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Development of a humanized mouse model for the study of primary myelofibrosis

Lillo Sierras, Alba (2024) MOBN02 20242
Degree Projects in Molecular Biology
Abstract
Primary myelofibrosis (PMF) is the most severe form of myeloproliferative neoplasms (MPNs), for which allogeneic stem cell transplantation (allo-HSCT) is the only curative option. However, only a small portion of patients are eligible for it, and even if they are, there is no guarantee of success due to the many comorbidities associated with the disease (older age, splenomegaly, anemia), risk of graft rejection and graft-versus-host disease (GvHD). Thus, there is an urgent need for new drug-based therapies that increase the survival and quality of life of PMF patients. We have developed a murine xenotransplantation model based on the generation of humanized ossicles from bone marrow-derived mesenchymal stem cells (MSCs). These ossicles... (More)
Primary myelofibrosis (PMF) is the most severe form of myeloproliferative neoplasms (MPNs), for which allogeneic stem cell transplantation (allo-HSCT) is the only curative option. However, only a small portion of patients are eligible for it, and even if they are, there is no guarantee of success due to the many comorbidities associated with the disease (older age, splenomegaly, anemia), risk of graft rejection and graft-versus-host disease (GvHD). Thus, there is an urgent need for new drug-based therapies that increase the survival and quality of life of PMF patients. We have developed a murine xenotransplantation model based on the generation of humanized ossicles from bone marrow-derived mesenchymal stem cells (MSCs). These ossicles mimic the human bone marrow (BM) environment, successfully supporting the long-term engraftment and function of hematopoietic stem cells (HSPCs). The transplantation of THPO-overexpressing HSPCs into the ossicles leads to the development of a PMF-like syndrome in mice that shares key features of the human disease. THPO-overexpressing ossicles showed dysregulated hematopoietic function compared to the controls, with a marked increase in myeloproliferation and abnormal megakaryocyte production. Moreover, we observed fibre deposition and enhanced bone formation, both characteristic of PMF. Overall, our findings support the use of our PMF model for uncovering the molecular mechanisms that contribute to disease progression. Although further validation is needed, we hope that our model will contribute to the identification of new therapeutic approaches and optimization of current ones. (Less)
Popular Abstract
Generation of a humanized mouse model for the study of primary myelofibrosis.

Primary myelofibrosis (PMF) is a hematopoietic disorder characterized by the uncontrolled deposition of fibres in the bone marrow, which hinder hematopoietic function. The only cure is performing an allogeneic stem cell transplantation to replace the diseased stem cells with healthy ones, but only a small portion of patients are eligible for it. Thus, there is an urgent demand for new drug-based therapies that increase the survival and quality of life of PMF patients. To that end, we must gain a deep understanding of the molecular mechanisms responsible for the progression of this disorder. However, current in vivo approaches do not fully recapitulate the... (More)
Generation of a humanized mouse model for the study of primary myelofibrosis.

Primary myelofibrosis (PMF) is a hematopoietic disorder characterized by the uncontrolled deposition of fibres in the bone marrow, which hinder hematopoietic function. The only cure is performing an allogeneic stem cell transplantation to replace the diseased stem cells with healthy ones, but only a small portion of patients are eligible for it. Thus, there is an urgent demand for new drug-based therapies that increase the survival and quality of life of PMF patients. To that end, we must gain a deep understanding of the molecular mechanisms responsible for the progression of this disorder. However, current in vivo approaches do not fully recapitulate the development of the disease, leaving an unmet need for a human-relevant PMF model. Our approach is based on the generation of humanized ossicles derived from bone marrow mesenchymal stem cells, which can support the engraftment and function of human hematopoietic stem cells (HSPCs). The injection of thrombopoietin-overexpressing HSPCs led to the development of a PMF-like syndrome that shared key features with the human PMF phenotype. Hopefully, our model will contribute to the identification and validation of new drugs that halt myelofibrosis development.

To establish the humanized bone marrow niche in mice, human MSCs isolated from the bone marrow of young donors were expanded, mixed with a synthetic matrix (Matrigel), and injected under the skin of immunodeficient mice. Once the MSCs had differentiated into hematopoietic-supporting cells, human HSPCs (overexpressing THPO or not) were transplanted into the ossicle cavities. After 4 to 10 weeks, mice were euthanized, and ossicles were retrieved for analysis.

We first confirmed that the humanized bone marrow niche successfully supported the long-term survival and hematopoietic function of the transplanted human HSPCs. Flow cytometry and histology analysis revealed the presence of human primitive hematopoietic stem cells and the corresponding descendant populations (myeloid and lymphoid lineages). Next, we assessed whether the model would faithfully recapitulate the most important aspects of PMF´s pathology. THPO-overexpressing ossicles showed a marked bias towards myeloid differentiation, abnormal megakaryopoiesis, and megakaryocyte clustering. Regarding their inner architecture, this group showed dense reticulin networks amidst the cell population that almost obliterated the marrow cavity. Moreover, we observed an increase in bone production but defective mineralization of the matrix, all clinical features found in PMF patients.

Although more research is needed, these results support the use of our xenotransplantation model to characterize new molecular targets in a human-relevant system.

Master’s Degree Project in Biology/Molecular Biology/Bioinformatics, 45 credits, 2025
Department of Biology, Lund University

Advisor: Stefan Scheding and Hongzhe Li
Division of Molecular Hematology, Lund Stem Cell Centre (BMC, B12) (Less)
Please use this url to cite or link to this publication:
author
Lillo Sierras, Alba
supervisor
organization
course
MOBN02 20242
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
9188147
date added to LUP
2025-05-07 16:16:39
date last changed
2025-05-07 16:16:39
@misc{9188147,
  abstract     = {{Primary myelofibrosis (PMF) is the most severe form of myeloproliferative neoplasms (MPNs), for which allogeneic stem cell transplantation (allo-HSCT) is the only curative option. However, only a small portion of patients are eligible for it, and even if they are, there is no guarantee of success due to the many comorbidities associated with the disease (older age, splenomegaly, anemia), risk of graft rejection and graft-versus-host disease (GvHD). Thus, there is an urgent need for new drug-based therapies that increase the survival and quality of life of PMF patients. We have developed a murine xenotransplantation model based on the generation of humanized ossicles from bone marrow-derived mesenchymal stem cells (MSCs). These ossicles mimic the human bone marrow (BM) environment, successfully supporting the long-term engraftment and function of hematopoietic stem cells (HSPCs). The transplantation of THPO-overexpressing HSPCs into the ossicles leads to the development of a PMF-like syndrome in mice that shares key features of the human disease. THPO-overexpressing ossicles showed dysregulated hematopoietic function compared to the controls, with a marked increase in myeloproliferation and abnormal megakaryocyte production. Moreover, we observed fibre deposition and enhanced bone formation, both characteristic of PMF. Overall, our findings support the use of our PMF model for uncovering the molecular mechanisms that contribute to disease progression. Although further validation is needed, we hope that our model will contribute to the identification of new therapeutic approaches and optimization of current ones.}},
  author       = {{Lillo Sierras, Alba}},
  language     = {{eng}},
  note         = {{Student Paper}},
  title        = {{Development of a humanized mouse model for the study of primary myelofibrosis}},
  year         = {{2024}},
}