Slobodanka Mitrovic

Protective role of IL-33/ST2 axis in Con A-induced hepatitis

BACKGROUND & AIMS We used Concanavalin A-induced liver injury to study the role of Interleukin 33 and its receptor ST2 in the induction of inflammatory pathology and hepatocellular damage. METHODS We tested susceptibility to Concanavalin A induced hepatitis in ST2 deficient and wild type BALB/c mice and analyzed the effects of single injection of Interleukin 33 as evaluated by liver enzyme test, quantitative histology, mononuclear cell infiltration, cytokine production, intracellular staining of immune cells, and markers of apoptosis in the liver. RESULTS ST2 deficient mice developed significantly more severe hepatitis and had significantly higher number of mononuclear cells in the liver, CD4+ and CD8+ T cells, NKp46+ and CD3+NKp46+ cells, and F4/80+ macrophages. The level of pro-inflammatory cytokines in the sera and number of TNF alpha, IFN gamma, and IL-17 producing cells was higher in ST2 deficient mice. In contrast, number of CD4+Foxp3+ cells was statistically higher in wild type mice. Additionally, treatment of wild type mice with single (1 μg) injection of Interleukin 33 led to attenuation of the liver injury and milder infiltration of mononuclear cells, increase in total number of liver CD4+Foxp3+ cells and IL-4 producing CD4+ T cells. Interleukin 33 also suppressed the activation of caspase 3, prevented the expression of BAX, and enhanced the expression of antiapoptotic Bcl-2 in the liver. CONCLUSIONS We concluded that Interleukin 33/ST2 axis downregulated Concanavalin A-induced liver injury and should be evaluated as potential target in fulminant hepatitis in humans.

Galectin-3 Plays an Important Pro-inflammatory Role in the Induction Phase of Acute Colitis by Promoting Activation of NLRP3 Inflammasome and Production of IL-1β in Macrophages

BACKGROUND AND AIMS Galectin-3 [Gal-3] is an endogenous lectin with a broad spectrum of immunoregulatory effects: it plays an important role in autoimmune/inflammatory and malignant diseases, but the precise role of Gal-3 in pathogenesis of ulcerative colitis is still unknown. METHODS We used a model of dextran sulphate sodium [DSS]-induced acute colitis. The role of Gal-3 in pathogenesis of this disease was tested by evaluating disease development in Gal-3 deficient mice and administration of Gal-3 inhibitor. Disease was monitored by clinical, histological, histochemical, and immunophenotypic investigations. Adoptive transfer was used to detect cellular events in pathogenesis. RESULTS Genetic deletion or pharmacological inhibition of Gal-3 significantly attenuate DSS-induced colitis. Gal-3 deletion suppresses production of pro-inflammatory cytokines in colonic macrophages and favours their alternative activation, as well as significantly reducing activation of NOD-like receptor family, pyrin domain containing 3 [NLRP3] inflammasome in macrophages. Peritoneal macrophages isolated from untreated Gal-3(-/-) mice and treated in vitro with bacterial lipopolysaccharide or DSS produce lower amounts of tumour necrosis factor alpha [TNF-α] and interleukin beta [IL-1β] when compared with wild type [WT] cells. Genetic deletion of Gal-3 did not directly affect total neutrophils, inflammatory dendritic cells [DCs] or natural killer [NK] T cells. However, the total number of CD11c+ CD80+ DCs which produce pro-inflammatory cytokines, as well as TNF-α and IL-1β producing CD45+ CD11c- Ly6G+ neutrophils were significantly lower in colons of Gal-3(-/-) DSS-treated mice. Adoptive transfer of WT macrophages significantly enhanced the severity of disease in Gal-3(-/-) mice. CONCLUSIONS Gal-3 expression promotes acute DSS-induced colitis and plays an important pro-inflammatory role in the induction phase of colitis by promoting the activation of NLRP3 inflammasome and production of IL-1β in macrophages.

Potential dual immunomodulatory role of VEGF in ulcerative colitis and colorectal carcinoma.

Objective. Progression from ulcerative colitis (UC) toward colorectal carcinoma (CRC) is multistep process that includes gene alterations of tumor suppressor genes, such as p53 and p16. The aim of this study was to investigate the expression patterns of p16, p53 and VEGF in affected tissue and serum levels of cytokines TNF-α, IFN-γ, IL-4, IL-6, IL-10 and IL-17 in patients with UC and CRC, respectively. Matherials and methods. Serum levels of cytokine in patients with UC (n=24) and CRC (n=75) and in a healthy group (n=37) were analyzed by ELISA. Endoscopic biopsies specimens of UC and CRC were studied by immunohistochemical staining for p16, p53 and VEGF. Results. Patients with UC with presence of extraintestinal manifestations, complications, and positive staining of p16, p53 and VEGF respectively had higher serum levels of pro-inflammatory cytokines. Higher percentage of CRC patients had positive staining of p16, p53 and VEGF. CRC patients with positive staining of VEGF had decreased systemic values of pro-inflammatory IFN-γ and increased values of immunosuppressive IL-10. Conclusions. Relatively low IL-10 in patients with severe UC is insufficient to compensate IL-6 secretion and subsequently enhanced type 1/17 immune response. In UC patients, p16 and p53 induce enhanced VEGF expression and subsequent production of pro-inflammatory TNF-α and IL-6. In CRC patients VEGF seems to have immunosuppressive role. It appears that tumor suppressor gene-VEGF axis have dual role on immune response in inflammation of UC and tumor growth and progression of CRC.

Ectopic mammary tissue in vulva.

BACKGROUND Ectopic mammary gland tissue is a residual tissue that persists during the embryologic development along ectodermal primitive milk streaks. Incomplete involution anywhere along the primitive milk streak can result in accessory or ectopic mammary tissue. CASE REPORT A woman, 27-year old, admitted to Obstetrics and Gynecology Clinic Kragujevac for surgery, of goose-egg size, vulva tumor, of elastic consistency. Menarche started in 12 years of age, with the regular menstrual cycle, without previous gyneocological diseases. The woman had one pregnancy terminated by cesarean section because of the multiple (twin) pregnancy. Excision of the tumor was completely done in the total endotracheal anesthesia. Pathohistologic (PH) findings was: Dysplasia fibrosa cystica simplex mammae, with focuses of sclerosing adenosis. Expression of estrogen (ER) and progesterone receptors (PR) were positive. CONCLUSION Ectopic mammary tissue in vulva in adult period is very rarely seen, and can be changed pathologically as well as normally positioned breast tissue into benign cystic changes, benign tumors, adenomas and fibroadenomas and tumors. Cells with low ER/PR receptor level grow independently of estrogene stimulation and they could be resistant to hormonal therapy effects.

In vitro and in vivo assessment of meadowsweet (Filipendula ulmaria) as anti-inflammatory agent

ETHNOPHARMACOLOGICAL RELEVANCE Meadowsweet (Filipendula ulmaria (L.) Maxim, Rosaceae) has been traditionally used in most European countries for the treatment of inflammatory diseases due to its antipyretic, analgesic, astringent, and anti-rheumatic properties. However, there is little scientific evidence on F. ulmaria anti-inflammatory effects regarding its impact on cyclooxygenases enzymatic activity and in vivo assessment of anti-inflammatory potential. This study aims to reveal the anti-inflammatory activity of methanolic extracts from the aerial parts (FUA) and roots (FUR) of F. ulmaria, both in in vitro and in vivo conditions. MATERIALS AND METHODS The characteristic phenolic compounds in F. ulmaria extracts were monitored via high performance thin layer chromatography (HPTLC). The in vitro anti-inflammatory activity of F. ulmaria extracts was evaluated using cyclooxygenase-1 (COX-1) and cyclooxygenase-2 (COX-2) enzyme assays, and an assay for determining COX-2 gene expression. The in vivo anti-inflammatory effect of F. ulmaria extracts was determined in two doses (100 and 200 mg/kg b.w.) with hot plate test and carrageenan-induced paw edema test in rats. Inflammation was also evaluated by histopathological and immunohistochemical analysis. RESULTS FUA extract showed the presence of rutoside, spiraeoside, and isoquercitrin. Both F. ulmaria extracts at a concentration of 50μg/mL were able to inhibit COX-1 and -2 enzyme activities, whereby FUA extract (62.84% and 46.43% inhibition, respectively) was double as effective as the root extract (32.11% and 20.20%, respectively). Extracts hardly inhibited the level of COX-2 gene expression in THP-1 cells at a concentration of 25μg/mL (10.19% inhibition by FUA and 8.54% by FUR). In the hot plate test, both extracts in two doses (100 and 200mg/kg b.w.), exhibited an increase in latency time when compared with the control group (p<0.05). In the carrageenan-induced acute inflammation test, FUA at doses of 100 and 200mg/kg b.w., and FUR at 200mg/kg, were able to significantly reduce the mean maximal swelling of rat paw until 6h of treatment. Indomethacin, FUA, and FUR extracts significantly decreased inflammation score and this effect was more pronounced after 24h, compared to the control group (p<0.05). CONCLUSIONS The observed results of in vitro and, for the first time, in vivo anti-inflammatory activity of meadowsweet extracts, provide support of the traditional use of this plant in the treatment of different inflammatory conditions. Further investigation of the anti-inflammatory compounds could reveal the mechanism of anti-inflammatory action of these extracts.

Systemic photochemotherapy decreases the expression of IFN-γ, IL-12p40 and IL-23p19 in psoriatic plaques

Psoriasis vulgaris (PV) is a chronic skin disease with unclear pathogenesis. In the present study we investigated the effect of systemic photochemotherapy (PUVA therapy- psoralen and UVA therapy) on the expression of IFN-γ, IL-12p40 and IL-23p19 in lesional psoriatic skin. Fifteen patients with chronic plaque type psoriasis selected to be treated with PUVA therapy were recruited for this study. Expression of IFN-γ, IL-12p40 and IL-23p19 in psoriatic lesions before and after twenty PUVA treatments was established by using immunohistochemistry (IHC). A significant decrease in expression (p < 0.05) of IFN-γ, IL-12p40 and IL-23p19 in epidermis and dermis of psoriatic lesions was observed. The immunosuppressive effect of PUVA therapy presented with decreased expression of biologically active IL-23 (IL-12/IL-23p40 + IL-23p19) as a part of the Th17 pathway, and IFN-γ (Th1 pathway) led, in our patients, to a marked clinical improvement as shown by PASI (before therapy 20.55 and after therapy 3.33).

The effects of N-acetylcysteine on cisplatin-induced changes of cardiodynamic parameters within coronary autoregulation range in isolated rat hearts.

The aim of this study was to evaluate the effects of chronic NAC administration along with cisplatin on cisplatin-induced cardiotoxicity by means of coronary flow (CF), cardiodynamic parameters, oxidative stress markers and morphological changes in isolated rat heart. Isolated hearts of Wistar albino rats (divided into four groups: control, cisplatin, NAC and cisplatin+NAC group) were perfused according to Langendorff technique at constant coronary perfusion pressure starting at 50 and gradually increased to 65, 80, 95 and 110 cm H2O to evaluate cardiodynamic parameters within autoregulation range. Samples of coronary venous effluent (CVE) were collected for determination of CF and biochemical assays, and heart tissue samples for biochemical assays and histopathological examination. Cisplatin treatment decreased CF and heart rate, and increased left ventricular systolic pressure and maximum left ventricular pressure development rate. Cisplatin increased H2O2 and TBARS, but decreased NO2(-) levels in CVE. In tissue samples, cisplatin reduced pathological alterations in myocardium and coronary vessels, with no changes in the amount of total glutathione, as well as in activity of glutathione peroxidase and glutathione reductase. NAC coadministration, by reducing oxidative damage, attenuated cisplatin-induced changes of cardiodynamic and oxidative stress parameters, as well as morphological changes in myocardium and coronary vasculature.

Actinic reticuloid presented as facies leonine

Kyriakos P. Kyriakis, MD, MPH Koumanoudi str 52 Athens 11474 Greece E-mail: fountou@otenet.gr References 1 Yeatman JM, Kilkenny M, Marks R. The prevalence of seborrhoeic keratoses in an Australian population: does exposure ton sunlight play a part in their frequency? Br J Dermatol 1997; 137: 411–414. 2 Rothman KJ. Stratified analysis. In: Modern Epidemiology. Boston, MA: Little Brown, 1986: 177–239. 3 Memon AA, Bothwell J, Tomenson J, Friedman PS. Prevalence of solar damage, keratosis and skin cancer in an English population. Br J Dermatol 1995; 132: 646.

Deletion of IL-33R attenuates VEGF expression and enhances necrosis in mammary carcinoma

Interleukin-33 (IL-33)/IL-33 receptor (IL-33R, ST2) signaling pathway promotes mammary cancer growth and metastasis by inhibiting anti-tumor immunity. However, the role of IL-33/IL-33R axis in neoangiogenesis and tumor necrosis is not elucidated. Therefore, the aim of this study was to investigate the role of IL-33/IL-33R axis in mammary tumor necrosis. Deletion of IL-33R (ST2) gene in BALB/c mice enhanced tumor necrosis and attenuated tumor growth in 4T1 breast cancer model, which was associated with markedly decreased expression of vascular endothelial growth factor (VEGF) and IL-33 in mammary tumor cells. We next analyzed IL-33, IL-33R and VEGF expression and microvascular density (MVD) in breast tumors from 40 female patients with absent or present tumor necrosis. We found significantly higher expression of IL-33, IL-33R and VEGF in breast cancer tissues with absent tumor necrosis. Both, IL-33 and IL-33R expression correlated with VEGF expression in tumor cells. Further, VEGF expression positively correlated with MVD in perinecrotic zone. Taking together, our data indicate that IL-33/IL-33R pathway is critically involved in mammary tumor growth by facilitating expression of pro-angiogenic VEGF in tumor cells and attenuating tumor necrosis. These data add an unidentified mechanism by which IL-33/IL-33R axis facilitates tumor growth.

ST2 deletion increases inflammatory bone destruction in experimentally induced periapical lesions in mice.

INTRODUCTION ST2 is a member of the interleukin (IL)-1 receptor family, and IL-33 is its natural ligand. ST2 signaling promotes Th2 immune response in allergy, autoimmunity, and chronic inflammatory disorders, but its role in the pathogenesis of periapical lesions is unknown. The purpose of this study was to investigate whether ST2 gene deletion affects the development of experimentally induced periapical lesions in mice. METHODS Pulps of mandibular molars from wild-type (WT) and ST2 knockout (ST2(-)/(-)) BALB/c mice were exposed and left open to the oral environment. After death, hemi-mandibles were isolated and prepared for histologic, immunohistochemical, and flow cytometric analysis. RESULTS The expression of IL-33 and its receptor ST2 was higher in periapical lesions in WT mice compared with normal root apices (both P < .05). The increased periapical bone loss observed in ST2(-)/(-) mice was associated with enhanced influx of neutrophils, CD3+ CXCR3+ Th1 cells, and CD3+ CCR6+ Th17 cells and increased number of tartrate-resistant acid phosphatase+ osteoclasts (all P < .05). Furthermore, periapical lesions in ST2(-)/(-) mice contained increased percentages of T cells expressing interferon-γ, IL-17, tumor necrosis factor-α, and IL-6 (all P < .05). In comparison with WT mice, CD3+ receptor activator of nuclear factor kappa B ligand+ T cells were increased, whereas CD3+ osteoprotegerin+ T cells were decreased in the lesions of ST2(-)/(-) mice (both P < .05). CONCLUSIONS ST2 deletion increases inflammatory bone loss in experimental periapical lesions in mice, which is associated with enhanced Th1/Th17 cell mediated periapical immune responses and increased osteoclastogenesis.