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could substantially reflect the cell adhesion ability of the designed PMs, indicating its potential

ability of tissue regeneration 50,52 .

3.2. Preparation and Characterizations of Cartilaginous and Endothelium-Osteoblastic

Microtissues

Among several constituents, chondrocytes and subchondral osteoblasts are key components

of the articular joints, in which the blood vessels are distributed in the subchondral bone, and the

endothelial cells play a pivotal role in the bone development and substantial repair of the articular

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body joints . Considering these aspects, MC3T3-E1 and MAEC were co-cultured in the PMs to
simulate the subchondral bone tissue. Different concentrations of PMs suspension were prepared

to treat the cells, and the live and dead staining fluorescence images at 24 h showed that almost no

dead cells existed, with a high cell survival rate, which indicated that PMs suspension had no

obvious toxicity to the cells at this time (Figure 3A). Furthermore, CCK-8 assay was used to detect

the cytotoxicity of PMs suspension, and the results showed that high cell viability was still observed

at the concentration of 1 mg/mL for 72 h (Figure 3B, C). Firstly, C-MTs and EO-MTs were

designed by static and dynamic co-culturing modes, respectively. The adhesion and growth of cells

on PMs were determined by nuclear staining and measurement of cell proliferation, respectively.

It was evident that the MC3T3-E1 and MAEC presented excellent proliferation efficacy with the

increased cell numbers in the initial 5 days, which could be attributed to the biocompatibility of the

PMs (Figure 4A). Additionally, the highly open and porous microstructure facilitates cellular

penetration into the PMs, while simultaneously enabling efficient substance exchange to meet the

nutritional requirements of cells within the PMs, thereby promoting cell filling within its interior.

Similarly, C-MTs also presented excellent proliferation efficacy of chondrocytes in the PMs

(Figure 4A), demonstrating that all three cell types could grow well in the PMs. Notably, the

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