羟基磷灰石/二氧化锆复合材料颗粒对骨髓间充质干细胞增殖及成骨分化的影响

Objective To evaluate the effect of the HA/ZrO2 composite particle on proliferation and osteogenesis of rabbit mesenchymal stem cells (MSCs) by using molecular biology methods.Methods The HA/ZrO2 composite particles were sintered at high temperature using the powder of HA and ZrO2 with different proportions.MSCs were isolated from rabbits and cultured.The effect of the composite particles on promoting cell proliferation of rabbit MSCs was detected using MTT method.Alkaline phosphatase activities were measured with ALP method.RT-PCR method was applied to measure the expression of collagen Ⅰ , osteocalcin and osteopontin mRNA.Results Pure HA pauicles and composite particles which contain HA promoted MSCs proliferation.Vonkossa staining showed that both pure HA particks and composite particles decreased the degree of positive stain of MSCs.ALP test showed that cell activity in the culture medium that contained pure HA particles or composite particles was significantly higher than that in the conventional culture medium and that contained pure ZrO2 particles (P ＜ 0.05＝.RT-PCR results showed that composite particles promoted the expression of collagen Ⅰ and osteocalcin gene.Conclusion The HA/ZrO2 composite particles promote proliferation and osteogenesis of MSCs.     

羟基磷灰石/二氧化锆复合材料颗粒对骨髓间充质干细胞增殖及成骨分化的影响

目的 采用分子生物学方法评价羟基磷灰石/二氧化锆(HA/ZrO2)复合材料颗粒对兔骨髓间充质干细胞(MSCs)增殖及成骨分化的影响.方法 按不同比例将纳米HA粉体与纳米ZrO2粉体混合在高温下烧结制备HA/ZrO2复合材料颗粒,分离和培养兔MSCs.MTT法检测复合材料颗粒悬液对兔MSCs增殖促进作用,ALP法测定碱性磷酸酶活性,RT-PCR检测细胞Ⅰ型胶原(collagen Ⅰ),骨钙蛋白(osteocalcin),骨桥蛋白(osteopontin)mRNA表达.结果 HA和含有HA的复合材料颗粒均对细胞增殖有一定促进作用.Vonkossa染色显示复合材料和纯HA颗粒可使细胞阳染程度降低.细胞在HA和含有HA的复合材料颗粒的成骨诱导培养液中的ALP活性均显著高于常规培养基质及纯ZrO2组(P＜0.05).RT-PCR结果显示复合材料颗粒能够促进collagen Ⅰ和osteocalcin基因的表达.结论 HA/ZrO2复合材料颗粒可促进骨髓间充质干细胞增殖及成骨分化.

Abstract：

Objective To evaluate the effect of the HA/ZrO2 composite particle on proliferation and osteogenesis of rabbit mesenchymal stem cells (MSCs) by using molecular biology methods.Methods The HA/ZrO2 composite particles were sintered at high temperature using the powder of HA and ZrO2 with different proportions.MSCs were isolated from rabbits and cultured.The effect of the composite particles on promoting cell proliferation of rabbit MSCs was detected using MTT method.Alkaline phosphatase activities were measured with ALP method.RT-PCR method was applied to measure the expression of collagen Ⅰ , osteocalcin and osteopontin mRNA.Results Pure HA pauicles and composite particles which contain HA promoted MSCs proliferation.Vonkossa staining showed that both pure HA particks and composite particles decreased the degree of positive stain of MSCs.ALP test showed that cell activity in the culture medium that contained pure HA particles or composite particles was significantly higher than that in the conventional culture medium and that contained pure ZrO2 particles (P ＜ 0.05＝.RT-PCR results showed that composite particles promoted the expression of collagen Ⅰ and osteocalcin gene.Conclusion The HA/ZrO2 composite particles promote proliferation and osteogenesis of MSCs.     

Effect of basic fibroblast growth factor and hyaluronic acid on proliferation of rabbit chondrocytes in vitro

Objective: To investigate the effect of basic fibroblast growth factor (bFGF) and hyaluronic acid (HA) on the proliferation of rabbit chondrocytes in vitro. Methods: Chondrocytes from the knee joints of New Zealand white rabbits were cultured, bFGF or HA or both were added into the culture medium respectively, and the proliferation of the ehondrocytes was measured with MTT 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl-tetra-zolium bromide. (MTT, Sigma, M2128). Results: Basic fibroblast growth factor (10ng/ml) with low concentration of fetal bovine serum in the culture medium promoted the proliferation of chondrocytes significantly, and this effect reached its maximum when concentration of bFGF reached 50ng/ml. HA itself had no effect on the proliferation of chondrocytcs. However, when bFGF was used in combination with HA, especially when the concentration of bFGF was 50-500ng/ml and that of HA was 10-50ng/ml, the effect on the proliferation of chondrocytes was much more than when bFGF or HA was used alone. Conclusions: bFGF can promote the proliferation of chondrocytes. HA, which has no effect on the proliferation of the cells, can maintain a normal growth of chondrocytcs.When bFGF is used in combination with HA, more proliferation is obtained.     

Difference of adherence, proliferation and osteogenesis of mesenchymal stem cells cultured on different HA/ZrO2 composites

Objective: To study the adherence,proliferation and osteogenesis of mesenchymal stem cells (MSCs) cultured on different HA/ZrO2 composites.Methods: The simplex and graded HA/ZrO2 composites were prepar...     

大鼠脂肪干细胞复合胶原-壳聚糖-硫酸软骨素三维支架构建组织工程软骨

Objective To evaluate the character of the collagen-chitosan-chondroitin sulfate scaffold seeded with rat adipose tissue-derived stromal cells. Methods A dipose tissue were harvested from 6 weeks old Wistar rats and the stromal cells were harvested by type Ⅰ collagenase and then cultured in vitro. Type Ⅰ collagen was fully mixed with chitosan, freeze-dried and cross-linked with chondroitin sulfate, then freeze-dried again and sterilized by ethylene oxide. The pore diameter, water content, porosity of the scaffold were tested. The adipose tissue-derived stromal cells were digested, seeded into the plates, scaffold, and cen-trifuged into pellet, and then induced into cartilage. MTT detection for cell proliferation was done. After 3 weeks, the cell morphology, and cell proliferation and adhesion were observed, and chondrngenic differenti-ation was also analyzed. Results The pore diameter, water content, porosity tested for the scaffold showed an appropriate form. Cell proliferation showed faster in the scaffold and pellet culture system after 5 day, there was still cell proliferation in the scaffold system after 14 days but no obvious changes in the pellet cul-ture system; ceils on the scaffold proliferated densely showed by histological staining, but there was a scaf-fold structure residues in the inner layer. The finding of type Ⅱ immunohistochemistry stain showed that cells express strong positive for type Ⅱ collagen in the scaffold and pellet culture system whereas it was weakly positive in the plate culture system; the specific mRNA for cartilage, type Ⅱ collagen, aggrecan and SOX-9 were expressed in all three systems showed by RT-PCR, but type X collagen was expressed continu-ously in the plate culture system and expressed after 21 days in the pellet culture system, whereas it was not detected in the collagen-chitosan-chondroitin sulfate scaffold system. Conclusion The parameters of the collagen-chitosan-chondroitin sulfate scaffold were suitable in our study. The results suggested that it can promote the adipose tissue-derived stromal cells proliferation and chondrogenic differentiation better than the plate and pellet culture systems and maintain the phenotype of chondrocytes well; it is the optimal choice for cartilage tissue engineering in the future.     

大鼠脂肪干细胞复合胶原-壳聚糖-硫酸软骨素三维支架构建组织工程软骨

Objective To evaluate the character of the collagen-chitosan-chondroitin sulfate scaffold seeded with rat adipose tissue-derived stromal cells. Methods A dipose tissue were harvested from 6 weeks old Wistar rats and the stromal cells were harvested by type Ⅰ collagenase and then cultured in vitro. Type Ⅰ collagen was fully mixed with chitosan, freeze-dried and cross-linked with chondroitin sulfate, then freeze-dried again and sterilized by ethylene oxide. The pore diameter, water content, porosity of the scaffold were tested. The adipose tissue-derived stromal cells were digested, seeded into the plates, scaffold, and cen-trifuged into pellet, and then induced into cartilage. MTT detection for cell proliferation was done. After 3 weeks, the cell morphology, and cell proliferation and adhesion were observed, and chondrngenic differenti-ation was also analyzed. Results The pore diameter, water content, porosity tested for the scaffold showed an appropriate form. Cell proliferation showed faster in the scaffold and pellet culture system after 5 day, there was still cell proliferation in the scaffold system after 14 days but no obvious changes in the pellet cul-ture system; ceils on the scaffold proliferated densely showed by histological staining, but there was a scaf-fold structure residues in the inner layer. The finding of type Ⅱ immunohistochemistry stain showed that cells express strong positive for type Ⅱ collagen in the scaffold and pellet culture system whereas it was weakly positive in the plate culture system; the specific mRNA for cartilage, type Ⅱ collagen, aggrecan and SOX-9 were expressed in all three systems showed by RT-PCR, but type X collagen was expressed continu-ously in the plate culture system and expressed after 21 days in the pellet culture system, whereas it was not detected in the collagen-chitosan-chondroitin sulfate scaffold system. Conclusion The parameters of the collagen-chitosan-chondroitin sulfate scaffold were suitable in our study. The results suggested that it can promote the adipose tissue-derived stromal cells proliferation and chondrogenic differentiation better than the plate and pellet culture systems and maintain the phenotype of chondrocytes well; it is the optimal choice for cartilage tissue engineering in the future.     

大鼠脂肪干细胞复合胶原-壳聚糖-硫酸软骨素三维支架构建组织工程软骨

Objective To evaluate the character of the collagen-chitosan-chondroitin sulfate scaffold seeded with rat adipose tissue-derived stromal cells. Methods A dipose tissue were harvested from 6 weeks old Wistar rats and the stromal cells were harvested by type Ⅰ collagenase and then cultured in vitro. Type Ⅰ collagen was fully mixed with chitosan, freeze-dried and cross-linked with chondroitin sulfate, then freeze-dried again and sterilized by ethylene oxide. The pore diameter, water content, porosity of the scaffold were tested. The adipose tissue-derived stromal cells were digested, seeded into the plates, scaffold, and cen-trifuged into pellet, and then induced into cartilage. MTT detection for cell proliferation was done. After 3 weeks, the cell morphology, and cell proliferation and adhesion were observed, and chondrngenic differenti-ation was also analyzed. Results The pore diameter, water content, porosity tested for the scaffold showed an appropriate form. Cell proliferation showed faster in the scaffold and pellet culture system after 5 day, there was still cell proliferation in the scaffold system after 14 days but no obvious changes in the pellet cul-ture system; ceils on the scaffold proliferated densely showed by histological staining, but there was a scaf-fold structure residues in the inner layer. The finding of type Ⅱ immunohistochemistry stain showed that cells express strong positive for type Ⅱ collagen in the scaffold and pellet culture system whereas it was weakly positive in the plate culture system; the specific mRNA for cartilage, type Ⅱ collagen, aggrecan and SOX-9 were expressed in all three systems showed by RT-PCR, but type X collagen was expressed continu-ously in the plate culture system and expressed after 21 days in the pellet culture system, whereas it was not detected in the collagen-chitosan-chondroitin sulfate scaffold system. Conclusion The parameters of the collagen-chitosan-chondroitin sulfate scaffold were suitable in our study. The results suggested that it can promote the adipose tissue-derived stromal cells proliferation and chondrogenic differentiation better than the plate and pellet culture systems and maintain the phenotype of chondrocytes well; it is the optimal choice for cartilage tissue engineering in the future.     

大鼠脂肪干细胞复合胶原-壳聚糖-硫酸软骨素三维支架构建组织工程软骨

Objective To evaluate the character of the collagen-chitosan-chondroitin sulfate scaffold seeded with rat adipose tissue-derived stromal cells. Methods A dipose tissue were harvested from 6 weeks old Wistar rats and the stromal cells were harvested by type Ⅰ collagenase and then cultured in vitro. Type Ⅰ collagen was fully mixed with chitosan, freeze-dried and cross-linked with chondroitin sulfate, then freeze-dried again and sterilized by ethylene oxide. The pore diameter, water content, porosity of the scaffold were tested. The adipose tissue-derived stromal cells were digested, seeded into the plates, scaffold, and cen-trifuged into pellet, and then induced into cartilage. MTT detection for cell proliferation was done. After 3 weeks, the cell morphology, and cell proliferation and adhesion were observed, and chondrngenic differenti-ation was also analyzed. Results The pore diameter, water content, porosity tested for the scaffold showed an appropriate form. Cell proliferation showed faster in the scaffold and pellet culture system after 5 day, there was still cell proliferation in the scaffold system after 14 days but no obvious changes in the pellet cul-ture system; ceils on the scaffold proliferated densely showed by histological staining, but there was a scaf-fold structure residues in the inner layer. The finding of type Ⅱ immunohistochemistry stain showed that cells express strong positive for type Ⅱ collagen in the scaffold and pellet culture system whereas it was weakly positive in the plate culture system; the specific mRNA for cartilage, type Ⅱ collagen, aggrecan and SOX-9 were expressed in all three systems showed by RT-PCR, but type X collagen was expressed continu-ously in the plate culture system and expressed after 21 days in the pellet culture system, whereas it was not detected in the collagen-chitosan-chondroitin sulfate scaffold system. Conclusion The parameters of the collagen-chitosan-chondroitin sulfate scaffold were suitable in our study. The results suggested that it can promote the adipose tissue-derived stromal cells proliferation and chondrogenic differentiation better than the plate and pellet culture systems and maintain the phenotype of chondrocytes well; it is the optimal choice for cartilage tissue engineering in the future.     

大鼠脂肪干细胞复合胶原-壳聚糖-硫酸软骨素三维支架构建组织工程软骨

Objective To evaluate the character of the collagen-chitosan-chondroitin sulfate scaffold seeded with rat adipose tissue-derived stromal cells. Methods A dipose tissue were harvested from 6 weeks old Wistar rats and the stromal cells were harvested by type Ⅰ collagenase and then cultured in vitro. Type Ⅰ collagen was fully mixed with chitosan, freeze-dried and cross-linked with chondroitin sulfate, then freeze-dried again and sterilized by ethylene oxide. The pore diameter, water content, porosity of the scaffold were tested. The adipose tissue-derived stromal cells were digested, seeded into the plates, scaffold, and cen-trifuged into pellet, and then induced into cartilage. MTT detection for cell proliferation was done. After 3 weeks, the cell morphology, and cell proliferation and adhesion were observed, and chondrngenic differenti-ation was also analyzed. Results The pore diameter, water content, porosity tested for the scaffold showed an appropriate form. Cell proliferation showed faster in the scaffold and pellet culture system after 5 day, there was still cell proliferation in the scaffold system after 14 days but no obvious changes in the pellet cul-ture system; ceils on the scaffold proliferated densely showed by histological staining, but there was a scaf-fold structure residues in the inner layer. The finding of type Ⅱ immunohistochemistry stain showed that cells express strong positive for type Ⅱ collagen in the scaffold and pellet culture system whereas it was weakly positive in the plate culture system; the specific mRNA for cartilage, type Ⅱ collagen, aggrecan and SOX-9 were expressed in all three systems showed by RT-PCR, but type X collagen was expressed continu-ously in the plate culture system and expressed after 21 days in the pellet culture system, whereas it was not detected in the collagen-chitosan-chondroitin sulfate scaffold system. Conclusion The parameters of the collagen-chitosan-chondroitin sulfate scaffold were suitable in our study. The results suggested that it can promote the adipose tissue-derived stromal cells proliferation and chondrogenic differentiation better than the plate and pellet culture systems and maintain the phenotype of chondrocytes well; it is the optimal choice for cartilage tissue engineering in the future.     

大鼠脂肪干细胞复合胶原-壳聚糖-硫酸软骨素三维支架构建组织工程软骨

Objective To evaluate the character of the collagen-chitosan-chondroitin sulfate scaffold seeded with rat adipose tissue-derived stromal cells. Methods A dipose tissue were harvested from 6 weeks old Wistar rats and the stromal cells were harvested by type Ⅰ collagenase and then cultured in vitro. Type Ⅰ collagen was fully mixed with chitosan, freeze-dried and cross-linked with chondroitin sulfate, then freeze-dried again and sterilized by ethylene oxide. The pore diameter, water content, porosity of the scaffold were tested. The adipose tissue-derived stromal cells were digested, seeded into the plates, scaffold, and cen-trifuged into pellet, and then induced into cartilage. MTT detection for cell proliferation was done. After 3 weeks, the cell morphology, and cell proliferation and adhesion were observed, and chondrngenic differenti-ation was also analyzed. Results The pore diameter, water content, porosity tested for the scaffold showed an appropriate form. Cell proliferation showed faster in the scaffold and pellet culture system after 5 day, there was still cell proliferation in the scaffold system after 14 days but no obvious changes in the pellet cul-ture system; ceils on the scaffold proliferated densely showed by histological staining, but there was a scaf-fold structure residues in the inner layer. The finding of type Ⅱ immunohistochemistry stain showed that cells express strong positive for type Ⅱ collagen in the scaffold and pellet culture system whereas it was weakly positive in the plate culture system; the specific mRNA for cartilage, type Ⅱ collagen, aggrecan and SOX-9 were expressed in all three systems showed by RT-PCR, but type X collagen was expressed continu-ously in the plate culture system and expressed after 21 days in the pellet culture system, whereas it was not detected in the collagen-chitosan-chondroitin sulfate scaffold system. Conclusion The parameters of the collagen-chitosan-chondroitin sulfate scaffold were suitable in our study. The results suggested that it can promote the adipose tissue-derived stromal cells proliferation and chondrogenic differentiation better than the plate and pellet culture systems and maintain the phenotype of chondrocytes well; it is the optimal choice for cartilage tissue engineering in the future.     

Faculty of Anaesthetists of the Royal College of Surgeons of Ireland

The Faculty of Anaesthetists of the Royal College of Surgeons of England, 35–43 Lincoln's Inn Fields, London WC2A 3PN. Telephone: 01-405 3474.     

深圳市某两所小学流行性腮腺炎突发疫情的流行病学分析

目的：通过对深圳市某两所小学发生的流行性腮腺炎突发疫情的流行病学特点及差异性进行分析,为制定科学、高效的防控策略提供科学依据。方法2013年5～7月深圳市大鹏新区某两所小学爆发流行性腮腺炎,以学校为整体研究对象,分别标记为学校A（24个班,学生1210例）和学校B（27个班,学生1274例）,对比两所小学的疫情流行病学差异性。结果分析发现,学校A流行性腮腺炎发病率为4.30%,发病班级所占比54.17%,均较学校B1.73%和29.63%高,对比差异有统计学意义（P＜0.05）;分析显示学校A学生出现疫病平均年龄为（11.2±1.1）岁,较学校B（9.34±1.0）岁,对比差异明显（P＜0.05）;且两组疫病患儿在接种疫苗率对比上差异无统计学意义（P＞0.05）;但疫情发生时,学校B疫苗紧急接种率明显高于学校A,对比差异有统计学意义（P＜0.05）。结论小学作为流行性腮腺炎爆发的主要场所之一,疫病爆发高峰季节前,针对易感染人群给予相应的疫苗接种等预防控制措施,同时加强流行性腮腺炎的监测,对于降低感染人群数量,减轻、遏制疫情有着积极的意义,值得相关防控部门重视。