Page 102 - Physico-Chemical Niche Conditions for Bone Cells
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PFF affects pre-osteoblast behavior
Cell orientation
Bin center and percentile were employed to describe potential similarity or differentia of cell orientation between static control and PFF-treated cells (Fig. 3J, K). Each angle (between the long axis of the cell and the vertical line) was approximately equally likely to occur, both in the static and PFF-treated cells. The direction of the length axis of the cells was randomly distributed in static cells as well as in cells exposed to PFF.
Short-term down-stream impact of PFF (hours)
Cell metabolic activity and ALP activity
Control cell metabolic activity did not change from 1 to 6 h post-incubation (Fig. 4A). One hour PFF-treatment followed by different post-incubation periods also did not alter cell metabolic activity compared to untreated controls at all time-points measured, i.e. at 0, 1, 3, and 6 h of post-incubation without mechanical loading (Fig. 4A). Two-way ANOVA analysis showed that there was a significant interaction effect between without and with PFF treatment (Table 1).
Treatment with 1 h PFF elicited a significant increase in ALP activity at 1 h post- incubation, but not thereafter (Fig. 4B). Two-way ANOVA analysis showed that there was a significant interaction effect between post-incubation time (0, 1, 3, and 6 h; Table 1).
Table 1. Two-way ANOVA analysis for short-term down-stream impact of PFF
Two-way ANOVA
Source of variation Interaction
Row factor
Time
Cell activity p value 0.8956 0.0001 0.8162
ALP activity p value 0.1308 0.3583 0.0306
Runx2     Ocn
Ki67
p value 0.6514 0.0001 0.9601
Fgf2
p value 0.0241 0.0088 0.0005
Dmp1
p value 0.0076 0.0201 0.0046
        p value 0.0663 0.1343 0.0319
p value 0.1634 0.0245 0.0553
              Note: Independent variables: time (post-incubation for 0, 1, 3 and 6 h) and treatment (without and with PFF).
Gene expression
The expression levels of proliferation marker gene Ki67, and osteogenesis-related genes including Runx2, Ocn, Fgf2, Mepe, Dmp1, and Col1a1 were assessed in static control and 1 h PFF-treated MC3T3-E1 pre-osteoblasts (Fig. 4). PFF increased gene expression of osteogenic markers Runx2 and Ocn at 3 h post-incubation (Fig. 4C, D). However, PFF did not change gene expression of the proliferation marker Ki67 after 1 h PFF and at 1, 3, and 6 h post-incubation (Fig. 4E). PFF significantly increased Fgf2 mRNA levels at 1 and 3 h post- incubation (Fig. 4F), but did not affect Mepe mRNA levels at 0, 1, 3, and 6 h post-incubation
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