Antisense oligonucleotide inhibition of acetylcholinesterase gene expression induces progenitor cell expansion and suppresses hematopoietic apoptosis ex vivo.

Abstract

To examine the role of acetylcholinesterase (EC 3.1.1.7) in hematopoietic cell proliferation and differentiation, we administered a 15-mer phosphorothioate oligonucleotide, antisense to the corresponding ACHE gene (AS-ACHE), to primary mouse bone marrow cultures. Within 2 hr of AS-ACHE addition to the culture, ACHE mRNA levels dropped by approximately 90%, as compared with those in cells treated with the "sense" oligomer, S-ACHE. Four days after AS-ACHE treatment, ACHE mRNA increased to levels 10-fold higher than in S-ACHE cultures or in fresh bone marrow. At this later time point, differential PCR display revealed significant differences between cellular mRNA transcripts in bone marrow and those in AS-ACHE- or S-ACHE-treated cultures. These oligonucleotide-triggered effects underlay considerable alterations at the cellular level: AS-ACHE but not S-ACHE increased cell counts, reflecting enhanced proliferation. In the presence of erythropoietin it also enhanced colony counts, reflecting expansion of progenitors. AS-ACHE further suppressed apoptosis-related fragmentation of cellular DNA in the progeny cells, and it diverted hematopoiesis toward production of primitive blasts and macrophages in a dose-dependent manner promoted by erythropoietin. These findings suggest that the hematopoietic role of acetylcholinesterase, anticipated to be inverse to the observed antisense effects, is to reduce proliferation of the multipotent stem cells committed to erythropoiesis and megakaryocytopoiesis and macrophage production and to promote apoptosis in their progeny. Moreover, these findings may explain the tumorigenic association of perturbations in ACHE gene expression with leukemia.

Novel Assay Utilizing Fluorochrome-Tagged Physostigmine (Ph-F) to In Situ Detect Active Acetylcholinesterase (AChE) Induced during Apoptosis

“…[4][5][6] AChE is also expressed in the hematopoietic system, within megakaryocytes and erythrocytes, where it is suspected to play a role in hematopoiesis. 7,8 While human megakaryocytes have low-and erythrocytes high-AChE expression, the reverse situation is present in rodents, where AChE is strongly expressed in megakarocytes but is essentially absent from erythrocyes. 9 It was recently reported that AChE was also expressed during apoptosis induced by a variety of agents in different cell types.…”

Morphogenetic roles of acetylcholine.

“…Inhibitory effects of acetylcholine on stem cell proliferation are indicated by the positive effects of antisense oligonucleotides for AChE on proliferation and expansion of progenitor cells in bone marrow cultures (27).…”

Morphogenetic Activities of Bendiocarb as Cholinesterase Inhibitor on Development of the Chick Embryo

“…But the full picture is more complex, since AChE also occurs in nonneural and embryonic tissues like red blood cells, megakaryocytes, and migrating neural crest cells (Brimijoin & Koenigsberger, 1999). Proposed noncholinergic roles for AChE range from neuromodulation by secreted forms to promotion of cell proliferation in tumor growth and hematopoiesis (Soreq et al, 1994). Prenatal exposure to organophosphate and carbamate pesticides could have adverse effects on neural development by interfering with the morphogenic function of AChE.…”