Screening method and application of direct mechanical response cell subpopulation

Abstract

The invention discloses a screening method and application of a direct mechanical response cell subpopulation. The screened cell subpopulation comprises a Fat <4+> subpopulation, a Fgfr <3 > subpopulation and a Scara <5+> subpopulation, and the invention also discloses application of the mechanical response cell subpopulation in preparation of drugs for local bone defect repair and regeneration and stress-related bone metabolic diseases. Direct mechanical response cells are screened through a single-cell sequencing technology, fine analysis of a cell gene expression profile under stress stimulation is carried out through a single-cell transcriptome level, a key cell subpopulation with fate change under stress regulation is searched for,and in-vitro flow type sorting and in-vitro stress loading are carried out through marker genes. In combination with construction of a CreER-loxP inducible reporter gene system, in-vivo lineage tracing is performed to verify the mechanical responsivenessof the cells, so that the purpose of identifying the direct mechanical response cells is achieved.

Description

technical field [0001] The invention belongs to the field of biology, and in particular relates to a screening method and application of direct mechanical response cell subgroups. Background technique [0002] Biomechanical phenomena widely exist in multiple organs and tissues of the body, such as skin, heart, alveoli, and bones. The skeletal system is the "reinforced iron bone" that protects and supports the human body, and is responsible for the body's tolerance, perception and transduction of stress stimuli. The skeletal system senses mechanical stress and adjusts its structure to environmental changes in response to mechanical stimuli. Many clinical phenomena such as distraction osteogenesis, fracture repair, orthodontic tooth movement, and weightlessness-induced osteoporosis indicate that mechanical stress is an important factor in regulating bone metabolism and remodeling. Therefore, revealing the mechanical response mechanism of bone tissue from the molecular level ...

AI Technical Summary

Problems solved by technology

①In vitro stress loading mostly adopts the method of direct in vitro stress loading on osteoblast cell lines such as C3H10T1 / 2 and MC3T3-E1, but the cell lines cannot reflect the mechanical response of cells in vivo

②Other studies used primary BMSCs to add stress, but BMSCs themselves are a mixed cell population containing multiple differentiation states, different lineage differentiation tendencies, and different functions. Under stress, different BMSCs subpopulations will have different response states and Functional effects, where true mechanoresponsive subpopulations could not be identified

③In addition to BMSCs, some scholars believe that mature osteoblasts or osteocytes in the skeletal system may also be mechanically sensitive cells, but it is difficult to directly obtain homogeneous primary osteoblasts and osteocytes by traditional methods, and it is still impossible to clearly identify the presence of mixed populations. The true mechano-responsive cells of

④ Model animal research uses the Cre-loxP system to construct specific fluorescent expression mice for lineage tracing to find mechanically responsive cells. However, due to the complexity of the Cre expression profile and the heterogeneity of most cells, the mechanical response cannot be accurately identified cell subpopulation

[0006] In summary, the existing technologies are essentially unable to screen and identify the true direct mechanical response cell subpopulations from mixed cell populations

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