Notch signaling and fluid shear stress in regulating osteogenic differentiation

Abstract

Osteoporosis is a common bone and metabolic disease that is characterized by bone density loss and microstructural degeneration. Human bone marrow-derived mesenchymal stem cells (hMSCs) are multipotent progenitor cells with the potential to differentiate into various cell types, including osteoblasts, chondrocytes, and adipocytes, which have been utilized extensively in the field of bone tissue engineering and cell-based therapy. Although fluid shear stress plays an important role in bone osteogenic differentiation, the cellular and molecular mechanisms underlying this effect remain poorly understood. Here, a locked nucleic acid (LNA)/DNA nanobiosensor was exploited to monitor mRNA gene expression of hMSCs that were exposed to physiologically relevant fluid shear stress to examine the regulatory role of Notch signaling during osteogenic differentiation. First, the effects of fluid shear stress on cell viability, proliferation, morphology, and osteogenic differentiation were investigated and compared. Our results showed shear stress modulates hMSCs morphology and osteogenic differentiation depending on the applied shear and duration. By incorporating this LNA/DNA nanobiosensor and alkaline phosphatase (ALP) staining, we further investigated the role of Notch signaling in regulating osteogenic differentiation. Pharmacological treatment is applied to disrupt Notch signaling to investigate the mechanisms that govern shear stress induced osteogenic differentiation. Our experimental results provide convincing evidence supporting that physiologically relevant shear stress regulates osteogenic differentiation through Notch signaling. Inhibition of Notch signaling mediates the effects of shear stress on osteogenic differentiation, with reduced ALP enzyme activity and decreased Dll4 mRNA expression. In conclusion, our results will add new information concerning osteogenic differentiation of hMSCs under shear stress and the regulatory role of Notch signaling. Further studies may elucidate the mechanisms underlying the mechanosensitive role of Notch signaling in stem cell differentiation.

Long non-coding RNA (lncRNA) MALAT1 in regulating osteogenic and adipogenic differentiation using a double-stranded gapmer locked nucleic acid nanobiosensor

“…This is different from the dsLNA/DNA probe, where the quenching efficiency was ∼97% when the quencher-to-donor ratio was 2. 7,14,46 This result indicates that the kinetic reaction between the LNA donor and the quencher depends on the length of the probe and the location of LNA monomers. It is noted that an overly abundant quantity of quencher can cause an elevation in background noise and a reduction in the signal-to-noise ratio.…”

“…Based on these efforts, a number of chemical stimuli (e.g., small bioactive molecules, growth factors, and genetic regulators) have been identified in regulating the lineage commitment of MSCs, including bone morphogenetic protein (BMP), Wnt, and Notch1-Dll4 signaling. [9][10][11] Moreover, several studies provide evidence that mechanical cues, both direct and indirect, including shear, stiffness, and topography, electrical stimulation, and acoustic tweezing cytometry (ATC), [12][13][14] play important roles in regulating stem cell fate. Moreover, it has been shown that ECM and topography enhance hMSC osteogenic differentiation by cellular tension and mechanotransduction of YAP activity.…”

Microphysiologically Engineered Vessel-Tumor Model to Investigate Vascular Transport Dynamics of Immune Cells

“…The vascular system is a natural mechanism in the immune response that heavily depends on its proper functioning. − It enables immune cells and signaling molecules to move through the blood vessels and reach sites of cancerous cell occurrence or injury, facilitating an effective immune response. Furthermore, the efficiency of immune therapy is influenced by the internal vessel network, which exhibits significant diversity between healthy and tumor tissues and the impact on immune cell recruitment. − The endothelial cell surface is typically coated with a layer of polysaccharides, which serves as the interface between the blood and the endothelium. − The endothelial glycocalyx (VGCX) influences the immune cell behavior during vascular transport, and the inherent features of VGCX are evident from experimental and computational models. − Since most leukocyte adhesion receptors (cell adhesion molecules, CAMs) are located in postcapillary venules, such as intercellular adhesion molecule (ICAM), endothelial-leukocyte adhesion molecule (E-Selectin), and P-Selectin, shedding of the venular glycocalyx may play an important role during the immune response. ,− Thus, the ability to model the structure and transport function of blood vessels in a comprehensive setting is important for understanding the physiological processes of immunotherapy. , …”

Microfluidic Droplet-Assisted Fabrication of Vessel-Supported Tumors for Preclinical Drug Discovery

“…With the continuous development of microfabrication technology, microfluidic tumor–vessel chips not only help to precisely arrange the spatial distribution of cellular components in vitro but can also apply biomechanical and biochemical stimuli, including blood flow and growth factor gradient, thereby reproducing the interaction between the heterotypic tumor and TME with high fidelity. − For example, Haase et al reported an on-chip 3D-vascularized tumor model for drug examination in a TME within a large bed of perfusable vasculature. , They demonstrated that the dose-dependent effect of anticancer drugs on tumor activity was significantly affected by the form of vascular infusion administration. Meanwhile, great strides have also been made in the field of biomaterials. , As a representative biomaterial, GelMA has been widely used in the regenerative medicine field for 3D cell culture. , Various organ models have been successfully established in GelMA, which demonstrate its good biocompatibility. ,, Antunes et al reported an in-air production of tumor spheroid-laden GelMA microgel to expedite drug screening, where GelMA was involved as tumor–ECM .…”