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.

Human umbilical cord mesenchymal stem cell-derived exosomes loaded into a composite conduit promote functional recovery after peripheral nerve injury in rats

Complete transverse injury of peripheral nerves is challenging to treat.Exosomes secreted by human umbilical cord mesenchymal stem cells are considered to play an important role in intercellular communication and regulate tissue regeneration.In previous studies,a collagen/hyaluronic acid sponge was shown to provide a suitable regeneration environment for Schwann cell proliferation and to promote axonal regeneration.This three-dimensional(3D)composite conduit contains a collagen/hyaluronic acid inner sponge enclosed in an electrospun hollow poly(lactic-co-glycolic acid)tube.However,whether there is a synergy between the 3D composite conduit and exosomes in the repair of peripheral nerve injury remains unknown.In this study,we tested a comprehensive strategy for repairing long-gap(10 mm)peripheral nerve injury that combined the 3D composite conduit with human umbilical cord mesenchymal stem cell-derived exosomes.Repair effectiveness was evaluated by sciatic functional index,sciatic nerve compound muscle action potential recording,recovery of muscle mass,measuring the cross-sectional area of the muscle fiber,Masson trichrome staining,and transmission electron microscopy of the regenerated nerve in rats.The results showed that transplantation of the 3D composite conduit loaded with human umbilical cord mesenchymal stem cell-derived exosomes promoted peripheral nerve regeneration and restoration of motor function,similar to autograft transplantation.More CD31-positive endothelial cells were observed in the regenerated nerve after transplantation of the loaded conduit than after transplantation of the conduit without exosomes,which may have contributed to the observed increase in axon regeneration and distal nerve reconnection.Therefore,the use of a 3D composite conduit loaded with human umbilical cord mesenchymal stem cell-derived exosomes represents a promising cell-free therapeutic option for the treatment of peripheral nerve injury.

Exosomes from umbilical cord mesenchymal stromal cells promote the collagen production of fibroblasts from pelvic organ prolapse

BACKGROUND Pelvic organ prolapse(POP)involves pelvic organ herniation into the vagina due to pelvic floor tissue laxity,and vaginal structure is an essential factor.In POP,the vaginal walls exhibit abnormal collagen distribution and decreased fibroblast levels and functions.The intricate etiology of POP and the prohibition of trans-vaginal meshes in pelvic reconstruction surgery present challenges in targeted therapy development.Human umbilical cord mesenchymal stromal cells(hucMSCs)present limitations,but their exosomes(hucMSC-Exo)are promising therapeutic tools for promoting fibroblast proliferation and extracellular matrix remodeling.suppressed inflammation in POP group fibroblasts,stimulated primary fibroblast growth,and elevated collagen I(Col1)production in vitro.High-throughput RNA-seq of fibroblasts treated with hucMSC-Exo and miRNA sequencing of hucMSC-Exo revealed that abundant exosomal miRNAs downregulated matrix metalloproteinase 11(MMP11)expression.CONCLUSION HucMSC-Exo normalized the growth and function of primary fibroblasts from patients with POP by promoting cell growth and Col1 expression in vitro.Abundant miRNAs in hucMSC-Exo targeted and downregulated MMP11 expression.HucMSC-Exo-based therapy may be ideal for safely and effectively treating POP.

Stem cell transplantation for treatment of cerebral ischemia in rats Effects of human umbilical cord blood stem cells and human neural stem cells

BACKGROUND： Exogenous neural stem cell transplantation promotes neural regeneration. However, various types of stem cells transplantation outcomes remain controversial. OBJECTIVE： To explore distribution, proliferation and differentiation of human neural stem cells （hNSCs） and human umbilical cord blood stem cells （hUCBSCs） following transplantation in ischemic brain tissue of rats, and to compare therapeutic outcomes between hNSCs and hUCBSCs. DESIGN, TIME AND SETTING： Randomized controlled animal studies were performed at the Experimental Animal Center of Nanjing Medical University and Central Laboratory of Second Affiliated Hospital of Nanjing Medical University of China from September 2008 to April 2009. MATERIALS： hNSCs were harvested from brain tissue of 10 13 week old fetuses following spontaneous abortion, and hUCBSCs were collected from umbilical cord blood of full-term newborns at the Second Affiliated Hospital of Nanjing Medical University of China. hNSCs and hUCBSCs were labeled by 5-bromodeoxyuridine （BrdU） prior to transplantation. METHODS： Rat models of cerebral ischemia were established by the suture method. A total of 60 healthy male Sprague Dawley rats aged 7-9 weeks were randomly assigned to hNSC transplantation, hUCBSC transplantation and control groups. The rat models in the hNSC transplantation, hUCBSC transplantation and control groups were infused with hNSC suspension, hUCBSC suspension and saline via the caudal vein, respectively. MAIN OUTCOME MEASURES： The distribution, proliferation and differentiation of hNSCs and hUCBSCs in ischemic brain tissue were observed using immunohistochemical methods. Neurological function in rats was assessed using the neurological severity score. RESULTS： The number of BrdU-positive cells was significantly greater in the hNSC transplantation group compared with hUCBSC transplantation group at 14 days following transplantation （P 〈 0.05） The number of BrdU-positive cells reached a peak at 28 days following transplantation. Nestin-positive, glial fibrillary acidic protein-positive, cyclic nucleotide 3＇ phosphohydrolase-positive and neuron specific enolase-positive cells were visible following transplantation. No significant difference was determined in the constituent ratio of various cells between hNSC and hUCBSC transplantation groups （P 〉 0.05）. The neurological severity score was significantly decreased in rats at 21 days following transplantation （P 〈 0.05）. No significant difference was detected in neurological severity score between hNSC and hUCBSC transplantation groups at various time points （P 〉 0.05）. CONCLUSION： The transplanted hNSCs and hUCBSCs can migrate into ischemic brain tissue, proliferate and differentiate into neuron-like, astrocyte-like and oligodendrocyte-like cells, and improve neurological function in rats with cerebral ischemia.

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.

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.

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.

Vein transplantation using human umbilical cord blood stem cells in the treatment of stroke sequela

BACKGROUND: Transplanted mononuclear cell (MNC) of umbilical blood can survive in central nervous system (CNS) of host through blood brain barrier, differentiate into nerve cells, migrate to damaged site and integrate morphological structure and function with nerve cells of host so as to improve deficiencies of sensatory function, motor function and cognitive function and influence on stroke sequela. OBJECTIVE: To observe the vein transplantation of human umbilical cord blood stem cells (HUCBSC) for improving neurological function, limb function and activity of daily living of patients with stroke and evaluate the reliability. DESIGN: Self-controlled study. SETTING: Department of Neurosurgery, the Second People's Hospital of Zhengzhou City; Red-crossed Blood Center of Henan Province; Department of Neurosurgery, the Fist Affiliated Hospital of Zhengzhou University. PARTICIPANTS: A total of 10 patients with stroke sequela were selected from Department of Cerebral Surgery, the Second People's Hospital of Zhengzhou City from April to December 2005. There were 9 males and 1 female aged from 35 to 75 years with the mean age of 56 years. All of them were diagnosed with CT and MRI examination and coincidence with diagnostic criteria of stroke established by the Fourth National Academic Meeting for Cerebrovascular Disease. All patients provided informed consent. METHODS: 80-140 mL umbilical blood of term birth of newborn was selected hermetically and maintained in sterile plastic bag. And then, the blood was centrifugated at the speed of 1 500 r/min for 30 minutes at 22 ℃ in order to separate MNC, i.e., HUCBSC. In addition, after final diagnosis during hospitalization, stroke patients were perfused with HUCBSC through superficial vein of back of the hand. Each patient was averagely perfused with 6 portions of HUCBSC (cellular numbers ≥ 1×108/portion) and the interval between each portion was 1-7 days with the mean interval of 4 days. MAIN OUTCOME MEASURES: ① Neurological function of stroke patients was evaluated with neurological function deficiency (NFD) before treatment and at 3 months after treatment. The scale includes consciousness, level fix function, facial paralysis, language, muscle force of upper limbs, muscle force of lower limb and step function. The total scores ranged from 0 to 45; meanwhile, the lower the scores were, the better the neurological function was. ② Motor function of injured limbs was evaluated with Fugl-Meyer Assessment (FMA), including motor function of upper limbs, motor function of lower limbs, balance ability, sensory function and motion of joint. The total scores ranged from 0 to 226; meanwhile, the higher the scores were, the better the motor function of limbs was. ③ Activities of daily living (ADL) was evaluated with Barthel Index (BI), including having meals, taking a bath, dressing oneself, putting on clothes, walking in balance and stair activity. The total scores ranged from 0 to 100; meanwhile, the higher the scores were, the stronger the ADL was. RESULTS: A total of 10 patients were involved in the final analysis. After treatment, NFD of stroke patients was (10.9±5.09) points, which was lower than that before treatment [(25.4±6.09) points, t =8.213, P < 0.01]. In addition, after treatment, FMA and BI of stroke patients were (80.9±25.00) points and (81.1±15.93) points, respectively, which were higher than those before treatment [(31.9±21.85) points, (36.2±19.41) points, t =13.024, 13.670, P < 0.01]. Immuno-suppressive drugs were not used during the whole therapeutic procedure; moreover, immunological rejection and allergic reaction were not observed during the same period. CONCLUSION: Transplanting HUCBSC through superficial vein of back of the hand is regarded as a simple and safe method for the treatment of stroke sequela.

Human umbilical cord blood-derived mesenchymal stem cells promote regeneration of crush-injured rat sciatic nerves

Several studies have demonstrated that human umbilical cord blood-derived mesenchymal stem cells can promote neural regeneration following brain injury. However, the therapeutic effects of human umbilical cord blood-derived mesenchymal stem cells in guiding peripheral nerve regeneration remain poorly understood. This study was designed to investigate the effects of human umbilical cord blood-derived mesenchymal stem cells on neural regeneration using a rat sciatic nerve crush injury model. Human umbilical cord blood-derived mesenchymal stem cells （1 ~ 106） or a PBS control were injected into the crush-injured segment of the sciatic nerve. Four weeks after cell injection, brain-derived neurotrophic factor and tyrosine kinase receptor B mRNA expression at the lesion site was increased in comparison to control. Furthermore, sciatic function index, Fluoro Gold-labeled neuron counts and axon density were also significantly increased when compared with control. Our results indicate that human umbilical cord blood-derived mesenchvmal stem cells promote the functinnal r~.RcJv^rv nf P.n I^h-inillr^4 ~r^i~tit, n^r~e

Transplantation of human umbilical cord blood mesenchymal stem cells to treat a rat model of traumatic brain injury

In the present study, human umbilical cord blood mesenchymal stem cells were injected into a rat model of traumatic brain injury via the tail vein. Results showed that 5-bromodeoxyuridine-labeled cells aggregated around the injury site, surviving up to 4 weeks post-transplantation. In addition, transplantation-related death did not occur, and neurological functions significantly improved. Histological detection revealed attenuated pathological injury in rat brain tissues following human umbilical cord blood mesenchymal stem cell transplantation. In addition, the number of apoptotic cells decreased. Immunohistochemistry and in situ hybridization showed increased expression of brain-derived neurotrophic factor, nerve growth factor, basic fibroblast growth factor, and vascular endothelial growth factor, along with increased microvessel density in surrounding areas of brain injury. Results demonstrated migration of transplanted human umbilical cord blood mesenchymal stem cells into the lesioned boundary zone of rats, as well as increased angiogenesis and expression of related neurotrophic factors in the lesioned boundary zone.

High tibial osteotomy with human umbilical cord blood-derived mesenchymal stem cells implantation for knee cartilage regeneration

BACKGROUND High tibial osteotomy(HTO)is a well-established method for the treatment of medial compartment osteoarthritis of the knee with varus deformity.However,HTO alone cannot adequately repair the arthritic joint,necessitating cartilage regeneration therapy.Cartilage regeneration procedures with concomitant HTO are used to improve the clinical outcome in patients with varus deformity.AIM To evaluate cartilage regeneration after implantation of allogenic human umbilical cord blood-derived mesenchymal stem cells(hUCB-MSCs)with concomitant HTO.METHODS Data for patients who underwent implantation of hUCB-MSCs with concomitant HTO were evaluated.The patients included in this study were over 40 years old,had a varus deformity of more than 5°,and a full-thickness International Cartilage Repair Society(ICRS)grade IV articular cartilage lesion of more than 4 cm2 in the medial compartment of the knee.All patients underwent second-look arthroscopy during hardware removal.Cartilage regeneration was evaluated macroscopically using the ICRS grading system in second-look arthroscopy.We also assessed the effects of patient characteristics,such as trochlear lesions,age,and lesion size,using patient medical records.RESULTS A total of 125 patients were included in the study,with an average age of 58.3±6.8 years(range:43-74 years old);95(76%)were female and 30(24%)were male.The average hip-knee-ankle(HKA)angle for measuring varus deformity was 7.6°±2.4°(range:5.0-14.2°).In second-look arthroscopy,the status of medial femoral condyle(MFC)cartilage was as follows:73(58.4%)patients with ICRS grade I,37(29.6%)with ICRS grade II,and 15(12%)with ICRS grade III.No patients were staged with ICRS grade IV.Additionally,the scores[except International Knee Documentation Committee(IKDC)at 1 year]of the ICRS grade I group improved more significantly than those of the ICRS grade II and III groups.CONCLUSION Implantation of hUCB-MSCs with concomitant HTO is an effective treatment for patients with medial compartment osteoarthritis and varus deformity.Regeneration of cartilage improves the clinical outcomes for the patients.

Protective Effect of Human Umbilical Cord Mesenchymal Stem Cells in Glucocorticoid-induced Osteonecrosis of Femoral Head

Objective:To evaluate the effect of human umbilical cord mesenchymal stem cells(hUC-MSCs)on preventing rats from glucocorticoid-induced osteonecrosis of femoral head(GC-ONFH)in the early stage in vivo and to investigate the possible mechanism of hUC-MSCs in regulating the balance of osteogenesis and adipogenesis.Methods:All rats were randomly divided into 3 groups:control group(C group),model group(M group),and intervention group(Ⅰ group).The model of GC-ONFH was developed by a sequential administration of lipopolysaccharide and methylprednisolone.The rats in the Ⅰ group were treated with caudal vein injection of hUC-MSCs.Six weeks later,the blood samples were obtained to measure the activity of alkaline phosphatase(ALP)and the content of triglyceride(TG)in serum,and the femoral heads were harvested and observed by hematoxylin-eosin staining,Micro-CT,Western blot and real-time quantitative polymerase chain reaction.Results:After intervention of hUC-MSCs,the necrosis rate of femoral head decreased from 83%(10/12)to 33%(4/12),the rate of empty bone lacuna was significantly decreased,the activity of ALP increased significantly,the content of TG decreased significantly,the bone density increased obviously,the expression of RUNX2 and Col Ⅰ increased significantly and the expression of PPARγ decreased significantly.Conclusion:These results revealed that caudal vein injection of hUC-MSCs can effectively reduce the incidence of GC-ONFH in rats by increasing ALP activity and reducing TG content in serum,increasing bone mineral density,promoting the expression of RUNX2 and Col I,and inhibiting the expression of PPARγ.

Genetically modified human umbilical cord blood cells as a promising strategy for treatment of spinal cord injury

Spinal cord injury （SCI） continues to be a pressing health and social problem. The injury leads to neuronal and glial cell death accompanied by degeneration of nerve fibers. There are currently no particularly effective treatments. SCI causes profound disabil- ity of people affected and has attracted increased attention in the international field of neuroregeneration. For the past two decades, much hope has been placed in cell therapies for the restoration of both structure and function of the injured spinal cord. Embryonic and neural stem cells, olfactory ensheathing cells, microglia-like cells, Schwann cells, mesenchymal stem cells.

Functional recovery and microenvironmental alterations in a rat model of spinal cord injury following human umbilical cord blood-derived mesenchymal stem cells transplantation

BACKGROUND： Transplantation of human umbilical cord blood-derived mesenchymal stem cells （MSCs） has been shown to benefit spinal cord injury （SCI） repair. However, mechanisms of microenvironmental regulation during differentiation of transplanted MSCs remain poorly understood. OBJECTIVE： To observe changes in nerve growth factor （NGF）, brain-derived neurotrophic factor （BDNF）, and interleukin-8 （IL-8） expression following transplantation of human umbilical cord-derived MSCs, and to explore the association between microenvironment and neural functional recovery following MSCs transplantation. DESIGN, TIME AND SETTING： A randomized, controlled, animal experiment was performed at the Department of Orthopedics, First Affiliated Hospital of Soochow University from April 2005 to March 2007. MATERIALS： Human cord blood samples were provided by the Department of Gynecology and Obstetrics, First Affiliated Hospital of Soochow University. Written informed consent was obtained. METHODS： A total of 62 Wister rats were randomly assigned to control （n = 18）, model （n = 22, SCI ＋ PBS）, and transplantation （n = 22, SCI ＋ MSCs） groups. The rat SCI model was established using the weight compression method. MSCs were isolated from human umbilical cord blood and cultured in vitro for several passages. 5-bromodeoxyuridine （BrdU）-Iabeled MSCs （24 hours before injection） were intravascularly transplanted. MAIN OUTCOME MEASURES： The rats were evaluated using the Basso, Beattie and Bresnahan （BBB） locomotor score and inclined plane tests. Transplanted cells were analyzed following immunohistochemistry. Enzyme-linked immunosorbant assay was performed to determine NGF, BDNF, and IL-8 levels prior to and after cell transplantation. RESULTS： A large number of BrdU-positive MSCs were observed in the SCI region of the transplantation group, and MSCs were evenly distributed in injured spinal cord tissue 1 week after transplantation. BBB score and inclined plane test results revealed significant functional improvement in the transplantation group compared to the model group （P 〈 0.05）, which was maintained for 2-3 weeks. Compared to the model group, NGF and BDNF levels were significantly increased in the injured region following MSCs transplantation at 3 weeks （P 〈 0.05）, but IL-8 levels remained unchanged （P 〉 0.05）. CONCLUSION： MSCs transplantation increased NGF and BDNF expression in injured spinal cord tissue. MSCs could promote neurological function recovery in SCI rats by upregulating NGF expression and improving regional microenvironments.

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.

Human umbilical cord blood stem cells and brainderived neurotrophic factor for optic nerve injury: a biomechanical evaluation

Treatment for optic nerve injury by brain-derived neurotrophic factor or the transplantation of human umbilical cord blood stem cells has gained progress, but analysis by biomechanical indicators is rare. Rabbit models of optic nerve injury were established by a clamp. At 7 days after injury, the vitreous body received a one-time injection of 50 μg brain-derived neurotrophic factor or 1 × 10^6 human umbilical cord blood stem cells. After 30 days, the maximum load, maximum stress, maximum strain, elastic limit load, elastic limit stress, and elastic limit strain had clearly improved in rabbit models of optical nerve injury after treatment with brain-derived neurotrophic factor or human umbilical cord blood stem cells. The damage to the ultrastructure of the optic nerve had also been reduced. These findings suggest that human umbilical cord blood stem cells and brain-derived neurotrophic factor effectively repair the injured optical nerve, improve biomechanical properties, and contribute to the recovery after injury.

Mesenchymal stem cells from the human umbilical cord ameliorate fulminant hepatic failure and increase survival in mice

BACKGROUND：Cell therapy has been promising for various diseases.We investigated whether transplantation of human umbilical cord mesenchymal stem cells（h UCMSCs）has any therapeutic effects on D-galactosamine/lipopolysaccharide（Gal N/LPS）-induced fulminant hepatic failure in mice.METHODS：h UCMSCs isolated from human umbilical cord were cultured and transplanted via the tail vein into severe combined immune deficiency mice with Gal N/LPS-induced fulminant hepatic failure.After transplantation,the localization and differentiation of h UCMSCs in the injured livers were investigated by immunohistochemical and genetic analy- ses. The recovery of the injured livers was evaluated histologi- cally. The survival rate of experimental animals was analyzed by the Kaplan-Meier method and log-rank test. RESULTS： hUCMSCs expressed high levels of CD29, CD73, CD13, CD105 and CD90, but did not express CD31, CD79b, CD133, CD34, and CD45. Cultured hUCMSCs displayed adip- ogenic and osteogenic differentiation potential. Hematoxylin and eosin staining revealed that transplantation of hUCMSCs reduced hepatic necrosis and promoted liver regeneration. Transplantation of hUCMSCs prolonged the survival rate of mice with fulminant hepatic failure. Polymerase chain reaction for human alu sequences showed the presence of human cells in mouse livers. Positive staining for human albumin, human alpha-fetoprotein and human cytokeratin 18 suggested the for- mation of hUCMSCs-derived hepatocyte-like cells in vivo.CONCLUSIONS： hUCMSC was a potential candidate for stem cell based therapies. After transplantation, hUCMSCs partially repaired hepatic damage induced by GalN/LPS in mice. hUC- MSCs engrafted into the injured liver and differentiated into hepatocyte-like cells.

Human umbilical cord blood-derived stem cells and brain-derived neurotrophic factor protect injured optic nerve:viscoelasticity characterization

The optic nerve is a viscoelastic solid-like biomaterial.Its normal stress relaxation and creep properties enable the nerve to resist constant strain and protect it from injury.We hypothesized that stress relaxation and creep properties of the optic nerve change after injury.Moreover,human brain-derived neurotrophic factor or umbilical cord blood-derived stem cells may restore these changes to normal.To validate this hypothesis,a rabbit model of optic nerve injury was established using a clamp approach.At 7 days after injury,the vitreous body received a one-time injection of 50 μg human brain-derived neurotrophic factor or 1 × 106 human umbilical cord blood-derived stem cells.At 30 days after injury,stress relaxation and creep properties of the optic nerve that received treatment had recovered greatly,with pathological changes in the injured optic nerve also noticeably improved.These results suggest that human brain-derived neurotrophic factor or umbilical cord blood-derived stem cell intervention promotes viscoelasticity recovery of injured optic nerves,and thereby contributes to nerve recovery.

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.