Effects of pulsed electromagnetic field frequencies on the osteogenic differentiation of human mesenchymal stem cells

The purpose of this study was to evaluate the effect of different frequencies of pulsed electromagnetic fields on the osteogenic differentiation of human mesenchymal stem cells. Third-generation human mesenchymal stem cells were irradiated with different frequencies of pulsed electromagnetic fields, including 5, 25, 50, 75, 100, and 150 Hz, with a field intensity of 1.1 mT, for 30 minutes per day for 21 days. Changes in human mesenchymal stem cell morphology were observed using phase contrast microscopy. Alkaline phosphatase activity and osteocalcin expression were also determined to evaluate human mesenchymal stem cell osteogenic differentiation.Different effects were observed on human mesenchymal stem cell osteoblast induction following exposure to different pulsed electromagnetic field frequencies. Levels of human mesenchymal stem cell differentiation increased when the pulsed electromagnetic field frequency was increased from 5 hz to 50 hz, but the effect was weaker when the pulsed electromagnetic field frequency was increased from 50 Hz to 150 hz. The most significant effect on human mesenchymal stem cell differentiation was observed at of 50 hz.The results of the current study show that pulsed electromagnetic field frequency is an important factor with regard to the induction of human mesenchymal stem cell differentiation. Furthermore, a pulsed electromagnetic field frequency of 50 Hz was the most effective at inducing human mesenchymal stem cell osteoblast differentiation in vitro.     

低强度脉冲超声对人脂肪干细胞成骨诱导影响的实验研究

[目的]探讨低强度脉冲超声在人脂肪干细胞成骨诱导中的影响。[方法]取手术患者腹部及腰背部处脂肪,采用胶原酶消化离心贴壁法分离培养脂肪间充质干细胞(adipose derived stem cells,ADSCs),流式细胞仪检测细胞表面标记。第6代细胞分为四组:(1)阴性对照组:ADSCs于完全培养基中培养,不接受LIPUS剌激;(2)LIPUS组:ADSCs于完全培养基中培养,接受LIPUS刺激;(3)成骨诱导组:ADSCs于成骨诱导培养基中培养;(4)LIPUS联合成骨诱导组:ADSCs在成骨诱导培养基中培养,接受LIPUS刺激。[结果]ADSCs形态传至10代时仍无变化。流式细胞仪分析第6代hADSCs细胞表面特异性标记结果显示,CD105:99.61%,CD29:97.54%,CD45:0.26%,CD44:97.4%,CD14:2.51%。LIPUS+诱导组的碱性磷酸酶(ALP)表达明显高于其他组,而单用LIPUS,并不能促进成人脂肪干细胞成骨分化。[结论]LIPUS可以促进成骨诱导培养基培养下的成人脂肪干细胞的成骨分化。     

脉冲电磁场促进人骨髓间充质干细胞成骨的研究

目的 :观察脉冲电磁场 (PEMFs)对人骨髓间充质干细胞 (hMSCs)成骨能力的影响。方法 :hMSCs受频率 12Hz、场强 1.1mT的PEMFs刺激 8h/d ,检测细胞增殖、细胞周期 ,观察细胞超微结构、骨钙素、Ⅰ型胶原免疫组化染色 ,碱性磷酸酶 (ALP)活性、核固红钙染色。结果 :脉冲电磁场作用后的细胞增殖能力提高 ,细胞形态发生变化 ,透射电镜观察提示细胞成熟 ,核固红钙染色、骨钙素、Ⅰ型胶原免疫组化染色均阳性 ,ALP活性增高。结论 :hMSCs经合理的PEMFs体外刺激后 ,细胞增殖加速 ,符合成骨细胞的形态特征和生物学特性。     

不同频率脉冲电磁场促人骨髓间充质干细胞增殖的实验研究

目的探求脉冲电磁场(pulsed electromagnetic fields,PEMF)促进人骨髓间充质干细胞(human mesenchymal stem cells,hMHCs)增殖的适合频率.方法选用场强1.1mT,频率分别为5、25、50、75、100、150Hz的PEMF刺激hMSCs,30min/d,光镜观察细胞形态,MTT法和流式细胞技术检测细胞增殖水平及生长周期.结果各频率组PEMF刺激均能促进MSCs增殖;频率不同,促增殖的效应不一,其中50Hz频段效果最为显著,生长曲线比对照组明显前移,S期细胞百分比(28.89±0.25)%较对照组(13.57±0.55)%增高107%.结论频率是PEMF促进hMSCs增殖的重要影响因素之一,50Hz为促进hMSCs增殖的适合频率.     

Prevention of osteoporosis by pulsed electromagnetic fields

Using an animal model, we examined the use of pulsed electromagnetic fields, induced at a physiological frequency and intensity, to prevent the osteoporosis that is concomitant with disuse. By protecting the left ulnae of turkeys from functional loading, we noted a loss of bone of 13.0 per cent compared with the intact contralateral control ulnae over an eight-week experimental period. Using a treatment regimen of one hour per day of pulsed electromagnetic fields, we observed an osteogenic dose-response to induced electrical power, with a maximum osteogenic effect between 0.01 and 0.04 tesla per second. Pulse power levels of more or less than these levels were less effective. The maximum osteogenic response was obtained by a decrease in the level of intracortical remodeling, inhibition of endosteal resorption, and stimulation of both periosteal and endosteal new-bone formation. These data suggest that short daily periods of exposure to appropriate electromagnetic fields can beneficially influence the behavior of the cell populations that are responsible for bone-remodeling, and that there is an effective window of induced electrical power in which bone mass can be controlled in the absence of mechanical loading.     

cAMP/PKA信号通路介导人脂肪干细胞成骨分化的体外试验

目的 研究环腺苷酸(cAMP)/蛋白激酶A(PKA)信号通路对脂肪干细胞(ADSCs)成骨分化的影响.方法 在成骨细胞培养基中加入cAMP/PKA信号通路的激动剂db-cAMP和抑制剂H-89,观察成骨细胞分化、PKA的活化及下游调节蛋白CREB的磷酸化和成骨基因表达的变化.结果 db-cAMP促进PKA的活化,上调CREB的磷酸化和相关基因表达的变化,促进ADSCs的成骨分化,同时以上作用可被PKA抑制剂H-89有效地阻断.结论 cAMP/PKA是介导人ADSCs成骨分化的重要信号通路.     

In vitro chondrogenic differentiation of human articular cartilage derived chondroprogenitors using pulsed electromagnetic field

BackgroundThe ability to grow new cartilage remains the standard goal of any treatment strategy directed at cartilage repair. Chondroprogenitors have garnered interest due to their applicability in cell therapy. Pulsed electromagnetic field (PEMF) favors chondrogenesis by possible upregulation of genes belonging to TGFβ superfamily. Since TGFβ is implicated in chondrogenic signalling, the aim of the study was to evaluate the ability of PEMF to induce chondrogenesis via endogenous TGFβ production in chondroprogenitors vs differentiation using chondrogenic medium inclusive of TGFβ.MethodsChondroprogenitors were harvested from three non-diseased human knee joints via fibronectin assay. Passage 3 pellets were subjected to four different culture conditions: a) negative control contained chondrogenic medium without TGFβ2, b) positive control contained medium with TGFβ2, c) PEMF 1 contained medium of negative control plus single exposure to PEMF and d) PEMF 2 contained medium of negative control plus multiple exposures to PEMF. Following differentiation (day 21), pellets were assessed for gene expression of ACAN, SOX9, COL2A1, TGFβ1, TGFβ2, and TGFβ3. Alcian blue staining to detect glycosaminoglycan deposition was also performed. Medium supernatant was used to detect endogenous latent TGF-β1 levels using ELISA.ResultsAll study arms exhibited comparable gene expression without any significant difference. Although positive control and PEMF study arms demonstrated notably better staining than negative control, the level of latent TGF-β1 was seen to be significantly high in supernatant from positive control (P < 0.05) when compared to other groups.ConclusionOur results indicate that PEMF induced chondrogenesis might involve other signalling molecules, which require further evaluation.     

脉冲电磁场对MC3T3-E1细胞增殖与分化的影响

目的研究不同强度50Hz脉冲电磁场(Pulse electromagnetic fields,PEMFs)对小鼠成骨细胞系MC3T3-E1增殖与分化的影响。方法 MC3T3-E1传代培养后分成七组,分别采用0.0 m T、0.6 m T、1.2 m T、1.8 m T、2.4 m T、3.0 m T和3.6 m T 50 Hz脉冲电磁场处理,90 min/d。MTT法检测细胞增殖情况,成骨性分化检测处理3 d、6 d、9 d、12 d后ALP活性和首次处理12 h后BMP-2、Runx-2、OPG基因表达情况。结果 6种强度的脉冲电磁场均促进细胞增殖,其中0.6 m T的促增殖效应最强(P<0.01)。在成骨性诱导培养条件下,细胞内ALP活性随电磁场处理时间的延长而逐渐增加,第9 d达到峰值,之后开始回落。0.6 m T、3.0 m T和3.6 m T均能提高ALP活性,其中0.6 m T始终处于最高水平(P<0.01)。三种基因的表达水平在首次处理12 h,在0.6 m T、1.2 m T、3 m T和3.6 m T组均显著提高,但仍然是0.6 m T的增强效应最为明显。结论 0.6 m T 50 Hz脉冲电磁场具有最佳的促进MC3T3-E1细胞增殖和成骨性分化活性,这可能为采用脉冲电磁场治疗骨质疏松症提供了理想的治疗参数。     

Treatment of nonunion using pulsed electromagnetic fields: a retrospective follow-up study

Pulsed electromagnetic fields (PEMF) are a useful means of treating cases of fracture nonunion. In 67.7% of nonunions with a disability time of at least 24 months, complete consolidation was obtained. This success rate is increased to 76.6% if we exclude nonunion, that presented contraindications for treatment with PEMF. The disability time had no effect on the success rate. Lesions of the humerus and atrophic nonunion had an unfavorable prognosis.     

Microcirculatory effects of pulsed electromagnetic fields

Purpose: Pulsed electromagnetic fields (PEMF) are used clinically to expedite healing of fracture non‐unions, however, the mechanism of action by which PEMF stimulation is effective is unknown. The current study examined the acute effects of PEMF stimulation on arteriolar microvessel diameters in the rat cremaster muscle. The study hypothesis was that PEMF would increase arteriolar diameters, a potential mechanism involved in the healing process. Methods: Local PEMF stimulation/sham stimulation of 2 or 60 min duration was delivered to the cremaster muscle of anesthetized rats. Arteriolar diameters were measured before and after stimulation/sham stimulation using intravital microscopy. Systemic hemodynamics also were monitored during PEMF stimulation. Results: Local PEMF stimulation produced significant (p < 0.001) vasodilation, compared to pre‐stimulation values, in cremasteric arterioles in anesthetized rats (n = 24). This dilation occurred after 2 min of stimulation (9% diameter increase) and after 1 h of stimulation (8.7% diameter increase). Rats receiving “sham” stimulation (n = 15) demonstrated no statistically significant change in arteriolar diameter following either “sham” stimulation period. PEMF stimulation of the cremaster (n = 4 rats) did not affect systemic arterial pressure or heart rate, nor was it associated with a change in tissue environmental temperature. Conclusions: These results support the hypothesis that local application of a specific PEMF waveform can elicit significant arteriolar vasodilation. Systemic hemodynamics and environmental temperature could not account for the observed microvascular responses. © 2003 Orthopaedic Research Society. Published by Elsevier Ltd. All rights reserved.     

促骨激素和生长因子对人骨髓基质细胞钙粘连素表达的调节

目的检测人骨髓基质细胞（hBMSCs）中钙粘连素是否连续表达，以及各种促骨激素和局部因子对其表达的调节。方法通过免疫定位和免疫印迹法检测原代hBMSCs和永生化骨母细胞系（hOP-7）中钙粘连素表达。结果钙粘连素表达与hOP-7细胞系的碱性磷酸酶（ALP）相关，多种促骨激素和局部因子对钙粘连素表达有调节作用。MoAb C-1821抗体阻断钙粘连素可增加ALP基础活性，并对1，25（OH）2D3诱导的ALP活性有增强作用。结论人骨母细胞中表达钙粘连素，并参与成骨分化。促骨因子对钙粘连素表达的差异性调节表明这些因子可以通过不同的钙     

葛根素诱导脂肪干细胞成骨分化的实验研究

目的观察葛根素诱导卵巢切除后骨质疏松大鼠模型的脂肪干细胞(adipose-derived stem cells,ADSCs)成骨分化的能力。方法分离、培养、传代及鉴定去卵巢骨质疏松模型大鼠ADSCs;含葛根素培养基诱导去卵巢骨质疏松大鼠ADSCs成骨分化,并测定最大有效浓度。结果去卵巢骨质疏松大鼠ADSCs成骨分化能力与加入含葛根素培养基的时间和培养基中葛根素的浓度呈正向相关,浓度为12 mmol/L时呈现无毒剂量下促进成骨分化最显著作用。结论葛根素具有较强促进去卵巢骨质疏松大鼠ADSCs成骨分化的诱导能力。     

不同年龄人骨髓间充质干细胞体外增殖及成骨分化的研究

目的 :观测年龄对人骨髓间充质干细胞 (humanmesenchymalstemcells ,hMSCs)体外增殖、成骨分化的影响。方法 :使用密度梯度离心法分离不同年龄人骨髓MSCs进行培养 ,保留贴壁细胞传代 ,观察细胞生长情况 ,检测其增殖活性、碱性磷酸酶活性 (ALP)、诱导后骨钙素定量测定。结果 :低龄hMSCs较高龄hMSCs体外生长快、MTT、ALP及骨钙素浓度高。结论 :hMSCs的增殖和成骨分化的能力和活性随着年龄的增加而降低。     

探讨电磁场影响骨髓间充质干细胞增殖分化的时间依赖性

目的 探究正弦波电磁场诱导大鼠骨髓间充质干细胞(BMSCs)成骨方向分化、促进其增殖的时间依赖性.方法 用差速贴壁法分离培养(bone marrow-derived mesenchymal stem cells,BMSCs),取用生长状态良好的第三代细胞进行实验.将细胞按照1×105个/孔接种于3.5 cm细胞培养皿中...     

The effect of pulsed electromagnetic fields on hindfoot arthrodesis: a prospective study

The aim of this study was to evaluate the effect of pulsed electromagnetic fields in a consecutive series of 64 patients undergoing hindfoot arthrodesis (144 joints). All patients who underwent elective triple/subtalar arthrodesis were randomized into control and pulsed electromagnetic field study groups. Subjects in the study group had an external pulsed electromagnetic fields device applied over the cast for 12 hours a day. Radiographs were taken pre- and postoperatively until radiographic union occurred. A senior musculoskeletal radiologist, blinded to the treatment scheme, evaluated the radiographic parameters. The average time to radiographic union in the control group was 14.5 weeks in 33 primary subtalar arthrodeses. There were 4 nonunions. The study group consisted of 22 primary subtalar arthrodeses and 5 revisions. The average time to radiographic union was 12.9 weeks (P =.136). The average time to fusion of the talonavicular joint in the control group was 17.6 weeks in 19 primary procedures. In the pulsed electromagnetic fields group of 20 primary and 3 revision talonavicular arthrodeses, the average time to radiographic fusion was 12.2 weeks (P =.003). For the 21 calcaneocuboid arthrodeses in control group, the average time to radiographic fusion was 17.7 weeks; it was 13.1 weeks (P =.010) for the 19 fusions in the study group. This study suggests that, if all parameters are equal, the adjunctive use of a pulsed electromagnetic field in elective hindfoot arthrodesis may increase the rate and speed of radiographic union of these joints.     

脉冲电磁场对接骨板性骨质疏松的影响

我们对钢板固定家兔胫骨用ＰＥＭＦｓ进行治疗 ,观察固定骨段的骨孔隙率 ,骨矿含量和矿化沉积率的变化 ,以期为临床应用ＰＥＭＦｓ防治接骨板性骨质疏松提供依据。1　材料与方法1 1　动物　健康成年大耳白家兔 18只 ,雌雄不限 ,体重在 2 5～ 3 0ｋｇ(河南医科大学实验动物中心提供 )。标准全价营养饲料分笼喂养。1 2　接骨板　采用天津产 4孔 316Ｌ不锈钢掌骨接骨板。1 3　仪器　ＴＦ ＶＩ型中医多功能治疗仪由郑州市颈肩腰腿痛医院研究所提供 (专利号 :992 385 4 2 3)。1 4　分组与处理　动物随机分为Ａ、Ｂ、Ｃ三组 ,每组各 6只。Ａ组为…     

不同强度低频正弦交变电磁场对大鼠骨髓间充质干细胞成骨性分化的影响

目的研究不同强度低频正弦交变电磁场(Sinusoidal electromagnetic fields,SEMFs)对体外培养大鼠骨髓间充质干细胞(rat Mesenchymal stem cells,rMSCs)成骨性分化的影响,从中筛选出最佳强度参数。方法原代培养大鼠骨髓间充质干细胞,每天在频率为50 Hz,强度为0.2 mT、0.6 mT、1.0mT、1.4 mT、1.8 mT、2.2 mT、2.6 mT、3.0 mT的磁场环境中照射30 min。检测细胞碱性磷酸酶活性、骨钙素分泌量和钙化结节数;并在照射后6、9、12、24 h提取细胞总RNA,用Real-time PCR法检测成骨性分化基因Runx2/Cbfal表达情况。结果磁场干预后细胞呈漩涡状排列和生长,1.8 mT磁场强度明显促进rMSCs的成骨性分化,表现在此组的ALP活性、骨钙素分泌量、钙化结节数和Runx2/Cbfal基因的表达量均高于其他组,亦显著高于对照组(P0.05)。结论在50 Hz、0.2～3.0 mT范围内,1.8 mT最能促进骨髓间充质干细胞的成骨性分化,是此范围内最佳的治疗用磁场参数。     

脉冲电磁场影响人骨髓基质干细胞缝隙连接通讯的实验研究

目的研究脉冲电磁场(PEMFs)对人骨髓基质干细胞(hBMSCs)缝隙连接所介导细胞间通讯(GJIC)的影响。方法透射电镜观察BMSCs超微结构,应用荧光漂白恢复(FRAP)技术,通过激光共聚焦显微镜检测hBM-SCs经PEMFs刺激后GJIC的功能变化。结果经PEMFs刺激后的hBMSCs,平均荧光漂白恢复率为(64.12±0.83)%,较对照组犤(35.26±0.76)%犦有显著性增加(P<0.05)。结论PEMFs刺激能促进hBMSCs的缝隙连接通讯功能。