Extracellular vesicles from human urine-derived stem cells delay aging through the transfer of PLAU and TIMP1

Aging increases the risks of various diseases and the vulnerability to death.Cellular senescence is a hallmark of aging that contributes greatly to aging and aging-related diseases.This study demonstrates that extracellular vesicles from human urine-derived stem cells(USC-EVs)efficiently inhibit cellular senescence in vitro and in vivo.The intravenous injection of USC-EVs improves cognitive function,increases physical fitness and bone quality,and alleviates aging-related structural changes in different organs of senescence-accelerated mice and natural aging mice.The anti-aging effects of USC-EVs are not obviously affected by the USC donors’ages,genders,or health status.Proteomic analysis reveals that USC-EVs are enriched with plasminogen activator urokinase(PLAU)and tissue inhibitor of metalloproteinases 1(TIMP1).These two proteins contribute importantly to the anti-senescent effects of USC-EVs associated with the inhibition of matrix metalloproteinases,cyclin-dependent kinase inhibitor 2A(P16INK4a),and cyclin-dependent kinase inhibitor 1A(P21cip1).These findings suggest a great potential of autologous USC-EVs as a promising anti-aging agent by transferring PLAU and TIMP1 proteins.

Therapeutic potential of urine-derived stem cells in renal regeneration following acute kidney injury:A comparative analysis with mesenchymal stem cells

BACKGROUND Acute kidney injury(AKI)is a common clinical syndrome with high morbidity and mortality rates.The use of pluripotent stem cells holds great promise for the treatment of AKI.Urine-derived stem cells(USCs)are a novel and versatile cell source in cell-based therapy and regenerative medicine that provide advantages of a noninvasive,simple,and low-cost approach and are induced with high multidifferentiation potential.Whether these cells could serve as a potential stem cell source for the treatment of AKI has not been determined.METHODS Stem cell markers with multidifferentiation potential were isolated from human amniotic fluid.AKI severe combined immune deficiency(SCID)mice models were induced by means of an intramuscular injection with glycerol.USCs isolated from human-voided urine were administered via tail veins.The functional changes in the kidney were assessed by the levels of blood urea nitrogen and serum creatinine.The histologic changes were evaluated by hematoxylin and eosin staining and transferase dUTP nick-end labeling staining.Meanwhile,we compared the regenerative potential of USCs with bone marrow-derived mesenchymal stem cells(MSCs).RESULTS Treatment with USCs significantly alleviated histological destruction and functional decline.The renal function was rapidly restored after intravenous injection of 5×105 human USCs into SCID mice with glycerol-induced AKI compared with injection of saline.Results from secretion assays conducted in vitro demonstrated that both stem cell varieties released a wide array of cytokines and growth factors.This suggests that a mixture of various mediators closely interacts with their biochemical functions.Two types of stem cells showed enhanced tubular cell prolif-eration and decreased tubular cell apoptosis,although USC treatment was not more effective than MSC treatment.We found that USC therapy significantly improved renal function and histological damage,inhibited inflammation and apoptosis processes in the kidney,and promoted tubular epithelial proliferation.CONCLUSION Our study demonstrated the potential of USCs for the treatment of AKI,representing a new clinical therapeutic strategy.

Application prospects of urine-derived stem cells in neurological and musculoskeletal diseases

Urine-derived stem cells(USCs)are derived from urine and harbor the potential of proliferation and multidirectional differentiation.Moreover,USCs could be reprogrammed into pluripotent stem cells[namely urine-derived induced pluripotent stem cells(UiPSCs)]through transcription factors,such as octamer binding transcription factor 4,sex determining region Y-box 2,kruppel-like factor 4,myelocytomatosis oncogene,and Nanog homeobox and protein lin-28,in which the first four are known as Yamanaka factors.Mounting evidence supports that USCs and UiPSCs possess high potential of neurogenic,myogenic,and osteogenic differentiation,indicating that they may play a crucial role in the treatment of neurological and musculoskeletal diseases.Therefore,we summarized the origin and physiological characteristics of USCs and UiPSCs and their therapeutic application in neurological and musculoskeletal disorders in this review,which not only contributes to deepen our understanding of hallmarks of USCs and UiPSCs but also provides the theoretical basis for the treatment of neurological and musculoskeletal disorders with USCs and UiPSCs.

Therapeutic utility of human umbilical cord-derived mesenchymal stem cells-based approaches in pulmonary diseases:Recent advancements and prospects

Pulmonary diseases across all ages threaten millions of people and have emerged as one of the major public health issues worldwide.For diverse disease con-ditions,the currently available approaches are focused on alleviating clinical symptoms and delaying disease progression but have not shown significant therapeutic effects in patients with lung diseases.Human umbilical cord-derived mesenchymal stem cells(UC-MSCs)isolated from the human UC have the capacity for self-renewal and multilineage differentiation.Moreover,in recent years,these cells have been demonstrated to have unique advantages in the treatment of lung diseases.We searched the Public Clinical Trial Database and found 55 clinical trials involving UC-MSC therapy for pulmonary diseases,including coronavirus disease 2019,acute respiratory distress syndrome,bron-chopulmonary dysplasia,chronic obstructive pulmonary disease,and pulmonary fibrosis.In this review,we summarize the characteristics of these registered clinical trials and relevant published results and explore in depth the challenges and opportunitiesfaced in clinical application.Moreover,the underlying mole-cular mechanisms involved in UC-MSC-based therapy for pulmonary diseases are also analyzed in depth.In brief,this comprehensive review and detailed analysis of these clinical trials can be expected to provide a scientific reference for future large-scale clinical application.

Transplantation of human placental chorionic plate-derived mesenchymal stem cells for repair of neurological damage in neonatal hypoxic-ischemic encephalopathy

Neonatal hypoxic-ischemic encephalopathy is often associated with permanent cerebral palsy,neurosensory impairments,and cognitive deficits,and there is no effective treatment for complications related to hypoxic-ischemic encephalopathy.The therapeutic potential of human placental chorionic plate-derived mesenchymal stem cells for various diseases has been explored.However,the potential use of human placental chorionic plate-derived mesenchymal stem cells for the treatment of neonatal hypoxic-ischemic encephalopathy has not yet been investigated.In this study,we injected human placental chorionic plate-derived mesenchymal stem cells into the lateral ventricle of a neonatal hypoxic-ischemic encephalopathy rat model and observed significant improvements in both cognitive and motor function.Protein chip analysis showed that interleukin-3 expression was significantly elevated in neonatal hypoxic-ischemic encephalopathy model rats.Following transplantation of human placental chorionic plate-derived mesenchymal stem cells,interleukin-3 expression was downregulated.To further investigate the role of interleukin-3 in neonatal hypoxic-ischemic encephalopathy,we established an in vitro SH-SY5Y cell model of hypoxic-ischemic injury through oxygen-glucose deprivation and silenced interleukin-3 expression using small interfering RNA.We found that the activity and proliferation of SH-SY5Y cells subjected to oxygen-glucose deprivation were further suppressed by interleukin-3 knockdown.Furthermore,interleukin-3 knockout exacerbated neuronal damage and cognitive and motor function impairment in rat models of hypoxic-ischemic encephalopathy.The findings suggest that transplantation of hpcMSCs ameliorated behavioral impairments in a rat model of hypoxic-ischemic encephalopathy,and this effect was mediated by interleukin-3-dependent neurological function.

Expansion of human umbilical cord derived mesenchymal stem cells in regenerative medicine

BACKGROUND Stem cells are undifferentiated cells that possess the potential for self-renewal with the capacity to differentiate into multiple lineages.In humans,their limited numbers pose a challenge in fulfilling the necessary demands for the regeneration and repair of damaged tissues or organs.Studies suggested that mesenchymal stem cells(MSCs),necessary for repair and regeneration via transplantation,require doses ranging from 10 to 400 million cells.Furthermore,the limited expansion of MSCs restricts their therapeutic application.AIM To optimize a novel protocol to achieve qualitative and quantitative expansion of MSCs to reach the targeted number of cells for cellular transplantation and minimize the limitations in stem cell therapy protocols.METHODS Human umbilical cord(hUC)tissue derived MSCs were obtained and re-cultured.These cultured cells were subjected to the following evaluation pro-cedures:Immunophenotyping,immunocytochemical staining,trilineage differentiation,population doubling time and number,gene expression markers for proliferation,cell cycle progression,senescence-associatedβ-galactosidase assay,human telomerase reverse transcriptase(hTERT)expression,mycoplasma,cytomegalovirus and endotoxin detection.RESULTS Analysis of pluripotent gene markers Oct4,Sox2,and Nanog in recultured hUC-MSC revealed no significant differences.The immunophenotypic markers CD90,CD73,CD105,CD44,vimentin,CD29,Stro-1,and Lin28 were positively expressed by these recultured expanded MSCs,and were found negative for CD34,CD11b,CD19,CD45,and HLA-DR.The recultured hUC-MSC population continued to expand through passage 15.Proliferative gene expression of Pax6,BMP2,and TGFb1 showed no significant variation between recultured hUC-MSC groups.Nevertheless,a significant increase(P<0.001)in the mitotic phase of the cell cycle was observed in recultured hUC-MSCs.Cellular senescence markers(hTERT expression andβ-galactosidase activity)did not show any negative effect on recultured hUC-MSCs.Additionally,quality control assessments consistently confirmed the absence of mycoplasma,cytomegalovirus,and endotoxin contamination.CONCLUSION This study proposes the development of a novel protocol for efficiently expanding stem cell population.This would address the growing demand for larger stem cell doses needed for cellular transplantation and will significantly improve the feasibility of stem cell based therapies.

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.

Self-assembly of differentiated dental pulp stem cells facilitates spheroid human dental organoid formation and prevascularization

BACKGROUND The self-assembly of solid organs from stem cells has the potential to greatly expand the applicability of regenerative medicine.Stem cells can self-organise into microsized organ units,partially modelling tissue function and regeneration.Dental pulp organoids have been used to recapitulate the processes of tooth development and related diseases.However,the lack of vasculature limits the utility of dental pulp organoids.AIM To improve survival and aid in recovery after stem cell transplantation,we demonstrated the three-dimensional(3D)self-assembly of adult stem cell-human dental pulp stem cells(hDPSCs)and endothelial cells(ECs)into a novel type of spheroid-shaped dental pulp organoid in vitro under hypoxia and conditioned medium(CM).METHODS During culture,primary hDPSCs were induced to differentiate into ECs by exposing them to a hypoxic environment and CM.The hypoxic pretreated hDPSCs were then mixed with ECs at specific ratios and conditioned in a 3D environment to produce prevascularized dental pulp organoids.The biological characteristics of the organoids were analysed,and the regulatory pathways associated with angiogenesis were studied.RESULTS The combination of these two agents resulted in prevascularized human dental pulp organoids(Vorganoids)that more closely resembled dental pulp tissue in terms of morphology and function.Single-cell RNA sequencing of dental pulp tissue and RNA sequencing of Vorganoids were integrated to analyse key regulatory pathways associated with angiogenesis.The biomarkers forkhead box protein O1 and fibroblast growth factor 2 were identified to be involved in the regulation of Vorganoids.CONCLUSION In this innovative study,we effectively established an in vitro model of Vorganoids and used it to elucidate new mechanisms of angiogenesis during regeneration,facilitating the development of clinical treatment strategies.

Human dental pulp stem/stromal cells in clinical practice

Dental pulp stem/stromal cells(DPSCs)are fibroblast-like,neural crest-derived,and multipotent cells that can differentiate into several lineages.They are relatively easy to isolate from healthy and inflamed pulps,with little ethical concerns and can be successfully cryopreserved and thawed.The therapeutic effects of DPSCs derived from animal or human sources have been extensively studied through in-vitro and in-vivo animal experiments and the findings indicated that DPSCs are effective not only for dental diseases but also for systemic diseases.Understanding that translational research is a critical step through which the fundamental scientific discoveries could be translated into applicable diagnostics and therapeutics that directly benefit humans,several clinical studies were carried out to generate evidence for the efficacy and safety of autogenous or allogeneic human DPSCs(hDPSCs)as a treatment modality for use in cell-based therapy,regenerative medicine/dentistry and tissue engineering.In clinical medicine,hDPSCs were effective for treating acute ischemic stroke and human exfoliated deciduous teeth-conditioned medium(SHED-CM)repaired vascular damage of the corpus cavernous,which is the main cause of erectile dysfunction.Whereas in clinical dentistry,autologous SHED was able to rege-nerate necrotic dental pulp after implantation into injured teeth,and micrografts enriched with autologous hDPSCs and collagen sponge were considered a treatment option for human intrabony defects.In contrast,hDPSCs did not add a significant regenerative effect when they were used for the treatment of post-extraction sockets.Large-scale clinical studies across diverse populations are still lacking to provide robust evidence on the safety and efficacy of hDPSCs as a new treatment option for various human diseases including dental-related problems.

One-step cell biomanufacturing platform:porous gelatin microcarrier beads promote human embryonic stem cell-derived midbrain dopaminergic progenitor cell differentiation in vitro and survival after transplantation in vivo

Numerous studies have shown that cell replacement therapy can replenish lost cells and rebuild neural circuitry in animal models of Parkinson’s disease.Transplantation of midbrain dopaminergic progenitor cells is a promising treatment for Parkinson’s disease.However,transplanted cells can be injured by mechanical damage during handling and by changes in the transplantation niche.Here,we developed a one-step biomanufacturing platform that uses small-aperture gelatin microcarriers to produce beads carrying midbrain dopaminergic progenitor cells.These beads allow midbrain dopaminergic progenitor cell differentiation and cryopreservation without digestion,effectively maintaining axonal integrity in vitro.Importantly,midbrain dopaminergic progenitor cell bead grafts showed increased survival and only mild immunoreactivity in vivo compared with suspended midbrain dopaminergic progenitor cell grafts.Overall,our findings show that these midbrain dopaminergic progenitor cell beads enhance the effectiveness of neuronal cell transplantation.

Human pluripotent stem cell-derived kidney organoids:Current progress and challenges

Human pluripotent stem cell(hPSC)-derived kidney organoids share similarities with the fetal kidney.However,the current hPSC-derived kidney organoids have some limitations,including the inability to perform nephrogenesis and lack of a corticomedullary definition,uniform vascular system,and coordinated exit path-way for urinary filtrate.Therefore,further studies are required to produce hPSC-derived kidney organoids that accurately mimic human kidneys to facilitate research on kidney development,regeneration,disease modeling,and drug screening.In this review,we discussed recent advances in the generation of hPSC-derived kidney organoids,how these organoids contribute to the understanding of human kidney development and research in disease modeling.Additionally,the limitations,future research focus,and applications of hPSC-derived kidney organoids were highlighted.

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.

Treatment of spinal cord injury with biomaterials and stem cell therapy in non-human primates and humans

Spinal cord injury results in the loss of sensory,motor,and autonomic functions,which almost always produces permanent physical disability.Thus,in the search for more effective treatments than those already applied for years,which are not entirely efficient,researches have been able to demonstrate the potential of biological strategies using biomaterials to tissue manufacturing through bioengineering and stem cell therapy as a neuroregenerative approach,seeking to promote neuronal recovery after spinal cord injury.Each of these strategies has been developed and meticulously evaluated in several animal models with the aim of analyzing the potential of interventions for neuronal repair and,consequently,boosting functional recovery.Although the majority of experimental research has been conducted in rodents,there is increasing recognition of the importance,and need,of evaluating the safety and efficacy of these interventions in non-human primates before moving to clinical trials involving therapies potentially promising in humans.This article is a literature review from databases(PubMed,Science Direct,Elsevier,Scielo,Redalyc,Cochrane,and NCBI)from 10 years ago to date,using keywords(spinal cord injury,cell therapy,non-human primates,humans,and bioengineering in spinal cord injury).From 110 retrieved articles,after two selection rounds based on inclusion and exclusion criteria,21 articles were analyzed.Thus,this review arises from the need to recognize the experimental therapeutic advances applied in non-human primates and even humans,aimed at deepening these strategies and identifying the advantages and influence of the results on extrapolation for clinical applicability in humans.

Exploring hematopoietic stem cell population in human milk and its benefits for infants:A scoping review

Objective:To comprehensively explore hematopoietic stem cells(HSCs)in human milk,understanding their molecular markers,isolation methods,benefits for infants,and potential medical applications.Methods:We conducted a scoping literature review following the PRISMA-ScR guidelines.This review included studies investigating HSCs in human milk,utilizing molecular markers such as CD34^(+),CD113^(+),and CD117^(+)for characterization.Both in vitro and in vivo studies exploring the morphology,function,and clinical implications of these cells were considered.The diverse range of papers reviewed were indexed in PubMed,Science Direct,Scopus,Sage Journals,and Google Scholar,published between 2010 and 2023.Results:This scoping review explored 577 articles and selected 13 studies based on our inclusion criteria,focusing on HSCs in human milk.Most studies dilute samples prior to HSC isolation,followed by detection using markers such as CD34^(+),CD113^(+),and CD117^(+),with flow cytometry serving as the primary analysis tool,focusing on their isolation and detection methods.While no definitive benefits have been conclusively established,there is a strong belief in the potential of HSCs to positively impact infant immunity,growth,and tissue repair.Conclusions:This review presents significant evidence supporting the presence of HSCs in human milk,identified by markers such as CD34^(+),CD113^(+),and CD117^(+).These cells show considerable potential in enhancing infant health,including immunity,tissue repair,cognitive development,and gastrointestinal health.Despite methodological variations in isolation and detection techniques,the collective findings underscore the potential clinical relevance of HSCs in human milk.Moreover,this review highlights the noninvasive accessibility of human milk as a source of HSCs and emphasizes the need for further research to unlock their therapeutic potential.

Patient-derived induced pluripotent stem cells with a MERTK mutation exhibit cell junction abnormalities and aberrant cellular differentiation potential

BACKGROUND Human induced pluripotent stem cell(hiPSC)technology is a valuable tool for generating patient-specific stem cells,facilitating disease modeling,and invest-igating disease mechanisms.However,iPSCs carrying specific mutations may limit their clinical applications due to certain inherent characteristics.AIM To investigate the impact of MERTK mutations on hiPSCs and determine whether hiPSC-derived extracellular vesicles(EVs)influence anomalous cell junction and differentiation potential.METHODS We employed a non-integrating reprogramming technique to generate peripheral blood-derived hiPSCs with and hiPSCs without a MERTK mutation.Chromo-somal karyotype analysis,flow cytometry,and immunofluorescent staining were utilized for hiPSC identification.Transcriptomics and proteomics were employed to elucidate the expression patterns associated with cell junction abnormalities and cellular differentiation potential.Additionally,EVs were isolated from the supernatant,and their RNA and protein cargos were examined to investigate the involvement of hiPSC-derived EVs in stem cell junction and differentiation.RESULTS The generated hiPSCs,both with and without a MERTK mutation,exhibited normal karyotype and expressed pluripotency markers;however,hiPSCs with a MERTK mutation demonstrated anomalous adhesion capability and differentiation potential,as confirmed by transcriptomic and proteomic profiling.Furthermore,hiPSC-derived EVs were involved in various biological processes,including cell junction and differentiation.CONCLUSION HiPSCs with a MERTK mutation displayed altered junction characteristics and aberrant differentiation potential.Furthermore,hiPSC-derived EVs played a regulatory role in various biological processes,including cell junction and differentiation.

Enhancing Erectile Function and Alleviating Andropause Symptoms: Clinical Efficacy of a Human Stem Cell Conditioned Medium Cream

Erectile dysfunction (ED) is increasingly prevalent in Japan, exceeding 30%, and increasing with age. Unhealthy lifestyle habits, obesity, insufficient exercise, and smoking have been implicated in its pathogenesis, along with endothelial dysfunction of the corpora cavernosa and impaired blood flow to the penis considered underlying factors. However, the current treatments are limited to Phosphodiesterase-5 (PDE5) inhibitors. ED is the primary symptom of andropathy. This study reports the clinical efficacy of human stem cell-conditioned medium cream for ED treatment. Ten men without underlying diseases suspected of andropause with ED (mean age 43.2 ± 4.4 y, Hb 15.2 ± 0.6 gm/dL, AST/ALT 30.2/37.9 ± 12.4/14.0, eGFR 82.7 ± 12.4 mL/min/1.73 m2) were targeted. The cream was applied twice daily to the genital and scrotal areas. The erectile hardness score (EHS), International Index of Erectile Function-5 (IIEF-5), and Aging Male Symptoms (AMS) scale were used to evaluate the participants before and 30 days after use, and the results were compared using paired t-tests. The post-use qualitative opinions were collected through interviews. Significant improvements were observed compared to baseline in the IIEF-5 (11.8 ± 4.6→17.2 ± 5.1, P < 0.001), and AMS (46.3 ± 6.7→37.6 ± 5.3, P < 0.001) scores post cream use. EHS did not show a statistically significant difference, but a trend towards improvement was observed. Qualitative feedback included increased morning erection, improved maintenance of erection during intercourse, and reduced post work fatigue. Human stem cell-conditioned medium contains endothelial growth factors that potentially contribute to the improvement of ED and andropause by enhancing corporal endothelial function. Future studies should include control groups to further investigate the efficacy of these treatments.

Safety evaluation of human umbilical cord-mesenchymal stem cells in type 2 diabetes mellitus treatment:A phase 2 clinical trial

BACKGROUND Progressive pancreaticβcell dysfunction is a fundamental aspect of the pathology underlying type 2 diabetes mellitus(T2DM).Recently,mesenchymal stem cell(MSC)transplantation has emerged as a new therapeutic method due to its ability to promote the regeneration of pancreaticβcells.However,current studies have focused on its efficacy,and there are few clinical studies on its safety.AIM To evaluate the safety of human umbilical cord(hUC)-MSC infusion in T2DM treatment.METHODS An open-label and randomized phase 2 clinical trial was designed to evaluate the safety of hUC-MSC transplantation in T2DM in a Class A hospital.Ten patients in the placebo group received acellular saline intravenously once per week for 3 wk.Twenty-four patients in the hUC-MSC group received hUC-MSCs(1×106 cells/kg)intravenously once per week for 3 wk.Diabetic clinical symptoms and signs,laboratory findings,and imaging findings were evaluated weekly for the 1st mo and then at weeks 12 and 24 post-treatment.RESULTS No serious adverse events were observed during the 24-wk follow-up.Four patients(16.7%)in the hUC-MSC group experienced transient fever,which occurred within 24 h after the second or third infusion;this did not occur in any patients in the placebo group.One patient from the hUC-MSC group experienced hypoglycemic attacks within 1 mo after transplantation.Significantly lower lymphocyte levels(weeks 2 and 3)and thrombin coagulation time(week 2)were observed in the hUC-MSC group compared to those in the placebo group(all P<0.05).Significantly higher platelet levels(week 3),immunoglobulin levels(weeks 1,2,3,and 4),fibrinogen levels(weeks 2 and 3),D-dimer levels(weeks 1,2,3,4,12,and 24),and neutrophil-to-lymphocyte ratios(weeks 2 and 3)were observed in the hUC-MSC group compared to those in the placebo group(all P<0.05).There were no significant differences between the two groups for tumor markers(alpha-fetoprotein,carcinoembryonic antigen,and carbohydrate antigen 199)or blood fat.No liver damage or other side effects were observed on chest X-ray.CONCLUSION Our study suggested that hUC-MSC transplantation has good tolerance and high safety in the treatment of T2DM.It can improve human immunity and inhibit lymphocytes.Coagulation function should be monitored vigilantly for abnormalities.

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.

Zinc enhances the cell adhesion,migration,and self-renewal potential of human umbilical cord derived mesenchymal stem cells

BACKGROUND Zinc(Zn)is the second most abundant trace element after Fe,present in the human body.It is frequently reported in association with cell growth and proliferation,and its deficiency is considered to be a major disease contributing factor.AIM To determine the effect of Zn on in vitro growth and proliferation of human umbilical cord(hUC)-derived mesenchymal stem cells(MSCs).METHODS hUC-MSCs were isolated from human umbilical cord tissue and characterized based on immunocytochemistry,immunophenotyping,and tri-lineage differentiation.The impact of Zn on cytotoxicity and proliferation was determined by MTT and Alamar blue assay.To determine the effect of Zn on population doubling time(PDT),hUC-MSCs were cultured in media with and without Zn for several passages.An in vitro scratch assay was performed to analyze the effect of Zn on the wound healing and migration capability of hUC-MSCs.A cell adhesion assay was used to test the surface adhesiveness of hUC-MSCs.Transcriptional analysis of genes involved in the cell cycle,proliferation,migration,and selfrenewal of hUC-MSCs was performed by quantitative real-time polymerase chain reaction.The protein expression of Lin28,a pluripotency marker,was analyzed by immunocytochemistry.RESULTS Zn at lower concentrations enhanced the rate of proliferation but at higher concentrations(>100μM),showed concentration dependent cytotoxicity in hUC-MSCs.hUC-MSCs treated with Zn exhibited a significantly greater healing and migration rate compared to untreated cells.Zn also increased the cell adhesion rate,and colony forming efficiency(CFE).In addition,Zn upregulated the expression of genes involved in the cell cycle(CDC20,CDK1,CCNA2,CDCA2),proliferation(transforming growth factorβ1,GDF5,hypoxia-inducible factor 1α),migration(CXCR4,VCAM1,VEGF-A),and self-renewal(OCT4,SOX2,NANOG)of hUC-MSCs.Expression of Lin28 protein was significantly increased in cells treated with Zn.CONCLUSION Our findings suggest that zinc enhances the proliferation rate of hUC-MSCs decreasing the PDT,and maintaining the CFE.Zn also enhances the cell adhesion,migration,and self-renewal of hUC-MSCs.These results highlight the essential role of Zn in cell growth and development.

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.