Beata Kaleta

Role of osteopontin in systemic lupus erythematosus.

Systemic lupus erythematosus (SLE) is a multisystemic disease, caused by a variety of factors, which lead to immunological abnormalities. Osteopontin (OPN) is a pleiotropic protein, important in bone remodeling and immune system signaling. OPN, produced by various cells, including immune cells, plays a key role in regulating T-helper 1/T-helper 2 balance, stimulating B lymphocytes to produce antibodies, regulating macrophages, neutrophils and inducing dendritic cells. OPN expression is influenced by genetic polymorphisms of its promoter, hormones and cytokines. Over expression of OPN has been associated with the pathogenesis of immune-mediated diseases. OPN has been implicated in the development of murine model of lupus and in humans with SLE. In this review, I will present current state of research on the role of OPN and OPN gene polymorphisms in pathogenesis and clinical course of SLE. A better understanding of the role of OPN in SLE will contribute to more precise diagnosis and treatment of the disease.

Vitamin D Receptor Gene BsmI Polymorphism in Polish Patients with Systemic Lupus Erythematosus

The hormonally active form of vitamin D3, 1,25(OH)2D3 (calcitriol), exerts actions through VDR receptor, which acts as a transcriptional factor. Calcitriol is an immunomodulator that affects various immune cells, and several studies link it to many autoimmune diseases. BsmI polymorphism affects the level of VDR gene transcription, transcript stability, and posttranscriptional modifications. It seems to be related to the systemic lupus erythematosus (SLE). Our study examined the characteristics of VDR gene BsmI polymorphism in Polish SLE patients and their relationship with clinical manifestations of the disease. We genotyped 62 patients with SLE and 100 healthy controls using the real-time PCR. There were no differences observed in the frequency of BsmI genotypes in SLE patients and in the control group. There was no significant correlation between BsmI genotypes and clinical symptoms of SLE, but the AA genotype correlates with higher levels of antinuclear antibodies (ANA) in this group (r = 0.438; P = 0.002). A larger study examining BsmI and other VDR gene polymorphisms is needed. It may allow explaining differences in the clinical picture of the disease and choosing a personalized therapy.

Vitamin D receptor gene polymorphism Fok I in the Polish population does not contribute to the risk of systemic lupus erythematosus

Sir, Recent studies have suggested that the vitamin D receptor (VDR) gene polymorphism may be involved in genetic predisposition toward susceptibility to development of autoimmune diseases such as rheumatoid arthritis. Fok I polymorphism is due to transition C instead of T nucleotide in a promoter region and results in greater transcriptional activity of VDR. Enhanced expression of the VDR gene may possibly affect the expression of other genes containing such a VDR response element, and in turn it may lead to dysregulation of the Th1/Th2 balance with subsequent greater risk of development of the autoimmune process. On the other hand, the VDR gene may not be responsible for the primary association with inflammation and autoimmunity but may be in linkage disequilibrium with a nearby novel disease-related locus. In systemic lupus erythematosus (SLE) reports concerning the role of Fok I are even more sparse than in rheumatoid arthritis. In the study by Huang et al., no essential differences in the frequency of Fok I between 52 patients with SLE and with the control group were revealed, though the patients were limited to an Asian population. In accordance with the latter in a recent study by Monticielo et al., no significant association was found in genotype and allelic frequencies of Fok I polymorphism between European-derived cases and controls in Brazilian patients with SLE. We studied 62 patients with SLE and 100 healthy individuals (Table 1). Genotyping of Fok I (rs2228570) VDR polymorphism was performed by real-time polymerase chain reaction with the simple probe. The genotype frequencies in the control group were consistent with the Hardy-Weinberg equilibrium; however, in SLE they were not ( 1⁄4 0.53; p1⁄4 0.46 and 1⁄4 4.14; p1⁄4 0.04, respectively). The comparison of the frequency of Fok I VDR genotypes based on the analysis with the 2 test did not reveal any essential difference between SLE patients and the control group. Our results corroborate both aforementioned studies, as the possible risk of SLE imputed to Fok I VDR gene polymorphism assessed by logistic regression analysis did not significantly differ in the group of Caucasian patients when compared to the healthy individuals. In assessing the association between VDR Fok I polymorphism and clinical and laboratory features of SLE,Monticielo et al. found no statistical significance. It remains in accordance with our results as no difference in the risk of development of a given clinical manifestation of SLE between genotypes of Fok I VDR gene was revealed. Several other factors such as variable sample size and ethnicity may also contribute toward contradictory results, therefore further and larger studies are required. In general, single nucleotide polymorphism in a given gene may not only lead to amino acid substitution altering protein function but also may cause altered splicing, and disruption of exonic splicing enhancer sequences or exonic mRNA stability/instability sequences. The VDR gene promoter directs the transcription of at least three VDR mRNA transcripts in the kidney that seem to arise from the differential splicing of 5’-noncoding exons. Thus VDR is regulated at both transcriptional and posttranscriptional levels. Alternative splicing of the retinoid receptor, thyroid hormone receptor, and other receptors is a common feature of this gene family of proteins. Interestingly, the VDR gene is induced with retinoic acid at a site that lies downstream of exon 1, and it is likely that this site mediates the recognition ability of induction of the transcription of VDR. Therefore additional studies require both genetic analysis in multiple ethnic groups and environmental influences in order to determine the overlapping and unique associations among genetic polymorphisms. In conclusion, in the Polish population of patients Fok I polymorphism appears not to increase the risk of SLE.

Toll-like receptor 4 gene polymorphism 1196 C/T does not influence the risk of neuropsychiatric systemic lupus erythematosus in Polish population – a preliminary report

Objective Recent data indicate that Toll-like receptors (TLRs) participate in various neuropathologic conditions, including ictogenesis, myelin disruptions associated with chronic alcohol abuse, behavioral and cognitive dysfunctions associated with alcohol-induced neuroinflammatory damage, and activation of microglia to reduce amyloid β deposits. As seizures and depression are well known neuropsychiatric syndromes in systemic lupus erythematosus (SLE) the aim of the study was to investigate whether TLR4 gene polymorphism 1196C/T (rs4986791, Thr399Ile) was a candidate for susceptibility of development of neuropsychiatric systemic lupus erythematosus (NPSLE). Methods The study covered 60 patients with SLE and 100 healthy individuals. TLR4 1196C/T genotyping was performed by real-time polymerase chain reaction with the SimpleProbe. Results The SLE group comprised 86.7% of patients with wild-type homozygotes CC and 13.3% heterozygotes CT and no homozygotes TT. The control group consisted of 85% wild-type homozygotes CC, 15% heterozygotes CT and no homozygotes TT. The frequencies of genotype and allele distribution in SLE patients did not differ significantly from those of the control subjects. The probability of describing the possible risk of SLE imputed to genotype did not significantly differ in comparison with the healthy individuals (p = 0.77, odds ratio = 0.87, 95% confidence interval 0.34–2.19). A significant genotype association of genotype CC with arthritis was found in SLE patients (p = 0.02). It was further confirmed by a significant association of a dominant allele C with arthritis (p = 0.02). No association between CC and CT genotypes of TLR4 1196C/T and NPSLE was found. Allele distribution of TLR4 1196C/T also was not associated with NPSLE. No other significant differences were found in genotype and allele frequencies regarding clinical manifestation of SLE patients. Conclusion In the Polish population of SLE patients, 1196C/T polymorphism of TLR4 gene does not increase the risk of development of NPSLE; however, genotype CC and a dominant allele C is associated with arthritis in the course of SLE.

Mesenchymal Stem Cells from Human Amniotic Membrane and Umbilical Cord Can Diminish Immunological Response in an in vitro Allograft Model

Background/Aim: Mesenchymal stem cells (MSCs) are gaining rising interest in gynecology and obstetrics. MSCs immunomodulatory properties are suitable enough to reduce perinatal morbidity caused by inflammation in premature neonates. The aim of this study was to evaluate and compare the ability to inhibit allo-activated lymphocytes proliferation by MSCs derived from different sources: amniotic membrane (AM), umbilical cord (UC) and adipose tissue (AT). Methods: MSCs were isolated from AM (n = 7) and UC (n = 6) and AT (n = 6) of healthy women. Cells were characterized by flow cytometry and differentiation assay. To evaluate the potential of fetal and adult MSCs to diminish immunological response, mixed lymphocytes reaction (MLR) was performed. Results: Amnion and UC-derived cells displayed typical MSCs characteristics. Addition of MSCs to MLR significantly inhibited the proliferation of stimulated lymphocytes. The effect was observed regardless of the MSCs type used (p < 0.01 in all groups). Comparative analysis revealed no significant differences in this action between tested MSCs types. Additionally, no type of MSCs significantly stimulated allogeneic lymphocytes. Conclusion: The results prove the immunosuppressive capacities of fetal-derived MSCs in vitro. In the future, they may be potentially used to treat premature newborn as well as in immunomodulation in post-transplant therapy.

Premature fetal tissues are possible source of valuable mesenchymal stem cells

OBJECTIVES Comparison of the ability to inhibit alloactivated lymphocytes proliferation of human Wharton Jelly (WJ) and amniotic membrane (AM) mesenchymal stem cells (MSCs) from preterm and term pregnancies. MATERIAL AND METHODS Term-WJ-MSCs (n = 5) and Preterm-WJ-MSCs (n = 1) were obtained from tissue explants by adherent method. Term-AM-MSCs (n = 5) and Preterm-AM-MSCs (n = 1) were obtained by tripsin and collagenase digestion method. Term and Preterm MSCs phenotype was confirmed in vitro by flow cytometry. To evaluate the potential of fetal and adult MSCs to diminish immunological response mixed lymphocytes reaction (MLR) has been performed. RESULTS Term and Preterm cells were positively identified as MSCs by the expression of CD73 and CD90 and CD105 with simultaneous absence of CD11b, CD14, CD19, CD34, CD45 and HLA-DR. The mean inhibition of allostimulated lymphocytes after addition of fetal derived MSCs amounted 64.8% for term AM-MSCs and 42.1% for term WJ-MSCs (for both populations the effect was statistically significant, p < 0.01). The addition of preterm-MSCs to MLR resulted in reduction of stimulated lymphocytes proliferation by 64.9% for AM-MSCs and 86.1% for WJ-MSCs. CONCLUSIONS Presented results suggest that preterm fetal tissues contain MSCs which posses similar immunosuppressive capacity as those from term pregnancies. In the future MSCs from the umbilical cord and amnion can be potentially used to prevent immuno-dependent injuries in premature newborns.

Toll-Like Receptor 4 Gene Polymorphism C1196T in Polish Women with Postmenopausal Osteoporosis - Preliminary Investigation.

BACKGROUND Postmenopausal osteoporosis is a systemic bone disease characterized by low bone mass after menopause. Bone remodeling is regulated by a number of factors, including the immune system. Toll-like receptors 4 (TLR4) are expressed on bone cells and modify the immune response. TLR4 gene polymorphism may take part in the development of chronic inflammation in women after menopause, which is the cause of severe bone resorption. OBJECTIVES To examine the frequency of TLR4 C1196T genotypes in postmenopausal osteoporotic and non-osteoporotic Polish women and to investigate the possible relationship between C1196T polymorphism, bone mineral density (BMD) and the incidence of osteoporotic fractures in this group of patients. MATERIAL AND METHODS The study involved 40 postmenopausal women with osteoporosis and 63 healthy postmenopausal non-osteoporotic women. BMD measurements were performed by dual-energy X-ray absorptiometry. DNA was extracted from peripheral blood. Genotyping was performed by real-time PCR using LightSNiP tests with SimpleProbe probes. Melting curve analysis of PCR amplicons enabled the identification of individual C1196T genotypes. RESULTS C1196T genotype frequencies in the osteoporotic group were 88% for CC and 12% for CT. In the control group, respectively 86% and 14%. We did not observe the TT genotype. There was no association of C1196T genotypes and BMD nor the incidence of fractures but there was a correlation between genotypes and body height (p=0.035, r=0.415). Homozygous subjects for the C-allele had a lower body height with respect to heterozygous subjects. CONCLUSIONS It is unlikely that TLR4 C1196T polymorphism is related to bone mineral density and fracture incidence in Polish osteoporotic women after menopause. However, our data suggests that the C allele may be associated with lower body height in this group. Due to the small number of participants, our observations should be considered as preliminary. Larger studies are needed to confirm our findings.