Different electromagnetic field waveforms have different effects on proliferation, differentiation and mineralization of osteoblasts in vitro.

Noninvasive electromagnetic fields (EMFs) have been known to be able to improve bone health; however, their optimal application parameters and action mechanisms remain unclear. This study compared the effects of different forms of EMFs (sinusoidal, triangular, square, and serrated, all set at 50 Hz frequency and 1.8 mT intensity) on proliferation, differentiation and mineralization of rat calvarial osteoblasts. Square EMFs stimulated osteoblast proliferation but sinusoidal EMFs inhibited it. Sinusoidal and triangular EMFs produced significantly greater alkaline phosphatase (ALP) activity, ALP staining areas, calcium deposition, mineralized nodule areas, and mRNA expression of Runx-2, osteoprotegerin and insulin-like growth factor-I than square and serrated EMFs (P < 0.01). Triangular EMFs had a greater effect than sinusoidal EMFs on every indices except for Runx-2 mRNA expression (P < 0.05). These results indicated that while square EMFs promoted proliferation and had no effect on the differentiation of osteoblasts, sinusoidal EMFs inhibited proliferation but enhanced osteogenic differentiation. Triangular EMFs did not affect cell proliferation but induced the strongest osteogenic activity among the four waveforms of EMFs. Thus, the effects of EMFs on proliferation and differentiation of osteoblasts in vitro were dependent on their waveforms.

[Comparison research with icariin and genistein by anti-inflammatory reaction and angiogenesis pathway to inhibit bone loss on ovariectomized rats].

OBJECTIVE: Through researching the relationship among osteoporosis and inflammatory reaction besides angiogenesis, to compare pharmacological differences between icariin and genistein to inhibit bone loss. METHODS: 6 months old female SD rats were randomly divided into SHAM group, model group, ICA group, GEN group and E group. The bone mineral density of total, femur and lumbar, serum OC, TRACP 5b, IL-6 and VEGF, biomechanics of femur and tibia microarchitecture were analyzed. RESULTS: Compared with SHAM group, model group of body weight, uterine weight, bone mineral density of total, femur and lumbar, serum OC, TRACP 5b, IL-6 and VEGF, biomechanics of femur and lumbar and tibia microarchitecture were significantly changed (P < 0.05). Compared with model group, ICA group of body weight, bone mineral density of total and femur, serum TRACP 5b and femural biomechanics were significantly changed (P < 0.05). GEN group of bone mineral density of total, femur and lumbar, serum OC, TRACP 5b, IL-6 and VEGF, biomechanics of femur and lumbar and tibia microarchitecture were significantly changed (P < 0.05). CONCLUSION: Icariin inhibits bone loss on model rat through suppressing bone resorption. Genistein prevents bone loss on model rat by the pathway of inhibiting inflammatory reaction, activating angiogenesis, enhancing bone formation and inhibiting bone resorption. Moreover, pharmacological activity of genistein is more potential than icariin.

Icariin induces osteoblast differentiation and mineralization without dexamethasone in vitro.

An effective method for preventing bone loss is by promoting osteoblast differentiation and bone formation. While dexamethasone has been routinely used as a classical inducer for osteoblast differentiation, limitations have been observed with its usage, including its varied effects on expression of osteoblast genes in different species and its potentials in suppressing osteoblastic differentiation and mineralization. In this study, we assessed the ability of flavonoid icariin in enhancing differentiation and mineralization of cultured rat primary osteoblasts in the absence of dexamethasone. It was found that, compared to the non-stimulated control, icariin at 10(-5) M produced a higher alkaline phosphatase activity, more and larger areas of alkaline phosphatase-positive colonies (CFU-FALP) and mineralized nodules, more osteocalcin secretion and calcium deposition, higher levels of mRNA expression of alkaline phosphatase, osteoblastic transcription factors osterix and runt-related transcription factor 2, and collagen 1α, higher levels of protein expression of collagen 1α, alkaline phosphatese, osterix, and runt-related transcription factor 2. In addition, icariin at 10(-5) M was always more potent than dexamethasone at its optimal concentration of 10(-8) M on the above osteoblast differentiation and mineralization markers. Taken together, our studies demonstrated that icariin has a pronounced ability in promoting osteoblast differentiation in vitro in the absence of dexamethasone.

[Effect of genistein on rat femoral bone metabolic activity in vitro].

This study is to investigate effects of genistein on rat femoral bone metabolic in vitro. Rat femoral tissues was isolated and randomly divided into two groups including control group and genistein (1 x 10(-5) mol x(-1)) group. Determinations of alkaline phosphatase (ALP) activity, calcium content and osteoprotegerin (OPG), type I-collagen (Collagen-I), RANKL, Runx-2 and bone morphogenetic protein (BMP-2) mRNA expression were done by real-time PCR. The results showed that 1 x 10(-5) mol x L(-1) genistein could increase the activity of ALP and contents of Ca, regulate bone metabolism activity of OPG, RANKL, BMP-2, Collagen-I and Runx-2 mRNA expression level. Genistein can significantly modulate bone metabolism related gene expression level of rat femoral tissue in vitro, and can increase calcium content and the activity of ALP.

[Effects of static magnetic field with different exposure time on the maturation of rat osteoblasts in vitro and the expression of the estrogen receptor gene].

OBJECTIVE: To investigate the effects of static magnetic fields (SMFs) with different exposure time on the maturation of rat osteoblasts in vitro and the expression of the estrogen receptor (ER) gene. METHODS: The calvarial osteoblasts were isolated from newborn rats by enzyme digestion and randomly divided into 9 groups after one passage based on the exposure time of the SMFs[0 (control), 0.5 h, 1.0 h, 1.5 h, 2.0 h, 2.5 h, 3.0 h, 3.5 h, and 4.0 h]. The intensity was 3.9 mT in all SMFs. Those without SMFs exposure were used as the controls. The oeteoblasts were observed under the contrast phase microscope on a daily basis. After 48 h, cell proliferation was assayed by MTT method. The osteocalcin contents were measured after exposure to SMFs for 3 d, 6 d, 9 d, and 12 d. ERΑ and ERΒ mRNA expressions were measured by real-time PCR after SMFs treatment for 0 h, 24 h, 48 h, and 72 h. RESULTS: Compared with the controls, the cell proliferation was significantly enhanced in the 2.0-h, 2.5-h, and 3.0-h groups (P<0.05). After SMFs treatment for 6 d, 9 d and 12 d, the 2.5-h group had significantly higher osteocalcin content than the control group did (P<0.05). After SMFs treatment for 0 h and 72 h, elevated ERΑ mRNA expression and reduced ERΒ mRNA expression were observed. CONCLUSION: Exposure to SMFs, regardless of exposure time, is associated with enhanced cell proliferation, increased osteocalcin contents, and altered ERΑ and ERΒ mRNA expressions in opposite directions.

[Effects of icariin and genistein on peak bone mass in rats].

OBJECTIVE: To compare the effects of icariin (ICA) and genistein (GEN) on rats bone peak mass and thus screen for a drug that can more effectively prevent osteoporosis. METHODS: Totally 36 one-month SD rats were randomly divided into three groups: ICA group [25 mg/(kg·d), intragastric administration], GEN group [10 mg/(kg·d), intragastric administration], and control group (fed with equal volume of distilled water). The body weight was monitored weekly and the bone mineral density of total body was measured monthly. All rats were sacrificed three months later. The femoral bone mineral density and the serum levels of osteocalcin and anti-tartaric acid phosphatase 5b, N-terminal propeptide of type 1 procollagen, and C-terminal propeptide of type 1collagen were measured. The bone microarchitectures were analyzed with micro-CT and the bone biomechanics properties were tested with universal material machine. RESULTS: The body weight and organ index showed no significant difference among these three groups(P>0.05). No obvious pathological change was found. The bone mineral density was also not significantly different in the first and second months; however, in the third months, the ICA group had significant higher bone mineral density for both total body and femur than those in the control and GEN group (P<0.05). The same trends were found for both femur bone mineral density and whole-body bone mineral density (P<0.05). The ICA group also had significantly higher serum levels of osteocalcin (P<0.05) and lower level of anti-tartaric acid phosphatase 5b(P<0.05). Besides, rats in the ICA group had significantly larger bone volume/tissue volume, trabecular thickness, and trabecular number than the control group, whereas the trabecular spacing and model coefficients were signicantly lower(all P<0.05), which, however, were not significantly different between ICA group and GEN group (P>0.05). Femoral maximum load, Youg's modulus, and yield load were significantly higher in these two groups than in the control group (P<0.05), which, again, were not significantly different between ICA group and GEN group (P>0.05). CONCLUSION: Orally administered ICA is more efficient than GEN in inhibiting resorption and promoting bone formation, and thus can dramatically improve the peak bone mineral density and bone quality.

[Mechanisms of icariin in regulating bone formation of osteoblasts and bone resorption of osteoclasts].

OBJECTIVE: To investigate the molecular mechanisms of icariin (ICA) in regulating the bone formation of osteoblasts and the bone resorption of osteoclasts. METHODS: Primary osteoblast cell cultures were obtained from newborn rat calvarial. Calcified nodules were stained by alizarin red. The mRNA levels of osterix (OSX), runt-related transcription factor 2 (Runx-2), alkaline phosphatase (ALP), Collagen1, osteoprotegerin (OPG), and receptor activator of nuclear factor-ΚB ligand (RANKL) were analyzed by quantitative real-time RT-PCR, the protein levels of OPG, RANKL, and Collagen1 were examined by Western blotting, and the intracellular Ca(2+) concentration of osteoblasts was measured on a flow cytometer using the Cellquest program. RESULTS: Compared with control group, ICA markedly promoted bone formation by significant up-regulating the gene expressions of OSX, Runx-2,ALP, and Collagen1, the protein expression of Collagen1(all P<0.01), and the Ca(2+) concentration. Furthermore, ICA remarkably inhibited bone resorption by significant up-regulating the mRNA and protein expressions of OPG as well as the OPG/RANKL ratio. CONCLUSIONS: ICA could promote bone formation of osteoblasts through inducting the gene expressions of OSX,Runx-2, ALP and Collagen1, and the protein expressions of Collagen1, and by increasing the Ca (2+) concentration. Moreover, ICA could inhibit bone resorption of osteoclasts through regulating OPG/RANKL signal pathway.

[Effect of osthole on bone metabolism in rat femoral tissues in vitro].

OBJECTIVE: To investigate the effect of osthole on bone metabolism in rat femoral tissues in vitro. METHODS: The rat femoral tissues were isolated in vitro. The optimal concentrations of ostehole (1×10(-5) mol/L) and estradiol (1×10(-8) mol/L) (the positive control) were selected by alkaline phosphatase activity (ALP). The ALP and calcium levels were detected by commmerical regents, and the expressions of osteoprotegerin, receptor activator of nuclear factor-κB ligand, runx-related gene 2, and bone morphogenetic protein-2 mRNA were determined by real-time reverse transcription-polymerase chain reaction. RESULT: The osthole (1×10(-5) mol/L) significantly increased the activity of ALP, calcium level as well as the expressions of osteoprotegerin, receptor activator of nuclear factor-κB ligand, runx-related gene-2 and bone morphogenetic protein-2 mRNA in rat femoral tissues in vitro. CONCLUSION: Osthole can improve calcium level and ALP activity and regulate the bone metabolism-related genes in rat femoral tissues.

[Comparative study on effect of 8-prenlynaringenin and narigenin on activity of osteoclasts cultured in vitro].

OBJECTIVE: To compare the effects of 8-prenylnaringenin (PNG) and naringenin (NG) on the activity and apoptosis of osteoclasts cultured in vitro, in order to study physiological activity of 8-prenyl perssad. METHOD: Osteoclasts were separated from long-limb bones of newly born rabbits, cultured in alpha-MEM containing 10% FBS, and then added with PNG and NG with the concentration of 1 x 10(-5) mol x L(-1). They were stained with TRAP and determined for enzymatic activity with TRAP after 4 d, and analyzed by toluidine blue staining after 7 d. The apoptotic osteoclasts were analyzed by Annexin V-FITC staining after 2, 4, 8, 12, 24, 36, and 48 hours, to observe their apoptosis. Their total RNAs were extracted, and analyzed for TRAP and Cathepsin K expressions by Real-time RT-PCR. RESULT: Compared with the control group, both of the PNG group and the NG group showed much less osteoclasts (TRAP positive cells), lower TRAP activity and TRAP and Cathepin K (CTSK) expression, and smaller number of bone resorption pits and areas. The PNG group show lower indexes than the NG group. Additionally, the PNG group reached the apoptotic peak of osteoclasts at 12 h after drug administration, whereas the NG group reached after 24 h. And the former had more apoptotic cells than the latter. CONCLUSION: 8-PNG is much more active than NG in inhibiting the resorption of osteoclasts and inducing apoptosis of osteoclasts. Their only difference lies in 8-prenyl perssad, which is proved to be able to enhance the anti-bone resorption activity of 8-prenylnarigenin.

[Comparison between icariin and genistein in osteogenic activity of marrow stromal cells].

OBJECTIVE: To compare the effect of icariin and genistein in the osteogenic differentiation of rat bone marrow stromal cells (rBMSC). METHOD: Rat marrow stromal cells were seperated in vitro, and the optimal concentration of genisten and icriin were screened. Genistein and icariin with the concentration of 1 x 10(-5) mol x L(-1) were adopted to intereven rBMSCs cultured in vitro. Alkaline phosphatase (ALP) was determined at 3, 6, 9, 12,15 d after intervention; calcified nodule was detected with alizarin red staining at 12 d; OXS, Runx-2, bone morphogenetic protein (BMP-2) and Collagen-I mRNA expression were observed with Real-time RT-PCR at 12, 24, 48, 72, 96 h. RESULT: Genistein and icariin with the concentration of 1 x 10(-5) mol x L(-1) could increase the activity of ALP and the content of Ca, regulate OXS, BMP-2, Runx-2 and Collagen-I mRNA expression. CONCLUSION: Icariin showed a stronger effect in improving the osteogenic differentiation of rat bone marrow stromal cells than genistein.

[Comparison of effects of kaempferide and anhydroicaritin on biomineralization of cultured osteoblasts].

This study is to compare the effects of kaempferide and anhydroicaritin on biomineralization of rat osteoblasts (ROB) in vitro. Calvarias were dissected aseptically from newborn SD rats, the osteoblasts were obtained by enzyme digestion and were cultured in MEM containing 10% FBS. The medium was changed every three days, and serial subculture was performed when cells covered with 90% of the dish. Kaempferide and anhydroicaritin were separately added with final concentrations of 1 x 10(-4), 1 x 10(-5), 1 x 10(-6) and 1 x 10(-7) mol x L(-1) under the conditions of osteogenic differentiation. The proliferation was measured by MTT, and the optimal concentration was detected by the ALP activity at the 9th day after osteogenic induction culture. The osteogenic indexes of kaempferide, anhydroicaritin and control group with the optimal concentration were compared. The result showed that the anhydroicaritin at concentration of 1 x 10(-5) mol x L(-1) had significantly promoted the activity of ALP, calcium content and osteocalcin content, increased the number of CFU-F(ALP) and mineralized nodules, enhanced the mRNA level of BMP-2, OSX and Runx-2, which are key genes of osteogenic differentiation, and raised the protein content of collagen-I. However, the kaempferide group had not significantly represented the ability that promoted osteogenic differentiation of ROB. The difference of osteogenic differentiation on ROB between kaempferide and anhydroicaritin was caused by the prenyl group on C-8 of icariin.

Effect of 3.6-mT sinusoidal electromagnetic fields on proliferation and differentiation of osteoblasts in vitro.

OBJECTIVE: To investigated the effect of 50-Hz 3.6-mT sinusoidal electromagnetic fields (SEMFs) on the proliferation and differentiation of osteoblasts in vitro. METHODS: The newborn rat calvarial osteoblasts were isolated by enzyme digestion and randomly divided into 6 groups after one passage. The treatment groups under 50-Hz 3.6-mT SEMFs and controls without SEMFs treatment. The cells were exposed in the SEMFs for 0.5 h, 1.0 h, 1.5 h, 2.0 h, and 2.5 h. They were observed under the contrast phase microscope each day. The calcified nodules were stained by alizarin red. The SEMFs were arranged in spiral appearance after 3 to 5 days. RESULTS: The SEMFs showed characteristic distribution 3 to 5 days after SEMFs treatment. On the 9(th) day after treatment, the activity of alkaline phosphatase (ALP) significantly increased in the 0.5-h group, whereas the ALP histochemical straining results and the area of calcified nodules were consistent with ALP activity. In the 48-h and 96-h groups, the genetic expression levels of osteoprotegerin and collagen-1 were significantly higher than that in the control group; particularly, the mRNA expression increased in the 0.5-h group. CONCLUSION: The SEMFs at 50-Hz 3.6-mT could suppress the proliferation of osteoblasts maturation but stimulate the differentiation and maturation of osteoblasts in vitro.

[Effect of sinusoidal electricity magnetic fields on the proliferation and differentiation of osteoblasts in vitro].

OBJECTIVE: To investigate the effect of sinusoidal electricity magnetic fields (SEMFs) on the proliferation and differentiation of osteoblasts in vitro. METHODS: Calvarial osteoblasts of newborn rats were isolated by enzyme digestion and randomly divided into 3 groups after subculture. Two groups of cells were exposed to 50 Hz 1.8 mT SEMFs for 30 min/d in parallel and vertical, respectively. Those without SEMFs exposure served as control. The cells were observed under the contrast phase microscope each day. After 48 h, cell proliferation was assayed by MTT method. The alkaline phasphatase (ALP) activities and calcium contents were measured after 3, 6, 9, and 12 days. The ALP positive colonies were histochemically stained after 10 days and the calcified nodules were stained by Alizarin Bordeaux after 12 days. Expressions of ALP, bone morphogenetic protein-2 (BMP-2) and Osterix (OSX) mRNA were measured at 0 h, 24 h, 48 h and 96 h. RESULTS: The cells exposed to the SEMFs were arranged in spiral appearance after 3 days. Compared with control, SEMFs inhibited cell proliferation (P < 0.01 or P < 0.05), but enhanced the maturation and mineralization of the osteoblasts. The results showed that SEMFs improved ALP activities, promoted calcium contents, increased calcified nodulues numbers, boosted expressions of ALP, BMP-2 and OSX mRNA. SEMFs with magnetic lines of force in parallel has stronger activities than those in vertical. CONCLUSION: The SEMFs at 1.8 mT and 50 Hz inhibit the proliferation of osteoblasts, but enhance the maturation and mineralization of osteoblasts.

[Study on mechanism of the anti hypoxia effect of genistein on osteoblasts in vitro].

OBJECTIVE: To study the protective effect of genistein on osteoblasts treated with hypoxia. METHODS: Rat osteoblasts were isolated from calvarias of newborn Sprague-Dawly rat by enzyme digestion and hypoxic environment was made by triple-gases incubator. Rat osteoblasts treated with hypoxia for 36 haurs. After 36 hours, cell viability, content of reactive oxygen species (ROS), analysis of cellular cycle and apoptosis, expression of proliferating cell nuclear antigen (PCNA), activity of iNOS and area of calcified nodules were detected. Total RNA was isolated and the gene expression of hypoxia inducible factor-1alpha (HIF-1alpha), BCL-2 and Caspase-3 was investigated by Real Time RT-PCR. RESULTS: Genistein could significantly improve cell viability, percentage of G1 phases, area of calcified nodules and decrease apoptosis rate, ROS content, expression of PCNA, activity of iNOS. Besides, mRNA levels of HIF-1alpha and BCL-2 were enhanced and that of Caspase-3 was inhibited. CONCLUSION: Genistein can protect osteoblasts from hypoxia and enhance osteogenic differentiation significantly.

Icariin is more potent than genistein in promoting osteoblast differentiation and mineralization in vitro.

There has been a strong interest in searching for natural therapies for osteoporosis. Genistein, an isoflavone abundant in soy, and icariin, a prenylated flavonol glycoside isolated from Epimedium Herb, have both been identified to exert beneficial effects in preventing postmenopausal bone loss. However, the relative potency in osteogenesis between the individual phytoestrogen flavonoids remains unknown. The present study compared ability of genistein and icariin in enhancing differentiation and mineralization of cultured rat calvarial osteoblasts in vitro. Dose-dependent studies in osteoblast differentiation measuring alkaline phosphatase (ALP) activity revealed optimal concentrations of genistein and icarrin for stimulating osteogenesis to be both at 10(-5) M. Time course studies comparing the two compounds both at 10(-5) M demonstrated that icariin treatment always produced higher ALP activity, more and larger areas of CFU-F(ALP) colonies and mineralized nodules, more osteocalcin secretion, and calcium deposition, and a higher level of mRNA expression of osteogenesis-related genes COL1α2, BMP-2, OSX, and RUNX-2. However, they inhibited the proliferation of osteoblasts to a similar degree. In conclusion, although future in vivo studies are required to investigate whether icariin is more efficient in improving bone mass and/or preventing bone loss, our in vitro studies have demonstrated that icariin has a stronger osteogenic activity than genistein. In addition, while the prenyl group on C-8 of icariin could be the active group that takes part in osteoblastic differentiation and explains its greater potency in osteogenesis, mechanisms of action, and reasons for the relative potency of icariin versus genistein need to be further studied.

Debridement and bone grafting with internal fixation via anterior approach for treatment of cervicothoracic tuberculosis.

The objective of this study was to investigate the outcome of the patients with cervicothoracic tuberculosis who underwent radical debridement, reconstruction with bone autograft or allograft, and plate internal fixation via anterior approach. From June 2000 to December 2010, 20 patients with tuberculosis in cervicothoracic junction underwent a standard cervical approach, which was combined with a partial median osteotomy and transverse osteotomy through the synostosis between the manubrium and body of the sternum to expose the lesion adequately. Radical debridement was performed, then tricortical iliac crest bone autograft or allograft was placed and internal fixation was done to reconstruct the spinal column. The pathologic change regions were as follows: 10 in the C7-T1 segment, 6 in the T1 segment, 3 in the T1-T3 segment, and 1 in the T2-T3 segment. The classifications of Frankel were as follows: 2 at grade A, 4 at grade B, 7 at grade C, 2 at grade D, and 5 at grade E. There was no injury of blood vessel, spinal cord, or recurrent nerve during the surgery. The follow-up period ranged approximately 16 to 39 months. Bony fusion was obtained in all patients, and there was no internal fixation failure and tuberculosis recurrence in any of these patients. The nerve function of the spinal cord recovered at different degrees: 2 at grade A, 1 at grade B, 1 at grade C, 3 at grade D, and 13 at grade E. The anterior approach can provide direct and safe access to the lesion. The structural iliac crest autograft or allograft and anterior instrumentation could work effectively to stabilize the cervicothoracic junction.

[The changes of iNOS and NO in the osteogenic differentiation process of rat bone marrow stromal cells promoted by icariside II].

This study is to investigate the effects on the expression of iNOS and production of NO in the osteogenic differentiation process of rat bone marrow stromal cells (rBMSCs) by icariside II. rBMSCs were cultured by adherence screening method. When the culture dishes were covered with 80% cells, the osteogenic induced cultures were adopted. Icariside II was supplemented into the culture at 1 x 10(-5) mol x L(-1). The activity of iNOS, content of NO and osteogenic differentiation markers including alkaline phosphatase (ALP) activity, CFU-FALP and mineralized bone nodules were compared among the icariside II-supplemented group, L-NMAE group, icariside II + L-NAME group and the control. Total RNA was isolated and the gene expression of iNOS, Osterix and Runx-2 was investigated by real-time PCR. Total protein was also isolated and the secretion of iNOS and collagen I was examined by Western blotting. Icariside II can significantly improved ALP activity, CFU-FALP amount and mineralized nodules. Besides, the mRNA level of factors related to the osteogenic differentiation includes Osterix and Runx-2 also enhanced. The secretion of collagen I also promoted significantly. But all of these effects can be inhibited by L-NAME which can specifically inhibit the activity of iNOS. Icariside II enhances the osteogenic differentiation of rBMSCs significantly, but if the activity of iNOS was blocked by L-NAME, the osteogenic differentiation markers decrease accompanied with iNOS and NO decrease, suggesting that icariside II stimulates the osteogenic differentiation via enhancing the activity of iNOS and promoting the generation of NO.

[Comparative study on the osteogenic differentiation of rat bone marrow stromal cells effected by icariin and icariside II].

OBJECTIVE: To investigate the effects of icariin and it's main metabolites-icariside II on the osteogenic differentiation of rat bone marrow stromal cells (rBMSCs). METHODS: rBMSCs were cultured by adherence screening method, icariin and icariside II were supplemented into the culture at 5 x 10(-5) mol/L respectively. The osteogenic differentiation markers including alkaline phosphatase (ALP) activity, CFU-F(ALp), osteocalcin secretion, calcium deposition and mineralized bone modulus were compared among the icariin-supplemented group, icariside II and the control. The gene expressions of bFGF, IGF-1, Osterix and Runx-2 were examined by RT-Real Time PCR. RESULTS: Both icariside II and icariin significantly improved ALP activity, CFU-F(ALP) amount, osteocalcin secretion, calcium deposition and mineralized modulus. Besides, they enhanced the gene expressions of bFGF, IGF-1, Osterix and Runx-2. Icariside II was obviously stronger than icariin at the above activities. CONCLUSION: Icariside II is stronger than icariin at enhancing the osteogenic differentiation of rBMSCs, suggesting that icariin can be administered via oral and it's metabolites are the effective constitutes for antiosteoporosis activity.

Reconstruction of degloved thumb with prefabricated flap.

The wrap around the toe flap from the great toe is considered to be a good reconstructive proce-dure for degloved injuries of the thumb. In this study, we used prefabricated flaps of the medial plantar skin to cover a degloved injury of the thumb of a patient unsuitable for application of this method and obtained satisfactory clinical results.

Culture of osteoblasts on bio-derived bones.

OBJECTIVE: To study the effect of bio-derived bones, as substitutes of autogenous bone grafts and demineralized cadaver bones, on the attachment, spreading and proliferation of isolated osteoblasts. METHODS: Osteoblasts were isolated from the calvaria of a fetal rabbit through sequential collagenase digestion. In the attachment study, the osteoblasts labeled with 3H-leucine were incubated with the bio-derived bone materials in sterile microcentrifugable tubes for 15, 90 and 180 minutes, and 24 hours, respectively. The attached cells were collected and the radioactivity was measured with liquid scintillation spectrometry. In the proliferation study, the osteoblasts were cultured with the bio-derived bone materials for 24 hours and 3H-thymidine was added during the last 2 hours of the incubation. The attached cells were collected and the radioactivity was measured with liquid scintillation spectrometry. Osteoblasts were seeded on the bone graft materials for 60 or 120 minutes, 24 or 48 hours, and 3 or 7 days, then the co-culture was processed for scanning electron microscopy to observe the interaction of osteoblasts and the bio-derived bone materials. RESULTS: Osteoblasts attached to the bio-derived bone materials in a time-dependent manner. There were significantly (P<0.05) more attached cells after 180 minutes than after 15 and 90 minutes of incubations (P<0.05). Osteoblasts were proliferated in a large amount on the surface and in the materials. Osteoblasts seeded onto 100 mg bio-derived bones resulted in significantly (P<0.05) more measurable proliferation than those seeded onto 10 mg bones. Osteoblasts appeared round as they attached to the materials, then flattened and spread over with time passing. CONCLUSIONS: Bio-derived bones can provide a good environment for the attachment and proliferation of osteoblasts.