Three-dimensional fibroblast aggregate and in vitro 3D skin dermis model comprising same
Inventors
Kim, Sang Heon • CHOI, Jong Hoon • Park, Kwi Deok
Assignees
Interested in licensing this patent?
MTEC can help explore whether this patent might be available for licensing for your application.
Publication Number
US-10725043-B2
Publication Date
2020-07-28
Expiration Date
Abstract
Provided are a three-dimensional (3D) fibroblast cluster, a method of preparing the same, an in vitro 3D skin dermis model including a fibroblast cluster cultured from a fibroblast, and a method of screening a drug by using the in vitro 3D skin dermis model.
Core Innovation
The invention relates to producing a fibroblast cluster without relying on an artificial porous ECM scaffold. A culture broth is used in a culture container having a surface coated with a protein having fibroblast-binding activity, and fibroblasts are cultured to obtain a culture including a fibroblast cluster formed by detaching the cultured fibroblasts from the surface. The protein having fibroblast-binding activity is weaker in binding between the protein and fibroblasts than binding between fibroblasts.
The invention further prepares an in vitro three-dimensional (3D) artificial skin model by culturing fibroblasts under the same coated-surface conditions to obtain a fibroblast cluster through detachment from the coated surface. The fibroblast cluster is then further cultured from the obtained culture for at least 12 hours. This enables the 3D artificial skin model to be formed by cluster growth under weaker protein-fibroblast binding conditions.
The described approach is characterized by reduced integrin-mediated adhesion and ECM-related remodeling associated with the formed 3D fibroblast cluster, including formation of spheroidal clusters together with ECM secretion. The 3D cluster shows decreased collagen type I and increased elastin, and increased MMP-1 expression and secretion compared with 2D cultures, with collagen type I fragmentation and lower collagen gene/protein readouts.
Claims Coverage
The partial content includes two independent claims. Across these independent claims, the inventive coverage centers on forming a fibroblast cluster by detaching fibroblasts from a surface coated with a weak fibroblast-binding protein, and using the resulting cluster for an in vitro 3D artificial skin model with further culturing for at least 12 hours.
Weak fibroblast-binding coated-surface cluster formation
Culturing fibroblasts in a culture broth in a culture container having a surface coated with a protein having fibroblast-binding activity; obtaining a culture including a fibroblast cluster formed by detaching the cultured fibroblasts from the surface; and separating the fibroblast cluster from the culture, wherein binding between the protein having fibroblast-binding activity and fibroblasts is weaker than binding between fibroblasts.
3D artificial skin model using further-cultured weak-binding clusters
Culturing fibroblasts in a culture broth container having a surface coated with a protein having fibroblast-binding activity to obtain a culture including a fibroblast cluster formed by detaching the cultured fibroblasts from the surface, wherein binding between the protein having fibroblast-binding activity and fibroblasts is weaker than binding between fibroblasts; and further culturing the fibroblast cluster from the obtained culture for at least 12 hours.
Together, the independent claims cover producing a fibroblast cluster via detachment from a container surface coated with a weakly binding fibroblast-binding protein, and using that cluster to prepare an in vitro 3D artificial skin model by additional culturing for at least 12 hours.
Stated Advantages
Enables well-plate-based high-throughput drug screening using a 3D skin dermis model.
Provides rapid, high-yield cluster production.
Provides effective in vitro skin-like mimicry.
3D system is more suitable for evaluating inhibitors compared with 2D.
Documented Applications
High-throughput drug screening using a 3D skin dermis model, including evaluation of MMP inhibitors and collagen enhancers.
Evaluation of inhibitors in a drug-screening device described as a well-plate-based drug-screening culture system.
In vitro artificial skin model preparation for use as the 3D skin-like mimicry system.
Interested in licensing this patent?