In vitro megakaryocyte culture from human bone marrow aspirates as a research and diagnostic tool

Abstract

Megakaryocytes (MKs) are relatively rare in bone marrow, comprising <0.05% of the nucleated cells, which makes direct isolation from human bone marrow impractical. As such, in vitro expansion of primary MKs from patient samples offers exciting fundamental and clinical opportunities. As most of the developed ex vivo methods require a substantial volume of biomaterial, they are not widely performed on young patients. Here we propose a simple, robust, and adapted method of primary human MK culture from 1 mL of bone marrow aspirate. Our technique uses a small volume of bone marrow per culture, uses straightforward isolation methods, and generates approximately 6 × 10⁵ mature MKs per culture. The relative high cell purity and yield achieved by this technique, combined with efficient use of low volumes of bone marrow, make this approach suitable for diagnostic and basic research of human megakaryopoiesis.

Keywords:

• Bone marrow
• human
• megakaryocyte
• proplatelet

Authorship Contribution

KRB performed experiments, analyzed data, worked with patients’ data, designed the research, and wrote the manuscript; EYO analyzed data and performed cell culture experiments; NBM performed cell culture experiments and light microscopy; NMT worked with patients, and performed bone marrow biopsies; MAP and KRM designed and supervised research, and wrote the manuscript.

Ethics Committee

The research was approved by the Center for Theoretical Problems of Physicochemical Pharmacology Ethics Committee. Prior to all collections, written informed consent was obtained from the parents.

Additional information

Funding

The work was supported by the grant from the endowment fund Doctors, Innovations, Science for Children to MAP. KRM is supported by the National Institute of Diabetes and Digestive and Kidney Diseases (K01DK111515) at the National Institutes of Health and is an American Society of Hematology Scholar.