Anti-HERV-WEnv antibodies are correlated with seroreactivity against Mycobacterium avium subsp. paratuberculosis in children and youths at T1D risk.

Recent evidence points at the role that human endogenous retroviruses (HERVs) may play through the activation of genes integrated across the human genome. Although a variety of genetic/epigenetic mechanisms maintain most HERVs silenced, independent environmental stimuli including infections may transactivate endogenous elements favoring pathogenic conditions. Several studies associated exposures to Mycobacterium avium subsp. paratuberculosis (MAP) with increased anti-MAP seroreactivity in T1D patients. Here, we assessed humoral responses against HERV envelope antigens (HERV-KEnv and HERV-WEnv) and four MAP-derived peptides with human homologs in distinct populations: Sardinian children at T1D risk (rT1D) (n = 14), rT1D from mainland Italy (n = 54) and Polish youths with T1D (n = 74) or obesity unrelated to autoimmunity (OB) (n = 26). Unlike Sardinian rT1D, youths displayed increased anti-HERV-WEnv Abs prevalence compared to age-matched OB or healthy controls (24.32 vs. 11.54%, p = 0.02 for Polish T1D/OB and 31.48 vs. 11.90%, p = 0.0025 for Italian rT1D). Anti-HERV-KEnv responses showed variable trends across groups. A strong correlation between Abs levels against HERV-WEnv and homologous peptides was mirrored by time-related Abs patterns. Elevated values registered for HERV-WEnv overlaped with or preceded the detection of T1D diagnostic autoantibodies. These results support the hypothesis of MAP infection leading to HERV-W antigen expression and enhancing the production of autoantibodies in T1D.

Gastric cancer increases transmigratory potential of peripheral blood monocytes by upregulation of ß1- and ß2-integrins.

INTRODUCTION: Immune responses within the tumor depend on the ability of leukocytes to migrate from peripheral circulation into the local microenvironment. This process is controlled by mechanisms that guide leukocytes to the side of inflammation, allowing them to cross vascular endothelial barrier. Monocytes/macrophages are the predominant population of leukocyte infiltrate of many tumors, including, gastric cancer. However, to date mechanisms that control monocyte trafficking to the side of tumor growth are not fully elucidated. AIM OF THE STUDY: It this study we aimed to evaluate transmigratory potential of peripheral blood monocytes from gastric cancer patients. MATERIAL AND METHODS: By using multicolor flow cytometry we assessed expression of ß1- and ß2-integrins on peripheral blood monocytes from gastric cancer patients. RESULTS: We found increased frequencies of VLA-4 and VLA-6 expressing monocytes and increased expression of analyzed ß2-integrins in gastric cancer patients when compared to age matched controls. CONCLUSIONS: In summary, this study revealed that gastric cancer increases transmigratory potential of peripheral blood monocytes.

Does the Mesenchymal Stem Cell Source Influence Smooth Muscle Regeneration in Tissue-Engineered Urinary Bladders?

A variety of tissue engineering techniques utilizing different cells and biomaterials are currently being explored to construct urinary bladder walls de novo, but so far no approach is clearly superior. The aim of this study was to determine whether mesenchymal stem cells (MSCs) isolated from different sources, (bone marrow [BM-MSCs] and adipose tissue [ADSCs]), differ in their potential to regenerate smooth muscles in tissue-engineered urinary bladders and to determine an optimal number of MSCs for urinary bladder smooth muscle regeneration. Forty-eight rats underwent hemicystectomy and bladder augmentation with approximately 0.8 cm2 graft. In the first and second groups, urinary bladders were reconstructed with small intestinal submucosa (SIS) seeded with 10 × 106 or 4 × 106 ADSCs/cm2, respectively. In the third and fourth groups, urinary bladders were augmented with SIS seeded with 10 × 106 or 4 × 106 BM-MSCs/cm2, respectively. In the fifth group, urinary bladders were augmented with SIS without cells. The sixth group (control) was left intact. Smooth muscle regeneration was evaluated by real-time polymerase chain reaction (RT-PCR) and histological examinations. Histologically, there were no significant differences between urinary bladders augmented with ADSCs and BM-MSCs, but there was a marked increase in smooth muscle formation in bladders augmented with grafts seeded with MSCs in higher density (10 × 106/cm2) compared to lower density (4 × 106/cm2). Molecular analysis revealed that bladders reconstructed with ADSC-seeded grafts expressed higher levels of smooth muscle myosin heavy chain, caldesmon, and vinculin. Bladders augmented with unseeded SIS were fibrotic and devoid of smooth muscles. ADSCs and BM-MSCs have comparable smooth muscle regenerative potential, but the number of MSCs used for graft preparation significantly affects the smooth muscle content in tissue-engineered urinary bladders.

Melanoma cell adhesion molecule as an emerging biomarker with prognostic significance in systolic heart failure.

BACKGROUND: Melanoma cell adhesion molecule (MCAM) is a marker of endothelial damage. MCAM diagnostic and prognostic value was assessed in chronic heart failure (CHF). MATERIALS & METHODS: 130 CHF patients and 32 controls were included in the study. Telephone follow-up lasted one year. End points were: death from all causes, and hospitalization with CHF exacerbation. RESULTS: MCAM was higher in patients than in controls (p = 0.01). Receiver operator curve analysis revealed that MCAM may serve as a predictor of death (area under the curve: 0.8404; p < 0.002). Patients with MCAM above 500 ng/ml had worse prognosis (p = 0.03). NT-proBNP and age were independent predictors of death in multivariate analysis. CONCLUSION: The increased MCAM indicates endothelial damage in CHF and may serve as a marker of worse prognosis in these patients.

Phenotype of NK Cells Determined on the Basis of Selected Immunological Parameters in Children Treated due to Acute Lymphoblastic Leukemia.

Acute lymphoblastic leukemia (ALL) is the most frequent pediatric malignancy. The chemotherapy for ALL is associated with a profound secondary immune deficiency.We evaluated the number and phenotype of natural killer (NK) cells at diagnosis, after the intensive chemotherapy and following the completion of the entire treatment for patients with ALL. The fraction, absolute number, and percentage of NK cells expressing interferon-γ were determined in full blood samples. The fraction of NK cells expressing CD158a, CD158b, perforin, A, B, and K granzymes was examined in isolated NK cells.We have shown that patients assessed at ALL diagnosis showed significantly lower values of the fraction of NK cells and percentage of NK cells with the granzyme A expression. Additionally, the absolute number of NK cells, the expression of CD158a, CD158b, perforin, and granzyme A were significantly lower in patients who completed intensive chemotherapy. Also, there was a significantly higher fraction of NK cells expressing granzyme K in patients who completed the therapy.Abnormalities of NK cells were found at all stages of the treatment; however, the most pronounced changes were found at the end of intensive chemotherapy.

Innate immunity components and cytokines in gastric mucosa in children with Helicobacter pylori infection.

PURPOSE. To investigate the expression of innate immunity components and cytokines in the gastric mucosa among H. pylori infected and uninfected children. Materials and Methods. Biopsies of the antral gastric mucosa from children with dyspeptic symptoms were evaluated. Gene expressions of innate immunity receptors and cytokines were measured by quantitative real-time PCR. The protein expression of selected molecules was tested by immunohistochemistry. RESULTS. H. pylori infection did not lead to a significant upregulation of MyD88, TLR2, TLR4, CD14, TREM1, and TREM2 mRNA expression but instead resulted in high mRNA expression of IL-6, IL-10, IFN-γ, TNF-α, and CD163. H. pylori cagA(+) infection was associated with higher IL-6 and IL-10 mRNA expression, as compared to cagA(-) strains. H. pylori infected children showed increased IFN-γ and TNF-α protein levels. IFN-γ mRNA expression correlated with both H. pylori density of colonization and lymphocytic infiltration in the gastric mucosa, whereas TNF-α protein expression correlated with bacterial density. CONCLUSION. H. pylori infection in children was characterized by (a) Th1 expression profile, (b) lack of mRNA overexpression of natural immunity receptors, and (c) strong anti-inflammatory activities in the gastric mucosa, possibly resulting from increased activity of anti-inflammatory M2 macrophages. This may explain the mildly inflammatory gastric inflammation often observed among H. pylori infected children.

Transdifferentiation of Bone Marrow Mesenchymal Stem Cells into the Islet-Like Cells: the Role of Extracellular Matrix Proteins.

Pancreatic islet implantation has been recently shown to be an efficient method of treatment for type 1 diabetes. However, limited availability of donor islets reduces its use. Bone morrow would provide potentially unlimited source of stem cells for generation of insulin-producing cells. This study was performed to evaluate the influence of extracellular matrix proteins like collagen, laminin, and vitronectin on bone marrow mesenchymal stem cells (BM-MSCs) transdifferentiation into islet-like cells (ILCs) in vitro. To our knowledge, this is the first report evaluating the importance of vitronectin in transdifferentiation of BM-MSCs into ILCs. Rat BM-MSCs were induced to ILCs using four-step protocol on plates coated with collagen type IV, laminin type I and vitronectin type I. Quantitative real-time PCR was performed to detect gene expression related to pancreatic ß cell development. The induced cells expressed islet-related genes including: neurogenin 3, neurogenic differentiation 1, paired box 4, NK homeobox factor 6.1, glucagon, insulin 1 and insulin 2. Laminin but not collagen type IV or vitronectin enhanced expression of insulin and promoted formation of islet-like structures in monolayer culture. Laminin triggered transdifferentiation of BM-MSCs into ILCs.

Collaborating with the enemy: function of macrophages in the development of neoplastic disease.

Due to the profile of released mediators (such as cytokines, chemokines, growth factors, etc.), neoplastic cells modulate the activity of immune system, directly affecting its components both locally and peripherally. This is reflected by the limited antineoplastic activity of the immune system (immunosuppressive effect), induction of tolerance to neoplastic antigens, and the promotion of processes associated with the proliferation of neoplastic tissue. Most of these responses are macrophages dependent, since these cells show proangiogenic properties, attenuate the adaptive response (anergization of naïve T lymphocytes, induction of Treg cell formation, polarization of immune response towards Th2, etc.), and support invasion and metastases formation. Tumor-associated macrophages (TAMs), a predominant component of leukocytic infiltrate, "cooperate" with the neoplastic tissue, leading to the intensified proliferation and the immune escape of the latter. This paper characterizes the function of macrophages in the development of neoplastic disease.

Hyperthermia induces cytoskeletal alterations and mitotic catastrophe in p53-deficient H1299 lung cancer cells.

Hyperthermia is used in cancer therapy, however much remains to be discovered regarding its mechanisms of action at the cellular level. In this study, the effects of hyperthermia on cell death, survival, morphology and the cytoskeleton were investigated in a non-small cell lung cancer cell line, H1299. Despite the fact that this cell line is widely used in research, it has not yet been tested for heat shock sensitivity. Cells were given a 30-min heat shock at 43.5°C and 45°C and left to recover at 37°C for 24 and 48 h. 24 h after heat shock treatment, we monitored changes in the organization of the cytoskeleton using immunofluorescence microscopy. The number of actin stress fibers was significantly reduced, microtubules formed a looser meshwork, a portion of the cells possessed multipolar mitotic spindles, whereas vimentin filaments collapsed into perinuclear complexes. 48 h following heat stress, most of the cells showed recovery of the cytoskeleton, however we observed a considerable number of giant cells that were multinucleated or contained one enlarged nucleus. The data obtained by MTT assay showed a dose-dependent decrease of cell viability, while flow cytometric analysis revealed an increase in the number of cells with externalized phosphatidylserine. The results suggest that one of the modes of heat-induced cell death in H1299 cells is mitotic catastrophe, which probably ends in apoptosis.

Peripheral blood T lymphocyte subsets in children with congenital asplenia.

The aim of the current study was to examine whether a congenital lack of the spleen changes distribution, state of activation and function of peripheral lymphocyte T subsets. Seven children with congenital asplenia (CA) aged 1.5-17 years and seven age-matched controls were tested. By triple-color flow cytometry we examined: (1) the expression of CD3(+), CD4(+), CD8(+), CD19(+), and CD56(+) on lymphocytes; (2) the distribution of CD45RA(+) and CD45RO(+) in CD4(+) and CD8(+); (3) the expression of CD27(+) in the CD4(+) and CD8(+) T-cell-bearing CD45RA(+), CD45RO(+), or CD45RB(+). Lymphocyte proliferative responses and cytokines production (IFN-gamma, IL-6, TNF-alfa, and IL-10) in anti-CD3-induced peripheral blood mononuclear cells were tested. The results indicate (1) a normal distribution of the basic lymphocyte subsets, (2) low CD3(+)/CD8(+) percentage but expressing CD8(+high) and non-significantly elevated CD4(+)/CD8(+) ratio, (3) CD45RA(+high) and CD27(+high) in the CD4(+) and CD8(+) T cell, and (4) CD45RB(+high) in the CD4(+) and CD45RO(+high) in the CD8(+). The distribution of CD27(+) in the CD45RA(+) and CD45RO(+) CD4(+) T cells remained unchanged. However, the percentage of CD8(+)/CD45RO(+)/CD27(+) T cells tended to be elevated. Altogether, these data indicate that CA is connected with (1) the presence CD4(+) T cells expressing the "naive" phenotype (CD45RA(+high) RB(+high) and CD27(+high)), (2) high numbers of activated CD8(+) T cells shifted toward the memory phenotype (CD45RO(+high)) but still showing high CD27(+) expression, which may indicate failure in T CD8(+) cytotoxic effectors differentiation, and (3) a tendency to the rather pro-inflammatory status of cells, low IL-10 expression, and suboptimal lymphocytes responses to mitogenic stimulation.

Generation and identification of thymic epithelial progenitor cells pTEC by in-vitro processing of human thymic fragments for allotransplantation.

The procedure of generation and identification of stromal progenitor cells derived from human thymic fragments (PL patent 378431) has been described in this article. Our aim was to prepare material for transplantation in elderly people. The method is based on in-vitro processing of thymic fragments to get rid of all immunogenic elements of lymphocytes, endothelial cells, macrophages, and fibroblasts. In the thymic culture process, this organ dies out in the incubation medium and epithelial cells emerge out of the organ. After about 4 weeks from the start of the culture, the population of various developmental forms of epithelial cells was generated, namely CK AE1/AE3+, SDF-1 alpha+ and a weak expression of FGF+ S-100+. Finally, we obtained approximately 3 million cells as a monolayer. The progenitor cells were experimentally transplanted into a 72-year-old volunteer in order to prove that they do not induce neither a local nor a systemic rejection response.

Zinc and iron concentration and SOD activity in human semen and seminal plasma.

The aim of the present study was to measure zinc (Zn) and iron (Fe) concentration in human semen and superoxide dismutase (SOD) activity in seminal plasma and correlate the results with sperm quality. Semen samples were obtained from men (N = 168) undergoing routine infertility evaluation. The study design included two groups based on the ejaculate parameters. Group I (n = 39) consisted of males with normal ejaculate (normozoospermia), and group II (n = 129) consisted of males with pathological spermiogram. Seminal Zn and Fe were measured in 162 samples (group I, n = 38; group II, n = 124) and SOD activity in 149 samples (group I, n = 37; group II, n = 112). Correlations were found between SOD activity and Fe and Zn concentration, and between Fe and Zn concentration. SOD activity was negatively associated with volume of semen and positively associated with rapid progressive motility, nonprogressive motility, and concentration. Negative correlation was stated between Fe concentration and normal morphology. Mean SOD activity in seminal plasma of semen from men of group I was higher than in seminal plasma of semen from men of group II. Fe concentration was higher in teratozoospermic males than in males with normal morphology of spermatozoa in group II. Our results suggest that Fe may influence spermatozoa morphology.

Inflammatory activation in children with primary hypertension.

Low-grade inflammation plays a role in the pathogenesis of primary hypertension (PH) and target organ damage (TOD). We evaluated the profile of inflammatory mediators (CRP, RANTES, MIP-1beta, MIP-1alpha, MCP-1, IL-6, angiogenin, adiponectin) in 30 healthy children (12.7 +/- 3.3 years) and 44 patients with untreated PH (13.7 +/- 2.7 years; n.s). Patients had greater concentrations of CRP, MIP-1beta, and RANTES than controls (all p < 0.05). Children with metabolic syndrome (MS) had greater CRP than children without MS (p = 0.007) and CRP correlated with number of MS criteria, body mass index (BMI), visceral fat, deep subcutaneous fat assessed by magnetic resonance imaging, carotid intima-media thickness (cIMT), left ventricular mass index, and markers of oxidative stress. RANTES correlated with cholesterol, LDL cholesterol, ApoB, and ApoB/ApoA1. Angiogenin correlated with BMI, waist circumference, visceral fat, uric acid, and patients with cIMT>2SD had greater concentration of angiogenin than those with normal cIMT (p = 0.03). Adiponectin was lower in patients with cIMT>2SD than in those with normal cIMT (p = 0.02). No model explaining variability of TOD has been built. Elevated RANTES and MIP-1beta and normal IL-6 and TNF-alpha levels indicate a vascular inflammatory process. Lack of correlation between CRP and chemokines suggests that vascular inflammation in PH precedes the systemic inflammatory changes.

Immunologic properties differ in preterm infants fed olive oil vs soy-based lipid emulsions during parenteral nutrition.

BACKGROUND: In the first period of life, premature infants need parenteral nutrition. Lipid emulsions (LEs), which are a part of parenteral nutrition, are known as potent immunological modulators and may therefore influence the immune status of parenterally fed infants. The aim of the study was to compare tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and IL-10 production in the peripheral blood mononuclear cells (PBMCs) of premature infants parenterally fed with 2 LEs: olive oil (OO) and soybean oil (SO). METHODS: Premature infants born at <32 weeks' gestation and with a birth weight <1500 g were randomized in a double-blind method within the first 48 hours of life to receive 1 of 2 LEs: OO based or SO based. At baseline and after 14 days, blood samples were collected, and PBMCs were isolated and then cultured for 48 hours in medium only and in the presence of anti-CD3 antibodies. RESULTS: Of 44 recruited infants, 38 completed the study, 18 in the OO group and 20 in the SO group. The cytokine synthesis profile before the LE introduction was the same in both groups (nonstimulated and anti-CD3-induced PBMC). In the succeeding 14 days of parenteral nutrition, TNF-alpha, IL-6, and IL-10 levels in nonstimulated PBMCs remained unchanged in both groups. In contrast, IL-6 production was significantly higher in the SO group. CONCLUSIONS: SO-based LE may promote an excess of IL-6 production, especially in the T cell-dependent way of PBMC activation (via anti-CD3). OO emulsion seems to be immunologically more neutral than SO emulsion.

Enhanced gastric IL-18 mRNA expression in Helicobacter pylori-infected children is associated with macrophage infiltration, IL-8, and IL-1 beta mRNA expression.

OBJECTIVE: To investigate IL-18 mRNA expression in the gastric mucosa in Helicobacter pylori-infected children and its association with macrophage infiltration, IL-8, and IL-1 beta mRNA expression. METHODS: From 39 children, blood samples were taken for IL-1 beta gene polymorphism analysis and antral biopsies were obtained for histology (including macrophage immunostaining), culture and semiquantitative analysis of IL-18, IL-8, IL-1 beta, and CD14 mRNA expression by reverse transcription-PCR (RT-PCR). RT-PCR was used for H. pylori ureA and cagA mRNA detection in gastric tissue. RESULTS: H. pylori-infected patients had significantly higher IL-18, IL-8, and IL-1 beta transcript levels and macrophage numbers in the antral mucosa than H. pylori-negative children. IL-1 beta-511/31 gene polymorphism had no impact on gastric IL-1 beta mRNA levels. IL-18 mRNA expression correlated with mRNA expression of IL-8 and IL-1 beta, and transcript levels of all three cytokines were associated with macrophage infiltration and CD14 mRNA expression in the gastric tissue. Significant correlation was also observed between macrophage numbers and histological parameters of gastritis. CONCLUSION: These results suggest that interleukin(IL)-18 and macrophages may have an important function in gastric inflammatory response to H. pylori infection in children. IL-18, and possibly CD14 receptor signalling pathway, may be involved in macrophage activation and subsequent IL-8 and IL-1 beta release.

[The influence of smoking of the cigarettes on the level of circulating immunological complexes in the serum of patients with the hypersensitivity of the type I]. / Wplyw palenia papierosów na poziom krazacych kompleksów immunologicznych w surowicy chorych z nadwrazliwoscia typu I.

Circulating immunological complexes are the answer of the immunological system on appearing in the organism of the substance about the character of antigenic. The growth of the concentration of immunological complexes is described in many diseases among others in allergies. The aim of the work was the opinion of levels of circulating immunological complexes serums smoke patients and no smokes with the hypersensitivity of I type. Patients serums given an examination with the hypersensitivity of the type I (n = 22) smoking (n = 11) and nonsmoking (n = 11). 45 persons been careful for healthy made up the supervisory group. The level of circulating immunological complexes was studied the simple method of PEG precipitation according to the Haskov method. Smoking patients step out higher levels of CIC in the hypersensitivity of the type I than at patients who don't smoke.

The presence and structure of circulating immune complexes in patients with prostate tumors.

BACKGROUND: The detection, measurement and characterization of circulating immune complexes (CICs) in tumor patients has shown both good and poor correlation with progression of the disease. The origin of the antigens making up the immune complexes in the sera of the cancer patients examined is unknown. They are perhaps antigens specific to the cancer's progress. MATERIAL/METHODS: The levels and molecular weights of circulating immune complexes were estimated in the sera of 48 patients with adenocarcinoma (grades G-1, G-2, and G-3) and 21 patients with benign prostate hyperplasia (BPH) by means of the polyethylene glycol (PEG) precipitation test. RESULTS: The results were compared with those of a group of 45 healthy blood donors. Elevated levels of CICs were observed in 66.7% of the patients with adenocarcinoma and in 38% of those with BPH. No seropositivity for CICs was observed in the control group. Increased seropositivity for CICs was observed in patients with the highness grade of adenocarcinoma: the proportion of seropositive patients in the group of patients with the G-1 phase of disease was 50%, with G-2 75%, and with G-3 71.3%. The molecular weights of the CIC proteins were determined by SDS/PAGE. The serum CICs of both BPH and prostate cancer patients consisted of proteins absent in the CICs of the sera of healthy persons. CONCLUSIONS: Determination and analysis of atypical proteins in CICs resulting from carcinogenesis may by useful in improving the diagnosis.

Schimke immuno-osseous dysplasia: two cases.

We report two patients with Schimke immuno-osseous dysplasia (SIOD). SIOD is characterised by growth retardation, renal failure, spondylo-epiphyseal dysplasia, specific phenotype and defective cellular immunity. These two children demonstrated a bone dysplasia with characteristic radiographic appearances. We postulate that SIOD should be considered in all cases of growth failure with an unclassifiable bone dysplasia. Repeated urine tests for proteinuria could be helpful in reaching the correct diagnosis.

[Biological activity of lipopolysaccharides from clinical Bacteroides fragilis strains isolated in Poland determined in reaction with limulus amoebocyte lysate]. / Biologiczna aktywnosc lipopolisacharydów klinicznych szczepów Bacteroides fragilis wyizolowanych w Polsce okreslona w reakcji z lizatem amebocytów Limulus (LAL).

The aim of this study was to determine a biological activity of lipopolysaccharides (LPS) from clinical Bacterioides fragilis strains isolated in Poland by means of quantitative, photometric BET (LAL) method with Limulus polyphemus amoebocyte lysate and chromogenic substrate S-2423. Lipopolysaccharides were extracted from nine clinical B. fragilis strains by the procedure of Westphal and Jann (1965). Crude LPS preparations were purified with ultracentrifugation. Biological activities of bacterial endotoxins were determined by quantitative BET method with chromogenic substrate S-2423 (ENDOCHROME kit). Tests were performed according to the recommendations of the producer (Charles River Endosafe Ltd., USA). E. coli O55:B5 LPS and LPS preparations from reference B. fragilis strains were applied to compare the results of examinations. Activities of endotoxins from clinical B. fragilis strains isolated in Poland determined in reaction with Limulus amoebocyte lysate were differentiated. Among endotoxins of clinical B. fragilis strains the most active was the preparation from strain cultured in the case of pancreatic ulcer (B. fragilis 80/81 LPS). Lipopolysaccharides of examined B. fragilis strains were less active in BET test than E. coli O55:B5 LPS.