Effects of areca nut consumption on cell differentiation of osteoblasts, myoblasts, and fibroblasts

Areca nut is used worldwide as a hallucinogenic addicting drug along the tropical belt.Arecoline,a toxic compound,is the most important alkaloid in areca nuts.The adverse effects of oral uptake and chewing of areca nut are well known.For example,the possibility of cancer caused by chewing areca nuts is widely discussed.Chewing areca nut has other adverse effects on other organs,including abnormal cell differentiation,oral cancer,and several other diseases.The use of areca nut is also associated with low birthweight.Skeletal musculature is the largest organ in the body and is attached to the bones.During embryo development,the differentiation of bone and muscle cells is critical.In this article,we reviewed the effects of areca nut and arecoline on embryonic cell differentiation,particularly osteoblasts,myoblasts,and fibroblasts.

Nanostructured Graphene Surfaces Promote Different Stages of Bone Cell Differentiation

Nanostructured graphene films were used as platforms for the differentiation of Saos-2 cells into bonelike cells. The films were grown using the plasma-enhanced chemical vapor deposition method, which allowed the production of both vertically and horizontally aligned carbon nanowalls(CNWs). Modifications of the techniqueallowed control of the density of the CNWs and their orientation after the transfer process. The influence of two different topographies on cell attachment, proliferation,and differentiation was investigated. First, the transferred graphene surfaces were shown to be noncytotoxic and were able to support cell adhesion and growth for over 7 days.Second, early cell differentiation(identified by cellular alkaline phosphatase release) was found to be enhanced on the horizontally aligned CNW surfaces, whereas mineralization(identified by cellular calcium production), a later stage of bone cell differentiation, was stimulated by the presence of the vertical CNWs on the surfaces. These results show that the graphene coatings, grown using the presented method, are biocompatible. And their topographies have an impact on cell behavior, which can be useful in tissue engineering applications.

Vascular smooth muscle cell differentiation－2010

Vascular smooth muscle cells have attracted considerable interest as a model for a flexible program of gene expression.This cell type arises throughout the embryo body plan via poorly understood signaling cascades that direct the expression of transcription factors and microRNAs which,in turn,orchestrate the activation of contractile genes collectively defining this cell lineage.The discovery of myocardin and its close association with serum response factor has represented a major break-through for the molecular understanding of vascular smooth muscle cell differentiation.Retinoids have been shown to improve the outcome of vessel wall remodeling following injury and have provided further insights into the molecular circuitry that defines the vascular smooth muscle cell phenotype.This review summarizes the progress to date in each of these areas of vascular smooth muscle cell biology.

Editor's Choice——Umbilical cord blood mesenchymal stem cell differentiation and transplantation in neural regeneration

Previous in vivo experiments have shown that human umbilical cord blood mesenchymal stem cells can promote the proliferation and differentiation of damaged celts, and help to repair damaged sites, Recent studies have reported that umbilical cord blood-derived mesenchymal stem cells can differentiate into neurons and glial cells. Recent studies have reported that the repair mechanisms underlying cord blood stern cells involve the replacement of damaged cells and mediation of the local micro-environment.

EFFECTS OF S86019, AN ACTIVE COMPONENT FROM PURALIA LOB AT A, ON CELL DIFFERENTIATION AND CELL CYCLE TRAVERSE OF HL-60 CELLS

The effects of S86019, an active component from Puralia lobata, on the induction of cell differentiation and cell cycle traverse of HL-60 cells were described. It was shown that cell proliferation of HL-60 cells was inhibited by S86019 in vitro. Under the action of S86019 the HL-60 cells were induced to differentiate into metamyelocytes, myelocytes and much matured cells with banded or segmented nucleus. Flow cytometry demonstrated that the cell population of HL-60 cells was blocked at G1 phase which resulted in the elevation of percentage of G1 cells and decrease of percentage of cells in S phase. Experimental results demonstrated that S86019 is an active inducer of cell differentiation in HL-60 cells.

Maintaining moderate levels of hypochlorous acid promotes neural stem cell proliferation and differentiation in the recovery phase of stroke

It has been shown clinically that continuous removal of ischemia/reperfusion-induced reactive oxygen species is not conducive to the recovery of late stroke.Indeed,previous studies have shown that excessive increases in hypochlorous acid after stroke can cause severe damage to brain tissue.Our previous studies have found that a small amount of hypochlorous acid still exists in the later stage of stroke,but its specific role and mechanism are currently unclear.To simulate stroke in vivo,a middle cerebral artery occlusion rat model was established,with an oxygen-glucose deprivation/reoxygenation model established in vitro to mimic stroke.We found that in the early stage(within 24 hours)of ischemic stroke,neutrophils produced a large amount of hypochlorous acid,while in the recovery phase(10 days after stroke),microglia were activated and produced a small amount of hypochlorous acid.Further,in acute stroke in rats,hypochlorous acid production was prevented using a hypochlorous acid scavenger,taurine,or myeloperoxidase inhibitor,4-aminobenzoic acid hydrazide.Our results showed that high levels of hypochlorous acid(200μM)induced neuronal apoptosis after oxygen/glucose deprivation/reoxygenation.However,in the recovery phase of the middle cerebral artery occlusion model,a moderate level of hypochlorous acid promoted the proliferation and differentiation of neural stem cells into neurons and astrocytes.This suggests that hypochlorous acid plays different roles at different phases of cerebral ischemia/reperfusion injury.Lower levels of hypochlorous acid(5 and 100μM)promoted nuclear translocation ofβ-catenin.By transfection of single-site mutation plasmids,we found that hypochlorous acid induced chlorination of theβ-catenin tyrosine 30 residue,which promoted nuclear translocation.Altogether,our study indicates that maintaining low levels of hypochlorous acid plays a key role in the recovery of neurological function.

Rheum palmatum L. and Salvia miltiorrhiza Bge. Alleviates Acute Pancreatitis by Regulating Th17 Cell Differentiation: An Integrated Network Pharmacology Analysis, Molecular Dynamics Simulation and Experimental Validation

Objective: To identify the core targets of Rheum palmatum L. and Salvia miltiorrhiza Bge.,(Dahuang-Danshen, DH-DS) and the mechanism underlying its therapeutic efficacy in acute pancreatitis(AP)using a network pharmacology approach and validate the findings in animal experiments. Methods: Network pharmacology analysis was used to elucidate the mechanisms underlying the therapeutic effects of DH-DS in AP. The reliability of the results was verified by molecular docking simulation and molecular dynamics simulation.Finally, the results of network pharmacology enrichment analysis were verified by immunohistochemistry,Western blot analysis and real-time quantitative PCR, respectively. Results: Sixty-seven common targets of DH-DS in AP were identified and mitogen-activated protein kinase 3(MAPK3), Janus kinase 2(JAK2), signal transducer and activator of transcription 3(STAT3), protein c-Fos(FOS) were identified as core targets in the protein interaction(PPI) network analysis. Gene ontology analysis showed that cellular response to organic substance was the main functions of DH-DS in AP, and Kyoto Encyclopedia of Genes and Genomes analysis showed that the main pathway included Th17 cell differentiation. Molecular docking simulation confirmed that DH-DS binds with strong affinity to MAPK3, STAT3 and FOS. Molecular dynamics simulation revealed that FOS-isotanshinone Ⅱ and STAT3-dan-shexinkum d had good binding capacity. Animal experiments indicated that compared with the AP model group, DH-DS treatment effectively alleviated AP by inhibiting the expression of interleukin-1β, interleukin-6 and tumor necrosis factor-α, and blocking the activation of Th17 cell differentiation(P<0.01). Conclusion: DH-DS could inhibit the expression of inflammatory factors and protect pancreatic tissues,which would be functioned by regulating Th17 cell differentiation-related m RNA and protein expressions.

An overview of autophagy in the differentiation of dental stem cells

Dental stem cells(DSCs)have attracted significant interest as autologous stem cells since they are easily accessible and give a minimal immune response.These properties and their ability to both maintain self-renewal and undergo multi-lineage differentiation establish them as key players in regenerative medicine.While many regulatory factors determine the differentiation trajectory of DSCs,prior research has predominantly been based on genetic,epigenetic,and molecular aspects.Recent evidence suggests that DSC differentiation can also be influenced by autophagy,a highly conserved cellular process responsible for maintaining cellular and tissue homeostasis under various stress conditions.This comprehensive review endeavors to elucidate the intricate regulatory mechanism and relationship between autophagy and DSC differentiation.To achieve this goal,we dissect the intricacies of autophagy and its mechanisms.Subsequently,we elucidate its pivotal roles in impacting DSC differentiation,including osteo/odontogenic,neurogenic,and angiogenic trajectories.Furthermore,we reveal the regulatory factors that govern autophagy in DSC lineage commitment,including scaffold materials,pharmaceutical cues,and the extrinsic milieu.The implications of this review are far-reaching,underpinning the potential to wield autophagy as a regulatory tool to expedite DSC-directed differentiation and thereby promote the application of DSCs within the realm of regenerative medicine.

Long noncoding RNA(lncRNA)H19:An essential developmental regulator with expanding roles in cancer,stem cell differentiation,and metabolic diseases

Recent advances in deep sequencing technologies have revealed that,while less than 2%of the human genome is transcribed into mRNA for protein synthesis,over 80%of the genome is transcribed,leading to the production of large amounts of noncoding RNAs(ncRNAs).It has been shown that ncRNAs,especially long non-coding RNAs(lncRNAs),may play crucial regulatory roles in gene expression.As one of the first isolated and reported lncRNAs,H19 has gained much attention due to its essential roles in regulating many physiological and/or pathological processes including embryogenesis,development,tumorigenesis,osteogen-esis,and metabolism.Mechanistically,H19 mediates diverse regulatory functions by serving as competing endogenous RNAs(CeRNAs),Igf2/H19 imprinted tandem gene,modular scaffold,cooperating with H19 antisense,and acting directly with other mRNAs or lncRNAs.Here,we summarized the current understanding of H19 in embryogenesis and development,cancer development and progression,mesenchymal stem cell lineage-specific differentiation,and metabolic diseases.We discussed the potential regulatory mechanisms underlying H19’s func-tions in those processes although more in-depth studies are warranted to delineate the exact molecular,cellular,epigenetic,and genomic regulatory mechanisms underlying the physiolog-ical and pathological roles of H19.Ultimately,these lines of investigation may lead to the development of novel therapeutics for human diseases by exploiting H19 functions.

Coactivator-associated arginine methyltransferase 1: A versatile player in cell differentiation and development

Protein arginine methylation is a common post-translational modification involved in the regulation of various cellular functions. Coactivator-associated arginine methyltransferase 1 (CARM1) is a protein arginine methyltransferase that asymmetrically dimethylates histone H3 and non-histone proteins to regulate gene transcription. CARM1 has been found to play important roles in cell differentiation and development, cell cycle progression, autophagy, metabolism, pre-mRNA splicing and transportation, and DNA replication. In this review, we describe the molecular characteristics of CARM1 and summarize its roles in the regulation of cell differentiation and development in mammals.

In vitro culture and differentiation of rat embryonic midbrain-derived neural stem cells

BACKGROUND： Midbrain-derived neural stem cells （mNSCs） can differentiate into functional mature dopaminergic neurons. The mNSCs are considered the ideal choice for cell therapy of Parkinson＇s disease. OBJECTIVE： To isolate rat embryonic mNSCs and to observe the differentiation characteristics of mNSCs induced by cell growth-promoting factors. DESIGN, TIME AND SETTING： An in vitro cell culture study based on the molecular biology of nerve cells was carried out at the Institute of Clinical Medicine, China-Japan Friendship Hospital （China） from March to November 2007. MATERIALS： Sprague Dawley rats at embryonic day 14 were used in this study. Nestin antibody, β-Ⅲ tubulin antibody, glial fibrillary acidic protein （GFAP） antibody and cyclic nucleotide 3＇-phosphohydrolase （CNPase） antibody were provided by Abcam; DMEM/F12 medium and N2 supplement were provided by Invitrogen; epidermal growth factor （EGF） and fibroblast growth factor-2 （FGF2） were provided by R＆D Systems. METHODS： The ventral mesencephalon was dissected from embryonic day 14 rat embryos. By trypsin digestion and mechanical separation, the brain tissue was triturated into a fine single-cell suspension. The cells were cultured in 5 mL serum-free medium containing DMEM/FI 2, 1% N： supplement, 20 ng/mL EGF and FGF2. The mNSCs at the third generation were coated with 10ug/mL polylysine and induced to differentiate in the DMEM/F12 supplemented with 1% fetal bovine serum and 1% N2. MAIN OUTCOME MEASURES： The neural spheres of the third passage were identified by nestin immunofluorescence; at the same time, the cells were induced to differentiate, and the types of differentiated cell were identified by immunofluorescence for β Ⅲ tubulin, GFAP and CNPase. RESULTS： Seven days after primary culture, a great many neurospheres could be obtained by successive pasage. Immunofluorescence assays showed that the neurospheres were nestin positive, and after differentiation, the cells expressed GFAP, CNPase and β -Ⅲ-tubulin. CONCLUSION： Embryonic day 14 rat mNSCs can differentiate into neuron-like cells and glial cells following induction by EGF, FGF2 and N： additive.

Experimental Study of Cell Migration and Functional Differentiation of Transplanted Neural Stem Cells Co-labeled with Superparamagnetic Iron Oxide and Brdu in an Ischemic Rat Model

Objective To explore the migration of transplanted neural stem cells co-labeled with superparamagnetic iron oxide （SPIO） and bromodeoxyuridine （Brdu） using the 4.7T MR system and to study the cell differentiation with immuno-histochemical method in ischemic rats. Methods Rat neural stem cells （NSCs） co-labelled with SPIO mediated by poly-L-lysine and bromodeoxyuridine （BrdU） were transplanted into the unaffected side of rat brain with middle cerebral artery occlusion （MCAO）. At weeks 1, 2, 3, 4, 5, and 6 after MCAO, migration of the labelled cells was monitored by MRI. At week 6 the rats were killed and their brain tissue was cut according to the migration site of transplanted ceils indicated by MRI and subjected to Prussian blue staining and immunohistochemical staining to observe the migration and differentiation of the transplanted NSCs. Results Three weeks after transplantation, the linear hypointensity area derived from the migration of labelled NSCs was observed by MRI in the corpus callosum adjacent to the injection site. Six weeks after the transplantation, the linear hypointensity area was moved toward the midline along the corpus callosum. MRI findings were confirmed by Prussian blue staining and immunohistochemical staining of the specimen at week 6 after the transplantation. Flourescence co-labelled immunohistochemical methods demonstrated that the transplanted NSCs could differentiate into astrocytes and neurons. Conclusion MRI can monitor the migration of SPIO-labelled NSCs after transplantation in a dynamical and non-invasive manner. NSCs transplanted into ischemic rats can differentiate into astrocytes and neurons during the process of migration.

Transfection of the glial cell line-derived neurotrophic factor gene promotes neuronal differentiation

Glial cell line-derived neurotrophic factor recombinant adenovirus vector-transfected bone marrow mesenchymal stem cells were induced to differentiate into neuron-like cells using inductive medium containing retinoic acid and epidermal growth factor. Cell viability, micro- tubule-associated protein 2-positive cell ratio, and the expression levels of glial cell line-derived neurotrophic factor, nerve growth factor and growth-associated protein-43 protein in the su- pernatant were significantly higher in glial cell line-derived neurotrophic factor/bone marrow mesenchymal stem cells compared with empty virus plasmid-transfected bone marrow mes- enchymal stem cells. Furthermore, microtubule-associated protein 2, glial cell line-derived neurotrophic factor, nerve growth factor and growth-associated protein743 mRNA levels in cell pellets were statistically higher in glial cell line-derived neurotrophic factor/bone marrow mesen- chymal stem cells compared with empty virus plasmid-transfected bone marrow mesenchymal stem cells. These results suggest that glial cell line-derived neurotrophic factor/bone marrow mesenchymal stem cells have a higher rate of induction into neuron-like cells, and this enhanced differentiation into neuron-like cells may be associated with up-regulated expression of glial cell line-derived neurotrophic factor, nerve growth factor and growth-associated protein-43.

Differentiation of Mesenchymal Stem Cells towards a Nucleus Pulposus-like Phenotype Utilizing Simulated Microgravity In Vitro

Mesenchymal stem cells （MSCs） were induced into a nucleus pulposus-like phenotype utilizing simulated microgravity in vitro in order to establish a new cell-based tissue engineering treatment for intervertebral disc degeneration. For induction of a nucleus pulposus-like phenotype, MSCs were cultured in simulated microgravity in a chemically defined medium supplemented with 0 （experimental group） and 10 ng/mL （positive control group） of transforming growth factor β1 （TGF-β1）. MSCs cultured under conventional condition without TGF-β1 served as blank control group. On the day 3 of culture, cellular proliferation was determined by WST-8 assay. Differentiation markers were evaluated by histology and reverse transcriptase-polymerase chain reaction （RT-PCR）. TGF-β1 slightly promoted the proliferation of MSCs. The collagen and proteoglycans were detected in both groups after culture for 7 days. The accumulation of proteoglycans was markedly increased. The RT-PCR revealed that the gene expression of Sox-9, aggrecan and type Ⅱ collagen, which were chondrocyte specific, was increased in MSCs cultured under simulated microgravity for 3 days. The ratio of proteoglycans/collagen in blank control group was 3.4-fold higher than positive control group, which denoted a nucleus pulposus-like phenotype differentiation. Independent, spontaneous differentiation of MSCs towards a nucleus pulposus-like phenotype in simulated microgravity occurred without addition of any external bioactive stimulators, namely factors from TGF-β family, which were previously considered necessary.

Human hair follicle-derived mesenchymal stem cells:Isolation,expansion,and differentiation

Hair follicles are easily accessible skin appendages that protect against cold and potential injuries.Hair follicles contain various pools of stem cells,such as epithelial,melanocyte,and mesenchymal stem cells(MSCs)that continuously self-renew,differentiate,regulate hair growth,and maintain skin homeostasis.Recently,MSCs derived from the dermal papilla or dermal sheath of the human hair follicle have received attention because of their accessibility and broad differentiation potential.In this review,we describe the applications of human hair follicle-derived MSCs(hHF-MSCs)in tissue engineering and regenerative medicine.We have described protocols for isolating hHF-MSCs from human hair follicles and their culture condition in detail.We also summarize strategies for maintaining hHF-MSCs in a highly proliferative but undifferentiated state after repeated in vitro passages,including supplementation of growth factors,3D suspension culture technology,and 3D aggregates of MSCs.In addition,we report the potential of hHF-MSCs in obtaining induced smooth muscle cells and tissue-engineered blood vessels,regenerated hair follicles,induced red blood cells,and induced pluripotent stem cells.In summary,the abundance,convenient accessibility,and broad differentiation potential make hHF-MSCs an ideal seed cell source of regenerative medical and cell therapy.

Elastic modulus affects the growth and differentiation of neural stem cells

It remains poorly understood if carrier hardness, elastic modulus, and contact area affect neural stem cell growth and differentiation. Tensile tests show that the elastic moduli of Tiansu and SMI silicone membranes are lower than that of an ordinary dish, while the elastic modulus of SMI silicone membrane is lower than that of Tiansu silicone membrane. Neural stem cells from the cerebral cortex of embryonic day 16 Sprague-Dawley rats were seeded onto ordinary dishes as well as Tiansu silicone membrane and SMI silicone membrane. Light microscopy showed that neural stem cells on all three carriers show improved adherence. After 7 days of differentiation, neuron specific enolase, glial fibrillary acidic protein, and myelin basic protein expression was detected by immunofluorescence. Moreover, flow cytometry revealed a higher rate of neural stem cell differentiation into astrocytes on Tiansu and SMI silicone membranes than on the ordinary dish, which was also higher on the SMI than the Tiansu silicone membrane. These findings con- firm that all three cell carrier types have good biocompatibility, while SMI and Tiansu silicone membranes exhibit good mechanical homogenization. Thus, elastic modulus affects neural stem cell differentiation into various nerve cells. Within a certain range, a smaller elastic modulus re- sults in a more obvious trend of cell differentiation into astrocytes.

Total saponins of Panax ginseng effects on proliferation and differentiation of human embryonic neural stem cells and in a Parkinson's disease mouse model

BACKGROUND： Total saponins of Panax ginseng （TSPG） exhibits neuroprotection against Parkinson＇s disease in the substantia nigra. OBJECTIVE： To investigate the effects of TSPG on human embryonic neural stem cells （NSCs） proliferation and differentiation into dopaminergic neurons using in vitro studies, and to observe NSC differentiation in a mouse model of Parkinson＇s disease, as well as behavioral changes before and after transplantation. DESIGN, TIME AND SETTING： In vitro neural cell biology trial and in vivo randomized, controlled animal trial were performed at the Institute of Basic Medical Sciences, Chongqing Medical University between September 2004 and December 2007. MATERIALS： TSPG （purity 〉 95%） was isolated, extracted, and identified by Chongqing Academy of Chinese Materia Medica. Recombinant human basic fibroblast growth factor （bFGF） and recombinant human epidermal growth factor （EGF） were purchased from PeproTech, USA. A total of 25 C57/BL6J mice, aged 18-20 weeks were included. Twenty were used to establish a Parkinson＇s disease model with i.p. injection of MPTP （1-methyl-4-phenyl-1, 2, 3, 6-tetrahydropyridine） and TSPG alone or combined with interleukin-1 （IL-1）-treated NSCs prior to transplantation into the corpus striatum. The remaining five mice were pretreated for 3 days with TSPG prior to MPTP injection, serving as the TSPG prevention group. METHODS： Primary NSCs were isolated, cultured and purified from embryonic cerebral cortex. Immunocytochemistry was employed to detect specific antigen expression in the NSCs. In vitro experiment： （1） to induce proliferation, NSCs were treated with TSPG, EGF＋bFGF, or TSPG＋EGF＋bFGF, respectively; （2） to induce dopaminergic neuronal differentiation, NSCs were treated with TSPG, IL-1, or TSPG＋IL-1, respectively. MAIN OUTCOME MEASURES： In vitro experiment： the effects of TSPG on NSCs proliferation were evaluated with flow cytometry and MTT assay. Tyrosine hydroxylase expression was determined by immunocytochemistry assay to observe effects of TSPG on dopaminergic neuronal differentiation. In vivo experiment： differentiation of grafted NSCs in the mouse brain was determined by immunohistochemical staining. Behavioral changes were evaluated by spontaneous activity frequency, memory function, and score of paralysis agitans. RESULTS： （1） NSCs were cultured and passaged for more than three passages. Immunocytochemistry revealed positive nestin staining, as well as neurofilament protein and glial fibrillary acidic protein. （2） TSPG significantly increased NSC proliferation, in particular when combined with EGF and bFGF, which was twice as effective as FGF or bFGF alone. TSPG also induced dopaminergic differentiation in NSCs, in particular when TSPG was added together with IL-1, resulting in an effect five times greater than that of IL-1 alone. （3） At day 30 following transplantation, most NSCs in the TSPG prevention group differentiated into dopaminergic neurons, and the scores of paralysis agitans, spontaneous activity, and memory function were significantly increased compared with TSPG alone or TSPG＋IL-1 groups （P 〈 0.05）. CONCLUSION： TSPG stimulated NSC proliferation, in particular when combined with FGF and bFGF. TSPG significantly induced dopaminergic neuronal differentiation of NSCs, and the effect was greater when combined with IL-1. In addition, TSPG greatly improved behavior in the Parkinson＇s disease mouse model following NSC transplantation. Following NSC transplantation, TSPG pretreatment exhibited superior efficacy over either TSPG alone or TSPG in combination with IL-1, in terms of behavioral improvements in the Parkinson＇s disease mouse model.

Engineering an enthesis-like graft for rotator cuff repair:An approach to fabricate highly biomimetic scaffold capable of zone-specifically releasing stem cell differentiation inducers

Rotator cuff(RC)attaches to humerus across a triphasic yet continuous tissue zones(bone-fibrocartilage-tendon),termed“enthesis”.Regrettably,rapid and functional enthesis regeneration is challenging after RC tear.The existing grafts bioengineered for RC repair are insufficient,as they were engineered by a scaffold that did not mimic normal enthesis in morphology,composition,and tensile property,meanwhile cannot simultaneously stimulate the formation of bone-fibrocartilage-tendon tissues.Herein,an optimized decellularization approach based on a vacuum aspiration device(VAD)was developed to fabricate a book-shaped decellularized enthesis matrix(O-BDEM).Then,three recombinant growth factors(CBP-GFs)capable of binding collagen were synthesized by fusing a collagen-binding peptide(CBP)into the N-terminal of BMP-2,TGF-β3,or GDF-7,and zone-specifically tethered to the collagen of O-BDEM to fabricate a novel scaffold(CBP-GFs/O-BDEM)satisfying the above-mentioned requirements.After ensuring the low immunogenicity of CBP-GFs/O-BDEM by a novel single-cell mass cytometry in a mouse model,we interleaved urine-derived stem cell-sheets into this CBP-GFs/O-BDEM to bioengineer an enthesis-like graft.Its high-performance on regenerating enthesis was determined in a canine model.These findings indicate this CBP-GFs/O-BDEM may be an excellent scaffold for constructing enthesis-like graft to patch large/massive RC tears,and provide breakthroughs in fabricating graded interfacial tissue.

Differentiation potential of human adipose tissue derived stem cells into photoreceptors through explants culture and enzyme methods

AIM： To investigate the retinal photoreceptor differentiation potential of human orbital adipose tissue-derived stem cells （ADSCs） generated by enzyme （EN） and explant （EX） culture methods.METHODS： We investigated potentials of human orbital ADSCs to differentiate into photoreceptors through EN and EX culture methods. EN and EX orbital ADSCs were obtained from the same donor during rehabilitative orbital decompression, and then were subject to a 3-step induction using Noggin, DKK-1, IGF-1 and b-FGF at different time points for 38d. Stem cell, eye-field and photoreceptor-related gene and protein markers were measured by reverse transcription-polymerase chain reaction （RT-PCR） and immunofluorescent （IMF） staining.RESULTS： Both EX and EN orbital ADSCs expressed CD133, a marker of cell differentiation. Moreover, PAX6 and rhodopsin, markers of the retinal progenitor cells, were detected from EX and EN orbital ADSCs. In EX orbital ADSCs, PAX6 mRNA was detected on the 17th day and then the rhodopsin mRNA was detected on the 24th day. In contrast, the EN orbital ADSCs expressed PAX6 and rhodopsin mRNA on the 31st day. EX orbital ADSCs expressed rhodopsin protein on the 24th day, while EN orbital ADSCs expressed rhodopsin protein on the 31st day. CONCLUSION： Orbital ADSCs isolated by direct explants culture show earlier and stronger expressions of markers towards eye field and retinal photoreceptor differentiation than those generated by conventional EN method.

Neural cell co-culture induced differentiation of bone marrow mesenchymal stem cells into neuronal-like cells

BACKGROUND： It has been previously demonstrated that the neural cell microenvironment has the ability to induce differentiation of bone marrow mesenchymal stem cells （BMSCs） into the neural cells. OBJECTIVE： To establish a co-culture system of human BMSCs and neural cells, and to observe effects of this co-culture system on differentiation of human BMSCs into neural cells. DESIGN, TIME AND SETTING： A comparative observation experiment, performed at the Center Labora-tory of the Affiliated Hospital of Medical College Qingdao University from October 2006 to December 2007. MATERIALS： Neural cells were obtained from human fetal brain tissue. BMSCs were harvested from fe-male patients that underwent autonomous stem cell transplantation. METHODS： BMSCs in the co-culture group consisted of BMSCs and third passage neural cells. BMSCs in the control group were solely cultured in vitro. MAIN OUTCOME MEASURES： Morphological changes of BMSCs were observed, and expression of the neuronal specific marker, neuron-specific enolase （NSE）, was analyzed by immunofluorescence staining after 4-5-day co-culture. RESULTS： The number of neural cells in the co-culture group increased and the cells spread on the culture bottle surface. Radial dendrite formed and connected with each other. NSE-immunoreactive cells were also detected. The positive ratio of NSE-positive cells reached （32.7±11.5）%, with morphological characteristics similar to neuronal cells. Human BMSCs did not express NSE in the control group. CONCLUSION： The microenvironment provided by neurons induced differentiation of BMSCs into neu-ronal-like cells.