Related Experiment Video
Updated: Jun 15, 2025
Study of Dendritic Cell Development by Short Hairpin RNA-Mediated Gene Knockdown in a Hematopoietic Stem and Progenitor Cell Line In vitro
Published on: March 7, 2022
Heterozygous missense CSF1R variants hamper in vitro CD34+-derived dendritic cell generation but not in vivo
Shanice Beerepoot1, Nicole I Wolf2, Marjo S van der Knaap3
1Department of Child Neurology, Amsterdam Leukodystrophy Center, Emma Children's Hospital, Amsterdam University Medical Center, Amsterdam, The Netherlands; Center for Translational Immunology, University Medical Center Utrecht, Utrecht, The Netherlands; Princess Máxima Center for Pediatric Oncology, Utrecht, The Netherlands.
Insights
Pathogenic variants in colony stimulating factor 1 receptor (CSF1R) impair in vitro dendritic cell (DC) development from stem cells in CSF1R-related disorder patients. However, DC development appears normal in vivo, suggesting compensatory mechanisms.
Area of Science:
- Immunology
- Neuroscience
- Genetics
Background:
- Colony stimulating factor 1 receptor (CSF1R) signaling is crucial for myeloid cell development, including monocytes and microglia.
- CSF1R-related disorder (CSF1R-RD), caused by pathogenic CSF1R variants, is a neurological condition characterized by microglia reduction and altered monocyte function.
- The impact of CSF1R variants on human dendritic cell (DC) lineage development is not well understood.
Purpose of the Study:
- To investigate the effect of heterozygous pathogenic CSF1R variants on the human DC lineage in patients with CSF1R-related disorder.
- To compare in vitro DC differentiation potential from CD34+ stem cells and CD14+ monocytes of patients versus healthy individuals.
Main Methods:
- Peripheral blood CD34+ stem cells and CD14+ monocytes were isolated from four unrelated patients with CSF1R-RD and healthy controls.
- Cells were cultured in vitro to differentiate into conventional DCs and monocyte-derived DCs.
- Flow cytometry was used to analyze DC differentiation and subset populations.
Main Results:
- Monocytes from CSF1R-RD patients could differentiate into monocyte-derived DCs in vitro.
- CD34+ stem cell differentiation into conventional DCs in vitro was impaired in patients.
- All naturally occurring DC subsets were present in the peripheral blood of patients.
Conclusions:
- In vitro impaired DC development from stem cells in CSF1R-RD patients does not necessarily reflect in vivo DC development.
- CSF1R signaling in vivo may be compensated, allowing for normal DC development despite pathogenic variants.
- Further research is needed to elucidate the compensatory mechanisms of CSF1R signaling in vivo.
Abstract:
Colony stimulating factor 1 receptor (CSF1R) is an essential receptor for both colony stimulating factor 1 (CSF1) and interleukin (IL) 34 signaling expressed on monocyte precursors and myeloid cells, including monocytes, dendritic cells (DC), and microglia. In humans, dominant heterozygous pathogenic variants in CSF1R cause a neurological condition known as CSF1R-related disorder (CSF1R-RD), typically with late onset, previously referred to as adult-onset leukoencephalopathy with axonal spheroids and pigmented glia (ALSP). CSF1R-RD is characterized by microglia reduction and altered monocyte function; however, the impact of pathogenic CSF1R variants on the human DC lineage remains largely unknown. We previously reported that cord blood CD34+ stem cell-derived DCs generated in vitro originate specifically from CSF1R expressing precursors. In this study, we examined the DC lineage of four unrelated patients with late-onset CSF1R-RD who carried heterozygous missense CSF1R variants (c.2330G>A, c.2375C>A, c.2329C>T, and c.2381T>C) affecting different amino acids in the protein tyrosine kinase domain of CSF1R. CD34+ stem cells and CD14+ monocytes were isolated from peripheral blood and subjected to an in vitro culture protocol to differentiate towards conventional DCs and monocyte-derived DCs, respectively. Flow cytometric analysis revealed that monocytes from patients with late-onset CSF1R-RD were still able to differentiate into monocyte-derived DCs in vitro, whereas the ability of CD34+ stem cells to differentiate into conventional DCs was impaired. Strikingly, the peripheral blood of patients contained all naturally occurring DC subsets. We conclude that the in vitro abrogation of DC-development in patients with heterozygous pathogenic missense CSF1R variants does not translate to an impairment in DC development in vivo and speculate that CSF1R signalling in vivo is compensated, which needs further study.
