Dysregulated angiogenesis in B-chronic lymphocytic leukemia: Morphologic, immunohistochemical, and flow cytometric evidence

John L Frater,Susan E Crawford,Charles L Goolsby,Neil E Kay,Loann C Peterson,Gordon W Dewald

doi:10.1186/1746-1596-3-16

Abstract

BackgroundThe extent of enhanced bone marrow angiogenesis in chronic lymphocytic leukemia (CLL) and relationship to proangiogenic factors and prognostic indicators is largely unexplored.MethodsTo further investigate the role of angiogenesis in CLL by evaluating the topography and extent of angiogenesis in a group of CLL bone marrow biopsies, to study the expression of pro and antiangiogenic vascular factors in CLL cells to more precisely document the cell types producing these factors, and to evaluate the role, if any, of localized hypoxia in upregulation of angiogenesis in CLL We used immunohistochemistry (IHC) (n = 21 pts) with antibodies to CD3 and CD20, proangiogenic (VEGF, HIF-1a) and antiangiogenic (TSP-1) factors, and VEGF receptors -1 and -2 to examine pattern/extent of CLL marrow involvement, microvessel density (MVD), and angiogenic characteristics; flow cytometry (FC) was performed on 21 additional cases for VEGF and TSP-1.ResultsCLL patients had higher MVD (23.8 vs 14.6, p~0.0002) compared to controls (n = 10). MVD was highest at the periphery of focal infiltrates, was not enhanced in proliferation centers, and was increased irrespective of the presence or absence of cytogenetic/immunophenotypic markers of aggressivity. By IHC, CLL cells were VEGF(+), HIF-1a (+), TSP-1(-), VEGFR-1(+), and VEGFR-2(+). By FC, CLL cells were 1.4–2.0-fold brighter for VEGF than T cells and were TSP-1(-).ConclusionCLL demonstrates enhanced angiogenesis, with increased MVD, upregulated VEGF and downregulated TSP-1. Upregulation of HIF-1a in all CLL cases suggests localized tissue hypoxia as an important stimulant of microvessel proliferation. The presence of VEGF receptors on CLL cells implies an autocrine effect for VEGF. Differences in MVD did not correlate with traditional genetic/immunophenotypic markers of aggressiveness.

Highlights

Angiogenesis, the branching of new microvessels from pre-existent larger blood vessels, is of major importance in normal embryogenesis and in physiologic processes such as ovulation and the menstrual cycle
If any, of localized hypoxia in upregulation of angiogenesis in chronic lymphocytic leukemia (CLL), we examined the expression of the vascular endothelial growth factor (VEGF) transcriptional regulator, HIF1α
Immunohistochemistry was performed on serial sections from the core biopsies using antibodies against CD3 and CD20, CD34, p53, VEGF, Hif-1α, TSP-1, and basic fibroblast growth factor (bFGF) and

Summary

Introduction

Angiogenesis, the branching of new microvessels from pre-existent larger blood vessels, is of major importance in normal embryogenesis and in physiologic processes such as ovulation and the menstrual cycle. Diagnostic Pathology 2008, 3:16 http://www.diagnosticpathology.org/content/3/1/16 anced in favor of proangiogenic molecules. This "angiogenic switch" [1] favors the production of new microvessels, facilitating tumor growth beyond 1–2 mm diameter, and metastasis of the malignant clone. A number of molecules, including vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and hypoxia-inducible factor 1α (HIF-1α) have been identified as positive regulators of angiogenesis. These are kept in balance by negative regulators of angiogenesis including thrombospondin-1 (TSP-1) [2] and interferon β (IFNβ) [3]. The extent of enhanced bone marrow angiogenesis in chronic lymphocytic leukemia (CLL) and relationship to proangiogenic factors and prognostic indicators is largely unexplored

Objectives

Methods

Results

Conclusion