Can FLAIR hyperintense vessel (FHV) signs be influenced by varying MR parameters and flow velocities? A flow phantom analysis.

BACKGROUND: Fluid-attenuated inversion recovery (FLAIR) hyperintense vessels (FHVs) have been used to assess leptomeningeal collateral flow in acute ischemic stroke. However, prior FHVs studies showed inconsistent results, which may be ascribable to different magnetic resonance (MR) parameters used. PURPOSE: To evaluate whether FHVs could be influenced by varying MR parameters and flow velocities, using a flow phantom. MATERIAL AND METHODS: A total of 512 sets of FLAIRs were performed with varying parameters and flow velocities, using a flow phantom. Flow phantom was manufactured with 3.5% agarose solution, an 8-mm inner diameter silicone tube and non-pulsatile pump. Varying MR parameters were repetition time (TR)/inversion time (TI), echo time (TE), flip angle (FA) of refocusing pulse, and periodically rotated overlapping parallel lines with enhanced reconstruction (PROPELLER). The signal intensity of flow were measured and regarded as the degree of FHVs. Simple and multiple linear regression analyses were applied to evaluate the association between the degree of FHVs and varying MR parameters as well as flow velocities. RESULTS: On univariate analysis, PROPELLER technique (R(2 )= 0.448) demonstrated strongest correlation with the degree of FHV, followed by flow velocities (R(2 )= 0.204), FA (R(2 )= 0.126), and TE (R(2 )= 0.031), whereas TR/TI showed no significant correlations. On multivariate analysis, TE, FA, PROPELLER technique, and flow velocities were independent factors influencing the degree of FHVs (<0.001). CONCLUSION: Flow velocities, FA of refocusing pulse, TE, and PROPELLER technique significantly affected the degree of FHVs. Optimized MR parameters should be used consistently in future studies, which may provide more reliable results.

Astroglial Activation by an Enriched Environment after Transplantation of Mesenchymal Stem Cells Enhances Angiogenesis after Hypoxic-Ischemic Brain Injury.

Transplantation of mesenchymal stem cells (MSCs) has paracrine effects; however, the effects are known to be largely limited. Here we investigated the combination effects of cell transplantation and enriched environment (EE) in a model of hypoxic-ischemic brain injury. Brain damage was induced in seven-day-old mice by unilateral carotid artery ligation and exposure to hypoxia (8% O2 for 90 min). At six weeks of age, the mice were randomly assigned to four groups: phosphate-buffered saline (PBS)-control (CON), PBS-EE, MSC-CON, and MSC-EE. Rotarod and grip strength tests were performed to evaluate neurobehavioral functions. Histologic evaluations were also performed to confirm the extent of astrocyte activation and endogenous angiogenesis. An array-based multiplex ELISA and Western blot were used to identify growth factors in vivo and in vitro. Two weeks after treatment, levels of astrocyte density and angiogenic factors were increased in MSC-EE mice, but glial scarring was not increased. Eight weeks after treatment, angiogenesis was increased, and behavioral outcomes were synergistically improved in the MSC-EE group. Astrocytes co-cultured with MSCs expressed higher levels of angiogenic factors than astrocytes cultured alone. The mechanisms of this synergistic effect included enhanced repair processes, such as increased endogenous angiogenesis and upregulation of angiogenic factors released from activated astrocytes.

The effects of AEB071 (sotrastaurin) with tacrolimus on rat heterotopic cardiac allograft rejection and survival.

BACKGROUND: AEB071 (sotrastaurin) is a specific inhibitor of protein kinase C that prevents T-cell activation. Our previous study demonstrated that AEB071 monotherapy could prevent acute cardiac allograft rejection in rats. Herein, we investigated the effects of AEB071 combined with various doses of tacrolimus (Tac) on the allograft rejection and survival in a rat heart transplantation model. MATERIALS AND METHODS: Heterotopic cardiac transplantation from Brown-Norway to Lewis rats was performed. Cardiac allograft survival was assessed by monitoring heartbeats in six recipients of each experimental group. Another four recipient rats were selectively sacrificed in each group at d 7 post-transplantation for histologic examination. Serum transaminases, blood urea nitrogen, and creatinine concentrations were measured. RESULTS: AEB071 monotherapy prolonged allograft mean survival time (MST) compared with the untreated control group. Also a combination of AEB071 and Tac prolonged MST compared with monotherapy groups with higher dose of Tac. In the cardiac graft histology, AEB071 combined with Tac 0.6 mg/kg/d significantly decreased the rejection grade as indicative of decreased inflammatory cell infiltration into the graft. No experimental group was found with any abnormal histologic or serologic evidence of liver and kidney toxicity. CONCLUSION: AEB071 combined with a smaller dosage of Tac may be clinically possible to establish calcineurin inhibitor (CNI) minimization protocol in solid organ transplantation.

Development and analysis of a collagen-based hemostatic adhesive.

BACKGROUND: Commercially available hemostatic-adhesive has the risk of disease transmission because it is derived from a plasma component. The purpose of this study is to manufacture a new hemostatic adhesive and evaluate its performances. METHODS: Two atelocollagen-based hemostatic adhesives were produced and named as Bleestop A (2% esterified atelocollagen in 75% ethanol + 1% CaCl(2) + 0.71% DOPA + 0.1% tranexamic acid in DW) and Bleestop B (2% esterified atelocollagen in DW + 1% CaCl(2) + 0.71% DOPA + 0.1% aminocaproic acid in DW). These are compared with the negative control group (no adhesives used group) and the positive control group (Tissucol Duo Quick group). The adult male Sprague-Dawley rat model was adopted and the hemostatic-adhesion activities of each group were assessed by the adhesion strength test and the morphologic features of adhesion. The liver tissues were used. Histologic assessment of adhesion was accessed using light microscopy. RESULTS: Bleeding was controlled immediately after application of Bleestop A and partial adhesion was observed after 30 s. More than 95% area of the resected surface attached after 1 min. In Bleestop B, partial adhesion was observed after 30 s. More than 95% area of the resected surface was attached after 45 s. Histologic evaluation showed that Bleestop A and B mediated organized adhesion rapidly between both resected tissue surfaces. The adhesion strength of Bleestop A and B was better than the negative control group and showed same adhesion strength as the positive control group after 3 min. CONCLUSION: Bleestop A and B showed significant hemostasis and adhesion capability within 1 min. They are comparable to the positive control group.

Dexamethasone increases angiopoietin-1 and quiescent hematopoietic stem cells: a novel mechanism of dexamethasone-induced hematoprotection.

Angiopoietin-1 (Ang-1) is known to have hematoprotective effects by increasing the quiescence of hematopoietic stem cells. However, it remains to be determined if the upregulation of Ang-1 and the subsequent increase in the quiescence of hematopoietic stem cells are also involved in the dexamethasone (Dex)-mediated bone marrow protection. Here Western blotting and flow cytometric analyses demonstrate that Dex increases the levels of Ang-1 in mouse bone marrow and the quiescence of hematopoietic stem cells. Our data for the first time suggest that the increased quiescence of hematopoietic stem cells provides a novel mechanism of Dex-induced hematoprotection.

Generation of insulin-producing human mesenchymal stem cells using recombinant adeno-associated virus.

The purpose of current experiment is the generation of insulin-producing human mesenchymal stem cells as therapeutic source for the cure of type 1 diabetes. Type 1 diabetes is generally caused by insulin deficiency accompanied by the destruction of islet beta-cells. In various trials for the treatment of type 1 diabetes, cell-based gene therapy using stem cells is considered as one of the most useful candidate for the treatment. In this experiment, human mesenchymal stem cells were transduced with AAV which is containing furin-cleavable human preproinsulin gene to generate insulin-producing cells as surrogate beta-cells for the type 1 diabetes therapy. In the rAAV production procedure, rAAV was generated by transfection of AD293 cells. Human mesenchymal stems cells were transduced using rAAV with a various multiplicity of infection. Transduction of recombinant AAV was also tested using beta-galactosidse expression. Cell viability was determined by using MTT assay to evaluate the toxicity of the transduction procedure. Expression and production of Insulin were tested using reverse transcriptase-polymerase chain reaction and immunocytochemistry. Secretion of human insulin and C-peptide from the cells was assayed using enzyme-linked immunosorbent assay. Production of insulin and C-peptide from the test group represented a higher increase compared to the control group. In this study, we examined generation of insulin-producing cells from mesenchymal stem cells by genetic engineering for diabetes therapy. This work might be valuable to the field of tissue engineering for diabetes treatment.

UV-irradiated parylene surfaces for proliferation and differentiation of PC-12 cells.

PC-12 cells originate from neuroblastic cells, which have an ability to differentiate into neuronlike cells. In this work, the purpose was to estimate the influence of microenvironments on cell attachment and neuritogenesis capacity of PC-12 cells on parylene-N and parylene-C films with and without ultraviolet (UV) light treatment. The estimate of total cell number after incubation for 72h, the ratio of adherent to suspended cells, counting of neurite outgrowths on parylene-N or parylene-C films after UV exposure suggested that these films were suitable for proliferation as well as differentiation of PC-12 cells. The differences in surface properties of parylene-N and parylene-C films with and without UV exposure were analyzed by contact angle measurement, Fourier-transform infrared spectrometry, and X-ray photoelectron spectroscopy. According to these analyses, introduction of oxygen-related chemical functional groups was presumed to result in increased hydrophilicity and efficiency of protein immobilization on parylene-N and parylene-C films after UV treatment. According to fluorescent staining, western blotting, and cell cycle analysis, UV-treated parylene-C and parylene-N films appear to effectively facilitate simultaneous proliferation and differentiation of PC-12 cells with neurite outgrowth.

Effects of sinusoidal electromagnetic field on structure and function of different kinds of cell lines.

This study investigated that whether a 2 mT, 60 Hz, sinusoidal electromagnetic field (EMF) alters the structure and function of cells. This research compared the effects of EMF on four kinds of cell lines: hFOB 1.19 (fetal osteoblast), T/G HA-VSMC (aortic vascular smooth muscle cell), RPMI 7666 (B lymphoblast), and HCN-2 (cortical neuronal cell). Over 14 days, cells were exposed to EMF for 1, 3, or 6 hours per day (hrs/d). The results pointed to a cell type-specific reaction to EMF exposure. In addition, the cellular responses were dependent on duration of EMF exposure. In the present study, cell proliferation was the trait most sensitive to EMF. EMF treatment promoted growth of hFOB 1.19 and HCN-2 compared with control cells at 7 and 14 days of incubation. When the exposure time was 3 hrs/d, EMF enhanced the proliferation of RPMI 7666 but inhibited that of T/G HA- VSMC. On the other hand, the effects of EMF on cell cycle distribution, cell differentiation, and actin distribution were unclear. Furthermore, we hardly found any correlation between EMF exposure and gap junctional intercellular communication in hFOB 1.19. This study revealed that EMF might serve as a potential tool for manipulating cell proliferation.

Influence of in vitro biomimicked stem cell 'niche' for regulation of proliferation and differentiation of human bone marrow-derived mesenchymal stem cells to myocardial phenotypes: serum starvation without aid of chemical agents and prevention of spontaneous stem cell transformation enhanced by the matrix environment.

Niche appears important for preventing the spontaneous differentiation or senescence that cells undergo during in vitro expansion. In the present study, it was revealed that human bone marrow-derived mesenchymal stem cells (hBM-MSCs) undergo senescence-related differentiation into the myocardial lineage in vitro without any induction treatment. This phenomenon occurred over the whole population of MCSs, much different from conventional differentiation with limited frequency of occurrence, and was accompanied by a change of morphology into large, flat cells with impeded proliferation, which are the representative indications of MSC senescence. By culturing MSCs under several culture conditions, it was determined that induction treatment with 5-azacytidine was not associated with the phenomenon, but the serum-starvation condition, under which proliferation is severely hampered, caused senescence progression and upregulation of cardiac markers. Nevertheless, MSCs gradually developed a myocardial phenotype under normal culture conditions over a prolonged culture period and heterogeneous populations were formed. In perspectives of clinical applications, this must be prevented for fair and consistent outcomes. Hence, the biomimetic 'niche' was constituted for hBM-MSCs by cultivating on a conventionally available extracellular matrix (ECM). Consequently, cells on ECM regained a spindle-shape morphology, increased in proliferation rate by two-fold and showed decreased expression of cardiac markers at both the mRNA and protein levels. In conclusion, the outcome indicates that progression of MSC senescence may occur via myocardial differentiation during in vitro polystyrene culture, and this can be overcome by employing appropriate ECM culture techniques.

Adipose tissue engineering using mesenchymal stem cells attached to injectable PLGA spheres.

The reconstruction of soft tissue defects remains a challenge in plastic and reconstructive surgery, and a real clinical need exists for an adequate solution. This study was undertaken in order to differentiate mesenchymal stem cells (MSCs) into adipocytes, and to then assess the possibility of constructing adipose tissue via the attachment of MSCs to injectable PLGA spheres. We also designed injectable PLGA spheres for scar-free transplantation. In this study, MSCs and adipo-MSCs (MSCs cultured in adipogenic medium for 7 days) were attached to PLGA spheres and cultured for 7 days, followed by injection into nude mice for 2 weeks. As a result, the difference between lipid accumulation in adipo-MSCs at 1 and 7 days was much higher in vitro than in the MSCs. Two weeks after injection, a massive amount of new tissue was formed in the APLGA group, whereas only a small amount was formed in the MPLGA group. We verified that the newly formed tissue originated from the injected MSCs via GFP testing, and confirmed that the created tissue was actual adipose tissue by oil red O staining and Western blot (PPAR(gamma) and C/EBP(alpha) were expressed only in APLGA groups). Therefore, this study presents an efficient model of adipose tissue engineering using MSCs and injectable PLGA spheres.

Evaluation of antibiotic-loaded collagen-hyaluronic acid matrix as a skin substitute.

The 1-ethyl-(3-3-dimethylaminopropyl) carbodiimide hydrochloride-crosslinked collagen-hyaluronic acid (HA) matrices containing tobramycin or ciprofloxacin as an antibiotic agent were fabricated for the control of wound contamination and characterized with respect to morphology, mechanical strength, in vitro release, antibacterial activity and cytotoxicity. For the tobramycin loaded matrix, the antibacterial capacity increased with the drug loading. Tobramycin and ciprofloxacin loaded matrices maintained their antibacterial effects for over 96 and 48 h, respectively. However, cell viability testing revealed that 0.4 mg/ml of ciprofloxacin has a cytotoxic effect on fetal human dermal fibroblasts. The effects of the bilayered collagen-HA matrices containing tobramycin and growth factors were also evaluated using an in vivo full thickness dermal defect model. Though the tobramycin incorporated collagen-HA matrix had no significant effect on wound healing compared with the control, the tobramycin incorporated matrix containing basic fibroblast growth factor or platelet-derived growth factor significantly enhanced wound healing. This study demonstrates the potential efficacy of crosslinked collagen-HA matrix containing antibiotics and growth factors for defective skin tissue replacement and infection prevention.

Sustained release of ascorbate-2-phosphate and dexamethasone from porous PLGA scaffolds for bone tissue engineering using mesenchymal stem cells.

The purpose of this research was to develop porous poly(D,L-lactide-co-glycolide) (PLGA) scaffolds from which ascorbate-2-phosphate (AsAP) and dexamethasone (Dex) are continuously released for a month for osteogenesis of mesenchymal stem cells for bone tissue engineering. Porous PLGA matrices containing AsAP and Dex were prepared by solvent casting/particulate leaching method. In vitro release and water uptake studies were performed in Dulbecco's phosphate buffered saline at 37 degrees C and 15 rpm. Drug loading and release rates were determined by high performance liquid chromatography. Release studies of Dex and AsAP showed that, after an initial burst release lasting 4 and 9 days, respectively, release rates followed zero order kinetics with high correlation coefficients at least until 35 days. Incorporation of AsAP into the scaffolds increased the release rates of Dex and AsAP, and the scaffold water uptake. When mesenchymal stem cells (MSCs) were cultured in the AsAP and Dex containing scaffolds in vitro, the amount of mineralization was significantly higher than in control scaffolds. In conclusion, AsAP and Dex were incorporated into porous PLGA scaffolds and continuously released over a month and osteogenesis of MSCs was increased by culture in these scaffolds.

Biological characterization of EDC-crosslinked collagen-hyaluronic acid matrix in dermal tissue restoration.

Porous collagen matrices crosslinked with various amounts of hyaluronic acid (HA) by 1-ethyl-3-(3-dimethyl aminopropyl)carbodiimide (EDC) were developed as scaffolds for dermal tissue regeneration. The effect of HA on cells in accordance with HA concentrations in the collagenous matrices was investigated using cultures of fetal human dermal fibroblasts, and the effect of EDC-crosslinked collagen-HA matrix on wound size reduction was also evaluated in vivo. Scanning electron microscopic views of the matrices demonstrated that all of the collagen-HA matrices had interconnected pores with mean diameters of 150-250 microm. An HA matrix retention test showed that the concentration of HA decreased slowly after an initial rapid decrease over 24h. Fetal human dermal fibroblasts adhered well to all of the collagen-based matrices as compared with the Porous polyurethane matrix used as a control. An 3-(4,5-dimethylthiazolyl)-2,5-diphenyltetrazolium bromide based proliferation test and the hematoxylin and eosin staining of a 2 week cultured matrix showed that the proliferation of fibroblasts was enhanced on a 9.6% HA contained collagen matrix. No significant difference was in terms of fibroblast migration into the various types of scaffolds as HA content was increased. In vivo testing showed that dermis treated with collagen or collagen-HA matrix was thicker than the control, and epithelial regeneration was accelerated, and collagen synthesis increased. However, no significant effect of HA on wound size reduction was found.

Characterization of porous collagen/hyaluronic acid scaffold modified by 1-ethyl-3-(3-dimethylaminopropyl)carbodiimide cross-linking.

In order to develop a scaffolding material for tissue regeneration, porous matrices containing collagen and hyaluronic acid were fabricated by freeze drying at -20 degrees C, -70 degrees C or -196 degrees C. The fabricated porous membranes were cross-linked using 1-ethyl-3-(3-dimethyl aminopropyl) carbodiimide (EDC) in a range of 1-100 mM concentrations for enhancing mechanical stability of the composite matrix. Scanning electron microscope (SEM) views of the matrices demonstrated that the matrices obtained before cross-linking process had interconnected pores with mean diameters of 40, 90 or 230 microm and porosity of 58-66% according to the freezing temperature, and also the porous structures after cross-linking process were retained. The swelling test and IR spectroscopic measurement of different cross-linked membranes were carried out as a measure of the extent of cross-linking. The swelling behavior of cross-linked membranes showed no significant differences as cross-linking degree increased. FT-IR spectra showed the increase of the intensity of the absorbencies at amide bonds (1655, 1546, 1458 cm(-1)) compared to that of CH bond (2930 cm(-1)). In enzymatic degradation test, EDC treated membranes showed significant enhancement of the resistance to collagenase activity in comparison with 0.625% glutaraldehyde treated membranes. In cytotoxicity test using L929 fibroblastic cells, the EDC-cross-linked membranes demonstrated no significant toxicity.

Development of collagenase-resistant collagen and its interaction with adult human dermal fibroblasts.

Collagen is regarded as one of the most useful biomaterials. The excellent biocompatibility and safety due to its biological characteristics, such as biodegradability and weak antigenecity, made collagen the primary source in biomedical application. Collagen has been widely used in the crosslinked form to extend the durability of collagen. The chemical treatment influences the structural integrity of collagen molecule resulting in the loss of triple helical characteristic. The structural characteristic of collagen is importantly related to its biological function for the interaction with cell. In this study, structural stability of collagen was enhanced thought EGCG treatment, resulting in high resistance against degradation by bacterial collagenase and MMP-1, which is confirmed by collagen zymography. The triple helical structure of EGCG-treated collagen could be maintained at 37 degrees C in comparison with collagen, which confirmed by CD spectra analysis, and EGCG-treated collagen showed high free-radical scavenging activity. Also, with fibroblasts culture on EGCG-treated collagen, the structural stability of EGCG-treated collagen provided a favorable support for cell function in cell adhesion and actin filament expression. These observations underscore the need for native, triple helical collagen conformation as a prerequisite for integrin-mediated cell adhesion and functions. According to this experiment, EGCG-treated collagen assumes to provide a practical benefit to resist the degradation by collagenase retaining its structural characteristic, and can be a suitable biomaterial for biomedical application.

Behavior of isolated rat oval cells in porous collagen scaffold.

The oval cell is regarded as a compensatory cell in liver injury, and is thought to be equivalent to liver stem/progenitor cells. Oval cells were induced by the 2-AAF/CCl(4) dietary method in Fischer 344 rats, and were isolated from excised liver by the collagenase perfusion, enzyme treatment, and cell cloning method. Transmission electron microscopy observation and double immunofluorescence methods were used to characterize the cells. We have developed an in vitro system consisting of three-dimensional collagen and hormonal and cytokine factors. Over 3 weeks, albumin secretion and urea detoxification rate were estimated to assess the biological function of the oval cells cultured in a scaffold. Oval cells cultured in the scaffold demonstrated higher biological functions than did those in a two-dimensional tissue culture plate. The pore structure and collagen in a scaffold may play an important role in fostering the biochemical functions of oval cells. The three-dimensional culture of oval cells could be considered in designing a cell-delivering tool for hepatic disease.

Ex vivo mechanical evaluation of carbonate apatite-collagen-grafted porous poly-L-lactic acid membrane in rabbit calvarial bone.

Appropriate mechanical recovery is an important parameter in successful restoration of skeletal defects. Carbonate apatite and type I atelocollagen mixture (CAp) was grafted on a porous poly-L-lactic acid (PLLA) membrane to produce a CAp bilayered PLLA membrane (CAp+PLLA). After implantation, regional mechanical change in the membrane was investigated in rabbit calvarial bone defects. Dynamic viscoelasticity and elastic modulus of the implanted specimen were measured after 2, 4, 8, 12, 16, 26, and 52 weeks. The modulus of the peripheral part was higher than that of the central part of the implanted area, whereas the central part demonstrated a gradual increase. This phenomenon indicates that regeneration initially occurs from the periphery of the calvarial bone. After 26 weeks, stiffness of the central part became similar to that of the periphery in the CAp+PLLA-implanted area. According to this result, measuring viscoelasticity of an implanted biodegradable material would be a useful method to determine degrees of regeneration and replacement of an implanted region.

Effects of green tea polyphenol on preservation of human saphenous vein.

The potential role of green tea polyphenol (GtPP) in preserving the human saphenous vein was investigated under physiological conditions. The vein segments were incubated for 1, 3, 5, 7 and 14 days, either after 4h of treatment with 1.0mg/ml GtPP or in the presence of GtPP at the same concentration. After incubation, the endothelial cell viability, endothelial nitric oxide synthase (eNOS) expression and the vein histology were evaluated. When the veins were not treated with GtPP, the viability of the endothelial cells was significantly reduced with the progress in the culture time, and none of the cells expressed eNOS after 5 days. Furthermore, severe histological changes and structural damage were observed in the non-treated veins. In contrast, incubating the veins after 4h of GtPP treatment significantly prevented these phenomena. The cellular viability of the GtPP-treated vein was approximately 64% after 7 days, and eNOS expression was maintained up to 40%, compared to that of the fresh vein. The histological observations showed that the vasculature was quite similar to that of the fresh vein. When incubated with GtPP, the vein could also be preserved for 1 week under physiological conditions retaining both its cellular viability (61%) and eNOS expression level (45%) and maintaining its venous structure without any morphological changes. These results demonstrate that GtPP treatment may be a useful method for preserving the HSV.

Behavior of fibroblasts on a porous hyaluronic acid incorporated collagen matrix.

A hyaluronic acid (HA) incorporated porous collagen matrix was fabricated at -70 degrees C by lyophilization. The HA incorporated collagen matrix showed increased pore size in comparison with collagen matrix. Biodegradability and mechanical properties of matrices were controllable by varying the ultraviolet (UV) irradiation time for cross-linking collagen molecules. Addition of HA to collagen matrix did not effect ultimate tensile stress after UV irradiation. HA incorporated collagen matrices demonstrated a higher resistance against the collagenase degradation than collagen matrix. In an in vitro investigation of cellular behavior using dermal fibroblasts on the porous matrix, HA incorporated collagen matrix induced increased dermal fibroblast migration and proliferation in comparison with collagen matrix. These results suggest that the HA incorporated collagen porous matrix assumes to enhance dermal fibroblast adaptation and regenerative potential.

Interaction of mesenchymal stem cells and osteoblasts for in vitro osteogenesis.

It has recently been reported that bone marrow-derived mesenchymal stem cells (MSCs), which are systemically administrated to different species, undergo site-specific differentiation. This suggests that the tissue specific cells may cause or promote the differentiation of the MSCs toward their cell type via a cell-to-cell interaction that is mediated not only by hormones and cytokines, but also by direct cell-to-cell contact. In this study, in order to assess the possible synergistic interactions for osteogenesis between the two types of cells, the MSCs derived from rabbit bone marrow were co-cultured with rat calvarial osteoblasts in direct cell-to-cell contact in a control medium (CM) and in an osteogenic medium (OM). The cell number, alkaline phosphatase activity, and amount of calcium deposition were assayed in the cultures of MSCs, osteoblasts, and co-cultures of them in either OM or CM for up to 40 days. The cell numbers and the alkaline phosphatase activities in the co-culture were somewhere in between those of the osteoblast cultures and the MSC cultures. The amounts of deposited calcium were lower in the co-culture compared to those of the other cultures. This suggests that there are little synergistic interactions during osteogenesis in vitro between the rat osteoblasts and rabbit MSCs.