Perfusion bioreactors have been an effective tool in bone tissue engineering. Improved nutrient delivery and the application of shear forces have stimulated osteoblast differentiation and matrix production, allowing for generation of large, clinically sized constructs. Differentiation of hypertrophic chondrocytes has been considered an alternative strategy for bone tissue engineering. We studied the effects of perfusion on hypertrophic chondrocyte differentiation, matrix production, and subsequent bone formation. Hypertrophic constructs were created by differentiation in chondrogenic medium (2 weeks) and maturation in hypertrophic medium (3 weeks). Bioreactors were customized to study a range of flow rates (0-1200 μm/s). During chondrogenic differentiation, increased flow rates correlated with cartilage matrix deposition and the presence of collagen type X. During induced hypertrophic maturation, increased flow rates correlated with bone template deposition and the increased secretion of chondroprotective cytokines. Following an 8-week implantation into the critical-size femoral defect in nude rats, nonperfused constructs displayed larger bone volume, more compact mineralized matrix, and better integration with the adjacent native bone. Therefore, although medium perfusion stimulated the formation of bone template in vitro, it failed to enhance bone regeneration in vivo. However, the promising results of the less developed template in the critical-sized defect warrant further investigation, beyond interstitial flow, into the specific environment needed to optimize hypertrophic chondrocyte-based constructs for bone repair.

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Background: Previously we demonstrated plaque lipid depletion by carotid MRI during lipid lowering. Effect of different lipid therapy on plaque lipid content was investigated. Methods: Subjects (n=182) with coronary or carotid artery disease, apoB ≥120 mg/dl, and lipid treatment history <1 year were randomized to: (1) single therapy (Rx) - atorvastatin (Atorva) (10-80 mg/day); or (2) double Rx - Atorva plus extended release-niacin (ERN) (2 g/day); or (3) triple Rx - Atorva, ERN plus colesevelam (3.8 g/day) with appropriate placebos for 2 years. These Rx lowered treatment-group average LDL-C by 38%, 29%, 45% and triglycerides by 17%, 22%, 39%, and raised HDL-C by 7%, 24%, 30%. Of the 182 subjects who underwent high-resolution, multi-contrast bilateral carotid MRI scans at baseline, 46 were identified with lipid-rich necrotic core (LRNC) and completed 2-year follow-up scans by July 2013. Cross-sectional images (10 slices on average) with 2 mm thickness were obtained and centered at the carotid bifurcation. All images were analyzed for quantifications of wall area and plaque composition while blinded to therapy, lab results and clinical course, using the previously published MRI criteria. LRNC-volume (LRNC-V) and %LRNC (LRNC area/wall area) were compared pre- and post-therapy in each of the 3 Rx groups using Mann-Whitney test. Results: Overall, the 46 subjects with measurable LRNC at baseline had a significant plaque lipid content reduction. LRNC-V decreased from 58.8±9.0 to 39.7±10.2 mm3 (p<0.001) and %LRNC from 13.3±1.0 to 8.1±1.2 (p<0.001) after 2 years of Rx. Single Rx (n=17) showed non-statistical significant reductions in LRNC-V (78.0±20.7 vs. 69.8±25.3 mm3, p=0.2) and %LRNC (14.3±1.9 vs. 11.2±2.8, p=0.2). However, double Rx (n=16) had significant reductions in LRNC-V (51.9±10.3 vs. 21.4±6.0 mm3, p<0.001) and %LRNC (13.3±1.5 vs. 6.1±1.0, p=0.001). Triple Rx (n=13) also showed significant reductions in LRNC-V (42.3±9.8 vs. 23.1±6.9 mm3, p=0.005) and %LRNC (12.1±1.9 vs. 6.6±1.9, p=0.002). Conclusions: Intensive combination lipid therapy of atorvastatin plus niacin or atorvastatin, niacin and colesevelam significantly depletes carotid plaque lipid content as assessed by MRI over 2 years.

Introduction: Postmortem coronary artery histopathology and post-surgical carotid endarterectomy studies demonstrate that women have less calcification and inflammatory cells, but more smooth muscle cells than men. Gender differences in atherosclerotic plaque composition are not well known. Yet, the age-adjusted incidence of stroke and myocardial infarction are higher in men than women. Hypothesis: We hypothesized that gender differences exist in carotid plaque composition (CPC), % lipid rich necrotic core (LRNC) and % wall volume (PWV), comparing living women and men. Methods: The CPC study is a prospective, randomized study evaluating the effect of 1) atorvastatin + placebo + placebo vs 2) atorvastatin + niacin ER + placebo 3) atorvastatin + niacin ER + colsevelam on CPC. Participants had coronary or carotid artery disease and ApoB levels ≥120 mg/dL. CPC was evaluated using MRI. Baseline PWV [(wall volume/total vessel volume) х 100%], a measure of plaque burden that adjusts for variation in artery size, and % LRNC volume (among slices with LRNC present) were evaluated. Statistical analysis used Wilcoxon rank sum test, chi-square, and multivariate linear regression. Results: There were 82 women and 118 men. Women vs. men were older, mean±SD age 58±8 vs. 55±8 yrs. (p=0.008), had higher HDL-C, 48±13 vs. 40±10 mg/dL (p<0.001), higher ApoA1 147±24 vs. 126±19 mg/dL (p<0.001), higher ApoE 5.5±3.4 vs. 4.5±1.6 mg/dL (p=0.04), lower % of prior myocardial infarction 22% vs. 52% (p=0.02) and higher % of metabolic syndrome 48% vs. 46% (p=0.01). There were no significant differences in ApoB levels, 123±32 vs. 120±29 (p=0.4). After adjusting for age, HDL-C (strongly correlated with ApoA1, r=0.89), ApoE, prevalence of myocardial infarction and metabolic syndrome, the men-women difference in PWV was small and statistically non-significant ([[Unable to Display Character: &#8710;]] 0.0%, 95% CI -2.5 to 2.6%, p=1.0). However, among participants with LRNC, men had more % LRNC than women ([[Unable to Display Character: &#8710;]] 5.9%, 95% CI 1.4 to 10.3%, p=0.01) after the same adjustments. Conclusions: Gender differences exist in CPC in that there is a higher volume of LRNC in men compared to women. Further studies are needed which delineate the mechanisms underlying the differences in prevalence of LRNC comparing men to women.

Introduction: Recent, age-adjusted stroke death rates declined greater in men than women. Whether this is related to gender differences in the atherosclerotic plaque response to therapy is not known. Postmortem coronary artery histopathology and post-surgical carotid endarterectomy studies demonstrate that women have less calcification and inflammatory cells, but more smooth muscle cells than men. Hypothesis: We hypothesized that gender differences exist in carotid plaque composition (CPC) in response to lipid lowering therapy (LLT) comparing living men and women. Methods: The CPC study is a prospective, randomized study evaluating the effect of LLT: 1) atorvastatin + placebo + placebo vs 2) atorvastatin + niacin ER + placebo 3) atorvastatin + niacin ER + colsevelam on CPC. Participants had coronary or carotid artery disease and ApoB levels ≥120 mg/dL. CPC was evaluated using MRI. The change over two years in % wall volume (PWV) [(wall volume/total vessel volume) х 100%], a measure of plaque burden that adjusts for variation in artery size, and % lipid rich necrotic core (LRNC) volume among slices with LRNC present were evaluated. Statistical analysis used Wilcoxon rank sum test, chi-square, and multivariate linear regression. Results: There were 40 women and 73 men in the study with both baseline and 2 year MRI scans. Women vs. men were older, mean±SD age 58±9 vs. 54±8 yrs. (p=0.009), had higher HDL-C, 49±14 vs. 40±11 mg/dL (p=0.002), and higher ApoA1 145±26 vs. 126±20 mg/dL (p<0.001). ApoB levels were not significantly different, 127±31 vs. 121±25 mg/dL (p=0.2). Adjusted for age, HDL-C (strongly correlated with ApoA1, r=0.89), ApoE, prevalence of MI and metabolic syndrome (statistically significant in the full baseline cohort), there were no statistically significant gender differences at 2 years with LLT in change in PWV, [[Unable to Display Character: &#8710;]] -0.1 (95% CI: -0.7, 0.6%) (p=0.8) or %LRNC among participants with LRNC, [[Unable to Display Character: &#8710;]] 0.5 (95% CI: -2.5, 3.5%) (p=0.7). Conclusions: We did not detect statistically significant gender differences in change in PWV and %LRNC in response to LLT. Although the results are consistent with no gender differences they remain inconclusive due to the small sample size. Further gender studies in the biology and treatment of arterial atherosclerosis are needed.