Investigation of Cell Chirality in Cell Differentiation and Morphogenesis

細胞分化及形態發生過程中的細胞手性研究

Student thesis: Doctoral Thesis

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Award date5 Oct 2022

Abstract

Cell chirality occurs in various forms and coordinates the left-right (LR) asymmetry of the organism. The consistency of chirality plays an essential role in embryonic development and function maintenance. Therefore, it is crucial to understand the influence of chirality on stem cells during differentiation and morphogenesis.

Firstly, using micropatterned human mesenchymal stem cells (hMSCs), we report an early committed clockwise (CW) cell chirality, which upregulates adipogenic differentiation. Firstly, hMSCs exhibited a positively biased chiral orientation on the microstrips. Additionally, single cells cultured on islands coated with fibronectin (FN) exhibited an anticlockwise (ACW)- rotational bias. Interestingly, after 3-6 days of adipogenic induction, the intrinsic chirality (positive/ACW) of hMSCs was reversed to a negative chiral orientation with a clockwise (CW) rotational bias. Furthermore, an up-regulation of adipogenesis was developed by forcing the CW-biased nuclear rotation with actin inhibitor treatments of Latrunculin A (Lat A), Cytochalasin D (CD), and Nocodazole (Noco). The results suggest that chirality in early differentiation may serve as a mechano-precursor involved in differentiation fate and regulating cell differentiation and LR morphogenesis.

Secondly, by differentiating C2C12 myoblasts on micro-groove patterns with different widths ranging from 0.4 to 200 μm, we observed that wide patterns could facilitate the formation of highly aligned myotubes with better maturity and contractility, but with the unavoidable biased alignment. For exploring the reason, we further examined cell orientation, rotation, and migration on the pattern before fusion. Results showed that cells were restricted to move along with their nuclei aligned to the ultrafine grooves (0.4 and 2 μm) during measurements, as such the lateral migration and chirality of cells were inhibited. Oppositely, myoblasts could maintain their lateral migration and chirality on wide grooves during differentiation. Additionally, regulation of cell chirality and lateral migration by Lat A treatment could also inhibit the myotube formation on the wide stripe. To be concluded, the lateral migration and intrinsic chiral alignment of C2C12 myoblasts are vital for developing functional myotubes. Hence, wide groove patterns could promote myotube formation by preserving the lateral migration and chirality during myogenesis. On basis of this, we successfully achieved different myotube patterns at a large scale by arranging wide patterns with corresponding compensation angles. Consequently, our results will have significant implications for tissue rebuilding studies.

In summary, our study showed that micropatterning could help to coordinate the early establishment of chiral morphogenesis during differentiation, providing new insight into stem cell research, morphogenesis, and mechanobiology.