Keratinocyte growth factor-2 and autologous serum potentiate the regenerative effect of mesenchymal stem cells in cornea damage in rats

AIM:To investigate the healing process after severe corneal epithelial damage in rats treated with mesenchymal stem cells(MSCs)cultured with or without keratinocyte growth factor(KGF-2)and autologous serum(AS)on amniotic membrane(AM).Many patients are blind and devastated by severe ocular surface diseases due to limbal stem cell deficiency.Bone marrow-derived MSCs are potential sources for cellbased tissue engineering to repair or replace the corneal tissue,having the potential to differentiate to epithelial cells.METHODS:The study included 5 groups each including 10 female'Sprague Dawley'rats in addition to20 male rats used as bone marrow donors.Group I rats received AM+MSCs,Group II rats AM+MSCs cultured with KGF-2,Group III rats AM+MSCs cultured with KGF-2+AS,Group IV rats only AM and Group V rats,none.AS was derived from blood drawn from male rats and bone marrow was obtained from the femur and tibia bones of the same animals.Therapeutic effect was evaluated with clinical,histopathological and immunohistochemical assessment.MSC engraftment was demonstrated via detection of donor genotype(Y+)in the recipient tissue(X)with polymerase chain reaction.RESULTS:Corneal healing was significantly better in Groups I-III rats treated with MSC transplantation compared to Group IV and Group V rats with supportive treatment only.The best results were obtained in Group III rats with 90%transparency,70%lack of neovascularization,and 100%epithelium damage limited to less than 1/4 of cornea.CONCLUSION:We suggest that culture of MSCs with KGF-2 and AS on AM is effective in corneal repair in case of irreversible damage to limbal stem cells.

Fetal bovine serum versus Chinese herbal formula Naoluoxintong serum supplementation for proliferation and differentiation of rat embryonic neural stem cells

BACKGROUND： How to induce endogenous neural stem cells （NSCs） to differentiate into needed neural cell types is a hot spot of current researches. OBJECTIVE： To compare differences between fetal bovine serum and Chinese herbal formula Naoluoxintong serum supplementation for inducing proliferation and differentiation in rat embryonic NSCs. DESIGN, TIME AND SETTING： An in vitro, serum pharmacology, comparative, observation study was performed from March to September in 2008 at the Laboratory of Neurodegenerative Diseases, College of Life Science in University of Science and Technology of China, the Key Laboratory Breeding Base of Acupuncture Foundation and Technology in Anhui University of Traditional Chinese Medicine, the Anhui Province Key Laboratory of R ＆ D of Chinese Medicine, and at the Level 3 Laboratory of Molecular Biology of the State Administration of Traditional Chinese Medicine. MATERIALS： The Chinese herbal formula Naoluoxintong was produced by Radix Astragali, Radix Notoginseng, Rhizoma Chuanxiong, Scolopendra at Anhui University of Traditional Chinese Medicine. Mouse anti-rat nestin, gliat fibrillary acidic protein, and galactocerebroside monoclonal antibodies, as well as rabbit anti-neuron-specific enolase polyclonal antibody were produced by Chemicon, Billerica, MA, USA. METHODS： Wistar rats aged 3 months were intragastrically infused with Naoluoxintong. Wistar rat embryonic NSCs （passage 8） were induced to proliferate and differentiate using 10% fetal bovine serum, 10% Naoluoxintong serum, and 10% rat serum. MAIN OUTCOME MEASURES： Phenotypic changes in cultured cells were detected using phase contrast microscopy, and cell proliferation and differentiation were observed using immunofluorescence staining. RESULTS： Proliferation and differentiation of embryonic NSCs was induced by three different types of blood serum. Although the differentiation time course with Nao/uoxintongserum was later than with the other two methods, the differentiated cells were morphologically similar to mature neurons to a greater extent. CONCLUSION： Nao/uoxintong serum supplementation induced differentiation of NSCs into neuronal-like cells and stimulated neuronal maturation.

Autologous serum can induce mesenchymal stem cells into hepatocyte-like cells

Objective： To investigate whether the rabbit serum after radiofrequency ablation to liver tumor can induce mesenchymal stem cells （MSCs） differentiating into hepatocyte-like cells in order to find a new source and culture process for repairing liver injury. Methods： A tumor piece of 1 mm× 1mm×1 mm was transplanted into a tunnel at right liver of rabbits. The model of liver tumor was established after 2-3 weeks. The serum was collected from rabbits 72 h after being subjected to radiofrequency ablation of the liver tumor. Mesenchymal stem cells were isolated from rabbit bone marrow and cultured in DMEM containing autologous rabbit serum. Three kinds of media （L-DMEM） were tested respectively： ① containing 10% fetal calf serum （FCS）; ② containing 30% rabbit autologous serum after radiofrequency ablation of the liver tumor （ASRF）; ③ containing 30% rabbit autologous serum （AS）. MSCs were cultured on 12-well plates until passage 2 and examined under the light and electron microscopy at indicted intervals. The expression of albumin and CKl8 was detected using immunofluorescence to identify the characteristics of differentiated cells. Results： MSCs performed differently in the presence of fetal calf serum, rabbit autologous serum and rabbit autologous serum after radiofrequency ablation of the liver tumor. Induced by the serum after radiofrequency ablation to liver tumor for 7 d, the spindle-shaped MSCs turned into round shaped and resembled hepatocyte-like cells. The reactions were not found in MSCs cultured in FCS and AS groups. After induction for 14 d, slender microvilli, cell-cell junction structure and cholangiole emerged, and the differentiated cells expressed albumin and CKl 8. All those could not been observed in 10% FCS and 30% autologous serum groups. Conclusion： Mesenchymal stem cells differentiate into hepatocyte-like cells in the serum after radiofrequency ablation of liver tumor, providing us a potential cell source and culture process for clinical application in liver injury repairing.

ALK Family Inhibitor A83-01 Promotes the Proliferation of Mouse Male Germline Stem Cells (mGSCs) Under Serum- and Feeder-Free Conditions

A83-01 is a selective inhibitor of the TGF-β type I receptor ALK,which inhibits the TGF-β-induced epithelial-to-mesenchymal transition（EMT） via the inhibition of Smad2 phosphorylation.Previous studies have showed that A83-01 promoted somatic cellular reprogramming significantly.Male germline stem cells（mGSCs）,as an alternative resource of pluripotent stem cells derived adult testis,have promising valuable in clinic medicine and regeneration,however,the derivation of mGSCs was complex and difficult.What the role A83-01 plays in promoting the proliferation of mGSCs is still unknown.In this study,combined with A83-01 and knockout serum replacement（KSR） medium,we obtained a relatively feeder-and serum-free system for mGSCs culturing in vitro and the optimal concentration of A83-01 was 0.25 μmol L-1.After continuous culturing,the proliferation efficiency of undifferentiated mGSCs and differentiation capacity of mGSC were examined as well.Results showed that,A83-01 dramatically increased the number of mGSCs and AP positive colonies,and the mitosis index according to the BrdU assay.A83-01 could also increase the expression of pluripotent markers including Oct4,Klf4,Nanog and c-Myc,analyzed byreal-time quantative PCR.mGSCs cultured in the optimal feeder-and serum-free system combined with A83-01 could form embryoid bodies（EBs）,which consisted of three embryonic layers detected by immunofluorescence and RT-PCR.Remarkably,the results demonstrated 0.25 μmol L-1A83-01 could promote the proliferation of mouse mGSC colonies and maintain their undifferentiated status under feeder-and serum-free systems.

Effect of endogenous insulin-like growth factor and stem cell factor on diabetic colonic dysmotility

AIM: To investigate whether the reduction of stem cell factor (SCF) is mediated by decreased endogenous insulin-like growth factor (IGF)-1 in diabetic rat colon smooth muscle. METHODS: Sixteen Sprague-Dawley rats were randomly divided into two groups: control group and streptozotocin-induced diabetic group. After 8 wk of streptozotocin administration, colonic motility function and contractility of circular muscle strips were measured. The expression of endogenous IGF-1 and SCF was tested in colonic tissues. Colonic smooth muscle cells were cultured from normal adult rats. IGF-1 siRNA transfection was used to investigate whether SCF expression was affected by endogenous IGF-1 expression in smooth muscle cells, and IGF-1 induced SCF expression effects were studied. The effect of high glucose on the expression of endogenous IGF-1 and SCF was also investigated. RESULTS: Diabetic rats showed prolonged colonic transit time (252 ± 16 min vs 168 ± 9 min, P < 0.01) and weakness of circular muscle contraction (0.81 ± 0.09 g vs 2.48 ± 0.23 g, P < 0.01) compared with the control group. Endogenous IGF-1 and SCF protein expression was significantly reduced in the diabetic colonic muscle tissues. IGF-1 and SCF mRNA expression also showed a paralleled reduction in diabetic rats. In the IGF-1 siRNA transfected smooth muscle cells, SCF mRNA and protein expression was significantly decreased. IGF-1 could induce SCF expression in a concentration and time-dependent manner, mainly through the extracellular-signal-regulated kinase 1/2 signal pathway. High glucose inhibited endogenous IGF-1 and SCF expression and the addition of IGF-1 to the medium reversed the SCF expression. CONCLUSION: Myopathy may resolve in colonic motility dysfunction in diabetic rats. Deficiency of endogenous IGF-1 in colonic smooth muscle cells leads to reduction of SCF expression.

Nutrient supplemented serum-free medium increases cardiomyogenesis efficiency of human pluripotent stem cells

AIM: To development of an improved p38 MAPK inhibitor-based serum-free medium for embryoid body cardiomyocyte differentiation of human pluripotent stem cells. METHODS: Human embryonic stem cells (hESC) differentiated to cardiomyocytes (CM) using a p38 MAPK inhibitor (SB203580) based serum-free medium (SB media). Nutrient supplements known to increase cell viability were added to SB medium. The ability of these supplements to improve cardiomyogenesis was evaluated by measurements of cell viability, total cell count, and the expression of cardiac markers via flow cytometry. An improved medium containing Soy hydrolysate (HySoy) and bovine serum albumin (BSA) (SupSB media) was developed and tested on 2 additional cell lines (H1 and Siu-hiPSC). Characterization of the cardiomyocytes was done by immunohistochemistry, electrophysiology and quantitative real-time reverse transcriptionpolymerase chain reaction. RESULTS: hESC cell line, HES-3, differentiating in SB medium for 16 d resulted in a cardiomyocyte yield of 0.07 ± 0.03 CM/hESC. A new medium (SupSB media) was developed with the addition of HySoy and BSA to SB medium. This medium resulted in 2.6 fold increase in cardiomyocyte yield (0.21 ± 0.08 CM/hESC). The robustness of SupSB medium was further demonstrated using two additional pluripotent cell lines (H1, hESC and Siu1, hiPSC), showing a 15 and 9 fold increase in cardiomyocyte yield respectively. The age (passage number) of the pluripotent cells did not affect the cardiomyocyte yields. Embryoid body (EB) cardiomyocytes formed in SupSB medium expressed canonical cardiac markers (sarcomeric α-actinin, myosin heavy chain and troponin-T) and demonstrated all three major phenotypes: nodal-, atrial- and ventricular-like. Electrophysiological characteristics (maximum diastolic potentials and action potential durations) of cardiomyocytes derived from SB and SupSB media were similar. CONCLUSION: The nutrient supplementation (HySoy and BSA) leads to increase in cell viability, cell yield and cardiac marker expression during cardiomyocyte differentiation, translating to an overall increase in cardiomyocyte yield.

Enhanced expression of epidermal growth factor receptor gene in gastric mucosal cells by the serum derived from rats treated with electroacupuncture at stomach meridian acupoints

AIM: To investigate the effect of serum derived from rats treated with electroacupuncture at stomach meridian acupoints on the expression of epidermal growth factor receptor (EGFR) gene in gastric mucosal cells. METHODS: The stress-induced gastric mucosal injury in rat model was established by water-immersion and restrained stress methods. 52 rats were randomly divided into: normal group (n = 8), model group (n = 8), model serum group (n = 12), stomach serum group (n = 12), and gallbladder serum group (n = 12). The gastric mucosal cells were separated by pronase-EDTA digestion method and incubated with serum. The EGFR gene expression in gastric mucosal cells was detected by reverse transcription-polymerase chain reaction (RT-PCR) method. RESULTS: Compared with normal group (0.6860 ± 0.0594), the serum derived from rats of the stomach group (1.2272 ± 0.0813, P = 0.00 < 0.01) and gallbladder group (0.9640 ± 0.0387, P = 0.00 < 0.01) had a tendency to enhance the EGFR gene expression in gastric mucosal cells. Such tendency existed in the model group (0.7104 ± 0.0457) but with no signifi cant difference (P = 0.495 > 0.05) and in model serum group (0.8516 ± 0.0409) with an extremely obvious difference (P = 0.001 < 0.01). Furthermore, the EGFR gene expression in stomach serum group was significantly higher than that in gallbladder serum group (P = 0.00 < 0.01). CONCLUSION: The present study shows that serumderived from rats treated with electroacupuncture at stomach meridian acupoints can distinctly increase the EGFR gene expression of gastric mucosal cells. Therefore, there is certain meridian specificity in the serum, which could provide a proof for the TCM theory “particular relation between meridian and internal organ”.

Novel Role of ER Stress and Mitochondria Stress in Serum-deprivation Induced Apoptosis of Rat Mesenchymal Stem Cells

The poor survival of mesenchymal stem cells （MSCs） compromises the efficacy of stem cell therapy. Growth factor deprivation is one of the important factors that have challenged the survival of donor MSCs in cell therapy. In this study, the aim was to evaluate the effect of serum deprivation on the cell death of MSCs and to investigate the underlying mechanisms. Apoptosis of MSCs was evaluated with Hoechst 33342/PI staining. Signaling pathways involved in serumdeprivation induced apoptosis were analyzed using Western blotting. The results revealed that serum deprivation induced apoptosis in MSCs within 72 h of treatment. Serum deprivation was shown to lead to protein expression alterations in Bax, Bcl-2, casepase-3, casepase-8, GRP78, and CHOP during experiments. The data suggested that the mitochondria death pathway, the extrinsic apoptotic pathway and the endoplastic reticulum（ER） stress pathway were all involved in MSCs apoptosis. The increase in expression of CHOP and the simultaneous decrease in Bcl- 2 expression suggest a synergistic effect in apoptosis induction in both the mitochondrion and the ER.

Role of Stem Cell Factor and Its Receptor in the Pathogenesis of Pediatric Aplastic Anemia

In order to investigate the levels of stem cell factor (SCF) and its receptor c-kit protein and mRNA in pediatric aplastic anemia (AA) and their relevance to the pathogenesis, immunocytochemical and in situ hybridization were utilized to detect the expression of SCF and its receptor c-kit gene protein and mRNA, respectively in 59 children with AA and 51 normal controls. The relationship between SCF and c-kit and the pathogenesis of AA was analyzed subsequently. The results showed that the positive rate of SCF protein and mRNA expression in children with AA was significantly lower than that in healthy controls (P<0.05). However, there was no significant difference in the positive rate of c-kit protein and mRNA expression between children with AA and control group (P>0.05). It was concluded that the expression of SCF is significantly decreased in children with AA, which may be closely associated with the pathogenesis of the AA. c-kit may be unrelated to the development of pediatric AA. Therefore, AA in children may have abnormalities at SCF/c-kit signal transduction levels.

Isolation and Identification of Cancer Stem Cells from Human Osteosarcom by Serum-free Three-dimensional Culture Combined with Anticancer Drugs

The cancer stem cells(CSCs)from human osteosarcoma by serum-free three-dimensional culture combined with anticancer drugs were isolated and identified.The primary cells derived from human osteosarcoma were digested by trypsin to prepare a single-cell suspension,and mixed homogeneously into 1.2% alginate gel.Single-cell alginate gel was cultured with serum-free DMEM/F12 medium.Epirubicin(0.8μg/mL)was added to the medium to enrich CSCs.After cultured conventionally for 7 to 10 days,most of cells suspended in ...

Mobilization Efficiency of Granulocyte Colony-stimulating Factor and Stem Cell Factor to Bone Marrow Mononuclear Cells and Mechanisms

The mobilization efficiency of granulocyte colony-stimulating factor (G-CSF) and stem cell factor (SCF) to bone marrow mononuclear cells (MNCs) in mice was observed, and the changes of CXCL12/CXCR4 signal were detected in order to find out the mobilization mechanism of stem cells. Kunming mice were randomly divided into two groups. The mice in treatment group were subjected to subcutaneous injection of G-CSF at a dose of 100 μg/kg and SCF at a dose of 25 μg/kg every day for 5 days, and those in control group were given isodose physiological saline. The MNCs were separated, counted and cultured, and the colony-forming unit-fibroblast (CFU-F) was evaluated. CD34+CXCR4+ MNCs were sorted by flow cytometry. The expression of CXCL12 protein in bone marrow extracellular fluid was detected by ELISA, and that of CXCL12 mRNA in bone marrow was measured by RT-PCR. The results showed that the counts of MNCs in peripheral blood and bone marrow were increased after administration of G-CSF/SCF (P<0.01). The factors had a dramatic effect on the expansion capability of CFU-F (P<0.05). Flow cytometric of bone marrow MNCs surface markers revealed that CD34+CXCR4+ cells accounted for 44.6%±8.7% of the total CD34+ MNCs. Moreover, G-CSF/SCF treatment induced a decrease in bone marrow CXCL12 mRNA that closely mirrored the fall in CXCL12 protein. In this study, it is evidenced that G-CSF/SCF can effectively induce MNCs mobilization by disrupting the balance of CXCL12/CXCR4 signaling pathway in the bone marrow and down-regulating the interaction of CXCL12/CXCR4.

Expression of stem cell factor in hair follicl epithelium

The interaction between flair folllcle dermis and its epidermis plays an important role in modulation of the growth and development of t he hair follicle. Stem cell factor (SCF ). which was found in recent years,is a cytokine related to the survival . growl h and development of the hemopoietic stem cells and can exert important biological effects on the development of keratinocytes and melanocytes. In this study, the expression of SCF in the hair follicle spithelium was invesstigated with immunohistochemistry and in situ hybridization. It was found that the gene of the encoded SCF was strongly expressed al a limited area in the middle of the hair follicle epithelium. The protein of SCF was evenly expressed in each part of the hair follicle epithelium. The findings suggest that the expression of SCF in the hair follicle epithelium at the level of molecule is different from thai at the level of protein.

Differentiation of Mesenchymal Stem Cells into Hepatocyte-Like Cells by Autologous Serum after Radiofrequency Ablation of Liver Tumor

Mesenchymal stem cells?(MSCs) have been shown to differentiate into liver cells in serum of part-resection liver, but it was hardly feasible in clinical use. Our studies revealed that MSCs could differentiate into hepatocyte-like cells in autologous serum after radiofrequency ablation (RFA) therapy of the liver tumor. Rabbits with liver tumor subsequently treated with RFA therapy. Serum was collected from those rabbits before RFA therapy and 72 hours after RFA therapy. MSCs were isolated from each rabbit’s bone marrow and cultured in DMEM medium containing the following different supplements: 30% fetal calf serum (FCS group), 30% rabbit autologous serum (AS group) or 30% autologous serum after RFA treatment of the liver tumor (ASRF group), observed by electron microscopy, flow cytometry, immunofluorescence. Seven days later, most of the spindle-shaped MSCs in the ASRF group transformed into polygon or round-shaped cells resembling hepatocytes, and the percentage in S/G2/M phase was higher than in the FCS or AS groups. Fourteen days later, slender microvilli, cell-cell junction structures and cholangiole emerged in the cells belonging to the ASRF group, the expression of albumin and CK18 was observed only in the differentiated cells from the ASRF group. These changes were not observed in the FCS group or the AS group. This study may provide a potential cell source and culture process for clinical application in liver injury treatment.

Shuyusan-containing serum protects SH-SY5Y cells against corticosterone-induced impairment

The Chinese herb Shuyusan, whose main constituent is jasminoidin, has been shown to protect SH-SY5Y cells against corticosterone-induced damage. SH-SY5Y cells injured by 400 μmol/L cor- ticosterone were treated with 5 and 30 μg/mL Shuyusan-containing serum. Results revealed that Shuyusan-containing serum elevated the survival rate of SH-SY5Y cells, reduced Bax expression, increased Bcl-2 expression, markedly elevated brain-derived neurotrophic factor mRNA expression, and blocked cell apoptosis. Moreover, the effect of high-dose （30 μg/mL） Shuyusan-containing se- rum was more remarkable. Therefore, Shuyusan-containing serum appears to protect SH-SY5Y cells against corticosterone-induced impairment by adjusting the expression of apoptosis-associ- ated proteins and brain-derived neurotrophic factor. Moreover, high-dose Shuyusan-containing se- rum has a protective effect on high-dose corticosterone-induced impairment.

Hepatocyte growth factor enhances the ability of dental pulp stem cells to ameliorate atherosclerosis in apolipoprotein E-knockout mice

BACKGROUND Atherosclerosis(AS),a chronic inflammatory disease of blood vessels,is a major contributor to cardiovascular disease.Dental pulp stem cells(DPSCs)are capable of exerting immunomodulatory and anti-inflammatory effects by secreting cytokines and exosomes and are widely used to treat autoimmune and inflam-mation-related diseases.Hepatocyte growth factor(HGF)is a pleiotropic cytokine that plays a key role in many inflammatory and autoimmune diseases.AIM To modify DPSCs with HGF(DPSC-HGF)and evaluate the therapeutic effect of DPSC-HGF on AS using an apolipoprotein E-knockout(ApoE-/-)mouse model and an in vitro cellular model.METHODS ApoE-/-mice were fed with a high-fat diet(HFD)for 12 wk and injected with DPSC-HGF or Ad-Null modified DPSCs(DPSC-Null)through tail vein at weeks 4,7,and 11,respectively,and the therapeutic efficacy and mechanisms were analyzed by histopathology,flow cytometry,lipid and glucose measurements,real-time reverse transcription polymerase chain reaction(RT-PCR),and enzyme-linked immunosorbent assay at the different time points of the experiment.An in vitro inflammatory cell model was established by using RAW264.7 cells and human aortic endothelial cells(HAOECs),and indirect co-cultured with supernatant of DPSC-Null(DPSC-Null-CM)or DPSC-HGF-CM,and the effect and mechanisms were analyzed by flow cytometry,RT-PCR and western blot.Nuclear factor-κB(NF-κB)activators and inhibitors were also used to validate the related signaling pathways.RESULTS DPSC-Null and DPSC-HGF treatments decreased the area of atherosclerotic plaques and reduced the expression of inflammatory factors,and the percentage of macrophages in the aorta,and DPSC-HGF treatment had more pronounced effects.DPSCs treatment had no effect on serum lipoprotein levels.The FACS results showed that DPSCs treatment reduced the percentages of monocytes,neutrophils,and M1 macrophages in the peripheral blood and spleen.DPSC-Null-CM and DPSC-HGF-CM reduced adhesion molecule expression in tumor necrosis factor-αstimulated HAOECs and regulated M1 polarization and inflammatory factor expression in lipopolysaccharide-induced RAW264.7 cells by inhibiting the NF-κB signaling pathway.CONCLUSION This study suggested that DPSC-HGF could more effectively ameliorate AS in ApoE-/-mice on a HFD,and could be of greater value in stem cell-based treatments for AS.

Microvesicles derived from mesenchymal stem cells inhibit acute respiratory distress syndrome-related pulmonary fibrosis in mouse partly through hepatocyte growth factor

BACKGROUND Pulmonary fibrosis is one of the main reasons for the high mortality rate among acute respiratory distress syndrome(ARDS)patients.Mesenchymal stromal cell-derived microvesicles(MSC-MVs)have been shown to exert antifibrotic effects in lung diseases.AIM To investigate the effects and mechanisms of MSC-MVs on pulmonary fibrosis in ARDS mouse models.METHODS MSC-MVs with low hepatocyte growth factor(HGF)expression(siHGF-MSC-MVs)were obtained via lentivirus transfection and used to establish the ARDS pulmonary fibrosis mouse model.Following intubation,respiratory mechanics-related indicators were measured via an experimental small animal lung function tester.Homing of MSC-MVs in lung tissues was investigated by near-infrared live imaging.Immunohistochemical,western blotting,ELISA and other methods were used to detect expression of pulmonary fibrosis-related proteins and to compare effects on pulmonary fibrosis and fibrosis-related indicators.RESULTS The MSC-MVs gradually migrated and homed to damaged lung tissues in the ARDS model mice.Treatment with MSC-MVs significantly reduced lung injury and pulmonary fibrosis scores.However,low expression of HGF(siHGF-MSC-MVs)significantly inhibited the effects of MSC-MVs(P<0.05).Compared with the ARDS pulmonary fibrosis group,the MSC-MVs group exhibited suppressed expression of type I collagen antigen,type III collagen antigen,and the proteins transforming growth factor-βandα-smooth muscle actin,whereas the siHGF-MVs group exhibited significantly increased expression of these proteins.In addition,pulmonary compliance and the pressure of oxygen/oxygen inhalation ratio were significantly lower in the MSC-MVs group,and the effects of the MSC-MVs were significantly inhibited by low HGF expression(all P<0.05).CONCLUSION MSC-MVs improved lung ventilation functions and inhibited pulmonary fibrosis in ARDS mice partly via HGF mRNA transfer.

High Glucose Promotes the CTGF Expression in Human Mesangial Cells via Serum and Glucocorticoid-induced Kinase 1 Pathway

The role of serum and glucocorticoid-induced kinase 1 （SGK1） pathway in the connective tissue growth factor （CTGF） expression was investigated in cultured human mesangial cells （HMCs） under high glucose. By using RT-PCR and Western blot, the effect of SGK1 on the CTGF expression in HMCs under high glucose was examined. Overexpression of active SGK1 in HMCs transfected with PIRES2-EGFP- S422D hSGK1 （SD） could increase the expression of phosphorylated SGK1 and CTGF as compared with HMCs groups transfected with PIRES2-EGFP （FP） under high glucose or normal glucose. Overexpression of inactive SGK1 in HMCs transfected with PIRES2-EGFP- K127N hSGK1 （KN） could decrease phosphorylated SGK1 and CTGF expression as compared with HMCs groups transfected with FP under high glucose. In conclusion, these results suggest that high glucose-induced CTGF expression is mediated through the active SGK1 in HMCs.

Multiple pretreatments can effectively improve the functionality of mesenchymal stem cells

In this editorial,we offer our perspective on the groundbreaking study entitled“Hypoxia and inflammatory factor preconditioning enhances the immunosup-pressive properties of human umbilical cord mesenchymal stem cells”,recently published in World Journal of Stem Cells.Despite over three decades of research on the clinical application of mesenchymal stem cells(MSCs),only a few therapeutic products have made it to clinical use,due to multiple preclinical and clinical challenges yet to be addressed.The study proved the hypoxia and inflammatory factor preconditioning led to higher immunosuppressive effects of MSCs without damaging their biological characteristics,which revealed the combination of inflammatory factors and hypoxic preconditioning offers a promising approach to enhance the function of MSCs.As we delve deeper into the intricacies of pretreat-ment methodologies,we anticipate a transformative shift in the landscape of MSC-based therapies,ultimately contributing to improved patient outcomes and advancing the field as a whole.

Overexpression of vascular endothelial growth factor enhances the neuroprotective effects of bone marrow mesenchymal stem cell transplantation in ischemic stroke

Although bone marrow mesenchymal stem cells(BMSCs)might have therapeutic potency in ischemic stroke,the benefits are limited.The current study investigated the effects of BMSCs engineered to overexpress vascular endothelial growth factor(VEGF)on behavioral defects in a rat model of transient cerebral ischemia,which was induced by middle cerebral artery occlusion.VEGF-BMSCs or control grafts were injected into the left striatum of the infarcted hemisphere 24 hours after stroke.We found that compared with the stroke-only group and the vehicle-and BMSCs-control groups,the VEGF-BMSCs treated animals displayed the largest benefits,as evidenced by attenuated behavioral defects and smaller infarct volume 7 days after stroke.Additionally,VEGF-BMSCs greatly inhibited destruction of the blood-brain barrier,increased the regeneration of blood vessels in the region of ischemic penumbra,and reducedneuronal degeneration surrounding the infarct core.Further mechanistic studies showed that among all transplant groups,VEGF-BMSCs transplantation induced the highest level of brain-derived neurotrophic factor.These results suggest that BMSCs transplantation with vascular endothelial growth factor has the potential to treat ischemic stroke with better results than are currently available.

Hypoxia and inflammatory factor preconditioning enhances the immunosuppressive properties of human umbilical cord mesenchymal stem cells

BACKGROUND Mesenchymal stem cells(MSCs)have great potential for the treatment of various immune diseases due to their unique immunomodulatory properties.However,MSCs exposed to the harsh inflammatory environment of damaged tissue after intravenous transplantation cannot exert their biological effects,and therefore,their therapeutic efficacy is reduced.In this challenging context,an in vitro preconditioning method is necessary for the development of MSC-based therapies with increased immunomodulatory capacity and transplantation efficacy.AIM To determine whether hypoxia and inflammatory factor preconditioning increases the immunosuppressive properties of MSCs without affecting their biological characteristics.METHODS Umbilical cord MSCs(UC-MSCs)were pretreated with hypoxia(2%O_(2))exposure and inflammatory factors(interleukin-1β,tumor necrosis factor-α,interferon-γ)for 24 h.Flow cytometry,polymerase chain reaction,enzyme-linked immunosorbent assay and other experimental methods were used to evaluate the biological characteristics of pretreated UC-MSCs and to determine whether pretreatment affected the immunosuppressive ability of UC-MSCs in coculture with immune cells.RESULTS Pretreatment with hypoxia and inflammatory factors caused UC-MSCs to be elongated but did not affect their viability,proliferation or size.In addition,pretreatment significantly decreased the expression of coagulationrelated tissue factors but did not affect the expression of other surface markers.Similarly,mitochondrial function and integrity were retained.Although pretreatment promoted UC-MSC apoptosis and senescence,it increased the expression of genes and proteins related to immune regulation.Pretreatment increased peripheral blood mononuclear cell and natural killer(NK)cell proliferation rates and inhibited NK cell-induced toxicity to varying degrees.CONCLUSION In summary,hypoxia and inflammatory factor preconditioning led to higher immunosuppressive effects of MSCs without damaging their biological characteristics.