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High School Poster

HS Poster

Poster 93 - Inducible Postnatal Connexin 43 Deficiency and Fracture Causes Significant Systemic Decreases in Bone Apposition in Mice

Saturday, October 26, 2024
10:00 AM - 12:00 PM EST
Location: Exhibit Hall E, F & G

Introduction: Connexin 43 (Cx43) is a gap junction protein in bone that allows for intracellular communication between osteocytes (1). Previous research has found that mice with constitutive Cx43 deficiency have decreased bone mass and/or impaired fracture healing (2). It has been shown that fracture causes systemic bone loss due to accelerated and imbalanced bone remodeling (3). Osteocyte Cx43 has been shown to attenuate catabolic bone remodeling; therefore, we hypothesized that Cx43 deficiency would attenuate systemic bone loss after fracture.

Materials and

Methods: All animal procedures were approved by VCU IACUC. Skeletal mature (6 month), DMP1 CRE-ERT2 Cx43fl/fl (4,5) (Cx43 icKO) or littermate controls (WT) were given tamoxifen (75mg/kg; oral) one week prior to bone injury and twice per week thereafter till sacrifice. Bone injuries were created in right limbs under anesthesia surgically and consisted of femoral fracture, tibial cortical defect or sham surgery. All mice were given Buprenorphine ER Lab (0.5 mg/kg) throughout the first week of healing. Baseline microCT scans were taken at D0 of the study (day before surgery). After 3 weeks, endpoint microCT scans of left femurs were taken. Changes in bone parameters (D21 – D0) were analyzed for effect of treatment (Sham, Defect, Fracture), genotype (WT, Cx43 icKO), and Sex (Male, Female) by 3-WAY repeated measures ANOVA with Tukey’s multiple comparisons test (*p < 0.05).

Results, Conclusions, and Discussions: Male mice showed significant decreases in percent cortical bone area (bone area/tissue area) and percent epiphyseal bone volume (bone volume/tissue volume) due to inducible Cx43 deficiency in the sham group (Figure 1A and Figure 1B). Although statistically insignificant, visual decreases in the above metrics were seen between treatments in WT mice, with fracture and defect groups showing less bone apposition after injury compared to sham controls. Cortical thickness and trabecular thickness also had visible differences between icKO and WT mice, although statistically insignificant (data not shown). Overall, male mice gained less epiphyseal bone volume compared to female mice over our three week experimental period (Figure 1B).
These results suggest that transient Connexin 43 deficiency, especially in Male C57BL/6J mice, may lead to reduced cortical and epiphyseal bone apposition over time. In addition, trends suggest that bone injury, especially in male mice, decreases bone anabolism but we didn’t observe a protective effect of Cx43 deficiency, refuting our hypothesis. Although differences were not yet statistically significant, trends in decreases in cortical B.Ar/T.Ar, cortical thickness, epiphyseal BV/TV, and epiphyseal trabecular thickness can be seen due to sex and genotype. Increased sample size may make these differences more evident. Minimal decreases in bone volume over time, regardless of injury type, may have been due to the acute continual dosing of tamoxifen, which is a known bone anabolic factor (6), used to induce Cx43 knockout. In the future, decreased doses of the drug tamoxifen or increased intervals of dosing could reduce these effects.

Acknowledgements (Optional): This work was supported by the VCU College of Engineering Foundation Research Endowment, NASA (Grant #: 80NSSC18KK1473) and the NIH (Grant # R01AR068132). Dr. Evan Buettmann was supported by the Translational Research Institute for Space Health (TRISH) through Cooperative Agreement with NASA (NNX16AO69A) and NIAMS K99 (AR082989).

References (Optional): 1. Riquelme MA et al., Int J Mol Sci. 2020 Feb; 21(3): 1146
2. Loiselle AE et al., J Orthop Res. 2013 Jan; 31(1): 147–154.
3. Osipov B et al., Clin Rev Bone Miner Metab. 2018 Dec;16(4): 116–130.
4. Kim SW et al. J Bone Miner Res. 2012 Oct;27(10):2075-84
5. Liao Y et al. Proc Natl Acad Sci USA. 2001 Aug 14;98(17):9989-94.
6. Zannit H.M. and Silva M.J., JBMR Plus. 2019 Nov; 3(11): e10227.