Marija Mostarica Stojković

Novel platinum(IV) complexes induce rapid tumor cell death in vitro

The anticancer activity of platinum complexes has been known since the discovery of classical Pt(II)‐based drug cisplatin. However, Pt(IV) complexes have greater inertness than corresponding Pt(II) complexes, thus allowing the oral administration and reducing the toxicity associated with platinum‐based chemotherapy. Here, we describe the in vitro antitumor activity of some novel Pt(IV)‐based agents against mouse fibrosarcoma L929 cells and human astrocytoma U251 cells. The cytotoxicity of 2 Pt(IV) complexes with bidentate ethylenediamine‐N,N′‐di‐3‐propanoato esters was found to be markedly higher than that of their Pt(II) counterparts and comparable to the antitumor action of cisplatin. In contrast to cisplatin, which caused oxidative stress‐independent apoptotic cell death of tumor cells, these Pt(IV) complexes induced oxygen radical‐mediated tumor cell necrosis. Importantly, the cytotoxic action of novel Pt(IV) complexes was markedly more rapid than that of cisplatin, indicating their potential usefulness in anticancer therapy.

Uric acid levels in sera from patients with multiple sclerosis

Abstract The levels of uric acid (UA), a natural peroxynitrite scavenger, were measured in sera from 240 patients with multiple sclerosis (MS) and 104 sex- and age-matched control patients with other neurological diseases (OND). The mean serum UA concentration was lower in the MS than in the OND group, but the difference did not reach the level of statistical significance (P=0.068). However, the mean serum UA level from patients with active MS (202.6+67.1 μmol/l) was significantly lower than that in inactive MS patients (226.5+78.6 μmol/l; P=0.046) and OND controls (P=0.007). We found a significant inverse correlation of serum UA concentration with female gender (P=0.0001), disease activity (P=0.012) and duration (P=0.017), and a trend towards an inverse correlation with disability as assessed by EDSS score, which did not reach statistical significance (P=0.067). Finally, multivariate linear regression analyses showed that UA concentration was independently correlated with gender (P=0.0001), disease activity (P=0.014) and duration of the disease (P=0.043) in MS patients. These findings suggest that serum UA might serve as a possible marker of disease activity in MS. They also provide support to the potential beneficial therapeutic effect of radical-scavenging substances in MS.

Interleukin-17 stimulates inducible nitric oxide synthase activation in rodent astrocytes

The effect of interleukin-17 (IL-17) on production of nitric oxide (NO) in rodent astrocytes was investigated. While IL-17 by itself did not induce NO production, it caused a dose-dependent enhancement of IFN-gamma-triggered NO synthesis in both mouse and rat primary astrocytes. In contrast, IL-17 was unable to stimulate NO synthesis in either murine or rat macrophages. IFN-gamma-triggered expression of mRNA for iNOS, but not for its transcription factor interferon regulatory factor-1 (IRF-1), was markedly elevated in IL-17-treated astrocytes. The induction of iNOS mRNA by IL-17 in IFN-gamma-pretreated astrocytes was abolished by antagonists of nuclear factor-kappaB (NF-kappaB) activation--a proteasome inhibitor MG132 and an antioxidant agent PDTC, as well as with specific p38 MAP kinase inhibitor SB203580. While IL-17 stimulated both IL-1beta and IL-6 production in astrocytes, only IL-1 was partly responsible for IL-17-induced NO release. Finally, IL-17 synergized with exogenous IL-1beta and TNF-alpha for astrocyte NO production. Having in mind a well-known neurotoxic action of NO, these results suggest a possible role for IL-17 in the inflammatory diseases of the CNS.

Astrocyte-induced regulatory T cells mitigate CNS autoimmunity.

Although astrocytes presumably participate in maintaining the immune privilege of the central nervous system (CNS), the mechanisms behind their immunoregulatory properties are still largely undefined. In this study, we describe the development of regulatory T cells upon contact with astrocytes. Rat T cells pre‐incubated with astrocytes completely lost the ability to proliferate in response to mitogenic stimuli. The cells were blocked in G0/G1 phase of the cell cycle, expressed less IL‐2R, and produced significantly lower amounts of interferon‐γ (IFN‐γ), but not interleukin‐2 (IL‐2), IL‐10, or tumor necrosis factor (TNF). These anergic cells completely prevented mitogen‐induced growth of normal T lymphocytes, as well as CNS antigen‐driven proliferation of autoreactive T cells. The suppressive activity resided in both CD4+ and CD8+ T‐cell compartments. Heat‐sensitive soluble T‐cell factors, not including transforming growth factor‐β (TGF‐β) or IL‐10, were solely responsible for the observed suppression, as well as for the transfer of suppressive activity to normal T cells. The administration of astrocyte‐induced regulatory T cells markedly alleviated CNS inflammation and clinical symptoms of CNS autoimmunity in rats with experimental allergic encephalomyelitis. Finally, the cells with suppressive properties were readily generated from human lymphocytes after contact with astrocytes. Taken together, these data indicate that astrocyte‐induced regulatory T cells might represent an important mechanism for self‐limitation of excessive inflammation in the brain.

Astrocytes in the tempest of multiple sclerosis

Astrocytes are the most abundant cell population within the CNS of mammals. Their glial role is perfectly performed in the healthy CNS as they support functions of neurons. The omnipresence of astrocytes throughout the white and grey matter and their intimate relation with blood vessels of the CNS, as well as numerous immunity‐related actions that these cells are capable of, imply that astrocytes should have a prominent role in neuroinflammatory disorders, such as multiple sclerosis (MS). The role of astrocytes in MS is rather ambiguous, as they have the capacity to both stimulate and restrain neuroinflammation and tissue destruction. In this paper we present some of the proved and the proposed functions of astrocytes in neuroinflammation and discuss the effect of MS therapeutics on astrocytes.

Expression of Th1 and Th17 cytokines and transcription factors in multiple sclerosis patients: does baseline T-bet mRNA predict the response to interferon-beta treatment?

We studied the effect of one-year interferon (IFN)-beta treatment on the in vivo mRNA expression of IFN-gamma, interleukin (IL)-17, T-bet and RoR-gammat, on peripheral blood mononuclear cells (PBMC) from 36 multiple sclerosis (MS) patients. In the total MS group, IFN-beta induced decrease in mRNA levels of IFN-gamma and T-bet (p<0.0001), while the levels of IL-17 and RoR-gammat remained similar. In both responders and non-responders, IFN-beta induced significant decrease of IFN-gamma (p<0.0001 and p=0.011, respectively), while decrease in T-bet was detected only in responders (p<0.0001). Higher pre-treatment T-bet allowed prediction of the clinical response in the first year (beta=0.601, p=0.036). Our preliminary findings suggest that T-bet expression might be a potential prognostic marker of treatment response to IFN-beta in MS.

Antibodies against myelin oligodendrocyte glycoprotein in the cerebrospinal fluid of multiple sclerosis patients

Antibodies against myelin oligodendrocyte glycoprotein (MOG) mediate demyelination in experimental autoimmune encephalomyelitis (EAE) in different animal species and are implicated in the immunopathogenesis of multiple sclerosis (MS). In order to evaluate the anti-MOG response, we have analyzed the cerebrospinal fluids (CSFs) from 44 MS patients and 51 controls, 11 with other inflammatory neurological disorders (OIND) and 40 with non-inflammatory neurological disorders (NIND). The frequency of anti-MOG antibodies positive patients in the MS group (30%) was significantly higher compared to the NIND (8%, p=0.02), but not compared to the OIND group (55%, p=0.228). Interestingly, all six patients with neurosarcoidosis had MOG-specific antibodies in their CSF. Frequency of anti-MOG antibodies was similar in patients with clinically active and stable MS (32% and 26%, respectively; p=0.921). However, in clinically active MS patients, antibody titers were higher in comparison with patients with stable disease, although the difference did not reach the level of statistical significance (p=0.06). These results further support the potential role of anti-MOG antibodies in the immunopathology of MS in the subset of patients with this disease. Furthermore, our findings suggest for the first time that anti-MOG antibodies could be an accessory diagnostic tool in neurosarcoidosis.

Decreased frequency of the tumor necrosis factor α-308 allele in Serbian patients with multiple sclerosis

Tumor necrosis factor (TNF) α has been considered the prototypic cytopathogenic cytokine in multiple sclerosis (MS), but recently this cytokine has been shown to possess significant anti-inflammatory and neuroprotective effects in demyelinating diseases. It has been reported that the TNFα –308 polymorphism influences levels of TNFα production, and that the rare allele, TNF2, is associated with high TNFα production. We investigated the TNFα –308 polymorphism in 143 unrelated Serbian patients with MS and 123 ethnically matched, healthy individuals using the allele-specific restriction fragment length polymorphism polymerase chain reaction technique. The frequency of the TNF2 allele was significantly decreased in MS patients (14%) in comparison with controls (24%; p = 0.044). The TNF2 allele had no influence on disease behavior, since it was not associated with the course and severity of MS in this group of patients. The result suggests that in the Serbian population polymorphism at position –308 of TNFα or at an adjacent locus might have a role in MS susceptibility.

Strain difference in susceptibility to experimental autoimmune encephalomyelitis in rats correlates with TH1 and TH17-inducing cytokine profiles

Albino Oxford (AO) rats are resistant to induction of experimental autoimmune encephalomyelitis (EAE), in contrast to susceptible Dark Agouti (DA) rats. We have previously shown that draining lymph node cells (DLNC) obtained from immunized DA rats before the onset of the clinical disease produced more interferon (IFN)-gamma and interleukin (IL)-17 (signature cytokines of T(H)1 and T(H)17 responses, respectively) compared to DLNC from AO rats. In this study, we extend our analysis to entire induction phase of EAE with the emphasis on the T(H)1 and T(H)17-inducing cytokines. As a result, we show that throughout the inductive phase of the disease DLNC of DA rats, not only expressed higher levels of IFN-gamma and IL-17, but also of T(H)1-inducing cytokine-IL-12. As for T(H)17-inducing cytokines, DLNC of DA rats expressed more mRNA for p19, specific subunit of IL-23, but the expression of transforming growth factor (TGF)-beta in both strains was similar. Interestingly, the analysis of IL-6 expression revealed striking difference: while all DA DLNC were positive for IL-6 mRNA, cells from none of AO rats expressed detectable levels of mRNA for this cytokine. Taken together, our data suggest that the differential regulation of production of T(H)1 and T(H)17 cytokines, and IL-6 in particular, during the induction phase of disease could be responsible for the discrepancy in susceptibility to EAE between these two rat strains.

Cerebrospinal fluid and serum uric acid levels in patients with multiple sclerosis

Abstract Background: Peroxynitrite was hypothesized to be involved in the pathogenesis of multiple sclerosis (MS) through its various neurotoxic effects. Uric acid (UA) was shown to be a strong peroxynitrite scavenger. Methods: We analyzed cerebrospinal fluid (CSF) and serum UA concentrations in 30 MS patients and 20 controls with non-inflammatory neurological diseases (NIND) and correlated these findings with demographic and clinical characteristics of MS patients. Disease activity was assessed by brain magnetic resonance imaging (MRI) and the CSF/serum albumin quotient as an indicator of the state of blood-brain-barrier (BBB). Results: Serum UA concentrations were found to be significantly lower in MS patients compared with controls (p=0.019). CSF UA concentrations were lower in MS patients as compared to controls, as well as in patients with active MS (clinical and/or MRI activity) in comparison to patients with inactive MS or controls, but these differences were not statistically significant. Significant correlation was found between CSF and serum UA concentrations (p=0.016) in MS patients, but not in controls; and between CSF UA concentrations and the CSF/serum albumin quotient in MS patients (p=0.043), but not in controls. Conclusions: Our results support the significance of UA in the pathogenesis of MS. Decreased serum UA concentrations in MS patients might be due to both intrinsically reduced antioxidant capacity and increased UA consumption in MS. CSF UA concentrations may not be a reliable marker of disease activity in MS since its concentration is dependent on leakage of UA molecules from serum through the damaged BBB and the balance between consumption/production within the central nervous system (CNS). Clin Chem Lab Med 2009;47:848–53.