Efficacy of serum-free cultured human umbilical cord mesenchymal stem cells in the treatment of knee osteoarthritis in mice

BACKGROUND We investigated the efficacy of intra-articular injection of human umbilical cord mesenchymal stem cells(hUC-MSCs)for the treatment of osteoarthritis(OA)progression in the knee joint.Although many experimental studies of hUC-MSCs have been published,these studies have mainly used fetal bovine serumcontaining cultures of hUC-MSCs;serum-free cultures generally avoid the shortcomings of serum-containing cultures and are not subject to ethical limitations,have a wide range of prospects for clinical application,and provide a basis or animal experimentation for clinical experiments.AIM To study the therapeutic effects of serum-free hUC-MSCs(N-hUCMSCs)in a mouse model of knee OA.METHODS Fifty-five male C57BL/6 mice were randomly divided into six groups:The blank control group,model control group,serum-containing hUC-MSCs(S-hUCMSC)group,N-hUCMSC group and hyaluronic acid(HA)group.After 9 weeks of modeling,the serum levels of interleukin(IL)-1β and IL-1 were determined.Hematoxylin-eosin staining was used to observe the cartilage tissue,and the Mankin score was determined.Immunohistochemistry and western blotting were used to determine the expression of collagen type II,matrix metalloproteinase(MMP)-1 and MMP-13.RESULTS The Mankin score and serum IL-1 and IL-1β and cartilage tissue MMP-1 and MMP-13 expression were significantly greater in the experimental group than in the blank control group(P<0.05).Collagen II expression in the experimental group was significantly lower than that in the blank control group(P<0.05).The Mankin score and serum IL-1 and IL-1β and cartilage tissue MMP-1 and MMP-13 levels the experimental group were lower than those in the model control group(P<0.05).Collagen II expression in the experimental group was significantly greater than that in the model control group(P<0.05).CONCLUSION N-hUCMSC treatment significantly alleviate the pathological damage caused by OA.The treatment effects of the ShUCMSC group and HA group were similar.

Human umbilical cord mesenchymal stem cells derivedexosomes on VEGF-A in hypoxic-induced mice retinal astrocytes and mice model of retinopathy of prematurity

AIM:To observe the effect of human umbilical cord mesenchymal stem cells(hUCMSCs)secretions on the relevant factors in mouse retinal astrocytes,and to investigate the effect of hUCMSCs on the expression of vascular endothelial growth factor-A(VEGF-A)and to observe the therapeutic effect on the mouse model of retinopathy of prematurity(ROP).METHODS:Cultured hUCMSCs and extracted exosomes from them and then retinal astrocytes were divided into control group and hypoxia group.MTT assay,flow cytometry,reverse transcription-polymerase chain reaction(RT-PCR)and Western blot were used to detect related indicators.Possible mechanisms by which hUCMSCs exosomes affect VEGF-A expression in hypoxia-induced mouse retinal astrocytes were explored.At last,the efficacy of exosomes of UCMSCs in a mouse ROP model was explored.Graphpad6 was used to comprehensively process data information.RESULTS:The secretion was successfully extracted from the culture supernatant of hUCMSCs by gradient ultracentrifugation.Reactive oxygen species(ROS)and hypoxia inducible factor-1α(HIF-1α)of mice retinal astrocytes under different hypoxia time and the expression level of VEGF-A protein and VEGF-A mRNA increased,and the ROP cell model was established after 6h of hypoxia.The secretions of medium and high concentrations of hUCMSCs can reduce ROS and HIF-1α,the expression levels of VEGF-A protein and VEGF-A mRNA are statistically significant and concentration dependent.Compared with the ROP cell model group,the expression of phosphatidylinositol 3-kinase(PI3K)/protein kinase B(AKT)/mammalian target of rapamycin(mTOR)signal pathway related factors in the hUCMSCs exocrine group is significantly decreased.The intravitreal injection of the secretions of medium and high concentrations of hUCMSCs can reduce VEGF-A and HIF-1αin ROP model tissues.HE staining shows that the number of retinal neovascularization in ROP mice decreases with the increase of the dose of hUCMSCs secretion.CONCLUSION:In a hypoxia induced mouse retinal astrocyte model,hUCMSCs exosomes are found to effectively reduce the expression of HIF-1αand VEGF-A,which are positively correlated with the concentration of hUCMSCs exosomes.HUCMSCs exosomes can effectively reduce the number of retinal neovascularization and the expression of HIF-1αand VEGF-A proteins in ROP mice,and are positively correlated with drug dosage.Besides,they can reduce the related factors on the PI3K/AKT/mTOR signaling pathway.

Effects of miR-214-5p and miR-21-5p in hypoxic endometrial epithelial-cell-derived exosomes on human umbilical cord mesenchymal stem cells

BACKGROUND Thin endometrium seriously affects endometrial receptivity,resulting in a significant reduction in embryo implantation,and clinical pregnancy and live birth rates,and there is no gold standard for treatment.The main pathophysiological characteristics of thin endometrium are increased uterine arterial blood flow resistance,angiodysplasia,slow growth of the glandular epithelium,and low expression of vascular endothelial growth factor,resulting in endometrial epithelial cell(EEC)hypoxia and endometrial tissue aplasia.Human umbilical cord mesenchymal stem cells(HucMSCs)promote repair and regeneration of damaged endometrium by secreting microRNA(miRNA)-carrying exosomes.However,the initiation mechanism of HucMSCs to repair thin endometrium has not yet been clarified.AIM To determine the role of hypoxic-EEC-derived exosomes in function of HucMSCs and explore the potential mechanism.METHODS Exosomes were isolated from normal EECs(EEC-exs)and hypoxia-damaged EECs(EECD-exs),before characterization using Western blotting,nanoparticletracking analysis,and transmission electron microscopy.HucMSCs were cocultured with EEC-exs or EECD-exs and differentially expressed miRNAs were determined using sequencing.MiR-21-5p or miR-214-5p inhibitors or miR-21-3p or miR-214-5p mimics were transfected into HucMSCs and treated with a signal transducer and activator of transcription 3(STAT3)activator or STAT3 inhibitor.HucMSC migration was assessed by Transwell and wound healing assays.Differentiation of HucMSCs into EECs was assessed by detecting markers of stromal lineage(Vimentin and CD13)and epithelial cell lineage(CK19 and CD9)using Western blotting and immunofluorescence.The binding of the miRNAs to potential targets was validated by dual-luciferase reporter assay.RESULTS MiR-21-5p and miR-214-5p were lowly expressed in EECD-ex-pretreated HucMSCs.MiR-214-5p and miR-21-5p inhibitors facilitated the migratory and differentiative potentials of HucMSCs.MiR-21-5p and miR-214-5p targeted STAT3 and protein inhibitor of activated STAT3,respectively,and negatively regulated phospho-STAT3.MiR-21-5p-and miR-214-5p-inhibitor-induced promotive effects on HucMSC function were reversed by STAT3 inhibition.MiR-21-5p and miR-214-5p overexpression repressed HucMSC migration and differentiation,while STAT3 activation reversed these effects.CONCLUSION Low expression of miR-21-5p/miR-214-5p in hypoxic-EEC-derived exosomes promotes migration and differentiation of HucMSCs into EECs via STAT3 signaling.Exosomal miR-214-5p/miR-21-5p may function as valuable targets for thin endometrium.

Preliminary study on the preparation of lyophilized acellular nerve scaffold complexes from rabbit sciatic nerves with human umbilical cord mesenchymal stem cells

BACKGROUND The gold standard of care for patients with severe peripheral nerve injury is autologous nerve grafting;however,autologous nerve grafts are usually limited for patients because of the limited number of autologous nerve sources and the loss of neurosensory sensation in the donor area,whereas allogeneic or xenografts are even more limited by immune rejection.Tissue-engineered peripheral nerve scaffolds,with the morphology and structure of natural nerves and complex biological signals,hold the most promise as ideal peripheral nerve“replacements”.AIM To prepare allogenic peripheral nerve scaffolds using a low-toxicity decellularization method,and use human umbilical cord mesenchymal stem cells(hUCMSCs)as seed cells to cultivate scaffold-cell complexes for the repair of injured peripheral nerves.METHODS After obtaining sciatic nerves from New Zealand rabbits,an optimal acellular scaffold preparation scheme was established by mechanical separation,varying lyophilization cycles,and trypsin and DNase digestion at different times.The scaffolds were evaluated by hematoxylin and eosin(HE)and luxol fast blue(LFB)staining.The maximum load,durability,and elastic modulus of the acellular scaffolds were assessed using a universal material testing machine.The acellular scaffolds were implanted into the dorsal erector spinae muscle of SD rats and the scaffold degradation and systemic inflammatory reactions were observed at 3 days,1 week,3 weeks,and 6 weeks following surgery to determine the histocompatibility between xenografts.The effect of acellular scaffold extracts on fibroblast proliferation was assessed using an MTT assay to measure the cytotoxicity of the scaffold residual reagents.In addition,the umbilical cord from cesarean section fetuses was collected,and the Wharton’s jelly(WJ)was separated into culture cells and confirm the osteogenic and adipogenic differentiation of mesenchymal stem cells(MSCs)and hUC-MSCs.The cultured cells were induced to differentiate into Schwann cells by the antioxidant-growth factor induction method,and the differentiated cells and the myelinogenic properties were identified.RESULTS The experiments effectively decellularized the sciatic nerve of the New Zealand rabbits.After comparing the completed acellular scaffolds among the groups,the optimal decellularization preparation steps were established as follows:Mechanical separation of the epineurium,two cycles of lyophilization-rewarming,trypsin digestion for 5 hours,and DNase digestion for 10 hours.After HE staining,no residual nuclear components were evident on the scaffold,whereas the extracellular matrix remained intact.LFB staining showed a significant decrease in myelin sheath composition of the scaffold compared with that before preparation.Biomechanical testing revealed that the maximum tensile strength,elastic modulus,and durability of the acellular scaffold were reduced compared with normal peripheral nerves.Based on the histocompatibility test,the immune response of the recipient SD rats to the scaffold New Zealand rabbits began to decline3 weeks following surgery,and there was no significant rejection after 6 weeks.The MTT assay revealed that the acellular reagent extract had no obvious effects on cell proliferation.The cells were successfully isolated,cultured,and passaged from human umbilical cord WJ by MSC medium,and their ability to differentiate into Schwann-like cells was demonstrated by morphological and immunohistochemical identification.The differentiated cells could also myelinate in vitro.CONCLUSION The acellular peripheral nerve scaffold with complete cell removal and intact matrix may be prepared by combining lyophilization and enzyme digestion.The resulting scaffold exhibited good histocompatibility and low cytotoxicity.In addition,hUC-MSCs have the potential to differentiate into Schwann-like cells with myelinogenic ability following in vitro induction.

Dose optimization of intrathecal administration of human umbilical cord mesenchymal stem cells for the treatment of subacute incomplete spinal cord injury

Human umbilical cord mesenchymal stem cells(hUC-MSCs)are a promising candidate for spinal cord injury(SCI)repair owing to their advantages of low immunogenicity and easy accessibility over other MSC sources.However,modest clinical efficacy hampered the progression of these cells to clinical translation.This discrepancy may be due to many variables,such as cell source,timing of implantation,route of administration,and relevant efficacious cell dose,which are critical factors that affect the efficacy of treatment of patients with SCI.Previously,we have evaluated the safety and efficacy of 4×10^(6) hUC-MSCs/kg in the treatment of subacute SCI by intrathecal implantation in rat models.To search for a more accurate dose range for clinical translation,we compared the effects of three different doses of hUC-MSCs-low(0.25×10^(6) cells/kg),medium(1×10^(6) cells/kg)and high(4×10^(6) cells/kg)-on subacute SCI repair through an elaborate combination of behavioral analyses,anatomical analyses,magnetic resonance imaging-diffusion tensor imaging(MRI-DTI),biotinylated dextran amine(BDA)tracing,electrophysiology,and quantification of mRNA levels of ion channels and neurotransmitter receptors.Our study demonstrated that the medium dose,but not the low dose,is as efficient as the high dose in producing the desired therapeutic outcomes.Furthermore,partial restoration of theγ-aminobutyric acid type A(GABAA)receptor expression by the effective doses indicates that GABAA receptors are possible candidates for therapeutic targeting of dormant relay pathways in injured spinal cord.Overall,this study revealed that intrathecal implantation of 1×10^(6) hUC-MSCs/kg is an alternative approach for treating subacute SCI.

Dissecting molecular mechanisms underlying ferroptosis in human umbilical cord mesenchymal stem cells:Role of cystathionineγ-lyase/hydrogen sulfide pathway

BACKGROUND Ferroptosis can induce low retention and engraftment after mesenchymal stem cell(MSC)delivery,which is considered a major challenge to the effectiveness of MSC-based pulmonary arterial hypertension(PAH)therapy.Interestingly,the cystathionineγ-lyase(CSE)/hydrogen sulfide(H_(2)S)pathway may contribute to mediating ferroptosis.However,the influence of the CSE/H_(2)S pathway on ferroptosis in human umbilical cord MSCs(HUCMSCs)remains unclear.AIM To clarify whether the effect of HUCMSCs on vascular remodelling in PAH mice is affected by CSE/H_(2)S pathway-mediated ferroptosis,and to investigate the functions of the CSE/H_(2)S pathway in ferroptosis in HUCMSCs and the underlying mechanisms.METHODS Erastin and ferrostatin-1(Fer-1)were used to induce and inhibit ferroptosis,respectively.HUCMSCs were transfected with a vector to overexpress or inhibit expression of CSE.A PAH mouse model was established using 4-wk-old male BALB/c nude mice under hypoxic conditions,and pulmonary pressure and vascular remodelling were measured.The survival of HUCMSCs after delivery was observed by in vivo bioluminescence imaging.Cell viability,iron accumulation,reactive oxygen species production,cystine uptake,and lipid peroxidation in HUCMSCs were tested.Ferroptosis-related proteins and S-sulfhydrated Kelchlike ECH-associating protein 1(Keap1)were detected by western blot analysis.RESULTS In vivo,CSE overexpression improved cell survival after erastin-treated HUCMSC delivery in mice with hypoxiainduced PAH.In vitro,CSE overexpression improved H_(2)S production and ferroptosis-related indexes,such as cell viability,iron level,reactive oxygen species production,cystine uptake,lipid peroxidation,mitochondrial membrane density,and ferroptosis-related protein expression,in erastin-treated HUCMSCs.In contrast,in vivo,CSE inhibition decreased cell survival after Fer-1-treated HUCMSC delivery and aggravated vascular remodelling in PAH mice.In vitro,CSE inhibition decreased H_(2)S levels and restored ferroptosis in Fer-1-treated HUCMSCs.Interestingly,upregulation of the CSE/H_(2)S pathway induced Keap1 S-sulfhydration,which contributed to the inhibition of ferroptosis.CONCLUSION Regulation of the CSE/H_(2)S pathway in HUCMSCs contributes to the inhibition of ferroptosis and improves the suppressive effect on vascular remodelling in mice with hypoxia-induced PAH.Moreover,the protective effect of the CSE/H_(2)S pathway against ferroptosis in HUCMSCs is mediated via S-sulfhydrated Keap1/nuclear factor erythroid 2-related factor 2 signalling.The present study may provide a novel therapeutic avenue for improving the protective capacity of transplanted MSCs in PAH.

Enhanced wound healing and hemostasis with exosome-loaded gelatin sponges from human umbilical cord mesenchymal stem cells

BACKGROUND Rapid wound healing remains a pressing clinical challenge,necessitating studies to hasten this process.A promising approach involves the utilization of human umbilical cord mesenchymal stem cells(hUC-MSCs)derived exosomes.The hypothesis of this study was that these exosomes,when loaded onto a gelatin sponge,a common hemostatic material,would enhance hemostasis and accelerate wound healing.AIM To investigate the hemostatic and wound healing efficacy of gelatin sponges loaded with hUC-MSCs-derived exosomes.METHODS Ultracentrifugation was used to extract exosomes from hUC-MSCs.Nanoparticle tracking analysis(NTA),transmission electron microscopy(TEM),and western blot techniques were used to validate the exosomes.In vitro experiments were performed using L929 cells to evaluate the cytotoxicity of the exosomes and their impact on cell growth and survival.New Zealand rabbits were used for skin irritation experiments to assess whether they caused adverse skin reactions.Hemolysis test was conducted using a 2%rabbit red blood cell suspension to detect whether they caused hemolysis.Moreover,in vivo experiments were carried out by implanting a gelatin sponge loaded with exosomes subcutaneously in Sprague-Dawley(SD)rats to perform biocompatibility tests.In addition,coagulation index test was conducted to evaluate their impact on blood coagulation.Meanwhile,SD rat liver defect hemostasis model and full-thickness skin defect model were used to study whether the gelatin sponge loaded with exosomes effectively stopped bleeding and promoted wound healing.RESULTS The NTA,TEM,and western blot experimental results confirmed that exosomes were successfully isolated from hUC-MSCs.The gelatin sponge loaded with exosomes did not exhibit significant cell toxicity,skin irritation,or hemolysis,and they demonstrated good compatibility in SD rats.Additionally,the effectiveness of the gelatin sponge loaded with exosomes in hemostasis and wound healing was validated.The results of the coagulation index experiment indicated that the gelatin sponge loaded with exosomes had significantly better coagulation effect compared to the regular gelatin sponge,and they showed excellent hemostatic performance in a liver defect hemostasis model.Finally,the full-thickness skin defect healing experiment results showed significant improvement in the healing process of wounds treated with the gelatin sponge loaded with exosomes compared to other groups.CONCLUSION Collectively,the gelatin sponge loaded with hUC-MSCs-derived exosomes is safe and efficacious for promoting hemostasis and accelerating wound healing,warranting further clinical application.

Protective effects of human umbilical cord mesenchymal stem cells on retinal ganglion cells in mice with acute ocular hypertension

AIM:To observe the protective effect of human umbilical cord mesenchymal stem cells(huc MSCs)on retinal ganglion cel s(RGCs)injury in mice with acute ocular hypertension(AOH).METHODS:Fifty-six adult male C57 BL/6 mice were randomly divided into four groups:normal group,AOH group,huc MSCs group,normal saline(NS)group.Left eye of mice was induced by 90 mm Hg intraocular pressure for 1 h to establish AOH model.huc MSCs 1×105/μL,1μL or NS 1μL was injected into the vitreous body the next day.CMDil fluorescent dye was used to label the 3 rd generation of huc MSCs,for tracing the cells in the vitreous cavity of mice.Seven days after the model established,hematoxylin-eosin(HE)staining was used to observe the thickness of the inner retina layer in four groups.Numbers and loss rate of RGCs were evaluated by counting Brn-3 a positive cells stained by immunofluorescencein.RESULTS:On the 7 th day after AOH established,labeled huc MSCs were found in the vitreous cavity.HE staining showed that the thickness of retinal inner layer in AOH group was significantly lower than that in normal group and huc MSCs group(P<0.05),same as that in NS group(P>0.05).Compared with AOH group,the RGCs in normal group was significantly higher;RGCs number increased in huc MSCs group and the loss rate was lower(P<0.05).Injection of NS had no protective effect on RGCs.CONCLUSION:In AOH mouse model,vitreous injection of hucMSCs have shown a protection for RGCs.

Effect of Human Umbilical Cord Mesenchymal Stem Cells on GRP78/ATF4 Pathway in Alzheimer s Disease Model Mice

[Objectives]To study the effect of human umbilical cord mesenchymal stem cells(hUC-MSCs)on GRP78/ATF4 pathway in APP/PS1 mice.[Methods]Twelve 6-month-old female APP/PS1 mice were randomly divided into model group(MOD,n=6)and human umbilical cord mesenchymal stem cell treatment group(MSC,n=6);six 6-month-old C57BL/6N mice were used as control group(CON,n=6).The mice in each group were treated with the fourth generation of human umbilical cord mesenchymal stem cells through tail vein.Four weeks later,the mice in each group were killed.The expression of GFP78 and ATF4 in the cortex of mice in each group was detected by Western blotting and real-time fluorescence quantitative PCR.[Results]The results of immunoblotting and real-time fluorescence quantitative PCR showed that the expression of GRP78 in MOD group was lower than that in CON group and the expression of ATF4 increased.The expression of GRP78 protein in MSC group was higher than that in MOD group,but the expression of ATF4 protein was lower.The results of real-time fluorescence quantitative PCR showed that the mRNA level of GRP78 decreased and the mRNA level of ATF4 increased in MOD group compared with CON group.The mRNA level of GRP78 in MSC group was higher than that in MOD group,while the mRNA level of ATF4 in MSC group was lower than that in MOD group.[Conclusions]Human umbilical cord mesenchymal stem cells can regulate the expression of GRP78/ATF4 pathway in APP/PSI mice,which may be related to the stress level of endoplasmic reticulum in the brain of APP/PS1 mice mediated by human umbilical cord mesenchymal stem cells.

Human umbilical cord mesenchymal stem cells as cell carriers in the treatment of gliomas: research status and prospects

Human umbilical cord mesenchymal stem cells(HuMSCs)have the multi-difFerentiation potential to differentiate into various types of cells without immune rejection.They are considered to be an ideal source of neural stem cells and also an ideal cell carrier for gene therapy.Because of the invasive growth of brain gliomas,most of them have no obvious boundaries with normal brain tissues.It is difficult to completely remove them by surgery and the remaining cells become the main source of tumor recurrence.In recent years,gene therapy has become a new method for the treatment of gliomas.The vector carrying the target gene is introduced into HuMSCs in a certain way to correct gene defects or play other roles.The differentiation potential of HuMSCs makes it an ideal source of nerve cells to play a greater role in gene therapy of glioma.Therefore,this article reviews the current status and prospects of HuMSCs as cell carriers in the treatment of glioma.

Influence of bone morphogenetic proteins-2 and strontium chloride on the human umbilical cord mesenchymal stem cells proliferation and osteogenic differentiation

Objective To study the effects of combination of bone morphogenetic protein-2 (BMP-2) and strontium chloride on proliferation and osteogenic differentiation of human umbilical cord mesenchymal stem cells(hUCM-SCs)in vitro culture.

Human umbilical cord mesenchymal stem cells to treat spinal cord injury in the early chronic phase: study protocol for a prospective, multicenter, randomized, placebo-controlled, single-blinded clinical trial

Human umbilical cord mesenchymal stem cells(hUC-MSCs)support revascularization,inhibition of inflammation,regulation of apoptosis,and promotion of the release of beneficial factors.Thus,they are regarded as a promising candidate for the treatment of intractable spinal cord injury(SCI).Clinical studies on patients with early chronic SCI(from 2 months to 1 year post-injury),which is clinically common,are rare;therefore,we will conduct a prospective,multicenter,randomized,placebo-controlled,single-blinded clinical trial at the Third Affiliated Hospital of Sun Yat-sen University,West China Hospital of Sichuan University,and Shanghai East Hospital,Tongji University School of Medicine,China.The trial plans to recruit 66 early chronic SCI patients.Eligible patients will undergo randomization at a 2:1 ratio to two arms:the observation group and the control group.Subjects in the observation group will receive four intrathecal transplantations of stem cells,with a dosage of 1×106/kg,at one calendar month intervals.Subjects in the control group will receive intrathecal administrations of 10 mL sterile normal saline in place of the stem cell transplantations.Clinical safety will be assessed by the analysis of adverse events and laboratory tests.The American Spinal Injury Association(ASIA)total score will be the primary efficacy endpoint,and the secondary efficacy outcomes will be the following:ASIA impairment scale,International Association of Neural Restoration-Spinal Cord Injury Functional Rating Scale,muscle tension,electromyogram,cortical motor and cortical sensory evoked potentials,residual urine volume,magnetic resonance imaging–diffusion tensor imaging,T cell subtypes in serum,neurotrophic factors and inflammatory factors in both serum and cerebrospinal fluid.All evaluations will be performed at 1,3,6,and 12 months following the final intrathecal administration.During the entire study procedure,all adverse events will be reported as soon as they are noted.This trial is designed to evaluate the clinical safety and efficacy of subarachnoid transplantation of hUC-MSCs to treat early chronic SCI.Moreover,it will establish whether cytotherapy can ameliorate local hostile microenvironments,promote tracking fiber regeneration,and strengthen spinal conduction ability,thus improving overall motor,sensory,and micturition/defecation function in patients with early chronic SCI.This study was approved by the Stem Cell Research Ethics Committee of the Third Affiliated Hospital of Sun Yat-sen University,China(approval No.[2018]-02)on March 30,2018,and was registered with ClinicalTrials.gov(registration No.NCT03521323)on April 12,2018.The revised trial protocol(protocol version 4.0)was approved by the Stem Cell Research Ethics Committee of the Third Affiliated Hospital of Sun Yat-sen University,China(approval No.[2019]-10)on February 25,2019,and released on ClinicalTrials.gov on April 29,2019.

Human umbilical cord mesenchymal stem cells promote peripheral nerve repair via paracrine mechanisms

Human umbilical cord-derived mesenchymal stem cells （hUCMSCs） represent a promising young-state stem cell source for cell-based therapy. hUCMSC transplantation into the transected sciatic nerve promotes axonal regeneration and functional recovery. To further clarify the para-crine effects of hUCMSCs on nerve regeneration, we performed human cytokine antibody array analysis, which revealed that hUCMSCs express 14 important neurotrophic factors. Enzyme-linked immunosorbent assay and immunohistochemistry showed that brain-derived neurotrophic factor, glial-derived neurotrophic factor, hepatocyte growth factor, neurotrophin-3, basic fibroblast growth factor, type I collagen, fibronectin and laminin were highly expressed. Treatment with hUCMSC-conditioned medium enhanced Schwann cell viability and proliferation, increased nerve growth factor and brain-derived neurotrophic factor expression in Schwann cells, and enhanced neurite growth from dorsal root ganglion explants. These ifndings suggest that paracrine action may be a key mechanism underlying the effects of hUCMSCs in peripheral nerve repair.

Differentiation of isolated human umbilical cord mesenchymal stem cells into neural stem cells

AIM：To investigate whether umbilical cord human mesenchymal stem cell（UC-MSC）was able to differentiate into neural stem cell and neuron.·METHODS：The umbilical cords were o btained from pregnant women with their written consent and the approval of the Clinic Ethnics Committee.UC-MSC were isolated by adherent culture in the medium contains 20%fetal bovine serum（FBS）,then they were maintained in the medium contain 10%FBS and induced to neural cells in neural differentiation medium.We investigated whether UC-MSC was able to differentiate into neural stem cell and neuron by using flow cytometry,reverse transcriptase-polymerase chain reaction（RT-PCR）and immunofluorescence（IF）analyzes.·R ESULTS：A substantial number of UC-MSC was harvested using the tissue explants adherent method at about 2wk.Flow cytometric study revealed that these cells expressed common markers of MSCs,such as CD105（SH2）,CD73（SH3）and CD90.After induction of differentiation of neural stem cells,the cells began to form clusters;RT-PCR and IF showed that the neuron specific enolase（NSE）and neurogenic differentiation 1-positive cells reached 87.3%±14.7%and 72.6%±11.8%,respectively.Cells showed neuronal cell differentiation after induced,including neuron-like protrusions,plump cell body,obviously and stronger refraction.RT-PCR and IF analysis showed that microtubule-associated protein 2（MAP2）and nuclear factor-M-positive cells reached 43.1%±10.3%and 69.4%±19.5%,respectively.·CONCLUSION：Human umbilical cord derived MSCs can be cultured and proliferated and differentiate into neural stem cells,which may be a valuable source for cell therapy of neurodegenerative eye diseases.

Human umbilical cord mesenchymal stem cells as treatment of adjuvant rheumatoid arthritis in a rat model

AIM:To investigate the effect of human umbilical cord stem cells,both mesenchymal and hematopoietic(CD34+),in the treatment of arthritis.METHODS:Mesenchymal stem cells(MSCs) and hematopoietic(CD34+) stem cells(HSC) were isolated from human umbilical cord blood obtained from the umbilical cord of healthy pregnant donors undergoing fullterm normal vaginal delivery.MSC,HSC,methotrexate(MTX) and sterile saline were injected intra-articularly into the rat hindpaw with complete freunds adjuvant(CFA) induced arthritis after the onset of disease(day 34),when arthritis had become well established(arthritis score ≥ 2).Arthritic indices were evaluated and the levels of interleukin(IL)-1,tumor necrosis factor(TNF)-α and interferon(IFN)-γ and anti-inflammatory cytokine IL-10 in serum were determined using enzyme-linked immunosorbent assay.Animals of all groups were sacrificed 34 d after beginning treatment,except positive control(PC) which was sacrificed at 10,21 and 34 d for microscopic observation of disease progression.We used hematoxylin,eosin and Masson's trichrome stains for histopathological examination of cartilage and synovium.RESULTS:The mean arthritis scores were similar in all groups at 12 and 34 d post immunization,with no statistical significant difference.Upon the injection of stem cells(hematopoietic and mesenchymal),the overall arthritis signs were significantly improved around 21 d after receiving the injection and totally disappeared at day 34 post treatment in MSC group.Mean hindpaw diameter(mm) in the MSC rats was about half that of the PC and MTX groups(P = 0.007 and P = 0.021,respectively) and 0.6 mm less than the HSC group(P = 0.047),as indicated by paw swelling.Associated with these findings,serum levels of TNF-α,IFN-γ and IL-1 decreased significantly in HSC and MSC groups compared to PC and MTX groups(P < 0.05),while the expression of IL-10 was increased.Histopathological examination with H and E stain revealed that the MTX treated group showed significant reduction of leucocytic infiltrate and hypertrophy of the synovial tissue with moderate obliteration of the joint cavity.Stem cells treated groups(both hematopoietic CD34+ and mesenchymal),showed significant reduction in leucocytic infiltrate and hypertrophy of the synovial tissue with mild obliteration of the joint cavity.With Masson's trichrome,stain sections from the PC group showed evidence of vascular edema of almost all vessels within the synovium in nearly all arthritic rats.Vacuoles were also visible in the outer vessel wall.The vessel became hemorrhagic and finally necrotic.In addition,there was extensive fibrosis completely obliterating the joint cavity.The mean color area percentage of collagen in this group was 0.324 ± 0.096,which was significantly increased when compared to the negative control group.The mean color area percentage of collagen in hematopoietic CD34+ and mesenchymal groups was 0.176 ± 0.0137 and 0.174 ± 0.0197 respectively,which showed a marked decrement compared to the PC group,denoting a mild increase in synovial tissue collagen fibers.CONCLUSION:MSC enhance the efficacy of CFAinduced arthritis treatment,most likely through the modulation of the expression of cytokines and amelioration of pathological changes in joints.

Experimental treatment of radiation pneumonitis with human umbilical cord mesenchymal stem cells

Objective:To evaluate of the curative effect of human umbilical cord mesenchymal stem cells(hUC-MSCs)on rat acute radiation pneumonitis.Methods:Fourty rats were randomly divided into control group,radiation group,stem cell prevention group,stem cell treatment group and prednisone treatment group.All rats except those in the control group were radiated with X ray to establish the acute radiation pneumonitis damage model.The hUC-MSCs cultured in vitro was administrated to the rats of the prevention group via tail vein(1×10~6 cells/kg BW)24 h before the radiation,while the same administration was performed in the rats of the treatment group 24 h after the radiation.After 24 h post the radiation,the rats in tbe radiation group were given 0.4 mL physiological saline,and those in the prednisone group were given 1 mg/kg prednisone.All rats were,observed and executed 72 h after the radiation to defect lung histological changes.Results:After the administration of hUC-MSCs,the survival status of the rats in the prevention group and treatment group was obviously better than that in the control group.As shown by the histological staining,the morphology,proliferation activity aad bronchial state of lung tissues were better in the prevention group and treatment group than in the control group.Conclusion:The hUC-MSCs have definite therapeutic effects on acute radiation pneumonitis in rats.

An experimental study of preventing and treating acute radioactive enteritis with human umbilical cord mesenchymal stem cells

Objective:To test the curative effect of human umbilical cord-derived mesenchymal stem cells on rat acute radioactive enteritis and thus in provide clinical therapeutic basis for radiation sickness.Methods:Human umbilical cord-derived mesenchymal stem cells were cultivated in vitro and the model of acute radioactive enteritis of rats was established.Then,the umbilical cord mesenchymal stem cells were injected into the rats via tail vein.Visual and histopathiological changes of the experimental rats were observed.Results:After the injection,the rats in the prevention group and treatment group had remarkably better survival status than those in the control group.The histological observations revealed that the former also had better intestinal mucosa structure,more regenerative cells and stronger proliferation activity than the latter.Conclusions:Human umbilical cord-derived mesenchymal stem cells have a definite therapeutic effect on acute radioactive enteritis in rats.

Ability of human umbilical cord mesenchymal stem cells to repair chemotherapy-induced premature ovarian failure

BACKGROUND Premature ovarian insufficiency(POI)and premature ovarian failure(POF)have become one of the major problems threatening women of childbearing age.Studies have shown that stem cells transplanted from bone marrow,umbilical cord,peripheral blood and amniotic fluid can migrate and proliferate to the ovary,promote ovarian function repair,increase the number of follicles and granulosa cells at all levels of ovary,improve endocrine function,and can differentiate into oocytes in specific ovarian environment to restore fertility to some extent.AIM To study the ability of human umbilical cord mesenchymal stem cells(hUCMSCs)to repair ovarian injury after chemotherapy.METHODS A total of 110 female BALB/c mice(aged 7-8 wk old)with body masses of 16.0-20.0 g were selected.The mice were fed until 12 wk of age,and cyclophosphamide was administered by intraperitoneal injection for 14 consecutive days to induce premature ovarian failure in mice.Seventy-five mice with estrous cycle disorder were screened and randomly divided into 3 groups according to their body weight:model group,positive control group and hUCMSC group,and each group had 25 mice.Another 25 mice were used as negative controls.The mice in the hUCMSC group were injected with hUCMSCs in the tail vein,and the mice in the positive control group were given an oestradiol valerate solution and a medroxyprogesterone acetate solution in the tail vein.On the 1^st,15^th,30^th,45^th,and 60^th days after intravenous administration,vaginal smears were made to monitor the estrous cycles of the mice.The ovaries were weighed,and pathological sections were made to observe the morphology of the follicles;blood samples were collected to monitor the concentration of sex hormones(oestradiol and follicle-stimulating hormone).RESULTS The estrous cycles of the model group mice were disrupted throughout the experiment.Mice in the hUCMSC group and the positive control group resumed normal estrous cycles.The ovarian weight of the model group mice continued to decline.The ovarian weight of the hUCMSC group mice and the positive control group mice decreased first and then gradually increased,and the ovarian weight of the hUCMSC group mice was heavier than that of the positive control group mice.The difference was statistically significant(P<0.05).Compared with the negative control group,the model group experienced a decrease in oestradiol and an increase in follicle-stimulating hormone,and the difference was statistically significant(P<0.05).Compared with the model group,the hUCMSC and positive control groups experienced a slight increase in oestradiol and a decrease in follicle-stimulating hormone;the difference was statistically significant(P<0.05).The pathological examination revealed that the mouse ovaries from the model group were atrophied,the volume was reduced,the cortical and medullary structures were disordered,the number of follicles at all stages was significantly reduced,the number of atretic follicles increased,the number of primordial follicles and corpus luteum significantly decreased,and the corpus luteum had an irregular shape.Compared with those of the model group,the lesions of the hUCMSC and positive control groups significantly improved.CONCLUSION hUCMSCs can repair ovarian tissue damaged by chemotherapy to a certain extent,can improve the degree of apoptosis in ovarian tissue,and can improve the endocrine function of mouse ovaries.

Human umbilical cord mesenchymal stem cells ameliorate liver fibrosis in vitro and in vivo: From biological characteristics to therapeutic mechanisms

Liver fibrosis is a wound-healing response to chronic injuries, characterized by the excessive accumulation of extracellular matrix or scar tissue within the liver;in addition, its formation is associated with multiple cytokines as well as several cell types and a variety of signaling pathways. When liver fibrosis is not well controlled, it can progress to liver cirrhosis, but it is reversible in principle. Thus far, no efficient therapy is available for treatment of liver fibrosis. Although liver transplantation is the preferred strategy, there are many challenges remaining in this approach, such as shortage of donor organs, immunological rejection, and surgical complications. Hence, there is a great need for an alternative therapeutic strategy. Currently, mesenchymal stem cell (MSC) therapy is considered a promising therapeutic strategy for the treatment of liver fibrosis;advantageously, the characteristics of MSCs are continuous self-renewal, proliferation, multipotent differentiation, and immunomodulatory activities. The human umbilical cord-derived (hUC)-MSCs possess not only the common attributes of MSCs but also more stable biological characteristics, relatively easy accessibility, abundant source, and no ethical issues (e.g., bone marrow being the adult source), making hUC-MSCs a good choice for treatment of liver fibrosis. In this review, we summarize the biological characteristics of hUC-MSCs and their paracrine effects, exerted by secretion of various cytokines, which ultimately promote liver repair through several signaling pathways. Additionally, we discuss the capacity of hUC-MSCs to differentiate into hepatocyte-like cells for compensating the function of existing hepatocytes, which may aid in amelioration of liver fibrosis. Finally, we discuss the current status of the research field and its future prospects.

Exosomes Derived from Human Umbilical Cord Mesenchymal Stem Cells Enhance the Osteoblastic Differentiation of Periodontal Ligament Stem Cells Under High Glucose Conditions Through the PI3K/AKT Signaling Pathway

Objective High glucose(HG)can influence the osteogenic differentiation ability of periodontal ligament stem cells(PDLSCs).Human umbilical cord mesenchymal stem cell-derived exosomes(hUCMSC-exo)have broad application prospects in tissue healing.The current study aimed to explore whether hUCMSC-exo could promote the osteogenic differentiation of hPDLSCs under HG conditions and the underlying mechanism.Methods We used a 30 mmol/L glucose concentration to simulate HG conditions.CCK-8 assay was performed to evaluate the effect of hUCMSC-exo on the proliferation of hPDLSCs.Alkaline phosphatase(ALP)staining,ALP activity,and qRT-PCR were performed to evaluate the pro-osteogenic effect of hUCMSC-exo on hPDLSCs.Western blot analysis was conducted to evaluate the underlying mechanism.Results The results of the CCK-8 assay,ALP staining,ALP activity,and qRT-PCR assay showed that hUCMSC-exo significantly promoted cell proliferation and osteogenic differentiation in a dosedependent manner.The Western blot results revealed that hUCMSC-exo significantly increased the levels of p-PI3K and p-AKT in cells,and the effect was inhibited by LY294002(PI3K inhibitor)or MK2206(AKT inhibitor),respectively.Moreover,the increases in osteogenic indicators induced by hUCMSC-exo were significantly suppressed by LY294002 and MK2206.Conclusion hUCMSC-exo promote the osteogenic differentiation of hPDLSCs under HG conditions through the PI3K/AKT signaling pathway.