Project 3

Lactosaminyl Glycans in Megakaryocytes and Thrombopoiesis

Abstract

Delayed platelet recovery, and attendant bleeding complications, is a major determinant of morbidity and mortality following myelosuppressive/myeloablative chemoradiotherapy. Hematopoiesis is critically dependent on the interplay of temporally expressed adhesion molecules that create relevant growth niches within the bone marrow. The stage-dependent display and function of distinct adhesion molecules is tightly controlled by hematopoietic cytokines and chemokines, which promote or inhibit hematopoietic stem cell (HSC) proliferation, differentiation, maturation and migration. These effects are mediated in part by modulation of cell surface lactosaminyl glycans, especially those containing terminal sialic acid and fucose modifications. Expression of these structures within hematopoietic marrow cells occurs in a stage-specific and lineage-specific fashion.  In studies to date using mice deficient in the (b1,4)-galactosyltransferase, b4GalT1, we have obtained direct evidence that lactosaminyl glycans are key regulators of thrombopoiesis. In this project, we seek to define the distinct lactosaminyl glycans expressed on mouse and human megakaryocyte progenitors (MKPs) and megakaryocytes (MKs), and the pertinent protein scaffolds and lipids that present these glycans critical for thrombopoiesis. We will define the changes in surface lactosaminyl glycans induced by thrombopoietic chemokines and cytokines or bone marrow endothelial cells (BMECs), and examine how these lactosaminyl structures mediate adhesive interactions with BMECs critical to hematopoiesis in vitro and in vivo.

To obtain knowledge translatable to practical therapeutic strategies, we will extend our studies to human cells to specifically determine how deficiency in b4GalT1 affects human thrombopoiesis and lactosaminoglycan surface expression. We will determine the pertinent protein scaffolds and lipids that present lactosaminoglycans glycans critical for thrombopoiesis in human MKs and analyze the effects of chemokines and cytokines or BMECs on lactosaminoglycan expression. We will examine how changes in surface lactosaminoglycans on human MKs affect adhesive functions with BMECs in vitro and in vivo.

These studies will address fundamental questions regarding the glycobiology of thrombopoiesis, and we will also investigate the capacity of platelets to serve as mediators of extrinsic glycosylation. The generated information will serve to develop strategies to modulate expression of key terminal lactosaminyl glycans to enhance thrombopoiesis, and, accordingly, to increase platelet production following myelosuppressive/myeloablative chemoradiotherapy and in other conditions of thrombocytopenia, including bone marrow failure states.

Delayed platelet recovery, and attendant bleeding complications, is a major determinant of morbidity and mortality following myelosuppressive/myeloablative chemoradiotherapy. Cell surface glycans, especially derivatives of the core Gal(β1,4)GlcNAc (LacNAc) such as the sialyl Lewis X (sLex) structure, are participants at key points in hematopoiesis and in hematopoietic cell function.  Studies by Dr. Sackstein (Project 1) have highlighted the importance of sLeX in migration of stem cells to the bone marrow, and others have provided evidence that sLex may be involved in hematopoietic stem cell quiescence (REF). However, contributions of glycans to thrombopoiesis remain poorly characterized. In Preliminary Results, we have observed that surface LacNAc is crucial for thrombopoiesis. In addition, we have obtained evidence that mature platelets are a rich source of glycosyltranserases and donor substrates suggesting that platelets may promote extrinsic glycosylation to regulate hematopoiesis and hematopoietic function. This study will address the gap of knowledge on the glycans displayed by MKPs, MKs and platelets, focusing on LacNAc-based structures, and their contribution to thrombopoietic regulation. We will use the mouse as a model for hematopoietic glycobiology, with validation on human cells in concert with Projects 1 and 2.

Dr. Karin Hoffmeister grew up in Poland which may have not entirely influenced her interest in medical sciences, but it certainly promoted her ability to adapt to random events as often encountered in science. She studied medicine in Düsseldorf/Germany, and trained as a cardiologist during her residency and fellowship at the Technical University Hospital in Aachen/Germany. Her clinical experience reinforced her interest in medical research, specifically in vascular biology and platelets. This interest led her to pursue a postdoctoral fellowship in the Department of Medicine at Brigham and Women’s Hospital. In 2001 she joined the medical faculty of Harvard Medical School. Since then, her research has focused on the mechanisms of platelet homeostasis, specifically related to glycans. She enjoys Yoga and biking, and dedicates the rest of her time to her husband and son, generating lots of art, books and LEGOs. 

Silvia GianniniSilvia Giannini received her Ph.D. in Biology and Molecular Biotechnology from the University of Perugia, Italy. Her post-doctoral research, under the supervision of Dr. Paolo Gresele, focused on the study of platelet pathophysiology and in particular in the study of rare congenital and acquired haemorrhagic disorders. She joined Dr. Hoffmeister’s lab in 2010 as a postdoctoral research fellow. Her current research focuses on the role of LacNAc in thrombopoiesis, specifically LacNAc structures generated by the Golgi galactosyltransferase, b4GalT1. During her free time, Silvia enjoys traveling, reading and hiking.

Dr. Melissa Lee-Sundlov is a post-doctoral research fellow with Dr. Hoffmeister interested in elucidating the role of platelet clearance in maintaining homeostasis of hematopoietic stem cells. Previously, under the tutelage of Dr. Joseph Lau in Project 2, she investigated the role of platelets and B cells in supplying key regulators of the extrinsic glycosylation process by ST6Gal-1. Her ultimate goal is to bridge the worlds of chemistry and biology: utilizing chemical and biochemical tools to study cell biology that has impact on human health. Melissa, who was born and raised in Malaysia, obtained her Ph.D. in Chemistry from the University of Buffalo under the supervision of Dr. Matthew Disney. Her doctoral research highlighted the advantage of modular assembly in enhancing small molecule-RNA affinity using a peptoid backbone to control the spacing between multivalent aminoglycosides. In her spare time, she works on improving her new ukulele skills and her tennis game.

 

Contact Information:
Karin Hoffmeister, M.D.
Brigham and Women’s Hospital
One Black Fan Circle, Karp 6
Boston, Massachusetts 02115
Office Number: 617-355-9010
Email: khoffmeister@rics.bwh.harvard.edu