不同持续时间的周期性压力对构建组织工程软骨的影响

目的 探讨在周期性压力条件下构建组织工程软骨每天加压的最佳持续时间.方法 构建自行没汁的生物反应器和往复式加压泵组成的"周期性压力场培养系统",将体外培养的第二代乳兔关节软骨细胞接种到聚乳酸-聚羟幕乙酸共聚物(PLGA)支架上,随机分成四组.第一、二、三组分别在每天持续时间为4、8、12 h的周期性压力(强度0～200 kPa,频率为0.1 Hz)下培养,第四组(对照组)为不加压的静态培养,各组培养时间均为2周.2周后肉眼人体观察,HE染色组织学观察工程软骨细胞增殖及分布;甲苯胺蓝染色法规察硫酸糖氨多糖(GAG)的分泌及分布,并用1,9.二甲基亚甲蓝法定量检测GAG的含接;采用Ⅱ型胶原免疫绀织化学法观察Ⅱ型胶原的分泌及分布,并用image-pro plus图像分析系统对Ⅱ型胶原染色面积行半定量分析.结果 在强度0～200 kPa,频率为0.1 Hz周期性压力作用下,8 h组支架-细胞复合体体积最大,表面光滑、有光泽、有弹性,支架内软骨细胞数量最多,排列最为规则,Ⅱ胶原和GAG含量也最高(P<0.01).结论 软骨细胞的新陈代谢受周期件压力持续时间的影响,在0～200 kPa、0.1 Hz频率作用下,每天持续8 h的周期性压力能更好地促进软骨细胞增殖,合成Ⅱ型胶原、GAG等细胞外基质.     

不同持续时间的周期性压力对构建组织工程软骨的影响

Objective To determine the best duration for exerting the cyclic pressure under which the tissue-engineered cartilage is constructed. Methods Free chondrocytes isolated from rabbit articular cartilage were seeded into polylactic acid-co-glycolic acid(PLGA) scaffolds after expansion in vitro, and ran-domized into 4 groups. In Groups 1 to 3, chondrocytes were cultured under daily cyclic pressure (0 ～ 200 kPa, 0.1Hz) for 4 hours, 8 hours, 12 hours respectively; Group 4 was a control in which no pressure was exerted. In each group, after 2 weeks of culture, the tissue engineered cartilages were observed in vitro and assessed by his-tological staining of liE. Next, the content of DNA and the secretion of type Ⅱ collagen and GAG in cartilages were detected quantitatively. Results Under the daily cyclic pressure (0 ～200 kPa, 0.1 Hz), the scaf-fold-chondrocytes complex in the group of 8 hours got the largest volume, smooth, lucidus, and elastic surface, the most queuing chondrocytes, and the highest content of type Ⅱ collagen and GAG (P < 0.01). Conclusions Since chondrecytes are baro-senstive, the metabolism of chondrocytes can be affected by the time of cyclic pressure. Under the effect of 0 ～200 kPa, 0.1Hz, the daily cyclic pressure of 8 hours may be optimal for chondrocytes to multiply and synthesize extracellular matrixes such as type Ⅱ collagen and GAG.     

不同持续时间的周期性压力对构建组织工程软骨的影响

Objective To determine the best duration for exerting the cyclic pressure under which the tissue-engineered cartilage is constructed. Methods Free chondrocytes isolated from rabbit articular cartilage were seeded into polylactic acid-co-glycolic acid(PLGA) scaffolds after expansion in vitro, and ran-domized into 4 groups. In Groups 1 to 3, chondrocytes were cultured under daily cyclic pressure (0 ～ 200 kPa, 0.1Hz) for 4 hours, 8 hours, 12 hours respectively; Group 4 was a control in which no pressure was exerted. In each group, after 2 weeks of culture, the tissue engineered cartilages were observed in vitro and assessed by his-tological staining of liE. Next, the content of DNA and the secretion of type Ⅱ collagen and GAG in cartilages were detected quantitatively. Results Under the daily cyclic pressure (0 ～200 kPa, 0.1 Hz), the scaf-fold-chondrocytes complex in the group of 8 hours got the largest volume, smooth, lucidus, and elastic surface, the most queuing chondrocytes, and the highest content of type Ⅱ collagen and GAG (P < 0.01). Conclusions Since chondrecytes are baro-senstive, the metabolism of chondrocytes can be affected by the time of cyclic pressure. Under the effect of 0 ～200 kPa, 0.1Hz, the daily cyclic pressure of 8 hours may be optimal for chondrocytes to multiply and synthesize extracellular matrixes such as type Ⅱ collagen and GAG.     

不同持续时间的周期性压力对构建组织工程软骨的影响

Objective To determine the best duration for exerting the cyclic pressure under which the tissue-engineered cartilage is constructed. Methods Free chondrocytes isolated from rabbit articular cartilage were seeded into polylactic acid-co-glycolic acid(PLGA) scaffolds after expansion in vitro, and ran-domized into 4 groups. In Groups 1 to 3, chondrocytes were cultured under daily cyclic pressure (0 ～ 200 kPa, 0.1Hz) for 4 hours, 8 hours, 12 hours respectively; Group 4 was a control in which no pressure was exerted. In each group, after 2 weeks of culture, the tissue engineered cartilages were observed in vitro and assessed by his-tological staining of liE. Next, the content of DNA and the secretion of type Ⅱ collagen and GAG in cartilages were detected quantitatively. Results Under the daily cyclic pressure (0 ～200 kPa, 0.1 Hz), the scaf-fold-chondrocytes complex in the group of 8 hours got the largest volume, smooth, lucidus, and elastic surface, the most queuing chondrocytes, and the highest content of type Ⅱ collagen and GAG (P < 0.01). Conclusions Since chondrecytes are baro-senstive, the metabolism of chondrocytes can be affected by the time of cyclic pressure. Under the effect of 0 ～200 kPa, 0.1Hz, the daily cyclic pressure of 8 hours may be optimal for chondrocytes to multiply and synthesize extracellular matrixes such as type Ⅱ collagen and GAG.     

不同持续时间的周期性压力对构建组织工程软骨的影响

Objective To determine the best duration for exerting the cyclic pressure under which the tissue-engineered cartilage is constructed. Methods Free chondrocytes isolated from rabbit articular cartilage were seeded into polylactic acid-co-glycolic acid(PLGA) scaffolds after expansion in vitro, and ran-domized into 4 groups. In Groups 1 to 3, chondrocytes were cultured under daily cyclic pressure (0 ～ 200 kPa, 0.1Hz) for 4 hours, 8 hours, 12 hours respectively; Group 4 was a control in which no pressure was exerted. In each group, after 2 weeks of culture, the tissue engineered cartilages were observed in vitro and assessed by his-tological staining of liE. Next, the content of DNA and the secretion of type Ⅱ collagen and GAG in cartilages were detected quantitatively. Results Under the daily cyclic pressure (0 ～200 kPa, 0.1 Hz), the scaf-fold-chondrocytes complex in the group of 8 hours got the largest volume, smooth, lucidus, and elastic surface, the most queuing chondrocytes, and the highest content of type Ⅱ collagen and GAG (P < 0.01). Conclusions Since chondrecytes are baro-senstive, the metabolism of chondrocytes can be affected by the time of cyclic pressure. Under the effect of 0 ～200 kPa, 0.1Hz, the daily cyclic pressure of 8 hours may be optimal for chondrocytes to multiply and synthesize extracellular matrixes such as type Ⅱ collagen and GAG.     

不同持续时间的周期性压力对构建组织工程软骨的影响

Objective To determine the best duration for exerting the cyclic pressure under which the tissue-engineered cartilage is constructed. Methods Free chondrocytes isolated from rabbit articular cartilage were seeded into polylactic acid-co-glycolic acid(PLGA) scaffolds after expansion in vitro, and ran-domized into 4 groups. In Groups 1 to 3, chondrocytes were cultured under daily cyclic pressure (0 ～ 200 kPa, 0.1Hz) for 4 hours, 8 hours, 12 hours respectively; Group 4 was a control in which no pressure was exerted. In each group, after 2 weeks of culture, the tissue engineered cartilages were observed in vitro and assessed by his-tological staining of liE. Next, the content of DNA and the secretion of type Ⅱ collagen and GAG in cartilages were detected quantitatively. Results Under the daily cyclic pressure (0 ～200 kPa, 0.1 Hz), the scaf-fold-chondrocytes complex in the group of 8 hours got the largest volume, smooth, lucidus, and elastic surface, the most queuing chondrocytes, and the highest content of type Ⅱ collagen and GAG (P < 0.01). Conclusions Since chondrecytes are baro-senstive, the metabolism of chondrocytes can be affected by the time of cyclic pressure. Under the effect of 0 ～200 kPa, 0.1Hz, the daily cyclic pressure of 8 hours may be optimal for chondrocytes to multiply and synthesize extracellular matrixes such as type Ⅱ collagen and GAG.     

不同持续时间的周期性压力对构建组织工程软骨的影响

Objective To determine the best duration for exerting the cyclic pressure under which the tissue-engineered cartilage is constructed. Methods Free chondrocytes isolated from rabbit articular cartilage were seeded into polylactic acid-co-glycolic acid(PLGA) scaffolds after expansion in vitro, and ran-domized into 4 groups. In Groups 1 to 3, chondrocytes were cultured under daily cyclic pressure (0 ～ 200 kPa, 0.1Hz) for 4 hours, 8 hours, 12 hours respectively; Group 4 was a control in which no pressure was exerted. In each group, after 2 weeks of culture, the tissue engineered cartilages were observed in vitro and assessed by his-tological staining of liE. Next, the content of DNA and the secretion of type Ⅱ collagen and GAG in cartilages were detected quantitatively. Results Under the daily cyclic pressure (0 ～200 kPa, 0.1 Hz), the scaf-fold-chondrocytes complex in the group of 8 hours got the largest volume, smooth, lucidus, and elastic surface, the most queuing chondrocytes, and the highest content of type Ⅱ collagen and GAG (P < 0.01). Conclusions Since chondrecytes are baro-senstive, the metabolism of chondrocytes can be affected by the time of cyclic pressure. Under the effect of 0 ～200 kPa, 0.1Hz, the daily cyclic pressure of 8 hours may be optimal for chondrocytes to multiply and synthesize extracellular matrixes such as type Ⅱ collagen and GAG.     

不同持续时间的周期性压力对构建组织工程软骨的影响

Objective To determine the best duration for exerting the cyclic pressure under which the tissue-engineered cartilage is constructed. Methods Free chondrocytes isolated from rabbit articular cartilage were seeded into polylactic acid-co-glycolic acid(PLGA) scaffolds after expansion in vitro, and ran-domized into 4 groups. In Groups 1 to 3, chondrocytes were cultured under daily cyclic pressure (0 ～ 200 kPa, 0.1Hz) for 4 hours, 8 hours, 12 hours respectively; Group 4 was a control in which no pressure was exerted. In each group, after 2 weeks of culture, the tissue engineered cartilages were observed in vitro and assessed by his-tological staining of liE. Next, the content of DNA and the secretion of type Ⅱ collagen and GAG in cartilages were detected quantitatively. Results Under the daily cyclic pressure (0 ～200 kPa, 0.1 Hz), the scaf-fold-chondrocytes complex in the group of 8 hours got the largest volume, smooth, lucidus, and elastic surface, the most queuing chondrocytes, and the highest content of type Ⅱ collagen and GAG (P < 0.01). Conclusions Since chondrecytes are baro-senstive, the metabolism of chondrocytes can be affected by the time of cyclic pressure. Under the effect of 0 ～200 kPa, 0.1Hz, the daily cyclic pressure of 8 hours may be optimal for chondrocytes to multiply and synthesize extracellular matrixes such as type Ⅱ collagen and GAG.     

不同持续时间的周期性压力对构建组织工程软骨的影响

Objective To determine the best duration for exerting the cyclic pressure under which the tissue-engineered cartilage is constructed. Methods Free chondrocytes isolated from rabbit articular cartilage were seeded into polylactic acid-co-glycolic acid(PLGA) scaffolds after expansion in vitro, and ran-domized into 4 groups. In Groups 1 to 3, chondrocytes were cultured under daily cyclic pressure (0 ～ 200 kPa, 0.1Hz) for 4 hours, 8 hours, 12 hours respectively; Group 4 was a control in which no pressure was exerted. In each group, after 2 weeks of culture, the tissue engineered cartilages were observed in vitro and assessed by his-tological staining of liE. Next, the content of DNA and the secretion of type Ⅱ collagen and GAG in cartilages were detected quantitatively. Results Under the daily cyclic pressure (0 ～200 kPa, 0.1 Hz), the scaf-fold-chondrocytes complex in the group of 8 hours got the largest volume, smooth, lucidus, and elastic surface, the most queuing chondrocytes, and the highest content of type Ⅱ collagen and GAG (P < 0.01). Conclusions Since chondrecytes are baro-senstive, the metabolism of chondrocytes can be affected by the time of cyclic pressure. Under the effect of 0 ～200 kPa, 0.1Hz, the daily cyclic pressure of 8 hours may be optimal for chondrocytes to multiply and synthesize extracellular matrixes such as type Ⅱ collagen and GAG.     

不同频率周期性压力对组织工程软骨的影响

目的观察不同频率周期性压力对组织工程软骨的影响。方法将兔关节软骨细胞接种到PLGA支架上，随机分成4组。A、B、C组分别在频率为0．01、0．1、1Hz的周期性压力（强度0～200kPa）下培养；D组不加压，为对照组。2周后采用组织化学，免疫组织化学法观察四组工程软骨的细胞增殖及基质分泌。并定量检测硫酸糖氨多糖（GAG）含量。结果四组之间Ⅱ型胶原染色面积百分比及GAG含量差异均有统计学意义（分别F=478．379，P〈0．01；F=67．402，P〈0．01）。其中B组工程软骨细胞最多，排列规则，Ⅱ胶原染色面积百分比[（83．95±2．23）％]和GAG含量[（476．33±25．66）μg]最高（P〈0．01）。结论周期性压力对软骨细胞的新陈代谢具有促进作用，其中0～200kPa、0．1Hz频率的周期性压力能最好的促进软骨细胞增殖，合成Ⅱ胶原、GAG等细胞外基质。     

不同强度周期性压力对组织工程软骨的影响

目的探讨不同强度的周期性压力对组织工程软骨的影响。方法将兔关节软骨细胞接种到聚乳酸-羟基乙酸（PLGA）支架上，随机分成四组，其中三组分别在压力强度为0～50kPa、0～100kPa、0～200kPa，频率0．1Hz的周期性压力下培养，另一组不加压，为对照组。2周后采用组织形态学、免疫组织化学法观察不同强度的周期性压力对组织工程软骨的结构及Ⅱ型胶原合成的影响．并定量检测组织工程软骨中氨基葡聚糖（GAG）的含量。结果在频率0．1Hz的周期性压力作用下，200kPa组支架内软骨细胞最多，排列规则，Ⅱ型胶原和GAG含量最高（P〈0．01）；100kPa组次之（P〈0．01）；50kPa组最差（P〈0．01）。三组加压组均优于对照组，差异有统计学意义（P〈0．01）。结论组织工程软骨细胞增殖和合成Ⅱ型胶原、GAG的能力受周期性压力强度的调控，其中频率0．1Hz、强度200kPa的周期性压力能更好地促进软骨细胞增殖，合成Ⅱ型胶原、GAG等细胞外基质。