Analysis of porcine preadipocytes differentiation by atomic force microscope / 生物工程学报

Abnormal changes during fat formation are closely related to the prevalence of many diseases. In order to understand the formation mechanism of fat, we used atomic force microscopy (AFM) to characterize the morphology and mechanical properties of porcine preadipocytes during the differentiation. Preadipocytes and adipocytes were different morphologically. The surface roughness of adipocytes was less than preadipocytes by detection of the ultrastructure. The mechanical properties of preadipocytes were changed during differentiation with AFM-based force spectroscopy. Preadipocytes were 20% higher than adipocytes in the adhesion force, stiffness and Young's modulus. Therefore, AFM analysis of membrane changes related to adipocytes formation provided quantitative data in the nanometer level for further studying the formation mechanism of the adipocytes.

Distribution and force spectroscopy of CD20 antigen-antibody binding on the B cell surface / 生物工程学报

The lower expression of CD20 antigen molecules on the B cell membrane is the primary characteristic of B-chronic lymphocytic leukemia (B-CLL). In this paper, we combined laser scanning confocal microscopy (LSCM) and quantum dots labeling to detect the expression and distribution of CD20 molecules on CD20+B lymphocyte surface. Simultaneously, we investigated the morphology and ultrastructure of the B lymphocytes that belonged to the normal persons and B-CLL patients through utilizing the atomic force microscope (AFM). In addition, we measured the force spectroscopy of CD20 antigen-antibody binding using the AFM tips modified with CD20 antibody. The fluorescent images indicated that the density of CD20 of normal CD20+B lymphocytes was much higher than that of B-CLL CD20+B cells. The AFM data show that ultrastructure of B-CLL CD20+B lymphocytes became more complicated. Moreover, the single molecular force spectroscopy data show that the special force of CD20 antigen-antibody was four times bigger than the nonspecific force between the naked AFM tip and cell surface. The force map showed that CD20 molecules distributed homogeneously on the normal CD20+B lymphocytes, whereas, the CD20 molecules distributed heterogenous on B-CLL CD20+B lymphocytes. Our data provide visualized evidence for the phenomenon of low-response to rituximab therapy on clinical. Meanwhile, AFM is possible to be a powerful tool for development and screening of drugs for pharmacology use.

New applications and the comparison between atomic force microscope and electron microscope in regenerative medicine / 生物医学工程学杂志

This article introduces the basic theories about atomic force microscope (AFM) and electron microscope (EM), respectively. New applications of each microscopic technology in regenerative medicine, covering both material science and life science, are discussed. The advantages or disadvantages of the kinds of microscopes in working conditions, sample preparation, resolution and the like, are discussed and compared systematically to make clear each scope of applications. This could be a useful guide for selecting the appropriate microscopic analysis in research work about regenerative medicine.

Optical Properties and Biomedical Application of Gold Nanorods / 生物化学与生物物理进展

Gold nanorods is a capsule-shaped gold nanoparticles. Gold nanorods give rise to two absorption peaks corresponding to their plasmonic modes, transverse mode and longitudinal mode, corresponding to light absorption and scattering along the short and long axis, respectively. The longitudinal surface plasmon resonance can be tuned by adjusting their aspect ratio from the visible to the NIR region and extremely sensitive to changes in the dielectric properties of the surroundings including solvents, adsorbates, and the interparticle distance of the gold nanorods. This unique optical property of gold nanorods opens up fascinating applications in biological and chemical sensors. Optical properties and biomedical application of gold nanorods are introduced, and its future research prospects are discussed.

Dynamic expression of E-cadherin in embryonic stem cell differentiation in vitro and correlation with cell adhesion / 中国组织工程研究

BACKGROUND:E-cadhedn plays an important role in development of liver tissue in the embryonic stage.Therefore,it is importance for investigating the feasibility of dynamic expression of E-cadherin in embryonic stern cell differentiation to in vitro development of liver tissue.OBJECTIVE:To observe the dynamic expression of E-cadherin in embryonic stem cell differentiation and the effect on cell adhesion.DESIGN,TIME AND SETTING:An in vitro cytological observation was performed at Surgical Laboratory of the First Affiliated Hospital of Sun Yat-sen University from December 2007 to December 2008.MATERIALS:Embryonic stem cells of BALB/c mice were obtained from Professor Huang (Department of Ophthalmology,Sun Yat-sen University).Twenty 13-day-old pregnant BALB/c mice of clean grade were provided by the Experimental Animal Centar of Sun Yat-sen University.METHODS:Following trypsinization,embryonic stem cells were suspend-incubated in DMEM culture medium containing fetal bovine serum,2-mercaptoethanol,HEPES,penicillin,and streptomycin.Embryoid body was formed 5 days after normal development and incubated in the culture plate at day 6.Liver tissue which was obtained from 13-day-old pregnant BALB/c mice was prepared for fetal liver cells which were frozen-sectioned as the controls.MAIN OUTCOME MEASURES:E-cadherin expression was detected using reverse transcriptase-polymerase chain reaction (RT-PCR) and immunocytochemistry st varying time points of 1,5,9,13,and 17 days in stages of initial differentiation of embryonic stem cells,formation of embryoid body,and formation of differentiated clusters.Additionally,the effect of E-cadherin expression on cell adhesion was also detected.RESULTS:RT-PCR showed that E-cadherin mRNA expression was not observed at day 1 but peaked at day 5;gradually,the expression was decreased until the expression was stopped at day 17.E-cadherin mRNA expression was strong in fetal liver cells in the control group.Immunocytochemistry showed a similar outcome.Morphologically,embryonic stem cells developed from unicells into compact three-embryonic layer embryoid body and into incompact cell population.CONCLUSION:E-cadherin expression correlates with differentiated cell adhesion;additionally,the lost expression in an in vitro environment may be an important cause for unable regularization of differentiated cells.

Stable transfection of E-cadherin gene into mouse embryonic stem cells and its effects on adhesive capacity of differentiated cells / 中国组织工程研究

BACKGROUND: During the embryonic stage, E-cadherin expression plays a critical role in the formation of hepatic tissue.OBJECTIVE: E-cadherin gene was transfected into mouse embryonic stem cells (ESCs) to observe its effects on adhesive capacity of differentiated cells.DESIGN, TIME AND SETTING: A cytological in vitro observation was performed at the Laboratory of Surgery, First Hospital Affiliated to Sun Yat-sen University between December 2007 and December 2008.MATERIALS: BALB/c mice at gestational 13 days, of clean grade, were provided by Laboratory Animal Center, Sun Yat-sen University. BALB/c mouse ESCs were preserved by professor Huang Bing from the Department of Ophthalmology, Sun Yat-sen University. CMV promoter-containing eukaryotic expression plasmid pEGFP-N1 was gifted by doctor Lu Zhi-yue from Medical College, Sun Yat-sen University.METHODS: Total RNA was extracted from BALB/c mouse fresh hepatic tissue and synthesized into cDNA by reverse transcription (RT). The synthesized cDNA was used as a template to perform a polymerase chain reaction (PCR) that amplifies a targeted fragment. Following double enzyme digestion, pEGFP-E-cadherin plasmids were reconstructed and transfected into mouse ESCs. In vitro differentiation of transfected mouse ESCs was performed.MAIN OUTCOME MEASURES: Detection of E-cadherin expression in the differentiation system using RT-PCR and immunocytochemistry and observation of adhesive capacity of differentiated cells.RESULTS: E-cadherin gene-transfected ESCs could stably express E-cadherin during differentiational 1-17 days, while non-transfected ESCs expressed a decreasing amount of E-cadherin. The adhesive capacity of differentiated cells that stably expressed E-cadherin was markedly enhanced. Compact cell connection and multi-layer growth state remained at 19 days. While non-transfected ESCs gradually changed from embryoid bodies into noncohesive cell populations.CONCLUSION: Differentiating E-cadherin ESCs exhibit markedly enhanced adhesive capacity and maintain multi-layer growth state.

Morphology and mechanical properties of normal lymphocyte and Jurkat revealed by atomic force microscopy / 生物工程学报

Alternations of lymphocyte in biophysical properties (e.g., morphology and viscoelasticity) are related to the human health, disease diagnosis and treatment. Here, we used atomic force microscopy (AFM) to characterize the morphology and mechanical properties of normal lymphocyte and Jurkat. The AFM images revealed that their cell shapes appeared similar. The mechanical properties of the two groups were tracked with AFM-based force spectroscopy. The normal lymphocyte cells had a high adhesion force distribution in (796.7 +/- 248.5) pN, whereas the Jurkat cells had a low force distribution in (158.5 +/- 37.5) pN. The adhesion force revealed that the Young's modulus of normal lymphocyte cells (0.471 kPa +/- 0.081 kPa) was nearly four times higher than that of Jurkat cells (0.0964 kPa +/- 0.0229 kPa) at the same loading rate. The stiffness of normal lymphocyte cells was (2.278 +/- 0.488) mN/m and that of Jurkat cells was (4.322 +/- 0.382) mN/m. The differences in mechanical properties of normal and cancerous cells were obvious that healthy and diseased states could be clearly distinguished. These results may be applied to the clinic disease diagnosis for distinguishing the normal cells from the cancer ones even when they show similar shapes.

Research on hyaluronic acid and pectin complex / 生物医学工程学杂志

Several complexes with different mass ratios of hyaluronic acid to pectin were studied using AFM and IR at the room temperature kept by air conditioning. The results showed that hyaluronic acid and pectin were in the state of being complex and were consistent when the concentrations of hyaluronic acid and pectin were above 1 mg/ml and 5 mg/ml respectively, and the mass ratio was 1 : 5. The complex self-assembled to composite grain-shaped film. In comparison with simple hyaluronic acid, the viscosity of hylauronic acid and pectin complex was stronger, and water-solubility was lower. The complex has the bio-function of both hylauronic acid and pectin and has wide application potential in the field of biomedical engineering.

Characterization and applications of self-assembled lipid films: atomic force microscopy studies / 生物医学工程学杂志

Self-assembled lipid films provide new insights into the structure-function relationships of biomolecules at the molecular level. It has potential applications in biology and bionics. In this paper, with regard to atomic force microscopy (AFM) characterization, the surface structures and growth kinetics of self-assembled lipid films as well as their applications in high-resolution AFM imaging of surface-immobilized biomolecules such as proteins, DNA and enzymes are reviewed.

Analysis of sodium benzoate biotoxicity by atomic force microscope / 生物工程学报

Atomic force microscope (AFM) was used to study biotoxicity of food preservative sodium benzoate (SB) at the single cellular level. Lymphocyte morphology and membrane ultrastructure treated with SB at different concentrations and time were analyzed visually. As compared to the normal lymphocyte, the cell morphology and membrane was significantly changed and its ultrastructure was also complicated. After treated with SB, the Rp-v, Rq, Ra and Z values were changed. The statistical analysis of lymphocytes after treated with SB was studied, and discussed its mechanism.

Embryonic stem cell-derived hepatocyte-like cell transplantation for acute liver failure / 中国组织工程研究

BACKGROUND: Effects of embryonic stem cell-derived hepatocyte-like cell transplantation on oncogenicity of differentiated hepatocyte-like cells and biochemical metabolism of liver should be further studied.OBJECTIVE: To evaluate the therapeutic efficacy of embryonic stem cell-derived hepatocyte-like cell transplantation on the acute liver failure.DESIGN: Randomized controlled study.SETTING: the First Affiliated Hospital of Sun Yat-sen University.MATERIALS: This study was performed at the Central Laboratory, the First Affiliated Hospital of Sun Yat-sen University from January 2005 to February 2006. D3-ES cells extracted from the mice which underwent transfection of green fluorescent protein were graciously presented by professor Huang, Ophthalmology Center of Sun Yat-sen University. Forty 6-week-old D3-129 mice of clean grade and irrespective of gender were provided by Experimental Animal Center of Sun Yat-sen University [certification: SCXK (yue) 2004-0011]. The experimentzl animals were disposed according to ethical criteria.Transforming growth factor, basic fibroblast growth factor, and hepatocyte growth factor were provided by Gibco BRL Company, USA.METHODS: Transforming growth factor, basic fibroblast growth factor, and hepatocyte growth factor were combined to differentiate D3-ES cells into hepatic cells. Cell suspension was poured into liver capsule of 20 mice with 2.0×10(6)cells per mouse. Another 20 mice that determined as the controls were injected with saline. Twenty-four hours later, intraperitoneal injection of 5 μL/20 g carbon tetrachloride was used to induce acute liver failure and to observe quality of life and mean survival time. Twenty-four hours after acute liver failure, vena cava posterior blood was drawn to detect total bilirubin,glutamate-pyruvate transaminase, albumin, blood glucose, pro-time prothrombin time, and other hepatic functional parameters. By scarification, hepatic samples were obtained to evaluate oncogenesis condition, and then HE staining and immunohistochemistry were adopted to detect growth of transplanted cells and albumin expression.MAIN OUTCOME MEASURES: Quality of life, average survival time, hepatic functional parameters, growth of transplanted cells, and oncogenesis condition.RESULTS: Quality of life and average survival time: After the onset of acute liver failure, mice in the control group had incoordination and other symptoms of central nervous system. In addition, 14 mice in the control group and 8 in the transplantation group had abdominal dropsy. Average survival time in the control group was significantly shorter than that in the transplantation group (23, 62 hours, P<0.05). Hepatic functional parameters: Levels of total bilirubin and glutamate-pyruvate transaminase in the control and transplantation groups were higher than those before modeling; levels of albumin and blood glucose were lower than those before modeling; pro-time prothrombin time was significantly longer than that before modeling(P<0.01). Furthermore, levels of total bilirubin and glutamate-pyruvate transaminase in the transplantation group were lower than those in the control group; blood glucose in the transplantation group was higher than that in the control group, and pro-time prothrombin time in the transplantation group was significantly shorter than that in the control group (P<0.05). Growth of transplanted cells and oncogenesis condition: Pathological section demonstrated that structure of liver tissue was not changed remarkably, and tumor was not formed. Moreover, transplanted cells and hepatocyte-like cell were well arranged and combined to express albumin.CONCLUSION: Embryonic stem cell-derived hepatocyte-like cell transplantation can improve quality of life, prolong survival time of model mice with acute liver failure; additionally, transplanted cells may well support biochemical metabolism of liver tissue.

Progress and application of atomic force microscope concerning mechanical properties of cells / 中国组织工程研究

Atomic force microscope(AFM) has one important feature that it is used to scan the samples with non-modified and modified probes to obtain sample appearance,and force-distance curve at certain point,based on which the adhesion,bond force and mechanical properties of the sample can be obtained.Until recently,the application of the AFM to measure the mechanical properties of biological sample is very extensive,which is significant in biomedicine and clinical medicine.This paper introduced the force curve theory of AFM,and reviewed the application of AFM to measure the mechanical properties of biological sample including elasticity,adhesion,stiffness,the interaction between antibody and antigen of the cell.In addition,we prospected the application and development of AFM to analyze cell mechanical properties.

Progress in the studies of DNA-protein interactions by atomic force microscopy / 生物医学工程学杂志

Atomic force microscopy (AFM) has been applied in many biological investigations in recent years, and this review focuses on the application of AFM in DNA-protein interactions. AFM images of static DNA-protein complexes, in air and in liquid, can be used to obtain quantitative and qualitative information on the structure of different complexes. And dynamic AFM images of DNA-protein complexation in real time under liquid conditions will help to understand biological processes and mechanisms at single molecule level. In addition, the measurement of intermolecular forces between biomolecules also provides new opportunities for studying mechanical properties of biomolecules and the interactions in their native environment. AFM has revealed many mechanisms of gene regulation, and will play a more and more important role in life science research.

Combined therapy of Super Lizer and Durogesic patch in elderly patients with herpes zoster and diabetes mellitus / 中国组织工程研究

BACKGROUND:The main symptoms of herpes zoster (HZ) manifest as pain and skin eruption and the pain, when treated inadequately, may proceed to become post herpetic neuralgia (PHN) which has progressively increasing incidence with age.OBJECTIVE: To observe the effect of combined therapy of super laser and Durogesic patch (transdermal fentanyl) in the treatment of elderly patients with herpes zoster and diabetes.DESIGN: Randomized controlled observation.SETTING: The First Affiliated Hospital of Jinan University.PARTICIPANTS: Thirty (14 males and 16 females) elderly patients of age 62-83 years with concurrent herpes zoster (duration 6-14 days) and diabetes who had received the conventional dermatological and medication treatment but still had persistent pain were selected form the First Affiliated Hospital of Jinan University from 2003 to 2006. All elderly patients were randomly divided into 3 groups with 10 in each group.METHODS: ① Super Lizer group (SL Group): Patients were received Super Lizer (linear polarized near-infrared light) therapy once a day for 15 days. ② Durogesic patch group (DR Group): Patients were received 2.5 mg Durogesic patch once for every 3 days. ③ Combined group (SL+ DR Group): Patients were received both the Super Lizer therapy and the Durogesic patch for 15 days.MAIN OUTCOME MASURES: All patients received the assigned treatment for 3 days and VAS was evaluated before and at the 3rd, 15th day during treatment, also at 7th day after termination of treatment. Visual analogue scales were used to assess the degree of pain and global evaluation were done With pain relief more than 70% rated as excellent, pain relief between 30%-70% rated as effective and pain relief less than 30% rated as ineffective. Adverse effects were also recorded.RESULTS: Thirty patients were all involved in the final analysis. VAS scores of all three groups were significantly decreased after 15-day treatment (P ＜ 0.05); after 3-day treatment, VAS scores of DR group (2.35±1.43) and SL+DR group (2.41 ±1.54) were significantly lower than their respective baselines and that of the SL group (7.00±0.82) (P ＜ 0.05) which showed no significant change from the baseline value. SL+DR group showed significantly higher excellent rating than that of the SL group and DR group (80%, 60%, 70%, P ＜ 0.05) at the end of the 15-day treatment. At 7th days after treatment, VAS scores of SL group (3.01±1.20) and SL+DR group (2.41±1.54) were still significantly lower than that of DR group (6.70±0.67) (P ＜ 0.05) which had returned to the pretreatment level. No serious adverse effects such as respiratory depression were observed in any of the patients. Mild side effects such as dizziness, nausea and vomiting, constipation were observed in the DR and SL+DR groups which usually subsided after a week.CONCLUSION: Combined therapy of SL and DR shows a better pain relief in the elderly patients with herpes zoster and diabetes without significant adverse effects. It provides the advantage of fast onset of effect with the Durogesic patch and the long-term effect of Super Lizer.

Hepatic differentiation from embryonic stem cells in vitro / 中华医学杂志(英文版)

<p><b>OBJECTIVE</b>To investigate an method for hepatic differentiation from embryonic stem cells (ES cells) in vitro and the resulting differentiation ratio, in order to develop a procedure for producing a new type of hepatocyte for hepatocyte replacement therapy in the treatment of liver failure.</p><p><b>METHODS</b>ES cells from Balb/C mice were cultured and maintained in an undifferentiated state in gelatin-coated dishes using Dulbecco's modified Eagle's medium (DMEM) containing 1000 U/ml leukemia inhibitory factor (LIF). Then, LIF was withdrawn from the DMEM to allow the ES cells to develop into embryonic bodies (EBs). EBs were plated onto tissue culture dishes, and growth factors such as acidicfibroblast growth factor (aFGF) and hepatocyte growth factor (HGF) were added to the medium to promote directional differentiation. The course of development and differentiation was observed dynamically using an inversion microscope. The expression of hepatic proteins, such as alpha-fetoprotein (AFP), albumin (ALB), cytokeratin 8 (CK8), cytokeratin 18 (CK18), in cytoplasm was analyzed by immunocytochemistry (ICC). The concentration of ALB in the medium was determined dynamically by radioimmunoassay (RIA).</p><p><b>RESULTS</b>ES cells replicated as clones, without differentiating, in DMEM containing LIF. They developed into EBs in medium without LIF. Our ICC assay showed that differentiating cells did not express hepatic proteins, such as AFP, ALB, CK8, and CK18 until day 7, day 9, day 11, and day 11, respectively (up to 2 days later when growth factors are not present). The concentration of AFP in the medium was first detected on day 8, at a concentration of 3.4 ng/ml, and increased to 22.8 ng/ml by day 15. The concentration of ALB in the medium was 0.2 micro g/ml on day 11, and increased to 2.2 micro g/ml by day 15. ALB-positive cells under ICC manifest morphological structures were consistent with normal mouse hepatocytes. The differentiation ratio of hepatocytes in the ES cell differentiation system was 30% on day 15 (significantly lower without the presence of growth factors).</p><p><b>CONCLUSIONS</b>ES cells can differentiate into mature hepatocytes. Growth factors, such as aFGF and HGF, can enhance this differentiation and produce sufficient numbers of functional hepatocytes. This method may be a reliable new way of differentiating ES cells into hepatocytes for use in replacement therapy in the treatment of liver failure.</p>

Analysis of chromosome aberration due to ethidium bromide using AFM / 中国病理生理杂志

AIM:To study chromosome aberration due to ethidium bromide (EB), a heterocyclic organic compound and an organic fluorescence dye commonly used in biochemical experiment, and to help further understanding the molecular mechanism of tumor or cancer induced by EB and other heterocyclic organic compounds. METHODS: The toxicity action of EB was evaluated from three aspects including DNA, chromosome and embryo stem cells (ESCs) using atomic force microscopy (AFM), and thereinto, the morphology structural difference of ESCs treated with two EB doses was also valuated. RESULTS: The morphological structures of DNA, chromosome and ESCs were dramatically damaged. The average height of DNA decreased 0.5 nm; chromosomal arms were ruptured from centromere location; molecules of cellular membrane congregated and loop-like structure formed, and ES cell masses were collapsed and became dead after large EB doses treatment and mesh-like morphological structure was discernable. CONCLUSION: The toxicity action of EB is strong and destroys the surface structure of DNA and chromosome. EB induces structural aberration of ES cellular membrane and cell death. The results indicate that the action of EB is externalized at gene level and cell level, which is important to study the carcinogenicity of EB.

The prevention of acute graft versus host disease by Rhodamine 123-mediated photodynamic therapy / 中国病理生理杂志

AIM: To evaluate the effect and safety of Rhodamine 123 (Rh123)-mediated photodynamic treatment (PDT) on acute graft versus host disease. METHODS: An acute graft versus host disease (aGVHD) mice model was established using C57B/6 mice as donors and BALB/c mice as recipients. Mixed lymphocytic cells were cultured with Rh123 (50 nmol/L) and irradiated by argon laser 30 mW/cm2 for 3 min, then transplanted to BALB/c recipient mice mixed with donor bone marrow. Hepatopoietic recovery, aGVHD occurrence, survival time after transplantation and pathological changes were observed. In addition, CD3+CD69+ positive rates of MLC were examined by flowcytometry. RESULTS: Occurrence of aGVHD decreased, degree of pathological manifestation became milder, survival rates were higher than non PDT groups. CD3+CD69+ rates of MLC cells treated with photodynamic therapy (PDT) and cultured for 24 h significantly decreased. CONCLUSION: Rh123-mediated PDT can effectively prevent aGVHD of allogeneic bone marrow transplantation in mice.

Conformational Changes of Hemoglobin Induced by Surfactants / 环境与健康杂志

Objective To study the effects of surfactants on conformation of hemoglobin and further to understand the potential impact of surfactants in environment on human health. Methods The changes of the conformation of hemoglobin induced by cation and anion surfactants were investigated with four methods, such as synchronous fluorescence spectra, UV, electrochemistry and atom force microscope. Results The changes of the conformation of hemoglobin induced by anion surfactant were obvious but were not by cation surfactant. Conclusion Sodium lauryl sulphate, an anion surfactant, may change the conformation of hemoglobin.

Cell ultrastructure and function studied by scanning near-field optics / 中国病理生理杂志

Scanning near-field optical microscopy (SNOM) is developing on the base of near-field optical theory and SPM technique, and producing the highest optical resolution to break through the diffraction limit. It is able to study cellular ultrastructure, function and interaction between biomolecules without destructing the living cells. Recent advances in applying SNOM to cellular and molecular biology were introduced in this review.

Ultrastructure and function of human bone marrow mesenchymal stem cells / 中国病理生理杂志

AIM: To study the cytoskeleton of mesench ymal stem cells (MSCs), the ultrastructure and function relationship by using atomic force microscope (AFM). METHODS: The ultrastructures and morphological feature of MSCs c ultured for 1 d and 5 d were studied by AFM. RESULTS: The special structures that possess peculiar morphologi cal characteristic of MSCs such as cytoskeleton, pseudopod, microfilament etc we re identified by AFM, and these special structures are difficult to observe unde r electronic microscopy or other conventional optical microscopy. CONCLUSIONS: AFM is a powerful tool to study ultrastructures, mo rphological features, and cytoskeleton of stem cells in near physiological condi t ions. Its application prospect in cellular biology is extensive. The special cyt oskeleton and other structures of MSCs observed above may represent the structur al base of multi-differentiation potential of MSCs.