In order to study the effects of electromagnetic fields (EMFs) on proliferation, differentiation and intercellular cyclic AMP (cAMP) in mouse bone marrow mesenchymal stem cells (MSCs) in vitro, the mouse bone MSCs were isolated and cultured in vitro. The third passage MSCs were divided into 4 groups and stimulated with EMFs. The cellular proliferation (MTT), the cellular differentiation (alkaline phosphatase activity, ALP), and the intercellular cAMP level were investigated at different time points. The results showed that EMF (50Hz pulse burst 2 mT peak) inhibited the cellular proliferation (P < 0.05), enhanced the cellular differentiation (P < 0. 05), and increased the intercellular cAMP level (P < 0.01) in the early time of the stimulation (1-3 days), but the intercellular cAMP level did not increased further in the later days. We are led to conclude that the cAMP may be involved in the mediation of the growth inhibitory and differentiation-inducing signals of specific EMFs in vitro.
Alkaline Phosphatase/metabolism ; Bone Marrow Cells/*cytology ; Cell Differentiation ; Cell Proliferation ; Cells, Cultured ; Cyclic AMP/metabolism ; Electromagnetic Fields ; Mesenchymal Stem Cells/*cytology

Objective To study the effects of electromagnetic field (EMF) exposure on osteoporosis in ovariectomized mice. Methods Sixty 8-week-old female Kunming mice were divided into four groups at random: a sham operation group (group A), an ovariectomized group (group B), an EMF and ovariectomized group (group C) and a nilestriol and ovariectomized group (group D). Bilateral ovariectomies were performed on all mice except those in group A. The mice of group C were exposured to a 15 Hz, 1.0 mT electromagnetic field. The mice of group D were given at nilestriol 1.5 mg/kg/week. The bone mineral density (BMD) of the lumbar vertebrae was measured before the mice were sacrificed at the 12th week. Blood specimens were collected every two weeks to measure the ac-tivity of alkaline phosphatase (ALP) and the concentration of bone gamma-carboxyglutamic-acid-containing proteins (BGP), calcium and estradiol in the serum. Histological sections were taken to examine and analyze the changes in bone trabeculae in the lumbar vertebrae after 6 and 12 weeks. Results EMF at 15 Hz and 1.0 mT intensity signifi-cantly increased the activity of ALP and the concentrations of BGP and calcium in the serum. In addition, the absorp-tion of bone trabeculae in the lumbar vertebrae was significantly restrained. Conclusions EMF at 15 Hz and 1.0 mT can restrain the development of osteoporosis in ovariectomized mice.

Objective To study the effects of an electromagnetic field (EMF) on the expression of fibro-blast growth factor (FGF-2) and it' s receptor (FGFR-2) mRNA in rat bone marrow mesenchymal stem cells (BMSCs) in vitro. Methods Rat BMSCs were isolated and cultured in vitro. The subcultured cells were divided into different groups to be EMF stimulated at 1.0 mT. The expression of FGF-2 and FGFR-2 mRNA were detected by reverse transcription-polymerase chain reaction (RT-PCR). Results Different frequencies and durations of 1.0 mT EMF exposure induced FGF-2 and FGFR-2 mRNA expression in comparison to blank controls. The expression of FGF-2 mRNA reached a peak after stimulation at 15 Hz for 10 min, 50 Hz for 60 min and 75 Hz for 30 min. And the expression of FGFR-2 mRNA reached a peak after 30 minutes at all frequencies. At 1.0 mT with 30 min exposure, the expression of FGF-2 mRNA peaked after 50 Hz stimulation, and the expression of FGFR-2 mRNA peaked after stimulation at 75 Hz. Conclusions Moderate EMF stimulation can significantly increase the expression of FGF-2 and FGFR-2 mRNA in rat BMSCs in vitro.

Objective To study the effects of pulsed electromagnetic field (PEMF) in promoting tibia and fibula fracture healing in rats. Methods Thirty rats were divided randomly into two groups after establishing the animal model of artificial tibia and fibula fracture: a PEMF treated group and a control group. Radiographs were taken immediately postoperatively and once a week thereafter until being sacrificed after week 5. The blood was sampled to measure the activity of alkaline phosphatase (ALP) and the concentration of bone gamma-carboxyglutamic-acid-con-taining proteins (BGP), calcium and phosphate in serum once a week, respectively. Histological sections were taken at weeks 3 and 5 to observe the pathological change of bony callus. Results PEMF of 15 Hz and 1.0 mT could sig-nificantly increase the amount of bony callus, significantly increase the amount of bony callus, promote the disappea-ring of fracture lines and the appearance of endochondral ossification and mature bone trabecula. The amount of colla-gen in bony callus raised and the process of callus moulding accelerated in PEMF treated group. In addition, the ac-tivity of ALP(P<0.04) and the concentration of BGP(P<0.05) in serum increased. Conclusions The PEMF of 15 Hz and 1.0 mT can obviously promote fracture healing of tibia and fibula in rats.

Objective To study the expressions of aggrecan (Agc) Ⅰ and Ⅱ collagen and Sox9 by bone mar-row mesenchymal stem cells exposed to electromagnetic fields (EMFs) and it's mechanisms involved. Methods Bone marrow mesenchymal stem cells were isolated from Sprague-Dawley rats and cultured in vitro. The third passage cells were harvested and exposed to 15 Hz 1 mT EMFs for 8 h/d. The semi-quantitative reverse transcription-polyme-rase chain reaction (RT-PCR) technique was used to measure parathyroid hormon receptor related peptide (PTHrp) ,Agc Ⅰ and Ⅱ collagen and Sox9 mRNA. Western blotting was used to measure type Ⅱ collagen expression. After the inhibitor of protein kinase A (PKA) H-89 and the inhibitor of protein kinase C (PKC) Go-6976 ( 12 μm) were added, the effects of EMFs on Agc Ⅰ and Ⅱ collagen and Sox9 mRNA expressions were measured again by using RT-PCR, and Western blotting technique. Results The EMFs induced significant increase of mRNA expressions of PTHrp, Agc Ⅰ and Ⅱ collagen and Sox9 in comparison to the controls, and promoted type Ⅱ collagen protein expres-sion. The Agc Ⅰ and Ⅱ collagen expressions decreased after PKA pathway inhibitor H-89 and PKC inhibitor Go-6976 were added, but the mRNA expression of Sox9 was not affected. Conclusion This study shows 15Hz 1mT EMFs can promote mRNA expressions of Agc Ⅰ and Ⅱ collagan and Sox9 of cbondrogenesis differentiation markers in bone marrow mesenchymal stem cells. The effect is correlated with PKA and PKC pathways.

Objective To study the influence of pulsed electromagnetic fields on the expression of type I collagen by bone marrow mesenchymal stem cells and it's mechanism. Methods The bone marrow mesenchymal stem cells of SD rats were isolated and cultured in vitro.The third passage cells were harvested and exposed to pulsed electromagnetic fields(PEMFs)at 15 Hz and 1 mT 8 h/d for 3 days.A semi-quantitative RT-PCR technique was used to measure the type I collagen mRNA expression;ELISA and immunohistochemitistry techniques were used to measure type I collagen expression.Inhibitors and promoters of the cAMP-dependent protein kinase A(cAMP-PKA)pathway were added.After the cAMP-PKA pathway had been inhibited or promoted,the effects of the PEMF on type I collagen expression were measured again using ELISA and immunohistoehemistry.Results PEMFs at 15 Hz and 1 mT induced significant promotion of the expression of type I collagen(P≤0.01)in comparison with the controls. The type I collagen expression was reduced when the cAMP-PKA pathway inhibitor H-89 was added,and raised when the promoter 8-Br-cAMP was added.Conclusion PEMFs at 15 Hz 1 mT can promote type I collagen expression of bone marrow mesenchymal stem cells.and the effect is correlated with the cAMP-PKA pathway.

Objective To investigate the effects of an electromagnetic field on the extra-cellularly regulated kinase(ERK)signalling pathway and to determine the impact of electromagnetic activation on osteogenic proliferation and differentiation in rat bone marrow mesenchymal stem cells.Methods Rat bone marrow mesenchymal stem cells were isolated and cultured in vitro.The third-passage cells were divided into 4 groups(Control,PD98059,EMF and EMF+PD98059).Western blotting Was used to detect the activation of the ERK signal pathway after exposure to an electromagnetic field.MTT assay Was used to determine the activation of proliferation in the celb in the different groups.The cells' alkaline phosphatase activities were also detected. Results (1)The ERK signal pathway in these rat bone marrow mesenchymal stem cells was activated after exposure to a 15 Hz.1 mT,sine wave form electromagnetic field for 5 min.Activation remained high for at least 1 h.PD98059 can effectively block the activation of the ERK signal pathway.(2)Cell proliferation was promoted after exposure to the electromagnetic field,and this effect could be significantly inhibited by PD98059.(3)Alkaline phosphatase was significantly elevated in these bone marrow mesenchymal stem cells after exposure to the electromagnetic field.The activation in the EMF+PD98059 group Was slightly greater than in the EMF group.Conclusion Electromagnetic fields of 15 Hz and 1 mT can activate the ERK signal pathway and alter proliferation and osteogenic differentiation in the bone marrow mesenchymal stem cells of rats.

Objective To evaluate whether immunization with △A146 Ply could confer protections against pneumococcal infections in murine models and to reveal the possible role of △A146 Ply-specific IgG in the protection elicited.MethodsBALB/c mice were immunized intraperitoneally with △A146 Ply or PBS plus alum.Fourteen days after the third immunization,mice were intranasally challenged with serotype 14 and 19F Streptococcus pneumoniae.Three days after inoculation,lungs were removed from mice and homogenized in PBS,followed by plated on red cell plates.Viable bacteria were counted after overnight incubation.As to the sepsis models,vaccinated mice were challenged intraperitoneally with different dosage bacteria of D39 and serotype 3 strain.The numbers of CFU log10 were compared by Mann Whitney U test and survival rates were analyzed using log-rank test.Passive protection was used to evaluate the role of △A146 Ply-specific IgG in the protection against otherwise lethal infection resulted from D39.Results ELISA analysis demonstrated higher titer specific antibody responses to △A146 Ply was produced after 3 times immunization.Mice boosted twice with △A146 Ply survived significantly longer than that for mice boosted once with △A146 Ply in Alum adjuvant,which were significantly longer than that of control group.Immunization with △A146 Ply was effective in reducing the numbers of pneumococcal strain 31614(serotype 14) and 31693(serotype 19F),which resulted in 50-and 20-fold decreases in bacterial load in the lungs respectively when compared to control protein-immunized mice.60% of vaccinated mice survived the infection with pneumococcal D39 of 1200 CFU.60% protection was achieved when mice intraperitoneally infected with D39 and received △A146 Ply-specific IgG,whereas no mice survived the infection when they were passively administered with △A146 Ply-specific IgG depleted antisera.Conclusion Immunization with △A146 Ply could confer protection against pneumococcal infections,and protection elicited was mediated by △A146 Ply-specific IgG.

Objective To screen the differential expression genes of bone marrow MSCs stimulated by electromagnetic field(EMF)with osteogenesis microarray analysis,and to study the underlying mechanism that EMF promotes the differentiation of bone marrow MSCs. Methods The Sprague-Dawley rat bone marrow MSCs were isolated and cultured in vitro.The third-passage cells who were stimulated by EMF and served as the stimulated group,and those who were not stimulated by the EMF served as the controls.Total RNA was extracted and purified,then it was used to synthesize cDNA and cRNA.The eRNA of stimulated group and the control group was hybridized with the rat oligo osteogenesis microarray,respectively.The hybridization signals were acquired by using X-ray film after chemiluminescent detection and the obtained data were analyzed using the web-based completely integrated GEArray Expression Analysis Suite.RT-PCR was used to identify the chosen genes BMP1,VDR and EGF. Results Nineteen differential expression genes were found between the stimulated group and the control group.There were 6 upregulated and 13 downregulated genes in the stimulated group.Semi-quantitative RT-PCR confirmed that the expression levels of BMP1,VDR mRNA in the stimulated group were upregulated and EGF downregulated. Conclusion The gene expression profiles about osteogenesis of the bone marrow MSCs were changed after EMF intervention(1 5 Hz,1 mT).These genes are involved in the difierentiation of bone marrow MSCs into osteoblast.These results provide deeper insight into the mechanism that EMF exposure facilitates the in vitro differentiation of bone marrow MSCs.

A preliminary experimental research on the proliferation and differentiation of rat bone marrow mesenchymal stem cells with exposure to 50 Hz magnetic fields is done. The experimental results can be well repeated, showing that 50 Hz magnetic fields can evidently affect the proliferation and differentiation of rat bone marrow mesenchymal stem cells (MSCs), besides, the results are not linear with the amplitude of magnetic fields, but existing therein a 'window' phenomenon. At the end of this paper, the domestic and overseas development of the research is discussed on with the emphasis how to cure the bone fracture with the help of magnetic fields, and our investigative thoughts are also presented.
Animals ; Bone Marrow Cells ; cytology ; Cell Differentiation ; Cell Proliferation ; Magnetic Fields ; Mesenchymal Stromal Cells ; cytology ; Rats