An improved explants culture method: Sustainable isolation of keloid fibroblasts with primary characteristics

Abstract

Background Keloid (KD) is a complex fibroproliferative disease, but the exact mechanisms underlying keloid pathogenesis remain to be elucidated. The primary keloid fibroblasts (KFs) culture in vitro has always been a fundamental measure to study the pathogenesis of keloid. However, the traditional primary culture methods have some limitations, such as a long culture cycle, low specimen utilization rate and so on. Aims Improve the keloid explants culture method sts. Materials & Methods We proposed an improved new “explants multiple culture method”—reusing keloid explants for primary culture and harvesting the primary KFs in specific culture times. Meanwhile, the purity, proliferation, apoptosis, migration, invasion, extracellular matrix synthesis, and some fibrosis and inflammation‐related proteins of KFs obtained from the first, fifth, and tenth explants cultures were detected. Results The results showed that the culture cycle of this new method (Cell Isolation: 2.67 ± 0.86 days, Explants removal: 8.83 ± 0.79 days, Cell Passage: 15.17 ± 1.39 days) was significantly shorter than that of the traditional method (Cell Isolation: 8.67 ± 1.84 days, Explants removal: 17.67 ± 2.17 days, Cell Passage: 22.67 ± 1.84 days). No significant difference was observed between the phenotypes of the fibroblasts obtained from the first explants culture and cultures less than 10 times (p > 0.05). Disscussion Taken together, this study provides an effective method for the primary culture of KFs with a higher specimen utilization rate and shorter culture cycle. Conclusion This method breaks through the limitation of traditional explants culture requiring a large number of keloid specimens and provides a rich source of KFs for the study of keloid.

Low‐Frequency Ultrasound Sensitive Piezo1 Channels Regulate Keloid‐Related Characteristics of Fibroblasts

“…Keloid tissues were obtained from the Department of Dermatology, Xiangya Hospital, Changsha, China (ID: 202 305 092). The protocol for PKF extraction was performed as described by He et al., [ 28 ] with several modifications (Figure 4A ). To identify the extracted cells type as fibroblasts, immunofluorescence staining of α‐SMA and F‐action were performed ( Figure 3 B,C ).…”