Dataset Information

Skeletal Site-Specific Variations in CD11b+ Cells: Insights from Single Cell RNA Sequencing of the Mandible and Femur [ATAC-Seq]

ABSTRACT: Osteoclasts of the craniofacial region facilitate processes such as tooth eruption and jawbone development. Evidence suggests that craniofacial osteoclasts exhibit distinct characteristics compared to their counterparts within the appendicular skeleton. A biological mechanism to explain the observable difference between craniofacial osteoclasts and osteoclasts in the long bones has not been previously explored. To further understand differences that exist between the two populations, here we perform an unbiased genetic survey via single-cell RNA sequencing of CD11b+ cells from mandibular- and the femur-derived bone marrow of 2-month-old C57BL/6 mice to further understand differences that exist between the two populations in an unbiased genetic survey. Our results reveal transcriptomic evidence that suggests a uniquely inflammatory genetic profile of the mandibular-derived CD11b+ cells. This correlates with an increase in select areas of open chromatin by ATAC-Sequencing and inflammatory gene expression by RT-qPCR. Further exploration into a specific upregulated genes determined KLF4 was both necessary and important for proper differentiation in mandibular derived cells. Suppression of KLF4 likewise limited gene expression of metabolic transporter MCT2. Because this transporter is well understood to impact lactate transportation into cells, we determined that concentration of intracellular lactate was increased in mandibular derived osteoclasts. Together these data suggest a potential metabolic mechanism for the observable differences seen between craniofacial and appendicular osteoclasts.

ORGANISM(S): Mus musculus

PROVIDER: GSE285313 | GEO | 2025/06/16

REPOSITORIES: GEO

ACCESS DATA

Dataset's files

Source:

			Action	DRS
		Other

Items per page:

1 - 1 of 1

Similar Datasets

Project description:Objective: Craniofacial bone defects caused by injuries and congenital diseases are a formidable challenge to clinicians. Research has shown promise in using bone marrow mesenchymal stem cells (BM-MSCs) from limb bones for craniofacial bone regeneration; yet little is known about the potential of BM-MSCs from craniofacial bones. This study compared BM-MSCs isolated from limb and craniofacial bones in pigs, a preclinical model closely resembling humans. Design: Bone marrow was aspirated from the tibia and mandible of four-month-old pigs (n=4), followed by BM-MSC isolation, culture-expansion and confirmation by flow cytometry. Proliferation rates were compared using population doubling times. Osteogenic differentiation was evaluated by quantifying alkaline phosphatase (ALP) activity. Total mRNA was extracted from freshly isolated BM-MSCs and analyzed to compare gene expressions of tibial and mandibular BM-MSCs using an Affymetrix GeneChip porcine genome array, followed by real-time RT-PCR evaluation of two neural crest markers. Results: BM-MSCs from both locations expressed MSC markers without expression of hematopoietic markers. Mandibular BM-MSCs proliferated significantly faster than tibial BM-MSCs. Without osteogenic inducers, mandibular BM-MSC alkaline phosphatase activities were 3.3-fold greater than those of tibial origin. Microarray analysis identified 383 differentially expressed genes in mandibular and tibial BM-MSCs, including higher expression of cranial neural crest-related genes nestin and BMP-4 in mandibular BM-MSCs, a trend also confirmed by real-time RT-PCR. Among differently expressed genes, only 47 showed greater than 1.5-fold differences in expression. Conclusions: These data indicate that despite many similarities in gene expression, mandibular BM-MSCs express of number of genes differently than tibial BM-MSCs and have a phenotypic profile that may make them advantageous for craniofacial bone regeneration. Bone marrow was aspirated from the mandibular symphyseal region and the tibia of 3 pigs. Mesenchymal stem cells were isolated from the bone marrow and cultured to 80% confluence. Cells were harvested for total RNA extraction and the RNA was analyzed by Affymetrix GeneChip porcine genome array.

Project description:The Dlx homeobox genes have central roles in controlling patterning and differentiation of the brain and craniofacial primordia. In the brain, loss of Dlx function results in defects in the production, migration and differentiation of GABAergic neurons, that can lead to epilepsy. In the branchial arches, loss of Dlx function leads to craniofacial malformations that include trigeminal axon pathfinding defects. To determine how these genes function, we wish to identify the transcriptional circuitry that lies downstream of these transcription factors by comparing gene expression in wild type with Dlx mutant CNS and craniofacial tissues. 1) Compare gene expression in the maxillay branch of the first branchial arch (BA) of E10.5 wild type and Dlx2 -/- mutants. 2) Compare gene expression in the maxillary branch of the first BA of E10.5 wild type and Dlx1/2 -/- mutants. 3) Compare gene expression in wild type maxillary and mandibular branchial arches. 4) Compare gene expressionin mandibular branch of Dlx5/6 -/- mutants with wild type mandibular branch. The Dlx transcription factors are essential for controlling patterning of the brain and craniofacial primordia. In the brain, they control differentiation of GABAergic neurons of the basal ganglia. In the branchial arches, they control regional patterning. I hypothesize that there will be some conserved and some divergent mechanisms that the Dlx genes use in controlling brain and craniofacial development. We have already performed array analyses on Dlx function in the developing basal ganglia (with TGEN) by comparing expressed genes in wild type and Dlx1/2 mutants. Here we will compare gene expression in the brachial arches of wild type and Dlx mutant mice. 1) Generate E10.5 mouse embryos that are either wild type, Dlx2-/-, Dlx1/2 -/- or Dlx5/6 -/-. 2) Determine genotype by PCR. 3) Dissect branchial arches from the different genotypes. 4) Separate maxillary and mandibular branch of each branchial arch. 5) Prepare total RNA from the specimens. Obtain sufficient tissue to obtain 10 ug of total RNA - based on previous experience we anticipate that this will require ~ 10 branchial arches. We will pool the tissue from different embryos of the same genotype. 6) Send total RNA to TGEN for probe preparation, hybridization and array result analysis.

Dataset Information

Skeletal Site-Specific Variations in CD11b+ Cells: Insights from Single Cell RNA Sequencing of the Mandible and Femur [ATAC-Seq]

Dataset's files

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets