The zonal organization of articular cartilage is crucial in providing the tissue with mechanical properties to withstand compression and shearing force. Current treatments available for articular cartilage injury are not able to restore the hierarchically organized architecture of the tissue. Implantation of zonal chondrocyte as a multilayer tissue construct could overcome the limitation of current treatments. However, it is impeded by the lack of efficient zonal chondrocyte isolation protocol and dedifferentiation of chondrocytes during expansion on tissue culture plate (TCP). This study aims to develop a protocol to produce an adequate number of high-quality zonal chondrocytes for clinical application via size-based zonal chondrocyte separation using inertial spiral microchannel device and expansion under dynamic microcarrier culture.

Full thickness (FT) chondrocytes isolated from porcine femoral condyle cartilage were subjected to two serial of size-based sorting into three subpopulations of different cell sizes, namely small (S1), medium (S2), and large (S3) chondrocytes. Zonal phenotype of the three subpopulations was characterised. To verify the benefit of stratified zonal chondrocyte implantation in the articular cartilage regeneration, a bilayer hydrogel construct composed of S1 chondrocytes overlaying a mixture of S2 and S3 (S2S3) chondrocytes was delivered to the rat osteochondral defect model. For chondrocyte expansion, two dynamic microcarrier cultures, sort-before-expansion and sort-after-expansion, which involved expansion after or before zonal cells sorting, were studied to identify the best sort-expansion strategy.

Size-sorted zonal chondrocytes showed zone-specific characteristics in qRT-PCR with a high level of PRG4 expression in S1 and high level of aggrecan, Type II and IX collagen expression in S2 and S3. Cartilage reformation capability of sorted zonal chondrocytes in three-dimensional fibrin hydrogel showed a similar trend in qRT-PCR, histology, extracellular matrix protein quantification and mechanical compression test, indicating the zonal characteristics of S1, S2 and S3 as superficial (SZ), middle (MZ) and deep (DZ) zone chondrocytes, respectively. Implantation of bilayered zonal chondrocytes resulted in better cartilage tissue regeneration in a rat osteochondral defect model than FT control group, with predominantly Type II hyaline cartilage tissue and significantly lower Type I collagen. Dynamic microcarrier expansion of sorted zonal chondrocytes was able to retain the zonal cell size difference that correlate to zonal phenotype, while maintaining the rounded chondrocyte morphology and F-actin distribution similar to that in mature articular cartilage. With the better retention of zonal cell size and zonal phenotype relation on microcarrier, zonal cells separation was achievable in the sort-after-expansion strategy with cells expanded on microcarrier, in comparison to cells expanded on TCP.

Inertial spiral microchannel device provides a label-free and high throughput method to separate zonal chondrocytes based on cell size. Stratified implantation of zonal chondrocytes has the potential to improve articular cartilage regeneration. Dynamic microcarrier culture allows for size-based zonal chondrocyte separation to be performed on expanded chondrocytes, thus overcoming the challenge of limited tissue availability from the patients. Our novel zonal chondrocyte isolation and expansion protocol provide a translatable strategy for stratified zonal chondrocyte implantation that could improve articular cartilage regeneration of critical size defects.

A functional total knee replacement has to be well aligned, which implies that it should lie along the mechanical axis and in the correct axial and rotational planes. Incorrect alignment will lead to abnormal wear, early mechanical loosening, and patellofemoral problems. There has been increased interest of late in total knee arthroplasty with robot assistance. This study was conducted to determine if robot-assisted total knee arthroplasty is superior to the conventional surgical method with regard to the precision of implant positioning.

Twenty knee replacements of ten robot-assisted and another ten conventional operations were performed on ten cadavers. Two experienced surgeons performed the surgery. Both procedures were undertaken by one surgeon on each cadaver. The choice of which was to be done first was randomized. After the implantation of the prosthesis, the mechanical-axis deviation, femoral coronal angle, tibial coronal angle, femoral sagittal angle, tibial sagittal angle, and femoral rotational alignment were measured via three-dimensional CT scanning. These variants were then compared with the preoperative planned values.

In the robot-assisted surgery, the mechanical-axis deviation ranged from −1.94 to 2.13° (mean: −0.21°), the femoral coronal angle ranged from 88.08 to 90.99° (mean: 89.81°), the tibial coronal angle ranged from 89.01 to 92.36° (mean: 90.42°), the tibial sagittal angle ranged from 81.72 to 86.24° (mean: 83.20°), and the femoral rotational alignment ranged from 0.02 to 1.15° (mean: 0.52°) in relation to the transepicondylar axis. In the conventional surgery, the mechanical-axis deviation ranged from −3.19 to 3.84°(mean: −0.48°), the femoral coronal angle ranged from 88.36 to 92.29° (mean: 90.50°), the tibial coronal angle ranged from 88.15 to 91.51° (mean: 89.83°), the tibial sagittal angle ranged from 80.06 to 87.34° (mean: 84.50°), and the femoral rotational alignment ranged from 0.32 to 4.13° (mean: 2.76°) in relation to the transepicondylar axis. In the conventional surgery, there were two cases of outlier outside the range of 3° varus or valgus of the mechanical-axis deviation. The robot-assisted surgery showed significantly superior femoral-rotational-alignment results compared with the conventional surgery (p=0.006). There was no statistically significant difference between robot-assisted and conventional total knee arthroplasty in the other variants. All the variants were measured with high intraobserver and interobserver reliability.

In conclusion, Robot-assisted total knee arthroplasty showed excellent precision in the sagittal and coronal planes of the three-dimensional CT. Especially, better accuracy in femoral rotational alignment was shown in the robot-assisted surgery than in the conventional surgery despite the fact that the surgeons who performed the operation were more experienced and familiar with the conventional surgery than with robot-assisted surgery. It can thus be concluded that robot-assisted total knee arthroplasty is superior to the conventional total knee arthroplasty.