bloodjourГематология и трансфузиологияRussian journal of hematology and transfusiology0234-57302411-3042ООО Издательский дом «Практика»10.18821/0234-5730/2016-61-3-126-133bloodjour-18Research ArticleОРИГИНАЛЬНЫЕ СТАТЬИORIGINAL ARTICLESИзменение уровней экспрессии генов в мультипотентных мезенхимных стромальных клетках, полученных из костного мозга больных острыми лейкозами в процессе терапииModification of gene expression in mesenchymal stromal cells of the acute myeloid leukemia patients during chemotherapyhttps://orcid.org/0000-0002-5665-7074СорокинаТ. В.SorokinaT. V.

Сорокина Тамара Викторовна, врач отделения высокодозной химиотерапии гемобластозов, депрессий кроветворения и трансплантации костного мозга

125167, г. Москва

 

Sorokina Tamara V., MD, Doctor of the Department of High-Dose Chemotherapy, Depressions of Hemopoiesis, and Bone Marrow Transplantation

Moscow, 125167

tamarasorokina0@yandex.ruhttps://orcid.org/0000-0003-1189-0283ШипуноваИ. Н.ShipunovaI. N.

125167, г. Москва

125167, Moscow

https://orcid.org/0000-0003-0215-9085БигильдеевА. Е.BigildeevA. E.

125167, г. Москва

125167, Moscow

https://orcid.org/0000-0002-7150-0403ДризеН. И.DrizeN. I.

125167, г. Москва

125167, Moscow

https://orcid.org/0000-0001-6201-6276КузьминаЛ. А.KuzminaL. A.

125167, г. Москва

125167, Moscow

https://orcid.org/0000-0001-6177-3566ПаровичниковаЕ. Н.ParovichnikovaE. N.

125167, г. Москва

125167, Moscow

https://orcid.org/0000-0001-8188-5557СавченкоВ. Г.SavchenkoV. G.

125167, г. Москва

125167, Moscow

ФГБУ «Гематологический научный центр» Минздрава РоссииРоссияNational Research Center for HematologyRussian Federation201626022019613126133Copyright © Сорокина Т.В., Шипунова И.Н., Бигильдеев А.Е., Дризе Н.И., Кузьмина Л.А., Паровичникова Е.Н., Савченко В.Г., 20192019Сорокина Т.В., Шипунова И.Н., Бигильдеев А.Е., Дризе Н.И., Кузьмина Л.А., Паровичникова Е.Н., Савченко В.Г.Sorokina T.V., Shipunova I.N., Bigildeev A.E., Drize N.I., Kuzmina L.A., Parovichnikova E.N., Savchenko V.G.Данная работа распространяется под лицензией Creative Commons Attribution 4.0.This work is licensed under a Creative Commons Attribution 4.0 License.https://www.htjournal.ru/jour/article/view/18

Цель исследования – охарактеризовать уровень экспрессии генов в мультипотентных мезенхимных стромальных клетках (ММСК), полученных из костного мозга больных острым лейкозом в дебюте заболевания и на фоне проводимого цитостатического воздействия.

Материал и методы

Материал и методы. ММСК были получены из КМ 54 больных ОЛ. Из клеток была выделена РНК и построена кДНК. Уровень экспрессии генов оценивали методом полимеразной цепной реакции в реальном времени.

Результаты

Результаты. Продемонстрировано повышение в дебюте заболевания уровней экспрессии генов, продукты которых способствуют пролиферации и миграции лейкозных клеток (IL-6, IL-8, IL-1b, CSF, JAG1, ICAM, VCAM). В ходе терапии они снижались, но повышалась экспрессии генов, продукты которых отвечают за пролиферацию и дифференцировку ММСК (IL-1R1, PDGERa, IGF, FGFR1, FGFR2, BGLAP). У больных вне ремиссии заболевания ингибирование стромы глубже.

Aim of the study

Aim of the study. To investigate the relevant expression level in multipotent mesenchymal stromal cells (MMCs) derived from the bone marrow (BM) of acute myeloid leukemia (AML) and acute lymphoblastic leukemia (ALL) patients before and over the course of chemotherapy.

Material and Methods

Material and Methods. BM derived MMSCs from 33 AML patients and 21 ALL patients were studied before and during chemotherapy. Total RNA was extracted from the MMSCs and the cDNA was synthesized. Gene expression levels were quantified by real-time quantitative PCR (RT-qPCR) with the use of gene-specific primers.

Results

Results. Before chemotherapy, the analysis of the gene expression of MMSCs from acute leukemia patients revealed a significant increase in the relative expression level (REL) of genes (IL-6, IL-8, IL-1b, CSF, JAG1, ICAM, VCAM) which regulate leukemic cell proliferation and migration. The REL of genes regulating MMSC proliferation and differentiation (IL-1R1, PDGERa, IGF, FGFR1, FGFR2, BGLAP) increased during chemotherapy. The alterations of bone marrow stroma were more pronounced in patients who didn’t achieve remission.

мультипотентные мезенхимные стромальные клеткиострый миелоидный лейкозострый лимфобластный лейкозmultipotent mesenchymal stromal cellsacute myeloid leukemiaacute lymphoblastic leukemiaРФФИ, грант 12-04-00457Russian Foundation of Basic Research, the Grant No 12-04-00457ReferencesKrampera M., Pizzolo G., Aprili G., Franchini M. Mesenchymal stem cells for bone, cartilage, tendon and skeletal muscle repair. Bone. 2006; 39(4): 678–83.Krampera M., Pizzolo G., Aprili G., Franchini M. Mesenchymal stem cells for bone, cartilage, tendon and skeletal muscle repair. Bone. 2006; 39(4): 678–83.Caplan A. Mesenchymal stem cells. J. Orthop. Res. 1991; 9(5): 641–50.Caplan A. Mesenchymal stem cells. J. Orthop. Res. 1991; 9(5): 641–50.Méndez-Ferrer S., Michurina T.V., Ferraro F., Mazloom A.R., MacArthur B.D., Lira S.A., et al. Mesenchymal and haematopoietic stem cells form a unique bone marrow niche. Nature. 2010; 466(7308): 829–34.Méndez-Ferrer S., Michurina T.V., Ferraro F., Mazloom A.R., MacArthur B.D., Lira S.A., et al. Mesenchymal and haematopoietic stem cells form a unique bone marrow niche. Nature. 2010; 466(7308): 829–34.Bruns I., Cadeddu R.P., Brueckmann I., Frobel J., Geyh S., Bust S., et al. Multiple myeloma-related deregulation of bone marrow-derived CD34(+) hematopoietic stem and progenitor cells. Blood. 2012; 120(13) : 2620–30.Bruns I., Cadeddu R.P., Brueckmann I., Frobel J., Geyh S., Bust S., et al. Multiple myeloma-related deregulation of bone marrow-derived CD34(+) hematopoietic stem and progenitor cells. Blood. 2012; 120(13) : 2620–30.Suragani R.N., Cadena S.M., Cawley S.M., Sako D., Mitchell D., Li R., et al. Transforming growth factor-β superfamily ligand trap ACE-536 corrects anemia by promoting late-stage erythropoiesis. Nat. Med. 2014; 20(4): 408–14. doi: 10.1038/nm.3512.Suragani R.N., Cadena S.M., Cawley S.M., Sako D., Mitchell D., Li R., et al. Transforming growth factor-β superfamily ligand trap ACE-536 corrects anemia by promoting late-stage erythropoiesis. Nat. Med. 2014; 20(4): 408–14. doi: 10.1038/nm.3512.Chertkov J.L., Gurevitch O.A., Udalov G.A. Role of bone marrow stroma in hemopoietic stem cell regulation. Exp. Hematol. 1980; 8(6): 770–8.Chertkov J.L., Gurevitch O.A., Udalov G.A. Role of bone marrow stroma in hemopoietic stem cell regulation. Exp. Hematol. 1980; 8(6): 770–8.Eppert K., Takenaka K., Lechman E.R., Waldron L., Nilsson B., van Galen P., et al. Stem cell gene expression programs influence clinical outcome in human leukemia. Nat. Med. 2011; 17(9): 1086–93.Eppert K., Takenaka K., Lechman E.R., Waldron L., Nilsson B., van Galen P., et al. Stem cell gene expression programs influence clinical outcome in human leukemia. Nat. Med. 2011; 17(9): 1086–93.Gentles A.J., Plevritis S.K., Majeti R., Alizadeh A.A. Association of a leukemic stem cell gene expression signature with clinical outcomes in acute myeloid leukemia. JAMA. 2010; 304(24): 2706–15.Gentles A.J., Plevritis S.K., Majeti R., Alizadeh A.A. Association of a leukemic stem cell gene expression signature with clinical outcomes in acute myeloid leukemia. JAMA. 2010; 304(24): 2706–15.Fiedler W., Graeven U., Ergün S., Verago S., Kilic N., Stockschlader M., et al. Vascular endothelial growth factor, a possible paracrine growth factor in human acute myeloid leukemia. Blood. 1997; 89(6): 1870–5.Fiedler W., Graeven U., Ergün S., Verago S., Kilic N., Stockschlader M., et al. Vascular endothelial growth factor, a possible paracrine growth factor in human acute myeloid leukemia. Blood. 1997; 89(6): 1870–5.Aguayo A., Estey E., Kantarjian H., Mansouri T., Gidel C., Keating M., et al. Cellular vascular endothelial growth factor is a predictor of outcome in patients with acute myeloid leukemia. Blood. 1999; 94(11): 3717–21.Aguayo A., Estey E., Kantarjian H., Mansouri T., Gidel C., Keating M., et al. Cellular vascular endothelial growth factor is a predictor of outcome in patients with acute myeloid leukemia. Blood. 1999; 94(11): 3717–21.Loges S., Heil G., Bruweleit M., Schoder V., Butzal M., Fischer U., et al. Analysis of concerted expression of angiogenic growth factors in acute myeloid leukemia: expression of angiopoietin-2 represents an independent prognostic factor for overall survival. J. Clin. Oncol. 2005; 23(6): 1109–17.Loges S., Heil G., Bruweleit M., Schoder V., Butzal M., Fischer U., et al. Analysis of concerted expression of angiogenic growth factors in acute myeloid leukemia: expression of angiopoietin-2 represents an independent prognostic factor for overall survival. J. Clin. Oncol. 2005; 23(6): 1109–17.Perez-Atayde A.R., Sallan S.E., Tedrow U., Connors S., Allred E., Folkman J. Spectrum of tumor angiogenesis in the bone marrow of children with acute lymphoblastic leukemia. Am. J. Pathol. 1997; 150(3): 815–21.Perez-Atayde A.R., Sallan S.E., Tedrow U., Connors S., Allred E., Folkman J. Spectrum of tumor angiogenesis in the bone marrow of children with acute lymphoblastic leukemia. Am. J. Pathol. 1997; 150(3): 815–21.Mirshahi P., Rafii A., Vincent L., Berthaut A., Varin R., Kalantar G., et al. Vasculogenic mimicry of acute leukemic bone marrow stromal cells. Leukemia.2009; 23(6): 1039–48.Mirshahi P., Rafii A., Vincent L., Berthaut A., Varin R., Kalantar G., et al. Vasculogenic mimicry of acute leukemic bone marrow stromal cells. Leukemia.2009; 23(6): 1039–48.Colmone A., Amorim M., Pontier A.L., Wang S., Jablonski E., Sipkins D.A. Leukemic cells create bone marrow niches that disrupt the behavior of normal hematopoietic progenitor cells. Science.2008; 322(5909): 1861–5.Colmone A., Amorim M., Pontier A.L., Wang S., Jablonski E., Sipkins D.A. Leukemic cells create bone marrow niches that disrupt the behavior of normal hematopoietic progenitor cells. Science.2008; 322(5909): 1861–5.Corre J., Mahtouk K., Attal M., Gadelorge M., Huynh A. FleuryCappellesso S., et al. Bone marrow mesenchymal stem cells are abnormal in multiple myeloma. Leukemia. 2007; 21(5): 1079–88.Corre J., Mahtouk K., Attal M., Gadelorge M., Huynh A. FleuryCappellesso S., et al. Bone marrow mesenchymal stem cells are abnormal in multiple myeloma. Leukemia. 2007; 21(5): 1079–88.Roela R.A., Carraro D.M., Brentani H.P., Kaiano J.H., Simao D.F., Guarnieiro R., et al. Gene stage-specific expression in the microenvironment of pediatric myelodysplastic syndromes. Leuk. Res. 2007; 31(5): 579–89.Roela R.A., Carraro D.M., Brentani H.P., Kaiano J.H., Simao D.F., Guarnieiro R., et al. Gene stage-specific expression in the microenvironment of pediatric myelodysplastic syndromes. Leuk. Res. 2007; 31(5): 579–89.Frisch B.J., Ashton J.M., Xing L., Becker M.W., Jordan C.T., Calvi L.M. Functional inhibition of osteoblastic cells in an in vivo mouse model of myeloid leukemia. Blood. 2012; 119(2): 540–50. doi: 10.1182/blood-2011-04-348151.Frisch B.J., Ashton J.M., Xing L., Becker M.W., Jordan C.T., Calvi L.M. Functional inhibition of osteoblastic cells in an in vivo mouse model of myeloid leukemia. Blood. 2012; 119(2): 540–50. doi: 10.1182/blood-2011-04-348151.Krevvata M., Silva B.C., Manavalan J.S., Galan-Diez M., Kode A., Matthews B.G., et al. Inhibition of leukemia cell engraftment and disease progression in mice by osteoblasts. Blood. 2014; 124(18): 2834–46.Krevvata M., Silva B.C., Manavalan J.S., Galan-Diez M., Kode A., Matthews B.G., et al. Inhibition of leukemia cell engraftment and disease progression in mice by osteoblasts. Blood. 2014; 124(18): 2834–46.Geyh S., Rodríguez-Paredes M., Jager P., Khandanpour C., Cadeddu R.P., Gutekunst J., et al. Functional inhibition of mesenchymal stromal cells in acute myeloid leukemia. Leukemia. 2016; 30(3): 683–91.Geyh S., Rodríguez-Paredes M., Jager P., Khandanpour C., Cadeddu R.P., Gutekunst J., et al. Functional inhibition of mesenchymal stromal cells in acute myeloid leukemia. Leukemia. 2016; 30(3): 683–91.Hanoun M., Zhang D., Mizoguchi T., Pinho S., Pierce H., Kunisaki Y., et al. Acute myelogenous leukemia-induced sympathetic neuropathy promotes malignancy in an altered hematopoietic stem cell niche. Cell Stem Cell. 2014; 15(3): 365–75.Hanoun M., Zhang D., Mizoguchi T., Pinho S., Pierce H., Kunisaki Y., et al. Acute myelogenous leukemia-induced sympathetic neuropathy promotes malignancy in an altered hematopoietic stem cell niche. Cell Stem Cell. 2014; 15(3): 365–75.Huan J., Hornick N.I., Shurtleff M.J., Skinner A.M., Goloviznina N.A., Roberts C.T., et al. RNA trafficking by acute myelogenous leukemia exosomes.Cancer Res. 2013; 73(2): 918–29.Huan J., Hornick N.I., Shurtleff M.J., Skinner A.M., Goloviznina N.A., Roberts C.T., et al. RNA trafficking by acute myelogenous leukemia exosomes.Cancer Res. 2013; 73(2): 918–29.Calkoen F.G., Vervat C., Eising E., Vijfhuizen L.S., ’t Hoen P.B., van den Heuvel-Eibrink M.M., et al. Gene-expression and in vitro function of mesenchymal stromal cells are affected in juvenile myelomonocytic leukemia. Haematologica. 2015; 100(11): 1434–41. doi: 10.3324/haematol.2015.126938.Calkoen F.G., Vervat C., Eising E., Vijfhuizen L.S., ’t Hoen P.B., van den Heuvel-Eibrink M.M., et al. Gene-expression and in vitro function of mesenchymal stromal cells are affected in juvenile myelomonocytic leukemia. Haematologica. 2015; 100(11): 1434–41. doi: 10.3324/haematol.2015.126938.Mudry R.E., Fortney J.E., York T., Hall B.M., Gibson L.F. Stromal cells regulate survival of B-lineage leukemic cells during chemotherapy. Blood. 2000; 96(5): 1926–32.Mudry R.E., Fortney J.E., York T., Hall B.M., Gibson L.F. Stromal cells regulate survival of B-lineage leukemic cells during chemotherapy. Blood. 2000; 96(5): 1926–32.Ayala F., Dewar R., Kieran M., Kalluri R. Contribution of bone microenvironment to leukemogenesis and leukemia progression. Leukemia. 2009; 23(12): 2233–41.Ayala F., Dewar R., Kieran M., Kalluri R. Contribution of bone microenvironment to leukemogenesis and leukemia progression. Leukemia. 2009; 23(12): 2233–41.Blau O. Bone marrow stromal cells in the pathogenesis of acute myeloid leukemia. Front. Biosci. (Landmark Ed.)2014; 19: 171–80.Blau O. Bone marrow stromal cells in the pathogenesis of acute myeloid leukemia. Front. Biosci. (Landmark Ed.)2014; 19: 171–80.Konopleva M., Zhao S., Hu W., Jiang S., Snell V., Weidner D., et al. The anti-apoptotic genes Bcl-X(L) and Bcl-2 are over-expressed and contribute to chemoresistance of non-proliferating leukaemic CD34+ cells. Br. J. Haematol. 2002; 118(2): 521–34.Konopleva M., Zhao S., Hu W., Jiang S., Snell V., Weidner D., et al. The anti-apoptotic genes Bcl-X(L) and Bcl-2 are over-expressed and contribute to chemoresistance of non-proliferating leukaemic CD34+ cells. Br. J. Haematol. 2002; 118(2): 521–34.Konopleva M., Konoplev S., Hu W., Zaritskey A.Y., Afanasiev B.V., Andreeff M. Stromal cells prevent apoptosis of AML cells by upregulation of anti-apoptotic proteins. Leukemia. 2002; 16(9): 1713–24.Konopleva M., Konoplev S., Hu W., Zaritskey A.Y., Afanasiev B.V., Andreeff M. Stromal cells prevent apoptosis of AML cells by upregulation of anti-apoptotic proteins. Leukemia. 2002; 16(9): 1713–24.

The authors declare that there are no conflicts of interest present.