Bv8 and endocrine gland-derived vascular endothelial growth factor stimulate hematopoiesis and hematopoietic cell mobilization - PubMed (original) (raw)

Bv8 and endocrine gland-derived vascular endothelial growth factor stimulate hematopoiesis and hematopoietic cell mobilization

Jennifer LeCouter et al. Proc Natl Acad Sci U S A. 2004.

Abstract

Bv8 and endocrine-gland-derived VEGF (EG-VEGF), or prokineticins, are two highly related, secreted proteins that we previously described as selective angiogenic mitogens. Here we describe the expression and functional characterization of Bv8 in peripheral blood cells, notably monocytes, neutrophils, and dendritic cells, and in the bone marrow. In human and mouse, the two Bv8 G protein-coupled receptors are expressed in hematopoietic stem cells and specific mature blood cells, including lymphocytes. Bv8 is highly expressed by neutrophils at sites of inflammation and can stimulate migration of monocytes, in a pertussis toxin-sensitive manner. Bv8, or EG-VEGF that shares the same receptors, increased numbers of colony-forming units granulocytic and monocytic in cultures of human or mouse hematopoietic stem cells. Systemic in vivo exposure to Bv8 or EG-VEGF resulted in significant increases in total leukocyte, neutrophil, and monocyte counts. Additionally, adenovirus (Av)Bv8 or AvEG-VEGF delivered just before 5-fluorouracil injury promoted the survival of hematopoietic cells and enhanced progenitor mobilization. In conclusion, Bv8 can promote survival and differentiation of the granulocytic and monocytic lineages. Bv8 potentially modulates growth, survival, and function of cells of the innate and adaptive immune systems, possibly through autocrine or paracrine signaling mechanisms.

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Figures

Fig. 1.

Fig. 1.

Expression of Bv8, VEGF, and receptors in hematopoietic and lymphocytic tissues, cells, and cell lines. (A) Bv8 mRNA levels relative to GAPDH transcript in HSC, within the total population of bone marrow-derived mononuclear cells (BM-MNC), tissues, primary cells, and hematopoietic lines. (B–D) Expression of EG-VEGFR-1 and EG-VEGFR-2 (B), VEGF (C), and VEGFR-1, and VEGFR-2 (D). Transcript data were confirmed in independently prepared RNA samples, and representative results are shown. Testis RNA was used to generate all standard curves, and data were normalized to GAPDH levels.

Fig. 2.

Fig. 2.

Human Bv8 is expressed by infiltrating cells at sites of inflammation. Light field (Left) and dark field (Right) micrographs of tonsillitis (A) and appendicitis (B) samples from in situ hybridization studies with a Bv8 antisense probe. Transcript is associated with infiltrating cells, predominantly neutrophils. Sense probe did not generate specific signal (data not shown).

Fig. 3.

Fig. 3.

EG-VEGFR-1 and EG-VEGFR-2 induction and activation in monocytes. (A) Expression of Bv8, EG-VEGFR-1, and EG-VEGFR-2 in freshly isolated human monocytes (C), and cells incubated without (–) or with (+) LPS for 3 h. (B) Migration of primary monocytes in response to SDF-1 (500 ng/ml), VEGF (ng/ml), Bv8, or EG-VEGF (nM). Heat-denatured proteins did not induce migration (data not shown). (C) Migration of monocytes pretreated in media alone (–) or 200 ng/ml Ptx (+), in response to SDF-1 (500 ng/ml), VEGF (500 ng/ml), and Bv8. (D) Bv8 stimulates ERK phosphorylation in freshly isolated monocytes. Cells were preincubated in RPMI or Ptx. Representative immunoblot for phosphorylated ERK(Upper); membrane after stripping and blotting for total ERK (Lower).

Fig. 4.

Fig. 4.

Bv8 or EG-VEGF increases granulocytic and monocytic colonies in cultures of human and mouse HSCs. (A) Colony formation increased in human CD34+ HSC cultures after the addition of 20 nM Bv8 or EG-VEGF with IL-3 and IL-6. Combining 10 nM Bv8 and 1 ng/ml G-CSF increased colony formation above either condition alone in human CD34+ cultures. G-CSF (5 ng/ml) served as control. Media control (–) does not contain cytokines. (B) Colony formation in mouse Sca+Kit+Linlo cultures treated with Bv8. Complete methylcellulose media contained stem cell factor, GM-CSF, IL-3, and Epo for human; IL-3, IL-6, stem cell factor, and Epo for mouse. n = 3 for each treatment. *, P ≤ 0.05.

Fig. 5.

Fig. 5.

Bv8 or EG-VEGF increases circulating leukocytes in mice and increases peripheral cells and spleen-derived colony forming units after 5-FU injury. Peripheral leukocyte (A) and absolute neutrophil (B) counts in animals treated with Bv8, EG-VEGF, GM-CSF, or control LacZ. Leukocyte (C) and neutrophil (D) counts in 5-FU-injured animals are shown. Bv8, EG-VEGF, GM-CSF, or control LacZ was administered 12 h before cytotoxin. (E) Spleen cell number in control, LacZ, Bv8, and Fltsel mice at day 10 after 5-FU. (F) Colony formation for 1 × 104 spleen cells from control, LacZ, Bv8, and Fltsel mice. Representative experiments are shown; n = 6–10 mice per group. *, P ≤ 0.05.

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