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Microstructure Analysis and Reconstruction of a Meniscus

To analyze the characteristics of meniscus microstructure and to reconstruct a microstructure‐mimicking 3D model of the meniscus.

Methods

Human and sheep meniscus were collected and prepared for this study. Hematoxylin-eosin staining (HE) and Masson staining were conducted for histological analysis of the meniscus. For submicroscopic structure analysis, the meniscus was first freeze‐dried and then scanned by scanning electron microscopy (SEM). The porosity of the meniscus was determined according to SEM images. A micro‐MRI was used to scan each meniscus, immersed in distilled water, and a 3D digital model was reconstructed afterward. A three‐dimensional (3D) resin model was printed out based on the digital model. Before high‐resolution micro‐CT scanning, each meniscus was freeze‐dried. Then, micro‐scale two‐dimensional (2D) CT projection images were obtained. The porosity of the meniscus was calculated according to micro‐CT images. With micro‐CT, multiple 2D projection images were collected. A 3D digital model based on 2D CT pictures was also reconstructed. The 3D digital model was exported as STL format. A 3D resin model was printed by 3D printer based on the 3D digital model.1

Results

As revealed in the HE and Masson images, a meniscus is mostly composed of collagen, with a few cells disseminated between the collagen fiber bundles at the micro‐scale. The SEM image clearly shows the path of highly cross‐linked collagen fibers, and massive pores exist between the fibers. According to the SEM images, the porosity of the meniscus was 34.1% (34.1% ± 0.032%) and the diameters of the collagen fibers were varied. In addition, the cross‐linking pattern of the fibers was irregular. The scanning accuracy of micro‐MRI was 50 μm. The micro‐MRI demonstrated the outline of the meniscus, but the microstructure was obscure. The micro‐CT clearly displayed microfibers in the meniscus with a voxel size of 11.4 μm. The surface layer, lamellar layer, circumferential fibers, and radial fibers could be identified. The mean porosity of the meniscus according to micro‐CT images was 33.92% (33.92% ± 0.03%). Moreover, a 3D model of the microstructure based on the micro‐CT images was built. The microscale fibers could be displayed in the micro‐CT image and the reconstructed 3D digital model. In addition, a 3D resin model was printed out based on the 3D digital model.1

Conclusion

It is extremely difficult to artificially simulate the microstructure of the meniscus because of the irregularity of the diameter and cross‐linking pattern of fibers. The micro‐MRI images failed to demonstrate the meniscus microstructure. Freeze‐drying and micro‐CT scanning are effective methods for 3D microstructure reconstruction of the meniscus, which is an important step towards mechanically functional 3D‐printed meniscus grafts.1


2 Scanning electron microscopy (SEM) images of freeze-dried meniscus. A freeze-dried human meniscus was submitted to SEM. (A) In the SEM images, massive aligned fibers are clearly visible. At a higher level of magnification (B), varied diameters of fibers can be distinguished, and the fibers are highly cross-linked with numerus pores. The porosity of the meniscus according to SEM images was 34.1% (34.1%  0.032%), Freeze-drying of a human meniscus. The shape of the freshly harvested human meniscus was freezedried. The fresh human meniscus (A) and freeze-dried meniscus (B) are almost the same.
SEM İmages and Freeze-drying of a human meniscus

1. Zhu, S., Tong, G., Xiang, J.‐p., Qiu, S., Yao, Z., Zhou, X. and Lin, L.‐j. (2021), Microstructure Analysis and Reconstruction of a Meniscus. Orthop Surg. https://doi.org/10.1111/os.12899