Small cracks, cavities, and notches in perfect material can dramatically reduce the tensile strength by producing stress concentration around them [ref]. We believe that in mammalian connective bone tissues, canaliculi, osteocyte lacunae, blood vessels and muscle insertions produce stress concentration to delay the progressive spread of minute cracks. Often times the cracks that are running against lacunae, for examples, stop on entering the lacunae, Hessian canals and blood vessels [ref]. We believe that the activity at the stress concentration can stimulate local reconstruction and remodeling, ultimately leading to the repairs in bone. We hypothesized that the human bone marrow osteoclasts (hBMOCl) can sense the mechanical stimulation.
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After ten consecutive days of differentiation conditions, the hBMØ differentiated hBMCl were plated onto the Collagen I coated PDMS uniform scaffolds. The PDMS uniform scaffolds expresses the uniform stress and strain throughout scaffold during 1% stretching. The PDMS uniform scaffold was produced by a mixture of 0.9 PDMS : 0.1 crosslinked agents, cured at 60C for 3 hours before submerging in xylene to get rid of the excess crosslinked agents for another hour and then dry under the vacuum desiccator at room temperature for overnight. To sterilize the scaffold, we submerged the scaffold in the 70% ethanol overnight and air dry under the biohood. We coated the scaffold surface with 1% Collagen I and allowed the collagen to coat to the surface overnight before plating the cells. The 10,000 hBMOCl cells were seeded on the ~ 20mm x 10mm collagen coated scaffold a day prior to undergo the 1% mechanical strain (i.e. 1% at 60 cycles per minute for 1 …show more content…
We found that the OA at the stress concentration (SC) is much higher than the area outside the SC (i.e. 0.504 OD at the SC and 0.24 OD at the outside area). Using speckle technique, we found that the Trapc stained pattern of the hBMOCl at the stress concentration [Figure 2 c] was matched with the theoretical simulation [Figure 2 a] and the speckle pattern by MatLab [Figure 2 b]. We found that the CD254, which can trace the TNF-related activation induced cytokine (TRANCE), and RANK ligand (RANKL) was also upregulated at the SC (i.e. 92.81% of CD254 positive cells) more than 2.6 folds higher than the outside area (i.e. 72.43% of CD254 positive cells) [Figure 3]. The gene expression showed that upregulated expression of the osteoclast differentiation related pathway like Traf6, Src, and NFKb were more than 2 folds at the stress concentration compared to the outside areas [Figure 4]. The PI3K at the SC, however, was upregulated with ~ 1.9 folds higher than the outside areas. The upregulated expression of Traf6, Src, PI3K, and NFKb corresponding well with the Trapc staining at the stress concentration could indicate that the osteoclast activity could most likely be related to the osteoclast differentiation. The upregulated PI3K and NFKb in osteoclasts have been found to relate with the bone activity like remodeling and homeostasis [ref]. The mechanical stimulation of
After ten consecutive days of differentiation conditions, the hBMØ differentiated hBMCl were plated onto the Collagen I coated PDMS uniform scaffolds. The PDMS uniform scaffolds expresses the uniform stress and strain throughout scaffold during 1% stretching. The PDMS uniform scaffold was produced by a mixture of 0.9 PDMS : 0.1 crosslinked agents, cured at 60C for 3 hours before submerging in xylene to get rid of the excess crosslinked agents for another hour and then dry under the vacuum desiccator at room temperature for overnight. To sterilize the scaffold, we submerged the scaffold in the 70% ethanol overnight and air dry under the biohood. We coated the scaffold surface with 1% Collagen I and allowed the collagen to coat to the surface overnight before plating the cells. The 10,000 hBMOCl cells were seeded on the ~ 20mm x 10mm collagen coated scaffold a day prior to undergo the 1% mechanical strain (i.e. 1% at 60 cycles per minute for 1 …show more content…
We found that the OA at the stress concentration (SC) is much higher than the area outside the SC (i.e. 0.504 OD at the SC and 0.24 OD at the outside area). Using speckle technique, we found that the Trapc stained pattern of the hBMOCl at the stress concentration [Figure 2 c] was matched with the theoretical simulation [Figure 2 a] and the speckle pattern by MatLab [Figure 2 b]. We found that the CD254, which can trace the TNF-related activation induced cytokine (TRANCE), and RANK ligand (RANKL) was also upregulated at the SC (i.e. 92.81% of CD254 positive cells) more than 2.6 folds higher than the outside area (i.e. 72.43% of CD254 positive cells) [Figure 3]. The gene expression showed that upregulated expression of the osteoclast differentiation related pathway like Traf6, Src, and NFKb were more than 2 folds at the stress concentration compared to the outside areas [Figure 4]. The PI3K at the SC, however, was upregulated with ~ 1.9 folds higher than the outside areas. The upregulated expression of Traf6, Src, PI3K, and NFKb corresponding well with the Trapc staining at the stress concentration could indicate that the osteoclast activity could most likely be related to the osteoclast differentiation. The upregulated PI3K and NFKb in osteoclasts have been found to relate with the bone activity like remodeling and homeostasis [ref]. The mechanical stimulation of