them and to demonstrate that HSV-1 is actively replicating in MS patients during clinical relapse, we searched both messenger RNA (mRNA) and DNA of HSV-1 in the PBMCs of 15 acute MS patients and 15 healthy controls. We found HSV-1 mRNA and DNA in a significant number of acute MS patients but not in the control group. On the whole these data indicate that HSV-1 reactivate in the peripheral blood of MS patients during clinical acute attack and probably play a role in the triggering of MS relapses.

The binding of fibronectin and fibronectin fragments to the enterotoxigenic strain E. coli B34289c was studied. E. coli cells bound to two distinct sites of fibronectin, one being the N-terminal domain, which also contains the binding sites for staphylococci and streptococci, and the other located within the central heparin binding region. In addition, the N-terminal and the heparin binding domain mediated the attachment of bacteria in a solid phase binding assay. E. coli cells expressed two classes of receptors, the first, a 17 kDa protein, recognized by the N-terminal fragment and the second, having a mol. mass of 55 kDa, which interacts with the internal heparin binding domain. Bacterial receptors, which bind the N-terminal end of fibronectin, may be structurally related.