21 results on '"Milenković, Pavle B."'
Search Results
2. Transcriptional profiling of erythroid progenitors from G-CSF mobilized and nonmobilized peripheral blood
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Čokić, Vladan, Diklić, Miloš, Subotički, Tijana, Beleslin-Čokić, Bojana, Marković, Dragana, Milenković, Pavle B., and Jovčić, Gordana
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G-CSF mobilized peripheral blood ,erythroid progenitors ,microarray analysis - Abstract
Purpose: The purpose of this study was to examine the gene expression profile of granulocyte colony stimulating factor (G-CSF)-mobilized peripheral blood (mPB)-derived progenitors, used in transplantation. Methods: We correlated gene expression patterns of highly enriched steady-state peripheral blood (PB)- and mPB-derived CD71(+) cells by microarray and ingenuity pathway analyses, to identify the transcriptional program during in vitro erythroid differentiation. Results: The gene expression was more than doubled in mPB-derived (4180 genes) compared to PB-derived erythroid progenitors (1667 genes) while PB-and mPB-derived erythroid progenitors shared 1534 common genes. Comparative analysis of transcript levels showed differential expression of 54 genes between cultured erythroid progenitors of PB and mPB origin, where we identified common 13 downregulated and 30 upregulated genes. The most significant genes in mPB-derived erythroid progenitors were P4HB, DDIA3, ARPC2 and ATP5G3. Regarding G-CSF stimulation the G-CSF receptor CSF2RB (1.1-fold) was linked via STAT3 to erythroid-specific ALAS2 (2.9-fold) and GATA2 (1.3-fold) factors, all upregulated in mPB-derived erythroid progenitors, coupled to common upregulated NUDC gene involved in the proliferation of erythroid cells. Conclusion: This report provides an extensive transcriptional profile of cultured erythroid progenitors and leads to a better understanding of diversity among the progenitor sources.
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- 2014
3. Gene expression profile of circulating CD34(+) cells and granulocytes in chronic myeloid leukemia
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Čokić, Vladan, Čokić, Vladan, Mojsilović, Slavko, Jauković, Aleksandra, Kraguljac-Kurtović, Nada, Mojsilović, Sonja, Sefer, Dijana, Mitrović-Ajtić, Olivera, Milošević, Violeta, Bogdanović, Andrija, Đikić, Dragoslava, Milenković, Pavle B., Puri, Raj K., Čokić, Vladan, Čokić, Vladan, Mojsilović, Slavko, Jauković, Aleksandra, Kraguljac-Kurtović, Nada, Mojsilović, Sonja, Sefer, Dijana, Mitrović-Ajtić, Olivera, Milošević, Violeta, Bogdanović, Andrija, Đikić, Dragoslava, Milenković, Pavle B., and Puri, Raj K.
- Abstract
Purpose: We compared the gene expression profile of peripheral blood CD34(+) cells and granulocytes in subjects with chronic myeloid leukemia (CML), with the accent on signaling pathways affected by BCR-ABL oncogene. Methods: The microarray analyses have been performed in circulating CD34(+) cells and granulocytes from peripheral blood of 7 subjects with CML and 7 healthy donors. All studied BCR-ABL positive CML patients were in chronic phase, with a mean value of 2012 +/- SD of CD34(+) cells/mu l in peripheral blood. Results: The gene expression profile was more prominent in CML CD34(+) cells (3553 genes) compared to granulocytes (2701 genes). The 41 and 39 genes were significantly upregulated in CML CD34(+) cells (HINT1, TXN, SERBP1) and granulocytes, respectively. BCR-ABL oncogene activated PI3K/AKT and MAPK signaling through significant upregulation of PTPN11, CDK4/6, and MYC and reduction of E2F1, KRAS, and NFKBIA gene expression in CD34(+) cells. Among genes linked to the inhibition of cellular proliferation by BCR-ABL inhibitor Imatinib, the FOS and STAT1 demonstrated significantly decreased expression in CML. Conclusion: The presence of BCR-ABL fusion gene doubled the expression quantity of genes involved in the regulation of cell cycle, proliferation and apoptosis of CD34(+) cells. These results determined the modified genes in PI3K/AKT and MAPK signaling of CML subjects.
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- 2015
4. Proinflammatory Cytokine IL-6 and JAK-STAT Signaling Pathway in Myeloproliferative Neoplasms
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Čokić, Vladan, Čokić, Vladan, Mitrović-Ajtić, Olivera, Beleslin-Čokić, Bojana, Marković, Dragana, Kovačić, Marijana, Diklić, Miloš, Kraguljac-Kurtović, Nada, Damjanović, Svetozar, Milenković, Pavle B., Gotić, Mirjana, Raj, Puri K., Čokić, Vladan, Čokić, Vladan, Mitrović-Ajtić, Olivera, Beleslin-Čokić, Bojana, Marković, Dragana, Kovačić, Marijana, Diklić, Miloš, Kraguljac-Kurtović, Nada, Damjanović, Svetozar, Milenković, Pavle B., Gotić, Mirjana, and Raj, Puri K.
- Abstract
The recent JAK1/2 inhibitor trial in myeloproliferative neoplasms (MPNs) showed that reducing inflammation can be more beneficial than targeting gene mutants. We evaluated the proinflammatory IL-6 cytokine and JAK-STAT signaling pathway related genes in circulating CD34(+) cells of MPNs. Regarding laboratory data, leukocytosis has been observed in polycythemia vera (PV) and JAK2V617F mutation positive versus negative primary myelofibrosis (PMF) patients. Moreover, thrombocytosis was reduced by JAK2V617F allele burden in essential thrombocythemia (ET) and PMF. 261 significantly changed genes have been detected in PV, 82 in ET, and 94 genes in PMF. The following JAK-STAT signaling pathway related genes had augmented expression in CD34(+) cells of MPNs: CCND3 and IL23A regardless of JAK2V617F allele burden; CSF3R, IL6ST, and STAT1/2 in ET and PV with JAK2V617F mutation; and AKT2, IFNGR2, PIM1, PTPN11, and STAT3 only in PV. STAT5A gene expression was generally reduced in MPNs. IL-6 cytokine levels were increased in plasma, as well as IL-6 protein levels in bone marrow stroma of MPNs, dependent on JAK2V617F mutation presence in ET and PMF patients. Therefore, the JAK2V617F mutant allele burden participated in inflammation biomarkers induction and related signaling pathways activation in MPNs.
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- 2015
5. Bradykinin stimulation of nitric oxide production is not sufficient for gamma-globin induction
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Čokić, Vladan, Čokić, Vladan, Subotički, Tijana, Beleslin-Čokić, Bojana, Diklić, Miloš, Milenković, Pavle B., Jovčić, Gordana, Čokić, Vladan, Čokić, Vladan, Subotički, Tijana, Beleslin-Čokić, Bojana, Diklić, Miloš, Milenković, Pavle B., and Jovčić, Gordana
- Abstract
Introduction Hydroxycarbamide, used in therapy of hemoglobinopathies, enhances nitric oxide (NO) production both in primary human umbilical vein endothelial cells (HUVECs) and human bone marrow endothelial cell line (TrHBMEC). Moreover, NO increases γ-globin and fetal hemoglobin levels in human erythroid progenitors. Objective In order to find out whether simple physiologic stimulation of NO production by components of hematopoietic microenvironment can increase γ-globin gene expression, the effects of NO-inducer bradykinin were examined in endothelial cells. Methods The study was performed in co-cultures of human erythroid progenitors, TrHBMEC and HUVECs by ozone-based chemiluminescent determination of NO and real-time quantitative RT-PCR. Results In accordance with previous reports, the endogenous factor bradykinin increased endothelial cell production of NO in a dose- and time-dependent manner (0.1-0.6 μM up to 30 minutes). This induction of NO in HUVECs and TrHBMEC by bradykinin was blocked by competitive inhibitors of NO synthase (NOS), demonstrating NOS-dependence. It has been shown that bradykinin significantly reduced endothelial NOS (eNOS) mRNA level and eNOS/s-actin ratio in HUVEC (by twofold). In addition, bradykinin failed to increase γ-globin mRNA expression in erythroid progenitors only, as well as in co-culture studies of erythroid progenitors with TrHBMEC and HUVEC after 24 hours of treatment. Furthermore, bradykinin did not induce γ/β globin ratio in erythroid progenitors in co-cultures with HUVEC. Conclusion Bradykinin mediated eNOS activation leads to short time and low NO production in endothelial cells, insufficient to induce γ-globin gene expression. These results emphasized the significance of elevated and extended NO production in augmentation of γ-globin gene expression., Uvod Hidroksikarbamid, koji se koristi u lečenju hemoglobinopatija, podstiče stvaranje azot-monoksida (NO) kako u primarnim ljudskim endotelnim ćelijama pupčane vene (HUVEC), tako i u izmenjenoj endotelnoj ćelijskoj liniji poreklom iz koštane srži (TrHBMEC). Štaviše, NO povećava stvaranje γ-globina i fetalnog hemoglobina u ljudskim progenitorima eritropoeze. Cilj rada Da bismo ustanovili da li jednostavna fiziološka stimulacija stvaranja NO od komponenti mikrosredine hematopoeze može povećati ekspresiju γ-globinskog gena, ispitivali smo efekte bradikinina, već poznatog stimulatora stvaranja NO. Metode rada Studija je izvedena u zajedničkim kulturama ljudskih progenitora eritropoeze sa TrHBMEC ili HUVEC i ispitivana hemiluminiscentnim merenjem NO posredstvom ozona, kao i primenom kvantitativnog RT-PCR na genskom nivou. Rezultati U skladu s prethodnim izveštajima, pokazali smo da endogeni faktor bradikinin povećava stvaranje NO u endotelnim ćelijama na dozno i vremenski zavisan način (0,1-0,6 μM do 30 minuta). Ovo stvaranje NO u HUVEC i TrHBMEC izazvano bradikininom blokirano je od strane konkurentskih inhibitora NO-sintaze (NOS), pokazujući NOS-zavisnost. Utvrdili smo da bradikinin značajno smanjuje stvaranje iRNK endotelne forme NOS (eNOS), kao i odnos eNOS i β-aktina u HUVEC (dvostruko manje). Pored toga, bradikinin ne povećava ekspresiju iRNK γ-globinskog gena ni u zasebnim progenitorima eritropoeze, niti u zajedničkim kulturama progenitora eritropoeze sa TrHBMEC ili HUVEC posle 24 sata tretmana. Bradikinin ne menja ni odnos γ i β globina u zajedničkim kulturama progenitora eritropoeze sa HUVEC. Zaključak Aktivacija eNO_ izazvana bradikininom dovodi do kratkog i malog povećanja NO u endotelnim ćelijama, što je nedovoljno da podstakne ekspresiju gena za γ-globin. Ovi rezultati naglašavaju važnost povećanog i produženog stvaranja NO radi stimulacije ekspresije γ-globina.
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- 2014
6. Concentrations of IL-17 and autoantibodies in sera of patients with primary Sjogren's syndrome
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Dokmanovic-Miletić, M. P., Stojanović, R.M., Pajić, O., Bugarski, Diana, Mojsilović, S., and Milenković, Pavle B.
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- 2007
7. Signaling pathways implicated in hematopoietic progenitor cell proliferation
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Bugarski, Diana, Petakov, Marijana, Vlaški, Marija, Krstić, Aleksandra, Čokić, Vladan, Jovčić, Gordana, Stojanović, Nevenka, and Milenković, Pavle B.
- Subjects
PI-3 kinase pathway ,MAP kinaze ,PI-3 kinazni put ,JAK/STAT pathway ,matične ćelije hematopoeze ,JAK/STAT put ,MAP kinase pathway ,hematopoietic progenitor cells - Abstract
Different biological functions of hematopoietic cells are regulated by signals cells receive from their environment that may come from interactions with other cells or from soluble growth factors and cytokines. Although the effects of hematopoietic regulators are well described, the exact intracellular signaling cascades leading to various cellular responses are not fully elucidated. All hematopoietic growth factors and cytokines seem able to activate all the major signal transduction pathways simultaneously, and it appears that the same network of signaling proteins may coordinate numerous cellular functions. However, the differences that lead to the unique biological events of the particular cytokine, as well as the differences that determine lineage-specific blood cell differentiation are only gradually being uncovered. With the help of specific inhibitors of known signal transduction pathways, we have examined the contribution of particular signaling molecules of Jak/Stat, MAP kinase and PI-3 kinase pathways, as well as the activation of nuclear factor kappaB (NFkB) transcription factor on the proliferation and differentiation of murine bone marrow granulocyte-macrophage (CFU- GM) and erythroid (BFU-E and CFU-E) progenitors. Preliminary results demonstrated that the effects of the inhibitors on the hematopoietic colony formation were lineage-dependent, as well as dependent on erythroid progenitors' stage of differentiation. The obtained differences suggest that different signal transduction intermediates regulate erythroid and myeloid progenitor cell proliferation and differentiation. Differences in the susceptibility of the progenitor cells to the inhibitors used were also observed. The data is consistent with other evidences indicating that different threshold levels are one of the mechanisms by which the signaling specificity may be achieved. Mnogobrojne funkcije hematopoetskih ćelija u toku procesa hematopoeze, regulisane su brojnim signalima koje ćelije primaju iz svoje okoline i to kako od drugih ćelija tako i od brojnih regulatornih molekula. Iako su biološki efekti regulatora hematopoeze uglavnom dobro opisani, putevi prenošenja signala unutar ćelija na koje deluju nisu još uvek u potpunosti razjašnjeni. Savremena istraživanja ukazuju da gotovo svi poznati hematopoetski regulatori mogu da aktiviraju gotovo sve bitne puteve prenošenja signala unutar ćelija, što sve upućuje da ista mreža signalnih komponenti može da koordinira brojne ćelijske funkcije. Međutim, razlike u transdukciji signala koje dovode do jedinstvenog biološkog dejstva svakog citokina, kao i razlike u prenosu signala kojom se određuje lozno-zavisna diferencijacija krvnih ćelija nisu još poznati. Naša istraživanja započela su određivanjem učešća pojedinih komponenti JAK/STAT puta signalizacije, MAP kinazne kaskade i PI-3 kinaznog puta, kao i aktivacije transkripcionog faktora NFKB, u proliferaciji i diferentovanju opredeljenih matičnih ćelija granulocitno-monocitne (CFU-GM) i eritrocitne (BFU- E i CFU-E) loze kostne srži normalnih miševa. Početni rezultati ovih ispitivanja su ukazali na postojanje kako lozno-zavisnih razlika u odgovoru matičnih ćelija hematopoeze na pojedine inhibitore, tako i na zavisnost odgovora od stepena zrelosti matičnih ćelija u okviru iste loze. Dobijene razlike upućuju da su različiti učesnici signalnih puteva uključeni u regulaciju proliferacije i diferencijacije eritrocitnih i granulocitno-monocitnih progenitora. Ovo je jedan od mogućih načina kojim se možda omogućava selektivna ekspanzija ćelija određenih krvnih loza u zavisnosti od potreba organizma. Takođe, uočene su i razlike u pragu osetljivosti različitih kategorija opredeljenih matičnih ćelija hematopoeze na ispitivane inhibitore signalnih puteva, što ide u prilog podacima da su različiti pragovi osetljivosti jedan od mogućih mehanizama kojim se najverovatnije ostvaruje specifičnost delovanja signalnih puteva.
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- 2004
8. Hematopoietic microenvironment
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Bugarski, Diana, Petakov, Marijana, Jovčić, Gordana, Stojanović, Nevenka, and Milenković, Pavle B.
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hematopoeza ,extracellular matrix ,regulatorni molekuli ,stromalne ćelije ,regulatory molecules ,ekstracelularni matriks ,hematopoiesis ,stromal cells - Abstract
Steady-state hematopoiesis takes place in the bone marrow permissive microenvironment, composed of stromal cells, as well as extracellular matrix (ECM) components and regulatory molecules, produced by both stromal and hematopoietic cells. Stromal cells are both mezenchymal (endothelial cells fibroblasts, adipocytes, osteoblasts, myocytes, adventitial reticular cells) and hematopoietic (macrophages) in origin, which together provide cell to cell interactions, matrix proteins and growth factors essential for the maintenance, growth and differentiation of hematopoietic stem and progenitor cells. The ECM components (collagen, laminin, fibronectin, thrombospondin proteoglycans, hemonectin and various proteinases participating in their remodeling) are involved in different biological functions such as cell adhesion, binding and presentation of various cytokines and regulation of cell growth. These components serve to localize hematopoietic cells in the specific bone marrow microenvironment and it is suggested that in combination with cytokines are crucial for their compartmentalization. The regulatory molecules of hematopoietic microenvironment are cytokines, which regulate the survival, proliferation and differentiation of hematopoietic cells and cell adhesion molecules, which are responsible for the localization of hematopoiesis to the bone marrow. The activity of cytokines may be greater when the cytokine is present in a membrane-bound or ECM-associated form. Hematopoietic cells could also regulate normal hematopoiesis in an autocrine/paracrine manner, since it was shown that these cells express and secrete various growth factors, cytokines and chemokines. Proces hematopoeze se odvija u definisanoj tkivnoj mikrosredini kostne srži koja ima ključnu ulogu u njegovoj regulaciji. Mikrosredina hematopoeze podrazumeva funkcionalno jedinstvo stromalnih ćelija i ćelijskih produkata (molekuli ekstracelularnog matriksa i regulatorni faktori), koji čine kompleksni molekularni milje u kome se ostvaruju specifične interakcije hematopoetskih ćelija i komponenti mikrosredine. Stromalne ćelije mezenhimalnog porekla (endotelne ćelije, fibroblasti, adipociti osteoblasti, miociti, adventicijske ćelije retikuluma) i nemezenhimalnog porekla (makrofage), deluju na matične ćelije hematopoeze direktno među ćelijskim interakcijama, kao i produkcijom i deponovanjem pojedinih komponenti kompleksnog ekstracelularnog matriksa, a takođe i produkcijom i koncentrovanjem lokalnih citokina i faktora rasta sa hematopoetskim efektima, obezbeđujući na taj način gotovo sve činioce neophodne za proliferaciju i diferencijaciju matičnih ćelija hematopoeze. Komponente ECM (kolagen, laminin, fibronektin, trombospondin, proteoglikani, hemonektin i drugi glikoproteini kao i proteolitički enzimi koji omogućavaju remodelovanje ECM) su uključene u ćelijsku adheziju, vezivanje i prezentaciju različitih citokina i regulaciju ćelijskog rasta. Smatra se da komponente ECM omogućavaju lokalizuju hematopoetskih ćelija u specifičnoj mikrosredini kostne srži, i da u kombinaciji sa citokinima imaju presudnu ulogu u formiranju specifičnih niša. Regulatorni faktori hematopoetske mikrosredine su citokini koji regulišu preživljavanje, proliferaciju i diferencijaciju hematopoetskih ćelija i ćelijski adhezivni molekuli koji su odgovorni za lokalizaciju hematopoeze u kostnoj srži i za posredovanje u fizičkoj vezi između hematopoetskih ćelija i stromalnog tkiva mikrosredine. Ovi molekuli se u mikrosredini kostne srži nalaze kao solubilni, ili vezani za membrane stromalnih ćelija ili za komponente ECM, što može da pojača njihovu aktivnost. Takođe i same hematopoetske ćelije učestviju u regulaciji procesa hematopoeze produkujući citokine koji ispoljavaju autokrino/parakrine efekte.
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- 2003
9. Application of non-equilibrium plasmas in medicine
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Petrović, Zoran Lj., Petrović, Zoran Lj., Puač, Nevena, Malović, Gordana, Lazović, Saša, Maletić, Dejan, Miletić, Maja, Mojsilović, Slavko, Milenković, Pavle B., Bugarski, Diana, Petrović, Zoran Lj., Petrović, Zoran Lj., Puač, Nevena, Malović, Gordana, Lazović, Saša, Maletić, Dejan, Miletić, Maja, Mojsilović, Slavko, Milenković, Pavle B., and Bugarski, Diana
- Abstract
The potential of plasma applications medicine, the connections to nanotechnologies and the results obtained by our group are reviewed. A special issue in plasma medicine is the development of the plasma sources that would achieve non-equilibrium at atmospheric pressure in an atmospheric gas mixture with no or only marginal heating of the gas, and with desired properties and mechanisms that may be controlled. Our studies have shown that control of radicals or chemically active products of the discharge, such as ROS (reactive oxygen species) and/or NO, may be used to control the growth of the seeds. Simultaneously, a specially designed plasma needle and other sources were shown to be efficient to sterilize not only colonies of bacteria but also plank- tonic samples (microorganisms protected by water) or bio films. Finally, it was shown that a plasma might induce differentiation of stem cells. Non-equilibrium plasmas may be used in detection of different specific markers in medicine. For example proton transfer mass spectroscopy may be employed in the detection of volatile organic compounds without their dissociation and thus as a technique for instantaneous measurement of the presence of markers for numerous diseases., U ovom radu dat je pregled primene plazme u medicini, povezanost sa nanotehnologijama i rezultate na ovom polju koje je postigla naša grupa. Poseban problem u plazma medicini je razvoj izvora plazme koji bi radili u neravnotežnim uslovima na atmosferskom pritisku i u smeši gasova kakva je u atmosferi uz zanemarljivo grejanje gasa i sa željenim karakteristikama koje se mogu podešavati po želji. Naša istraživanja su pokazala da se kontrola prisustva radikala i drugih hemijski aktivnih čestica kao što su reaktivne kiseonične čestice (ROS) i/ili NO, može koristiti za kontrolu klijanja semenki. U isto vreme je dokazano za posebno konstruisanu plazma iglu da može efikasno da steriliše ne samo kolonije bakterija već i planktonske uzorke (mikroorganizme zaštićene vodom) pa i biofilmove. Na kraju, mi smo pokazali da plazma može da indukuje diferencijaciju matičnih ćelija. Neravnotežna plazma se može koristiti za detekciju raznih specifičnih markera u medicini. Na primer masena spektroskopija na bazi izmene protona može da se koristi za detekciju isparivih organskih jedinjenja bez njihove disocijacije i na taj način se može ostvariti trenutna detekcija markera za brojne bolesti iz daha.
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- 2012
10. Serum interleukin-17 & nitric oxide levels in patients with primary Sjogren's syndrome
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Miletić, Maja, Miletić, Maja, Stojanović, Roksanda M., Pajić, O., Bugarski, Diana, Mojsilović, Slavko, Čokić, Vladan, Milenković, Pavle B., Miletić, Maja, Miletić, Maja, Stojanović, Roksanda M., Pajić, O., Bugarski, Diana, Mojsilović, Slavko, Čokić, Vladan, and Milenković, Pavle B.
- Abstract
Background & objectives: The interleukin (IL)-17 producing T-helper cells have been linked to pathogenesis of autoimmunity and mostly investigated in rheumatoid arthritis (RA). In this study we tested the IL-17 levels, as well as the levels of nitric oxide (NO) as possible IL-17-induced product, in patients with primary Sjogren's syndrome (pSS), an intricate and complex chronic autoimmune disorder of exocrine glands. Methods: Serum IL-17 levels and nitrite concentrations determined in patients with pSS (n=30) were compared with the values obtained in patients with RA (n=10) and healthy controls (n=15). The values obtained for 11,17 in pSS patients were also associated with the patients' clinical characteristics, particularly the rheumatoid factor (RF) and total antinuclear antibodies (tANA) levels. Results: Serum concentrations of IL-17 were significantly (P lt 0.01) higher in patients with pSS (12.9 +/- 28.0 pg/ml) as compared to those obtained in healthy individuals (0.2 +/- 0.6 pg/ml), but not as high as the values obtained for the patients with RA (34.5 +/- 56.2 pg/ml). The mean IL-17 levels were significantly (P lt 0.05) higher in the pSS patients positive for rheumatoid factor (20.3 +/- 33.3 pg/ml) than in RF-negatives (0.3 +/- 0.6 pg/ml). Mean serum concentrations of IL-17 were also higher in antinuclear antibody (ANA)positive samples (19.8 +/- 33.5 pg/ml) in comparison to ANA-negative sera (1.1 +/- 3.1 pg/ml) (P lt 0.05). The NO levels also showed elevated values in both pSS and RA patients, as compared to the healthy controls, since mean nitrite levels in patients with pSS and RA were 38.2 +/- 29.2 mu M and 41.7 +/- 21.1 mu M, respectively, while those in healthy controls were significantly lower, at 19.2 +/- 10.5 mu M. Interpretation & conclusions: The findings of this study showed that there was increased IL-17 and NO production in patients with primary SS, especially if they had associated elevated rheumatoid factor and antinuclear antibody values.
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- 2012
11. The effect of a plasma needle on bacteria in planktonic samples and on peripheral blood mesenchymal stem cells
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Lazović, Saša, Lazović, Saša, Puač, Nevena, Miletić, Maja, Pavlica, Dusan, Jovanović, Milena, Bugarski, Diana, Mojsilović, Slavko, Maletić, Dejan, Malović, Gordana, Milenković, Pavle B., Petrović, Zoran Lj., Lazović, Saša, Lazović, Saša, Puač, Nevena, Miletić, Maja, Pavlica, Dusan, Jovanović, Milena, Bugarski, Diana, Mojsilović, Slavko, Maletić, Dejan, Malović, Gordana, Milenković, Pavle B., and Petrović, Zoran Lj.
- Abstract
In this paper, we study the application of a plasma needle to induce necrosis in planktonic samples containing a single breed of bacteria. Two different types of bacteria, Staphylococcus aureus (ATCC 25923) and Escherichia coli (ATCC 25922), were covered in this study. In all experiments with bacteria, the samples were liquid suspensions of several different concentrations of bacteria prepared according to the McFarland standard. The second system studied in this paper was human peripheral blood mesenchymal stem cells (hPB-MSC). In the case of hPB-MSC, two sets of experiments were performed: when cells were covered with a certain amount of liquid (indirect) and when the cell sample was in direct contact with the plasma. Most importantly, the study is made with the aim to see the effects when the living cells are in a liquid medium, which normally acts as protection against the many agents that may be released by plasmas. It was found that a good effect may be expected for a wide range of initial cell densities and operating conditions causing destruction of several orders of magnitude even under the protection of a liquid. It was established independently that a temperature increase could not affect the cells under the conditions of our experiment, so the effect could those hPB-MSC that were not protected by a liquid, gas flow proved to produce a considerable effect, presumably due to poor adhesion of the cells, but in a liquid the effect was only due to the plasma. Further optimization of the operation may be attempted, opening up the possibility of localized in vivo sterilization.
- Published
- 2010
12. Syphacia obvelata modifies mitogen-activated protein kinases and nitric oxide synthases expression in murine bone marrow cells
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Ilić, Vesna, Ilić, Vesna, Krstić, Aleksandra, Katić-Radivojević, Sofija, Jovčić, Gordana, Milenković, Pavle B., Bugarski, Diana, Ilić, Vesna, Ilić, Vesna, Krstić, Aleksandra, Katić-Radivojević, Sofija, Jovčić, Gordana, Milenković, Pavle B., and Bugarski, Diana
- Abstract
Syphacia obvelata is a rodent nematode parasite with high prevalence in laboratory mice. In our previous work we have demonstrated that this gut-dwelling helminth induces significant hematopoietic changes, characterized by increased myelopoiesis and erythropoiesis in infected animals, and accompanied with altered reactivity of bone marrow hematopoietic progenitors to interleukin (IL)-17. In this study we extended these investigations by demonstrating that naturally acquired S. obvelata infection induces significant alterations in murine bone marrow cells manifested at the molecular level. Namely, S. obvelata infection induced Sustained phosphorylation of the members of three major groups of distinctly regulated mitogen-activated protein kinases (MAPKs), the p38, the c-Jun amino-terminal kinase (JNK) and the extracellular signal-regulated kinase (ERK), as well as enhanced expression of mRNA for the inducible nitric oxide synthase (iNOS) in the bone marrow cells of infected animals. Furthermore, the infection interfered with the IL-17-mediated effects in bone marrow cells, since in normal mice IL-17 significantly enhanced phosphorylation of p38 MAPK and upregulated the expression of NOS and the constitutive, endothelial (e)NOS mRNA, while in S. obvelata-infected animals IL-17 did not influence the MAPKs activation, but markedly down-regulated the expression of both NOS isoforms. The data obtained demonstrating that S. obvelata is able to manipulate signal transduction pathways in the hosts' bone marrow cells, pointed to the multiple layers of immuno modulatory ability of this pinworm parasite and highlighted the importance of working under pinworm-free conditions when using experimental murine models for immunohematopoietic investigations.
- Published
- 2010
13. Cultivation of hamster bone marrow haematopoietic stem and progenitor cells
- Author
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Kovačević-Filipović, Milica, Kovačević-Filipović, Milica, Okić, Ivana, Petrićević, Tanja, Mojsilović, S., Krstić, Aleksandra, Jovčić, Gordana, Bugarski, Diana, Milenković, Pavle B., Petakov, Marijana, Radovanović, Anita, Božić, Tatjana, Ivanović, Z., Kovačević-Filipović, Milica, Kovačević-Filipović, Milica, Okić, Ivana, Petrićević, Tanja, Mojsilović, S., Krstić, Aleksandra, Jovčić, Gordana, Bugarski, Diana, Milenković, Pavle B., Petakov, Marijana, Radovanović, Anita, Božić, Tatjana, and Ivanović, Z.
- Abstract
Hamster, a hibernating animal, is an important experimental model in research on the influence of hypothermia on different physiological processes. A simple procedure for cultivation and identification of hamster hematopoetic stem cells (HSC) and hematopoetic progenitor cells (HPC) is a premise for a successful investigation upon hypothermia effects on hematopoiesis. The aim of this work was to evaluate the utilization of commercially available methylcellulose media (MC) and recombinant mouse and human cytokines for hamster HSC and HPC assays, in order to enable further studies on these cells. Hamster bone marrow mononuclear cells (BMMNC) were plated in MC containing cytokines that support mouse or human HPC growth. Also, BMMNC were resuspended in cytokine supplemented liquid media and incubated for 5 weeks with a four day monitoring of viable cell number. We demonstrated that hamster hematopoietic progenitor cells committed for erythroid lineage and myeloid lineage successfully formed recognizable colonies in both mouse and human MC, while multipotent progenitor cells formed colonies only in mouse MC. We also defined conditions for the evaluation of hamster HSC activity in liquid cultures, based on continuous 5 weeks HSC proliferation. The obtained results verify the utilization of mouse specific MC for further research on hamster HPC biology during hypothermia., Fiziološka hibernacija u koju hrčci ulaze prilikom izlaganja niskim temperaturama, čini ove životinje zanimljivim eksperimentalnim modelom za ispitivanje hematopoeze u uslovima hipotermije. Preduslov za ovo ispitivanje je postojanje jednostavne metode za kultivaciju i identifikaciju hematopoetskih ćelija hrčka. Cilj ovog rada je bio da se ispita mogućnost kultivacije progenitorskih ćelija hematopoeze hrčka u kompletnoj metil celulozi dizajniranoj za kultivaciju mišijih i humanih hematopoetskih ćelija, kao i da se odrede optimalni uslovi za kultivaciju matičnih ćelija hematopoeze hrčka u tečnoj kulturi. Mononuklearne ćelije kostne srži hrčka su posađene u metil celulozu i u tečnu kulturu. Oba medijuma su sadržala kombinacije rekombinantnih mišijih i/ili humanih citokina. Kolonije progenitorskih ćelija opredeljenih za mijelopoezu i opredeljenih za eritropoezu su se formirale u metil celulozi dizajniranoj za kultivaciju mišijih i humanih hematopoetskih ćelija, dok su se primitivnije kolonije sastavljene od oba tipa ćelija (mijeloidna i eritrocitna loza) formirale samo u metil celulozi dizajniranoj za kultivaciju mišijih hematopoetskih ćelija. Osim toga, populacija matičnih ćelija hematopoeze hrčka je proliferisala u tečnim kulturama tokom 5 nedelja bez znakova opadanja proliferativnog potencijala. Ova istraživanja pokazuju da se primenjene metode mogu uspešno koristiti za ispitivanje hematopoeze kod hrčka.
- Published
- 2010
14. p38 MAPK signaling mediates IL-17-induced nitric oxide synthase expression in bone marrow cells
- Author
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Krstić, Aleksandra, Krstić, Aleksandra, Ilić, Vesna, Mojsilović, Slavko, Jovčić, Gordana, Milenković, Pavle B., Bugarski, Diana, Krstić, Aleksandra, Krstić, Aleksandra, Ilić, Vesna, Mojsilović, Slavko, Jovčić, Gordana, Milenković, Pavle B., and Bugarski, Diana
- Abstract
The effects of interleukin (IL)-17 on nitric oxide (NO) synthase (NOS) expression, as well as the participation of mitogen-activated protein kinases (MAPKs) in IL-17-mediated effects were examined in murine bone marrow cells. The results demonstrated the ability of IL-17 to upregulate the expression of mRNA for both inducible NOS and constitutive, endothelial NOS isoforms, as well as to enhance the phosphorylation of p38 MAPK. Moreover, both the NOS-inducing effect of IL-17 and the in vitro IL-17-mediated inhibition colony forming unit-erythroid (CFU-E) growth were dependent on p38 MAPK activity. The data demonstrating that the in vivo reducing effect of IL-17 on bone marrow CFU-E was prevented by co-treatment with the NOS inhibitor Nw-nitro-l-arginine methyl ester hydrochloride (L-NAME), implied that this effect is mediated through NOS activation. Besides revealing a link between the IL-17, NO, and haematopoiesis, data presented gave an insight into the mechanisms by which IL-17 exerts its modulatory effects on bone marrow cells.
- Published
- 2009
15. Low O-2 concentrations enhance the positive effect of IL-17 on the maintenance of erythroid progenitors during co-culture of CD34+and mesenchymal stem cells
- Author
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Krstić, Aleksandra, Krstić, Aleksandra, Vlaški, Marija, Hammoud, Mohammad, Chevaleyre, Jean, Duchez, Pascale, Jovčić, Gordana, Bugarski, Diana, Milenković, Pavle B., Bourin, Philippe, Boiron, Jean-Michel, Praloran, Vincent, Ivanović, Zoran, Krstić, Aleksandra, Krstić, Aleksandra, Vlaški, Marija, Hammoud, Mohammad, Chevaleyre, Jean, Duchez, Pascale, Jovčić, Gordana, Bugarski, Diana, Milenković, Pavle B., Bourin, Philippe, Boiron, Jean-Michel, Praloran, Vincent, and Ivanović, Zoran
- Abstract
Co-culture of haematopoietic cells with a stromal cell layer does not mimic the physiological, micro-environmental niche, whose major feature is a low oxygen (O-2) concentration. Thus, in order to study the effects of IL-17 in a context which better approximates the physiological state, we investigated its effects on cell expansion, colony-forming ability, and the phenotypical profile of normal, human blood CD34(+) cells co-cultured for five days with MSC layers at various O-2 concentrations (20%, 12.5% and 3% O-2). We demonstrated that IL-17 enhances CD34(+) and total CFC production during the five days of MSC/CD34(+) co-culture. This effect depends upon the O-2 concentration, reaching its maximum at 3% O-2, and is more pronounced on erythroid progenitors (BFU-E). In addition, the stimulation of IL-6 production by IL-17 in MSC cultures and co-cultures is enhanced by low O-2 concentration. The expression of some differentiation markers (CD34, CD13 and CD41) on haematopoietic cells in co-cultures also depends upon the oxygen concentration. Our results strengthen the concept that physiological levels of O-2 (mistakenly called hypoxia), should be considered as an important environmental factor that significantly influences cytokine activity.
- Published
- 2009
16. Code of ethics in science and research good scientific practice
- Author
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Vučković-Dekić, Ljiljana, Vučković-Dekić, Ljiljana, Radulović, Siniša, Stanojević-Bakić, Nevenka, Jelić, Svetislav, Borojević, Nenad, Stojanović, Nevenka, Đurković-Đaković, Olgica, Jovović, Đurđica, Milenković, Pavle B., Vučković-Dekić, Ljiljana, Vučković-Dekić, Ljiljana, Radulović, Siniša, Stanojević-Bakić, Nevenka, Jelić, Svetislav, Borojević, Nenad, Stojanović, Nevenka, Đurković-Đaković, Olgica, Jovović, Đurđica, and Milenković, Pavle B.
- Published
- 2007
17. Signaling pathways implicated in hematopoietic progenitor cell proliferation and differentiation
- Author
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Bugarski, Diana, Bugarski, Diana, Krstić, Aleksandra, Mojsilović, Slavko, Vlaški, Marija, Petakov, Marijana, Jovčić, Gordana, Stojanović, Nevenka, Milenković, Pavle B., Bugarski, Diana, Bugarski, Diana, Krstić, Aleksandra, Mojsilović, Slavko, Vlaški, Marija, Petakov, Marijana, Jovčić, Gordana, Stojanović, Nevenka, and Milenković, Pavle B.
- Abstract
The objective of this study was to investigate the signal transduction pathways associated with the clonal development of myeloid and erythroid progenitor cells. The contribution of particular signaling molecules of protein tyrosine kinases (PTKs), mitogen-activated protein (MAP) kinase, and PI-3 kinase signaling to the growth of murine bone marrow colony forming unit-granulocyte-macrophage (CFU-GM) and erythroid (burst forming unit-erythroid [BFU-E] and colony forming unit-erythroid [CFU-E]) progenitors was examined in studies performed in the presence or absence of specific signal transduction inhibitors. The results clearly pointed to different signal transducing intermediates that are involved in cell proliferation and differentiation depending on the cell lineage, as well as on the progenitors' maturity. Lineage-specific differences were obtained when chemical inhibitors specific for receptor- or nonreceptor-PTKs, as well as for the main groups of distinctly regulated MAPK cascades, were used because all of these compounds suppressed the growth of erythroid progenitors, with no major effects on myeloid progenitors. At the same time, differential involvement of MEK/extracellular signal-regulated kinase (ERK) MAPK transduction pathway was observed in the proliferation and/or differentiation of early, BFU-E, and late, CFU-E, erythroid progenitor cells. The results also demonstrated that phosphatydylinositol (PI)-3 kinase and nuclear factor kappaB (NF-kappa B) transcriptional factor were required for maintenance of both myeloid and erythroid progenitor cell function. Overall, the data obtained indicated that committed hematopoietic progenitors express a certain level of constitutive signaling activity that participates in the regulation of normal steady-state hematopoiesis and point to the importance of evaluating the impact of signal transduction inhibitors on normal bone marrow when used as potential therapeutic agents.
- Published
- 2007
18. Interleukin-6 (IL-6) and low O-2 concentration (1%) synergize to improve the maintenance of hematopoietic stem cells (Pre-CFC)
- Author
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Kovačević-Filipović, Milica, Kovačević-Filipović, Milica, Petakov, Marijana, Hermitte, Francis, Debeissat, Christelle, Krstić, Aleksandra, Jovčić, Gordana, Biligarski, Dijana, Lafarge, Xavier, Milenković, Pavle B., Praloran, Vincent, Ivanović, Zoran, Kovačević-Filipović, Milica, Kovačević-Filipović, Milica, Petakov, Marijana, Hermitte, Francis, Debeissat, Christelle, Krstić, Aleksandra, Jovčić, Gordana, Biligarski, Dijana, Lafarge, Xavier, Milenković, Pavle B., Praloran, Vincent, and Ivanović, Zoran
- Abstract
Low O-2 concentration (1%) favors the self-renewal of hematopoietic stem cells and inhibits committed progenitors (CFC). Since IL-6 influences both stem cells and committed progenitors at 20% O-2, we studied its effects in cultures at 1% O-2. The pre-CFC activity in Lin- population of mouse bone marrow was analyzed following 10 days of serum-free culture in medium (LCI) supplemented with IL-3 with and without IL-6, at 20 and 1% O-2 and phenotypic differentiation and proliferative history monitored. The IL-6 receptor expression and initiation of VEGF-A synthesis were also investigated. At 20% O-2, the effects of IL-6 on pre-CFC were negligible but effects on CFC were apparent; conversely, at 1% O-2, the IL-6 enhances activity of pre-CFC but not of CFC. Unlike at 20% O-2, at 1% O-2 a subpopulation of cells remained Lin- in spite of extensive proliferation. However, the absolute number of Lin- cells, did not correlate with pre-CFC activity. A relative increase in VEGF transcripts at 1% O-2 in presence of IL-3 alone was enhanced by the addition of IL-6. IL-6 enhanced pre-CFC activity at 1% O-2 and this was correlated to the induction of VEGF. These data reinforce the concept that physiologically low oxygenation of bone marrow is a regulator of stem cell maintenance. Since the 20% O-2 does not exist in tissues in vivo, further studies in vitro at lower O-2 concentrations should revise our knowledge relating to cytokine effects on stem and progenitor cells.
- Published
- 2007
19. Hematopoietic changes and altered reactivity to IL-17 in Syphacia obvelata-infected mice
- Author
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Bugarski, Diana, Bugarski, Diana, Jovčić, Gordana, Katić-Radivojević, Sofija, Petakov, Marijana, Krstić, Aleksandra, Stojanović, Nevenka, Milenković, Pavle B., Bugarski, Diana, Bugarski, Diana, Jovčić, Gordana, Katić-Radivojević, Sofija, Petakov, Marijana, Krstić, Aleksandra, Stojanović, Nevenka, and Milenković, Pavle B.
- Abstract
Pinworm parasites commonly infect laboratory mice with high prevalence even in well-managed animal colonies. Although often considered as irrelevant, these parasites if undetected may significantly interfere with the experimental settings and alter the interpretation of final results. There are a few reports documenting the effects of pinworms on research and the effects of pinworms on the host hematopoiesis have not yet been investigated. In this study we examined the changes within various hematopoietic cell lineages in the bone marrow, spleen, peripheral blood and peritoneal space during naturally acquired Syphacia obvelata infection in inbred CBA mice. The data obtained showed significant hematopoietic alterations, characterized by increased myelopoiesis and erythropoiesis in S. obvelata-infected animals. In order to additionally evaluate if this pinworm infection modifies hematopoietic cells' reactivity, we examined the effect of murine interleukin-17, T cell-derived cytokine implicated in the regulation of hematopoiesis and inflammation, on the growth of bone marrow progenitor cells and demonstrated that bone marrow myeloid and erythroid progenitors from S. obvelata-infected mice displayed altered sensitivity to IL-17 when compared to non-infected controls. Taken together the alterations presented pointed out that this rodent pinworm is an important environmental agent that might significantly modify the hosts' hematopoietic response, and therefore interfere with the experimental settings and alter the interpretation of the final results. However, the results obtained also contributed new data concerning the activity of IL-17 on bone marrow hematopoietic cells, supporting our previous reports that depending on physiological/pathological status of the organism IL-17 exerts differential effects on the growth of progenitor-cells.
- Published
- 2006
20. Incidence of anti-alfa fodrin antibodies in primary sjogren syndrome and its correlation with anti ro/SS-A and anti La/SS-B antibodies
- Author
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Dokmanovic-Miletić, M. P., Dokmanovic-Miletić, M. P., Stojanović, R.M., Pajić, O., Bugarski, Diana, Marković, D., Milenković, Pavle B., Dokmanovic-Miletić, M. P., Dokmanovic-Miletić, M. P., Stojanović, R.M., Pajić, O., Bugarski, Diana, Marković, D., and Milenković, Pavle B.
- Published
- 2006
21. Effects of IL-17 on functional activity of peripheral blood cells
- Author
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Vlaški, Marija, Vlaški, Marija, Krstić, Aleksandra, Jovčić, Gordana, Bugarski, Diana, Petakov, Marijana, Stojanović, Nevenka, Milenković, Pavle B., Vlaški, Marija, Vlaški, Marija, Krstić, Aleksandra, Jovčić, Gordana, Bugarski, Diana, Petakov, Marijana, Stojanović, Nevenka, and Milenković, Pavle B.
- Abstract
Interleukin-17 (IL-17) is a proinflammatory cytokine produced mainly by activated CD4+ and CD8+ T cells, while its specific receptor is ubiquitously distributed. The inflammatory capacity of IL-17 is based on its ability to stimulate a wide range of stromal cells to produce and release a number of proinflammatory mediators, some with a known impact on hematopoiesis particularly granulopoiesis. Recent data indicate a role for IL-17 in the pathogenesis of several inflammatory diseases, transplant rejection and tumor growth. The purpose of this study was to determine functional responses including the respiratory burst, nitric oxide (NO) production, adhesiveness and metabolical activity/viability of human peripheral blood leukocytes (total white blood cells, mononuclear cells and granulocytes) from healthy donors in the presence of recombinant human (rh)IL-17. The obtained results showed that rhIL-17 did not induce significant changes in the respiratory burst, NO production, and metabolical activity of each peripheral blood cell fraction the tested, while a slight increase in phorbol-12-myristate-13-acetate (PMA) stimulated adhesiveness of granulocytes and mononuclear cells was noted. The absence of significant changes in tested functional activities of various peripheral blood cells suggests that IL-17 does not express its proinflammatory ability in steady-state, since the requirement for its action really does not exist., Interleukin 17 (IL-17) je proinflamatorni citokin koga produkuju aktivirane CD4+ i CD8+ T ćelije, dok je njegov receptor ubikvitarno distribuiran. Inflamatorni kapacitet IL-17 se zasniva na njegovoj sposobnosti da stimuliše širok spektar stromalnih ćelija da produkuju i oslobađaju različite proinflamatorne medijatore, među kojima neki imaju efekte na hematopoezu posebno granulopoezu. Dosadašnji podaci ukazuju na ulogu IL-17 u patogenezi različitih inflamatornih bolesti, odbacivanju transplanta i razvoju tumora. Cilj ovog rada je bio da se odrede funkcionalni odgovori, uključujući respiratorni prasak, produkciju azot monoksida (NO), adhezivnost i metaboličku aktivnost/vijabilnost različitih ćelija periferne krvi (ukupnih leukocita, mononuklearnih ćelija i granulocita) zdravih donora, u prisustvu IL-17. Dobijeni rezultati su ukazali da IL-17 ne dovodi do značajnih promena respiratornog praska, produkcije NO i metaboličke aktivnosti ćelija periferne krvi, ali da uzrokuje blago povećanje forbol-12-miristat-13-acetat (PMA) stimulisane adhezivnosti granulocita i mononuklearnih ćelija. Odsustvo značajnih promena u ispitivanim funkcionalnim aktivnostima različitih ćelija periferne krvi, ukazuje da IL-17 ne eksprimira proinflamatorno dejstvo kod zdravih osoba, jer najverovatnije i ne postoji potreba za njegovim delovanjem.
- Published
- 2004
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