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Bone marrow derived stem cells in normal physiology and in cancer. An introduction.

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Presentation on theme: "Bone marrow derived stem cells in normal physiology and in cancer. An introduction."— Presentation transcript:

1 Bone marrow derived stem cells in normal physiology and in cancer. An introduction

2 Hill, J. M. et al. NEJM 2003;348:593-600 Association between Cardiovascular Risk Factors and Endothelial-Progenitor-Cell Colony Counts

3 Survival analyzed for cardiovascular death compared to levels of CD34+, KDR+, circulating endothelial progenitor cells Werner N, Kosiol S. et al. NEJM 2005;353:999-1007

4 Intra-coronary autologous bone marrow cell transfer after myocardial infarction. The BOOST randomised controlled trial. Wollert KC, Meyer GP, et al. Lancet 2004; 364: 141-148 60 patients randomized 1:1 Given approximately 9x10 6 CD34+ cells via injection in coronary artery

5 Intra-coronary autologous bone marrow cell transfer after myocardial infarction. The BOOST randomised controlled trial. Wollert KC, Meyer GP, et al. Lancet 2004; 364: 141-148

6 Intra-coronary autologous bone marrow cell transfer after myocardial infarction. The BOOST randomised controlled trial. Wollert KC, Meyer GP, et al. Lancet 2004; 364: 141-148 4 of 5 outcome measure indicators showed no change.

7 REVIVAL2: Stem cell mobilization by granulocyte- colony stimulating factor for patients with acute myocardial infarction. A randomized, controlled trial. 114 patients randomized 1:1 Given G-CSF + standard care 10 fold increase in circulating endothelial cells after 1 week Zohlnhoefer D, Ott I, et al. JAMA 2006; 295: 1003-1010

8 REVIVAL2: Stem cell mobilization by granulocyte- colony stimulating factor for patients with acute myocardial infarction. A randomized, controlled trial. Zohlnhoefer D, Ott I, et al. JAMA 2006; 295: 1003-1010

9 Stem cells and differentiation in the hematopoietic lineage Zipori D. Stem Cells 2005; 23: 719-726

10 “Stem state: Plasticity is essential, … self renewal and hierarchy are optional.” Zipori D. Stem Cells 2005; 23: 719-726

11 Mezey, Vogelsang, et al. PNAS 2003; 100: 1364-1369 Transplanted bone marrow generates new neurons in human brains Autopsy material was studied from 4 female patients who had bone marrow transplant from male donors for leukemia and died months to years after the transplant. Each 6 micron section of brain was stained with 3 immunofluorescent reagents marking Y chromosome, neural differentiation, and cell nucleus.

12 Mezey, Vogelsang, et al. PNAS 2003; 100: 1364-1369 A 6-µm-thin section from somatosensory cortex of patient 2 demonstrates the presence of the Y chromosome depicted as red dots and viewed through a rhodamine filter. The immunostaining for the neuronal marker NeuN in green (B), and the UV filter shows all cell nuclei in blue after staining with 4',6-diamidino-2-phenylindole, a chromosomal stain (C). (D) The overlay of the three filters, where arrows point to cells that carry all markers, indicating that they derived from the donor bone marrow (Y chromosome positive) and bear the specific neuronal marker NeuN. Arrowheads point at nonneuronal donor-derived cells. (Scale bars, 10 µm.)

13 Liver from bone marrow in humans Theise ND, Krause DS, et al. Hepatology 2000; 32: 11 - 16 Liver biopsies of male recipients of liver transplants from female donors Slides examined and photographed with IHC to identify hepatocytes and cholangiocytes, then restained with FISH to identify X and Y chromosomes

14 Theise ND, Krause DS, et al. Hepatology 2000; 32: 11 - 16 Immuno for CK 8, 18, 19Red = X chromosome Turquoise = Y chromosome Blue is DAPI Liver from bone marrow in humans

15 Theise ND, Krause DS, et al. Hepatology 2000; 32: 11 - 16 Immuno for CK 8, 18, 19Red = X chromosome Turquoise = Y chromosome Blue is DAPI Liver from bone marrow in humans

16 Increased chimerism of bronchial and alveolar epithelium in human lung allografts undergoing chronic injury Kleeberger W, Versmold A, et al. Am J Path 2003; 162: 1487 - 1494 Laser-capture microdissected samples of tissue from transplanted lung in gender- mismatched cases. PCR used to quantitate X and Y chromosome to measure % chimerism.

17 Transplanted lung with chronic inflammation and squamous metaplasia Recipient genotype by short tandem repeat PCR Kleeberger W, Versmold A, et al. Am J Path 2003; 162: 1487 - 1494 Donor and recipient genotype detected Transplanted lung with normal bronchial glands Lesser degree of chimerism

18 Progenitor cells from bone marrow can migrate into blood vessels, liver, myocardium, and brain. BMDC’s can differentiate and incorporate at site of implantation. This process is more extensive if there is chronic inflammation at the site. Homing and implantation may be controlled by expression of specific integrins and may require the the stimulus of VEGF. This physiologic process has not yet been used therapeutically.

19 Biofluorescence in Aequorea victoria

20 Fluorescent proteins from Aequorea and Anthozoa do not require cofactors or ATP Intrinsic luminescence is 1,000 fold greater than luciferin Verkuosha, Lukyanov. Nature Biotechnology 2004; 22: 289-296

21 Fluorescent proteins can be transfected into a wide variety of tissues and organisms with genetic control of expression. Verkuosha, Lukyanov. Nature Biotechnology 2004; 22: 289-296

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24 Hoffman, R. Nature Reviews Cancer 2005; 5: 796-806 A metastasis in lung composed of cells from 2 different clones of fibrosarcoma

25 Lyden D, Hattori K, et al. Nature Medicine 2001; 7: 1194-1201 Id protein deficient mice are resistant to the growth of transplanted tumors Susceptibility returns with transplantation of normal bone marrow or VEGF-mobilized circulating endothelial precursor cells Id mouse model of CEP cells in tumor angiogenesis

26 Lyden D, Hattori K, et al. Nature Medicine 2001; 7: 1194-1201 Id mouse model of CEP cells in tumor angiogenesis

27 Lyden D, Hattori K, et al. Nature Medicine 2001; 7: 1194-1201 Id mouse model of CEP cells in tumor angiogenesis Transplanted LacZ + bone marrow cells forming a blood vessel in a tumor in an Id1 +/- Id3 -/- mouse

28 Lyden D, Hattori K, et al. Nature Medicine 2001; 7: 1194-1201 The effect of transplanting Id - animal with normal bone marrow is blocked if treated with antibody against VEGF receptor. Id mouse model of CEP cells in tumor angiogenesis

29 Lyden D, Hattori K, et al. Nature Medicine 2001; 7: 1194-1201 Id mouse model of CEP cells in tumor angiogenesis Tumor growth requires participation of endothelial precursor cells mobilized from the bone marrow by VEGF (vascular endothelial growth factor)

30 Jin, Aiyer, et al. J Clin Invest Mar 2006; 116: 652-662 Homing of bone-marrow derived cells in tumor angiogenesis Sections of N202 breast cancer growing in a nu/nu mouse with injected with GFP+, CD34+ bone marrow cells

31 Jin, Aiyer, et al. J Clin Invest Mar 2006; 116: 652-662 Homing of bone-marrow derived cells in tumor angiogenesis In this system, homing of bone marrow cells depends on expression of  4  1 integrin (VLA-4). The bone marrow-derived cells differentiate into endothelial cells within the growing tumor.

32 Homing of bone marrow- derived precursors cells before tumor metastasis Kaplan R N, Lyden D, et al. Nature Dec 2005; 438: 820-827

33 Homing of bone marrow- derived precursors cells before tumor metastasis Kaplan R N, Lyden D, et al. Nature Dec 2005; 438: 820-827 Lung following bone marrow transplant, before injection of melanoma cells Lung day 14 after injection of melanoma cells

34 Homing of bone marrow- derived precursors cells before tumor metastasis Kaplan R N, Lyden D, et al. Nature Dec 2005; 438: 820-827 Lung day 23 after injection of melanoma cells Lung day 18 after injection of melanoma cells

35 C57BL/6 ROSA26 eGFP Cells express GFP and non-mammalian beta- galactosidase Radiation Bone marrow transplant with male donor, female recipient

36 C57BL/6 mouse transplanted with GFP+,  -gal+ bone marrow or wild-type marrow Infected with Helicobacter Gastric muscosa assayed for GFP, beta-galactosidase

37 Immunohistochemical stain for beta-galactosidase in in dysplastic gastric glands with chronic Helicobacter infection. In C note beta-galactosidase in cytoplasm of adipocyte. Li, Houghton, et al. W. J. Gastroenterology. 2006; 12(3): 363-371

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39 Houghton, Stoicov, et al. Science. 2004; 306: 1568-71 Gastric glands with intra-epithelial neoplasia with immunofluorescence for cytokeratin (green) and beta-galactosidase (red). Wild-type marrow ROSA26,eGFP marrow

40 Houghton, Stoicov, et al. Science. 2004; 306: 1568-71 Tumor cells in gastric carcinoma (black nuclei) with Y chromosome by FISH (green) in cells with cytokeratin immunofluorescence (red).

41 » Stem cells exist in the normal bone marrow that can engraft in endothelial, mesenchymal, and epithelial tissues. » The cytokine and integrin signals that control mobilization, tissue homing, engraftment, proliferation, and differentiation are not fully known.

42 » Engraftment of normal tissues by bone marrow derived stem cells may be permissive or required for tumor metastasis. » Sites of engraftment following chronic inflammation may be vulnerable to malignant transformation. » Tumors in solid tumors may arise in stem cells in tissue derived from bone marrow.

43 Thank you. James.Strauss@usoncology.com


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