Functional Evaluation of a Bioartificial Liver Support System Using Immobilized Hepatocyte Spheroids in a Porcine Model of Acute Liver Failure.

Bioartificial livers (BAL) may offer acute liver failure (ALF) patients an opportunity for cure without liver transplantation. We evaluated the efficacy of a spheroid-based BAL system, containing aggregates of porcine hepatocytes, in a porcine model of ALF. ALF pigs were divided into three groups. The control group consisted of treatment naïve pigs (n = 5), blank group consisted of pigs that were attached to the BAL system not containing hepatocytes for 12 hours (n = 5) and BAL group consisted of pigs that were attached to the BAL containing hepatocytes for 12 hours (n = 5). Increase in serum ammonia levels were significantly greater in the blank group (P < 0.01) and control group (P < 0.01), compared to the BAL group during the treatment period. Increase in ICP was significantly greater in the control group compared to the BAL group (P = 0.01). Survival was significantly prolonged in the BAL group compared to the blank group (P = 0.03). A BAL system with a bioreactor containing hepatocyte spheroids showed effective clearance of serum ammonia, preservation of renal function and delayed ICP increase in a porcine model of ALF.

Combined effect of pulsed electromagnetic field and sound wave on In vitro and In vivo neural differentiation of human mesenchymal stem cells.

Biophysical wave stimulus has been used as an effective tool to promote cellular maturation and differentiation in the construction of engineered tissue. Pulsed electromagnetic fields (PEMFs) and sound waves have been selected as effective stimuli that can promote neural differentiation. The aim of this study was to investigate the synergistic effect of PEMFs and sound waves on the neural differentiation potential in vitro and in vivo using human bone marrow mesenchymal stem cells (hBM-MSCs). In vitro, neural-related genes in hBM-MSCs were accelerated by the combined exposure to both waves more than by individual exposure to PEMFs or sound waves. The combined wave also up-regulated the expression of neural and synaptic-related proteins in a three-dimensional (3-D) culture system through the phosphorylation of extracellular signal-related kinase. In a mouse model of photochemically induced ischemia, exposure to the combined wave reduced the infarction volume and improved post-injury behavioral activity. These results indicate that a combined stimulus of biophysical waves, PEMFs and sound can enhance and possibly affect the differentiation of MSCs into neural cells. Our study is meaningful for highlighting the potential of combined wave for neurogenic effects and providing new therapeutic approaches for neural cell therapy. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 33:201-211, 2017.

Neuroprotective Effect of Low Frequency-Pulsed Electromagnetic Fields in Ischemic Stroke.

Low frequency-pulsed electromagnetic fields (LF-PEMFs) affect many biological processes; however, the fundamental mechanisms responsible for these effects remain unclear. Our study aimed to investigate the effect of LF-PEMFs on neuroprotection after ischemic stroke. C57B6 mice were exposed to LF-PEMF (F = 60 Hz, Bm = 10 mT) after photothrombotic occlusion. We measured the BDNF/TrkB/Akt signaling pathway, pro-apoptotic and pro-survival protein and gene expressions, and the expression of inflammatory mediators and performed behavioral tests in both LF-PEMF-treated and untreated ischemic stroke mice. Our results showed that LF-PEMF treatment promotes activation of the BDNF/TrkB/Akt signaling pathway. Subsequently, pro-survival proteins were significantly increased, while pro-apoptotic proteins and inflammatory mediators were decreased in ischemic stroke mice after LF-PEMF treatment. The results demonstrated that LF-PEMF exposure has a neuroprotective effect after ischemic stroke in mice during the recovery process.

Effect of human mesenchymal stem cell transplantation on cerebral ischemic volume-controlled photothrombotic mouse model.

Various animal models of stroke have been developed to simulate the human stroke with the development of the ischemic method facilitates preclinical stroke research. The photothrombotic ischemia model, based on the intravascular photochemical reaction, is widely used for in vivo studies. However, this study has limitations, which generated a relatively small-sized infarction model on superficial cortex compared to that of the MCAO stroke model. In this study, the photothorombosis mouse model is adapted and the optimum conditions for generation of cell death and deficits with high reproducibility is determined. The extent of damage within the cortex was assessed by infarct volume and cellular/behavioral analyses. In this model, the neural cell death and inflammatory responses is detected; moreover, the degree of behavioral impairment is correlated with the brain infarct volume. Further, to enhance the understanding of neural repair, the effect of neural differentiation by transplantation of human bone marrow-derived mesenchymal stem cells (BM-MSCs) is analyzed. The authors demonstrated that transplantation of BM-MSCs promoted the neural differentiation and behavioral performance in their photothrombosis model. Therefore, this research was meaningful to provide a stable animal model of stroke with low variability. Moreover, this model will facilitate development of novel MSC-based therapeutics for stroke.

Sound Waves Induce Neural Differentiation of Human Bone Marrow-Derived Mesenchymal Stem Cells via Ryanodine Receptor-Induced Calcium Release and Pyk2 Activation.

Mesenchymal stem cells (MSCs) have shown considerable promise as an adaptable cell source for use in tissue engineering and other therapeutic applications. The aims of this study were to develop methods to test the hypothesis that human MSCs could be differentiated using sound wave stimulation alone and to find the underlying mechanism. Human bone marrow (hBM)-MSCs were stimulated with sound waves (1 kHz, 81 dB) for 7 days and the expression of neural markers were analyzed. Sound waves induced neural differentiation of hBM-MSC at 1 kHz and 81 dB but not at 1 kHz and 100 dB. To determine the signaling pathways involved in the neural differentiation of hBM-MSCs by sound wave stimulation, we examined the Pyk2 and CREB phosphorylation. Sound wave induced an increase in the phosphorylation of Pyk2 and CREB at 45 min and 90 min, respectively, in hBM-MSCs. To find out the upstream activator of Pyk2, we examined the intracellular calcium source that was released by sound wave stimulation. When we used ryanodine as a ryanodine receptor antagonist, sound wave-induced calcium release was suppressed. Moreover, pre-treatment with a Pyk2 inhibitor, PF431396, prevented the phosphorylation of Pyk2 and suppressed sound wave-induced neural differentiation in hBM-MSCs. These results suggest that specific sound wave stimulation could be used as a neural differentiation inducer of hBM-MSCs.

Enhancement of osseointegration of artificial ligament by nano-hydroxyapatite and bone morphogenic protein-2 into the rabbit femur.

The MTT assay showed that the cell proliferation on hydroxyapatite (HAp) and HAp/bone morphogenic protein (BMP) coated group was better than the control and BMP coated groups at 5 days. And after 7 days of culture, the mRNA expression levels of type I collagen, osteonectin, osteopontin, bonesialoprotein, BMP-2, alkaline phosphatase (ALP) and Runx-2 in the HAp/BMP coated group were significantly higher than the other groups. Also, in this group showed the most significant induction of osteogenic gene expression compared to mesenchymal stem cells (MSCs) grown on the other groups. In addition, the cells in the HAp/BMP coated group delivered higher levels of ALP than the other three groups. Also, silk scaffolds were implanted as artificial ligaments in knees of rabbits, and they were harvested 1 and 3 months after implantation. On gross examination, HE staining showed that new bone tissue formation was more observed in the HAp/BMP coated group 3 weeks postoperatively. And masson staining showed that in the HAp/BMP coated group, the silk fibers were encircled by osteoblast, chondrocyte, and collagen. Furthermore, the analysis showed that the width of the graft-bone interface in the HAp and HAp/BMP coated group was narrower than that in the other two groups 3 weeks postoperatively. So, it is concluded that BMP incorporated HAp coated silk scaffold can be enhanced osseointegration and osteogenesis in bone tunnel. As a result, these experimental designs have been demonstrated to be effective in the acceleration of graft-to-bone healing by increasing new bone or fibrocartilage formation at the interface between graft and bone.

Pigmentation effect of electromagnetic fields at various intensities to melanocytes.

Melanogenesis is the biological process that results in the synthesis of skin pigment of melanin and it has various functions in living systems and is synthesized by the melanosome within the melanocytes. A variety of physical treatments are used to promote melanin production in the melanocytes for pigmentation control. The purpose of this study was to evaluate the intensity-dependent effect of extremely low-frequency electromagnetic fields (ELF-EMFs) on melanogenesis by melanocytes in vitro. Melanocytes were exposed to ELF-EMFs at a frequency of 50 Hz and at intensities in the range of 0.5-20 G over 4 days. The results of lactate dehydrogenase assay showed that there were no significant differences between cells exposed to 0.5 G or 2 G groups and the controls. The melanin contents increased 1.2-1.5-fold in cells exposed to ELF-EMFs and tyrosinase activity increased 1.3-fold in cells exposed to ELF-EMFs, relative to the controls. Also, exposure to ELF-EMFs was associated with activation in cyclic-AMP response element binding protein and microphthalmia-associated transcription factor (MITF) was up-regulated. Up-regulation of MITF induces the expression of melanogenesis-related markers, such as tyrosinase, tyrosinase-related protein (TRP)-1, TRP-2. In conclusion, the present study showed that the exposure to ELF-EMFs at low intensities can stimulate melanogenesis in melanocyte, and these results may be used to a therapeutic devices for inducing repigmentation in vitiligo patients.

Regeneration of rabbit calvarial defects using cells-implanted nano-hydroxyapatite coated silk scaffolds.

BACKGROUND: The aim of this study was to characterize the efficacy of nano-hydroxyapatite-coated silk fibroin constructs as a scaffold for bone tissue engineering and to determine the osteogenic effect of human dental pulp and periodontal ligament derived cells at an early stage of healing in rabbits. 3D silk fibroin constructs were developed and coated using nano-hydroxyapatite crystals. Dental pulp and periodontal ligament cells from extracted human third molars were cultured and seeded onto the silk scaffolds prior to in vivo implantation into 8 male New Zealand White rabbits. Four circular windows 8 mm in diameter were created in the calvarium of each animal. The defects were randomly allocated to the groups; (1) silk scaffold with dental pulp cells (DPSS), (2) silk scaffold with PDL cells (PDLSS), (3) normal saline-soaked silk scaffold (SS), and (4) empty control. The animals were sacrificed 2 (n = 4) or 4 weeks (n = 4) postoperatively. The characteristics of the silk scaffolds before and after cell seeding were analyzed using SEM. Samples were collected for histologic and histomorphometic analysis. ANOVA was used for statistical analysis. RESULT: Histologic view of the experimental sites showed well-maintained structure of the silk scaffolds mostly unresorbed at 4 weeks. The SEM observations after cell-seeding revealed attachment of the cells onto silk fibroin with production of extracellular matrix. New bone formation was observed in the 4 week groups occurring from the periphery of the defects and the silk fibers were closely integrated with the new bone. There was no significant difference in the amount of bone formation between the SS group and the DPSS and PDLSS groups. CONCLUSION: Within the limitations of this study, silk scaffold is a biocompatible material with potential expediency as an osteoconductive scaffold in bone tissue engineering. However, there was no evidence to suggest that the addition of hDPCs and hPDLCs to the current rabbit calvarial defect model can produce an early effect in augmenting osteogenesis.

Enhanced Cellular Uptake of Silica-Coated Magnetite Nanoparticles Compared with PEG-Coated Ones in Stem Cells.

Monodispersed magnetite (Fe3O4) nanoparticles (NPs) were prepared through the thermal decomposition method. The obtained NPs were surface modified with silica (SiO2) and polyethylene glycol (PEG), to enhance their stability in aqueous environment and their cellular uptake efficiency for biomedical applications. The NPs were characterized by X-ray diffraction (XRD), high-resolution transmission electron microscopy (HR-TEM), Fourier transform infrared (FT-IR) spectroscopy, and dynamic light scattering (DLS). The cytotoxicity of these NPs on bone marrow mesenchymal stem cells (BM-MSCs) was measured by MTT assay (cell viability test) at various concentrations (2, 5, 12.5, 25, and 50 µg/mL). The cells remained more than 90% viable at concentrations as high as 50 µg/mL. To compare the cellular uptake efficiency, these NPs were treated in BM-MSCs and the Fe concentration within the cells was measured by inductively coupled plasma-atomic emission spectrometry (ICP-AES) analysis. The uptake process displayed a time- and dose-dependency. The uptake amount of SiO2-coated Fe3O4 (Fe3O4@SiO2) NPs was about 10 times higher than that of the PEG-coated ones (Fe3O4@PEG).

Overexpressed Calponin3 by Subsonic Vibration Induces Neural Differentiation of hUC-MSCs by Regulating the Ionotropic Glutamate Receptor.

In this study, we used proteomics to investigate the effects of sonic vibration (SV) on mesenchymal stem cells derived from human umbilical cords (hUC-MSCs) during neural differentiation to understand how SV enhances neural differentiation of hUC-MSCs. We investigated the levels of gene and protein related to neural differentiation after 3 or 5 days in a group treated with 40-Hz SV. In addition, protein expression patterns were compared between the control and the 40-Hz SV-treated hUC-MSC groups via a proteomic approach. Among these proteins, calponin3 (CNN3) was confirmed to have 299 % higher expression in the 40-Hz SV stimulated hUC-MSCs group than that in the control by Western blotting. Notably, overexpression of CNN3-GFP in Chinese hamster ovary (CHO)-K1 cells had positive effects on the stability and reorganization of F-actin compared with that in GFP-transfected cells. Moreover, CNN3 changed the morphology of the cells by making a neurite-like form. After being subjected to SV, messenger RNA (mRNA) levels of glutamate receptors such as PSD95, GluR1, and NR1 as well as intracellular calcium levels were upregulated. These results suggest that the activity of glutamate receptors increased because of CNN3 characteristics. Taken together, these results demonstrate that overexpressed CNN3 during SV increases expression of glutamate receptors and promotes functional neural differentiation of hUC-MSCs.

Illite improves memory impairment and reduces Aß level in the Tg-APPswe/PS1dE9 mouse model of Alzheimer׳s disease through Akt/CREB and GSK-3ß phosphorylation in the brain.

ETHNOPHARMACOLOGICAL RELEVANCE: The use of illite in Korean medicine has a long history as a therapeutic agent for various cerebrovascular diseases. According to Dongui Bogam, illite can be used for Qi-tonifying, phlegm dispersing and activation of blood circulation which is an important principle for the treatment of brain-associated diseases. AIM OF THE STUDY: This study was undertaken to evaluate beneficial effects of illite on the neurodegenerative diseases such as Alzheimer׳s disease (AD). MATERIAL AND METHODS: The transgenic mice of AD, Tg-APPswe/PS1dE9, were fed with 1% or 3% of illite for 3 months. Behavioral, immunological and ELISA analyses were used to assess memory impairment with additional measurement of Aß accumulation and plaque deposition in the brain. Other in vitro studies were performed to examine whether illite inhibits the Aß-induced neurotoxicity in human neuroblastoma cell line, SH-SY5Y cells. RESULTS: Illite treatment rescued Aß-induced neurotoxicity on SH-SY5Y cells, which was dependent on the PI3K/Akt activation. Intake of illite improved the Aß-induced memory impairment and suppressed Aß levels and plaque deposition in the brain of Tg-APPswe/PS1dE9 mice. Illite increased CREB, Akt, and GSK-3ß phosphorylation and suppressed tau phosphorylation in the AD-like brains. Moreover, 1% of illite reduced weight gain and suppressed glucose level in the blood. CONCLUSION: The present study suggests that illite has the potential to be a useful adjunct as a therapeutic drug for the treatment of AD.

Electromagnetic fields and nanomagnetic particles increase the osteogenic differentiation of human bone marrow-derived mesenchymal stem cells.

Human bone marrow-derived mesenchymal stem cells (hBM-MSCs) are widely used in a number of cell therapies and have osteogenic differentiation capacity. Exposure to electromagnetic fields (EMFs) increases the osteogenic differentiation of hBM-MSCs. Nanomagnetic particles (MPs) also promote the differentiation potential of stem cells. In the present study, we investigated the effects of EMFs and MPs on the osteogenic differentiation of hBM-MSCs. hBM-MSCs were treated with 50 µg/ml of Fe3O4 MPs or exposed to a frequency of 45 Hz and an intensity of 1 mT EMF twice every 8 h per day for 7 days. MP incorporation, EMF exposure and MP incorporation with exposure to EMFs did not induce cytotoxic effects. A strong expression of osteogenic markers (osteocalcin, osteopontin and osteonectin) and von Kossa staining intensity was observed in the cells treated with MPs, the cells exposed to EMFs and in the cells treated with MPs and exposed to EMFs compared with the control group, as shown by immunohistochemical staining. Quantitative RT-PCR revealed that the mRNA expression levels of osteoblast markers [osteocalcin, osteopontin, osteonectin, collagen Ⅰ, collagen Ⅲ, bone morphogenetic protein 2 (BMP-2), bone sialoprotein (BSP) and runt-related transcription factor 2 (runx-2)] were markedly increased in the cells treated with MPs and exposed to EMFs. Furthermore, the mRNA expression of calcium channels (CACNA1C, CACNA1E, CACNA1G and CACNA1I) was activated during osteogenic differentiation. The expression levels of osteogenesis-related proteins (BSP, BMP-2, osteopontin and osteonectin) and phosphorylated extracellular signal-regulated kinase (p-ERK) were increased in the cells treated with MPs, those exposed to EMFs and in the cells treated with MPs and exposed to EMFs compared with the control group, as shown by western blot analysis. Fluorescence-activated cell sorting (FACS) analysis was performed for the hBM-MSC markers, CD73, CD90 and CD105. The expression levels of hBM-MSC surface antigens were decreased in the cells treated with MPs, those exposed to EMFs and in the cells treated with MPs and exposed to EMFs compared with the control group. The cell numbers were determined to be approximately 3.4 x 10(5) cells in the control group, 3.7 x 10(5) cells in the MP-treated group, 3.1 x 10(5) cells in the group exposed to EMFs and 3.9 x 10(5) cells in the group treated with MPs and exposed to EMFs. The cell mitochondrial activity among the 4 experimental groups was similar. The hBM-MSCs treated with MPs and exposed to EMFs showed an increase in alkaline phosphatase (ALP) activity. Taken together, these results suggest that the treatment of hBM-MSCs with MPs or exposure to EMFs increases osteogenic differentiation, and that treatment with MPs in conjunction with EMF exposure is more effective in increasing osteogenic differentiation.

Enhancing proliferation and ECM expression of human ACL fibroblasts by sonic vibration.

Effects of mechanical vibration on cell activity and behavior remain controversial: There has been evidence on both positive and negative effects. Furthermore, research on the anterior cruciate ligament (ACL) has as yet been limited and the frequency-related effects remain unknown, even though ACL injury is common and an injured ACL hardly spontaneously recovers. The object of this work was to address the influence of mechanical vibration on ACL fibroblasts, to determine the effects of frequencies, and to further study this effect at the cellular level. We found that sonic vibration affected ACL fibroblasts' proliferation and metabolism in a frequency-dependent manner, and 20 Hz gave rise to the most ACL cell activity and comprehensively increased extracellular matrix (ECM) contents, including collagen type I, collagen type III, fibronectin, elastin, tenascin, glycosaminoglycan (GAG), and the cytoskeleton protein vimentin. Thus, our results indicate that sonic vibration possesses frequency-dependent effects on proliferation and productivity of ACL fibroblast with an optimal frequency of 20 Hz under the present stimulation conditions, providing further information for future research in how vibrational stimulation manipulates ACL cellular behavior.

Electromagnetic fields mediate efficient cell reprogramming into a pluripotent state.

Life on Earth is constantly exposed to natural electromagnetic fields (EMFs), and it is generally accepted that EMFs may exert a variety of effects on biological systems. Particularly, extremely low-frequency electromagnetic fields (EL-EMFs) affect biological processes such as cell development and differentiation; however, the fundamental mechanisms by which EMFs influence these processes remain unclear. Here we show that EMF exposure induces epigenetic changes that promote efficient somatic cell reprogramming to pluripotency. These epigenetic changes resulted from EMF-induced activation of the histone lysine methyltransferase Mll2. Remarkably, an EMF-free system that eliminates Earth's naturally occurring magnetic field abrogates these epigenetic changes, resulting in a failure to undergo reprogramming. Therefore, our results reveal that EMF directly regulates dynamic epigenetic changes through Mll2, providing an efficient tool for epigenetic reprogramming including the acquisition of pluripotency.

Stimulation of neural differentiation in human bone marrow mesenchymal stem cells by extremely low-frequency electromagnetic fields incorporated with MNPs.

Human bone marrow-derived mesenchymal stem cells (hBM-MSCs) have been investigated as a new cell-therapeutic solution due to their capacity that could differentiate into neural-like cells. Extremely low-frequency electromagnetic fields (ELF-EMFs) therapy has emerged as a novel technique, using mechanical stimulus to differentiate hBM-MSCs and significantly enhance neuronal differentiation to affect cellular and molecular reactions. Magnetic iron oxide (Fe3O4) nanoparticles (MNPs) have recently achieved widespread use for biomedical applications and polyethylene glycol (PEG)-labeled nanoparticles are used to increase their circulation time, aqueous solubility, biocompatibility, and nonspecific cellular uptake as well as to decrease immunogenicity. Many studies have used MNP-labeled cells for differentiation, but there have been no reports of MNP-labeled neural differentiation combined with EMFs. In this study, synthesized PEG-phospholipid encapsulated magnetite (Fe3O4) nanoparticles are used on hBM-MSCs to improve their intracellular uptake. The PEGylated nanoparticles were exposed to the cells under 50 Hz of EMFs to improve neural differentiation. First, we measured cell viability and intracellular iron content in hBM-MSCs after treatment with MNPs. Analysis was conducted by RT-PCR, and immunohistological analysis using neural cell type-specific genes and antibodies after exposure to 50 Hz electromagnetic fields. These results suggest that electromagnetic fields enhance neural differentiation in hBM-MSCs incorporated with MNPs and would be an effective method for differentiating neural cells.

Silk and collagen scaffolds for tendon reconstruction.

In this study, silk thread (Bombyx mori) was braided to a tube-like shape and sericin was removed from the silk tube. Thereafter, collagen/chondroitin-6-sulfate solution was poured into the silk tube, and the lyophilization process was performed. To assess the inflammatory response in vivo, raw silk and sericin-free silk tubes were implanted in the subcutaneous layer of mice. After 10 days of in vivo implantation, mild inflammatory responses were observed around the sericin-free silk tubes, and severe inflammation with the presence of neutrophils and macrophages was observed around the raw silk tubes. At 24 weeks post implantation, the regenerated tendon had a thick, cylindrical, grayish fibrous structure and a shiny white appearance, similar to that of the native tendon in the rabbit model of tendon defect. The average tensile strength of the native tendons was 220 ± 20 N, whereas the average tensile strength of the regenerated tendons was 167 ± 30 N and the diameter of the regenerated tendon (3 ± 0.2 mm) was similar to that of the native tendons (4 ± 0.3 mm). Histologically, the regenerated tendon resembled the native tendon, and all the regenerated tissues showed organized bundles of crimped fibers. Masson trichrome staining was performed for detecting collagen synthesis, and it showed that the artificial tendon was replaced by new collagen fibers and extracellular matrix. However, the regenerated tendon showed fibrosis to a certain degree. In conclusion, the artificial tendon, comprising a braided silk tube and lyophilized collagen sponge, was optimal for tendon reconstruction. Thus, this study showed an improved regeneration of neo-tendon tissues, which have the structure and tensile strength of the native tendon, with the use of the combination of collagen and silk scaffold.

Comparison of light-emitting diode wavelength on activity and migration of rabbit ACL cells.

The purpose of this study was to evaluate the biological response and gene expression of New Zealand White Rabbit anterior cruciate ligament (ACL) fibroblasts for different wave lengths of light-emitting diode (LED) irradiation. In other words, this study was undertaken to evaluate the effects of different wavelengths of LED irradiation on cell growth, expression of extracellular matrix and growth factors, migration, and expression of actin and integrin. Proliferation assay showed that red (630 nm, 9.5 J/cm(2)) and green LED (530 nm, 9.8 J/cm(2)) irradiated cells were more increased than control group but there was no difference between the control group and the blue LED (460 nm, 27 J/cm(2)) irradiated group. Moreover, the expression of insulin-like growth factor, transforming growth factor-beta (TGF-ß1), and collagen I were significantly increased in the red and green LED-irradiated group, but the expression of collagen was decreased in the blue LED-irradiated group. The results of staining showed that collagen and TGF-ß1 were weaker in the control group and blue LED-irradiated cells, but stronger in the red and green LED-irradiated cells. Also, in the red and green LED-irradiated group, the expression of actin and integrin was not changed compared to the control group, but the expression of actin and integrin was decreased in the blue irradiated group. This study revealed that irradiation with a wavelength of 460 nm (blue LED) is cytotoxic to ACL cells, but irradiation with nontoxic fluencies of 530 (green LED) and 630 nm (red LED) wavelengths induced cell growth in cultured ACL cells.

Effects of magnetic nanoparticle-incorporated human bone marrow-derived mesenchymal stem cells exposed to pulsed electromagnetic fields on injured rat spinal cord.

Transplanting mesenchymal stem cells into injured lesions is currently under study as a therapeutic approach for spinal cord injury. In this study, the effects of a pulsed electromagnetic field (PEMF) on injured rat spinal cord were investigated in magnetic nanoparticle (MNP)-incorporated human bone marrow-derived mesenchymal stem cells (hBM-MSCs). A histological analysis revealed significant differences in MNP-incorporated cell distribution near the injured site under the PEMF in comparison with that in the control group. We confirmed that MNP-incorporated cells were widely distributed in the lesions under PEMF. The results suggest that MNP-incorporated hBM-MSCs were guided by the PEMF near the injured site, and that PEMF exposure for 8 H per day over 4 weeks promoted behavioral recovery in spinal cord injured rats. The results show that rats with MNP-incorporated hBM-MSCs under a PEMF were more effective on the Basso, Beattie, and Bresnahan behavioral test and suggest that the PEMF enhanced the action of transplanted cells for recovery of the injured lesion.

Immobilization of BMP-2 on a nano-hydroxyapatite-coated titanium surface using a chitosan calcium chelating agent.

We conducted experiments to determine the most effective calcium chelating agents for use in enhancing adhesion of human bone marrow mesenchymal stem cells (BM-MSCs) on nano-hydroxyapatite (nHAp)-coated titanium substrates by covalently immobilizing bone morphogenetic protein-2 (BMP-2). The quantity of amine groups on the chitosan chelated surface was 7 µg/surface area, and it was 1.4 µg/surface area on the alendronate chelated surface. The quantity of BMP-2 on the BMP-2 immobilized surface chelated with chitosan (4 ng/surface area) was higher than that on BMP-2 immobilized surface chelated with alendronate (2.2 ng/surface area). Contact angles of the nHAp-coated titanium, alendronate chelated, chitosan chelated, and BMP-2 immobilized surfaces chelated with alendronate were 68.8 ± 3.6°, 78.2 ± 1.9°, 74.8 ± 5.2°, and 76.0 ± 2.5°, respectively. The contact angle of the BMP-2 immobilized surface chelated with chitosan was significantly lower (56.2 ± 2.0°) than that of any of the other groups. BM-MSCs on the chitosan surface and BMP-2 immobilized on the surface chelated with chitosan appeared to be healthy and showed a spindle-like fibroblastic morphology. In addition, BM-MSCs on these surfaces appeared to have the ability to differentiate into bone-forming cells. We suggest that chitosan can be used as an effective calcium chelating agent for implants.

Electromagnetic fields induce neural differentiation of human bone marrow derived mesenchymal stem cells via ROS mediated EGFR activation.

Even though the inducing effect of electromagnetic fields (EMF) on the neural differentiation of human bone marrow mesenchymal stem cells (hBM-MSCs) is a distinctive, the underlying mechanism of differentiation remains unclear. To find out the signaling pathways involved in the neural differentiation of BM-MSCs by EMF, we examined the CREB phosphorylation and Akt or ERK activation as an upstream of CREB. In hBM-MSCs treated with ELF-EMF (50 Hz, 1 mT), the expression of neural markers such as NF-L, MAP2, and NeuroD1 increased at 6 days and phosphorylation of Akt and CREB but not ERK increased at 90 min in BM-MSCs. Moreover, EMF increased phosphorylation of epidermal growth factor receptor (EGFR) as an upstream receptor tyrosine kinase of PI3K/Akt at 90 min. It has been well documented that ELF-MF exposure may alter cellular processes by increasing intracellular reactive oxygen species (ROS) concentrations. Thus, we examined EMF-induced ROS production in BM-MSCs. Moreover, pretreatment with a ROS scavenger, N-acetylcystein, and an EGFR inhibitor, AG-1478, prevented the phosphorylation of EGFR and downstream molecules. These results suggest that EMF induce neural differentiation through activation of EGFR signaling and mild generation of ROS.