Altered in vitro chondrogenic properties of chondrocytes harvested from unaffected cartilage in osteoarthritic joints.

OBJECTIVE: In vitro models of chondrogenesis often depart from chondrocytes harvested from less-affected areas of osteoarthritic joints. However, there are indications that these chondrocytes are phenotypically different from chondrocytes from healthy joints and thus might differ in their capacity to generate hyaline cartilage. The goal of this study was to compare the chondrogenic capacity of chondrocytes from healthy and OA joints. DESIGN: Chondrocytes isolated from nine healthy and nine OA knee joints were expanded in monolayer for two passages. Chondrocytes from passages 1 and 2 were analyzed for expression of (de)differentiation and hypertrophy markers and were seeded at passage 2 on collagen-coated filters for redifferentiation culture to study cartilage matrix formation. RESULTS: The collagen II/I mRNA ratio, reflecting differentiation, decreased from passage 1 to 2 in both chondrocytes from OA joints and chondrocytes from healthy joints (P<0.05), without a significant difference between the two donor types. At passage 1, levels of the cartilage transcription factors Sox-5, Sox-6 and Sox-9 appeared to be higher in chondrocytes from OA joints (n.s.), but this was not seen at passage 2. However, a clear difference was observed in collagen type X expression, which was high in chondrocytes from OA joints at both passages, while undetectable in chondrocytes from healthy joints (P<0.01). Tissue generated by chondrocytes from healthy joints redifferentiated for 28 days, showed a significantly better morphology, as assessed by histological scoring (P<0.01) and higher proteoglycan content (P<0.05), compared to chondrocytes from OA joints. Matrix turnover parameters, i.e., proteoglycan synthesis and degradation rate, were not significantly affected by donor tissue origin. CONCLUSIONS: These results suggest that clear differences between chondrocytes from healthy and OA joints exist and that these are not completely abolished during the process of de- and redifferentiation. Therefore, in vitro cartilage regeneration models, which use chondrocytes from OA joints, should be interpreted with care.

Nonmuscle myosin heavy chain-B expression in balloon-dilated and stented arteries: a study in the atherosclerotic Yucatan micropig.

BACKGROUND: Restenosis after balloon angioplasty is in part due to remodelling, whereas restenosis after stenting is entirely due to neointima formation. Nonmuscle myosin heavy chain-B (NMMHC-B) is expressed by vascular smooth muscle cells and because of its overexpression in restenotic lesions after balloon angioplasty, NMMHC-B is proposed as a potential therapeutic target. Because the mechanisms underlying restenosis after balloon angioplasty or after stenting are different we hypothesised that the expression of NMMHC-B would differ in balloon-dilated versus stented arteries. METHODS: To study the localisation and time course of expression of NMMHC-B, we performed stenting or balloon dilation in peripheral arteries of 16 atherosclerotic Yucatan micropigs and used serial intravascular ultrasound (IVUS) and angiography to measure geometric dimensions following balloon angioplasty or stenting. In situ hybridisation techniques were used to detect NMMHC-B mRNA. 5'-bromo-2'-deoxyuridine (BrdU) was administered to detect proliferating cells. By counting the number of silver grains in the different layers of the artery, we could compare the amount of expression at the different time points between the groups. RESULTS: In intima and media, NMMHC-B expression increased after balloon dilation and stenting and peaked at 7 days. In stented arteries, the expression of NMMHC-B remained high for up to 42 days after injury, whereas in balloon-dilated arteries it had normalised. In the adventitia of balloon-dilated arteries, but not of stented arteries, NMMHC-B expression peaked at 7 days. NMMHC-B expression was not limited to proliferating cells. CONCLUSION: NMMHC-B is expressed near sites of active repair after arterial injury, but not limited to proliferating cells. The different pattern of NMMHC-B expression after balloon dilation compared with stenting may be related to arterial remodelling, because stented arteries that do not remodel lack this conspicuous adventitial expression at 7 days.

Effects of increasing doses of simvastatin on fasting lipoprotein subfractions, and the effect of high-dose simvastatin on postprandial chylomicron remnant clearance in normotriglyceridemic patients with premature coronary sclerosis.

Postprandial hyperlipidemia has been linked to premature coronary artery disease (CAD) in fasting normotriglyceridemic patients. We investigated the effects of increasing doses of simvastatin up to 80 mg/day on fasting and postprandial lipoprotein metabolism in 18 normotriglyceridemic patients with premature CAD. Fasting lipoprotein subfractions and cholesteryl ester transfer protein (CETP) activity were determined after each 5-week dose titration (0, 20, 40 and 80 mg/day). At baseline and after treatment with simvastatin 80 mg/day, standardised Vitamin A oral fat loading tests (50 g/m2; 10 h) were carried out. Ten normolipidemic healthy control subjects matched for gender, age and BMI underwent tests without medication. Treatment with simvastatin resulted in dose-dependent reductions of fasting LDL-cholesterol, without changing cholesterol levels in the VLDL-1, VLDL-2 and IDL fractions. In addition, simvastatin decreased CETP activity dose-dependently, although HDL-cholesterol remained unchanged. Simvastatin 80 mg/day decreased fasting plasma triglycerides (TG) by 26% (P < 0.05), but did not decrease significantly TG levels in any of the subfractions. The TG/cholesterol ratio increased in all subfractions. The plasma TG response to the oral fat loading test, estimated as area under the curve (TG-AUC), improved by 30% (from 21.5 +/- 2.5 to 15.1 +/- 1.9 mmol h/L; P < 0.01). Treatment with simvastatin 80 mg/day improved chylomicron remnant clearance (RE-AUC) by 36% from 30.0 +/- 2.6 to 19.2 +/- 3.3 mg h/L (P < 0.01). After therapy, remnant clearance in patients was similar to controls (19.2 +/- 3.3 and 20.3 +/- 2.7 mg h/L, respectively), suggesting a normalization of this potentially atherogenic process. In conclusion, high-dose simvastatin has beneficial effects in normotriglyceridemic patients with premature CAD, due to improved chylomicron remnant clearance, besides effective lowering of LDL-cholesterol. In addition, the lipoprotein subfractions became more cholesterol-poor, as reflected by the increased TG/cholesterol ratio, which potentially makes them less atherogenic.