Qualitative analysis of trabecular bone shows inferior bone quality resulting from multi-deficiencies diet combined with bilateral ovariectomy in rats after 3

<p> <b>M.</b> (A) BV/TV showed how bone tissue is affected within the total volume; these results suggest that at 3 M the treatment results in less mineralized tissue in OVX+Diet when compared to the 3 M Sham. (B) Structure Model Index (SMI) indicated the trabecular shape change at 3 M in the OVX+Diet group. (C) Tb. N was lower in the 3 M OVX+Diet group compared to the Sham group. (D) Higher trabecular separation in the OVX+Diet group when compared to the Sham after 3 M of treatment. (* = p≤0.05, one-way ANOVA with bonferroni correction, 0 M, n = 8; 1 M, n = 3 per group; 3 M n = 8 per group).</p

µCT analysis of cortical bone parameters show affected porosity and thickness in the OVX+Diet treatment after 3 months.

<p>(A) Porosity measurement showed that both groups; the 0 M control and the 3 M Sham have significantly lower porosity than the 3 MOVX+Diet. (B) Cortical thickness was significantly higher in the 3 M Sham group when compared to either 3 M OVX+Diet or 0 M control group. (* = p≤0.05, Mann Whitney U with bonferroni correction, n = 8 per group).</p

Unbalanced cellular populations contribute in 3 M OVX+Diet bone alteration.

<p>(A+B) 3 M OVX+Diet showed less activity of osteoblasts at the trabecular area when compared with 3 M Sham animals suggesting a lower bone formation. (C+D) Osteoclasts number at trabecular surface of OVX+Diet showed a higher trend than in the Sham group at 3 M indicating an increased bone resorption caused by the treatment. (* = (p≤0.05), Mann Whitney U with bonferroni correction, n = 3 per group 15 microscopic fields, scale bar = 5 mm, Tb = trabecular bone, OB = osteoblasts, OC = osteoclasts).</p

Biochemical markers reflect the bone metabolism.

<p>(A–D) Opn, BGP, RANKL, and PTH concentration in serum was higher in the 3 M OVX+Diet than the 3 M Sham. (D -F) lower concentrations of TRAP5b and leptin were seen in the OVX+Diet at 3 M compared to the Sham group. (* = (p≤0.05), Mann Whitney U with bonferroni correction, n = 8 per group).</p

A chart depicting the work plan and experimental design.

<p>Female Sprague-Dawley rats were utilized for this experiment, 10 of which were sacrificed, analysis on these animals were carried out to obtain an initial bone status. Eleven animals were then ovariectomized (OVX), and another eleven rats were Sham operated (Sham) and animals were left to recover for two weeks. After recovery OVX animals were given a multi-deficiencies-diet where Sham animals received a standard diet. One month (1 M) and three months (3 M) after dietary treatment begun, 3 and 8 animals were sacrificed, respectively. Rats were scanned by DEXA at every time point and DEXA results were reported elsewhere. Besides DEXA, at 1 M only µCT analysis was performed on the left tibia. At both 0 M and 3 M Left tibia was analyzed in µCT before performing undecalcified histology; right femur and right tibia were tested biomechanically, and right femur was used for decalcified histology.</p

Biomechanical testing of tibia and femur of the ovariectomized rat after three months of multi-deficient diet treatment.

<p>(A) Ultimate torque at failure shows no significance between the groups at 3 M. (B) Tibia of treated rats showed lower torsional stiffness compared with the Sham operated rats. (C) Breaking load of the femur shows an increased needed load to break the femur of treated animals at 3 M. (D) Bending stiffness of the femur showed no significant difference between the treatment group and the Sham group. (* = p≤0.05, Mann Whitney U with bonferroni correction, n = 8 per group).</p

Weight and bone length of different groups at all time points.

<p>Weight and bone length of different groups at all time points.</p