Human Amniotic Mesenchymal Stem Cells Research Articles

The improvement of inflammatory infiltration and pregnancy outcome in mice with recurrent spontaneous abortion by human amniotic mesenchymal stem cells.

Recurrent spontaneous abortion (RSA) is thought to be mostly triggered by immune-related causes. Mesenchymal stem cells (MSCs), which exhibit the traits of multi-directional differentiation capacity and low immunogenicity, have recently been recommended as a viable treatment for spontaneous abortion-prone mice to increase the success of pregnancy. Amniotic membrane tissue is a byproduct of pregnancy and delivery that has a wide range of potential uses due to its easy access to raw materials and little ethical constraints. To construct an abortion-prone mouse model for this investigation, CBA/J female mice were coupled with male DBA/2 mice, while CBA/J female mice were paired with male BALB/c mice as a control. The identical volume of hAMSCs or PBS was injected intraperitoneally on the 4.5th day of pregnancy. CBA/J female mice were sacrificed by cervical dislocation on the 13.5th day of pregnancy, the embryo absorption rate was calculated, and the uterus, decidua tissues and placenta were gathered for examination. Through detection, it was discovered that hAMSCs significantly increased the expression of interleukin 10 (IL-10) and transforming growth factor beta (TGF-β), while significantly decreased the expression of interleukin 1 beta (IL-1β) and interleukin 6 (IL-6), improved vascular formation and angiogenesis, minimized the embryo absorption rate and inflammatory cell infiltration in the RSA + hAMSCs group. In any case, hAMSCs regulate inflammatory factors and cell balance at the maternal-fetal interface, which result in a reduction in the rate of embryo absorption and inflammatory infiltration and provide an innovative perspective to the clinical therapy of RSA.

Human amniotic mesenchymal stem cells-derived conditioned medium and exosomes alleviate oxidative stress-induced retinal degeneration by activating PI3K/Akt/FoxO3 pathway

Age-related macular degeneration (AMD) is the leading cause of vision loss among the elderly, which is primarily attributed to oxidative stress-induced damage to the retinal pigment epithelium (RPE). Human amniotic mesenchymal stem cells (hAMSC) were considered to be one of the most promising stem cells for clinical application due to their low immunogenicity, tissue repair ability, pluripotent potential and potent paracrine effects. The conditional medium (hAMSC-CM) and exosomes (hAMSC-exo) derived from hAMSC, as mediators of intercellular communication, play an important role in the treatment of retinal diseases, but their effect and mechanism on oxidative stress-induced retinal degeneration are not explored. Here, we reported that hAMSC-CM alleviated H2O2-induced ARPE-19 cell death through inhibiting mitochondrial-mediated apoptosis pathway in vitro. The overproduction of reactive oxygen species (ROS), alteration in mitochondrial morphology, loss of mitochondrial membrane potential and elevation of Bax/Bcl2 ratio in ARPE-19 cells under oxidative stress were efficiently reversed by hAMSC-CM. Moreover, it was found that hAMSC-CM protected cells against oxidative injury via PI3K/Akt/FoxO3 signaling. Intriguingly, exosome inhibitor GW4869 alleviated the inhibitory effect of hAMSC-CM on H2O2-induced decrease in cell viability of ARPE-19 cells. We further demonstrated that hAMSC-exo exerted the similar protective effect on ARPE-19 cells against oxidative damage as hAMSC-CM. Additionally, both hAMSC-CM and hAMSC-exo ameliorated sodium iodate-induced deterioration of RPE and retinal damage in vivo. These results first indicate that hAMSC-CM and hAMSC-exo protect RPE cells from oxidative damage by regulating PI3K/Akt/FoxO3 pathway, suggesting hAMSC-CM and hAMSC-exo will be a promising cell-free therapy for the treatment of AMD in the future.

IGF-1 Genome-Edited Human MSCs Exhibit Robust Anti-Arthritogenicity in Collagen-Induced Arthritis.

Stem cell therapy stands out as a promising avenue for addressing arthritis treatment. However, its therapeutic efficacy requires further enhancement. In this study, we investigated the anti-arthritogenic potential of human amniotic mesenchymal stem cells (AMM) overexpressing insulin-like growth factor 1 (IGF-1) in a collagen-induced mouse model. The IGF-1 gene was introduced into the genome of AMM through transcription activator-like effector nucleases (TALENs). We assessed the in vitro immunomodulatory properties and in vivo anti-arthritogenic effects of IGF-1-overexpressing AMM (AMM/I). Co-culture of AMM/I with interleukin (IL)-1β-treated synovial fibroblasts significantly suppressed NF-kB levels. Transplantation of AMM/I into mice with collagen-induced arthritis (CIA) led to significant attenuation of CIA progression. Furthermore, AMM/I administration resulted in the expansion of regulatory T-cell populations and suppression of T-helper-17 cell activation in CIA mice. In addition, AMM/I transplantation led to an increase in proteoglycan expression within cartilage and reduced infiltration by inflammatory cells and also levels of pro-inflammatory factors including cyclooxygenase-2 (COX-2), IL-1β, NF-kB, and tumor necrosis factor (TNF)-α. In conclusion, our findings suggest that IGF-1 gene-edited human AMM represent a novel alternative therapeutic strategy for the treatment of arthritis.

Effects of Mesenchymal Stem Cells-Derived Extracellular Vesicles on Inhibition of Hepatic Fibrosis by Delivering miR-200a.

Hepatic fibrosis (HF) is a common pathological feature of chronic hepatic diseases. We aimed to illuminate the significance of amniotic mesenchymal stem cells (AMSCs)-derived extracellular vesicles (AMSCs-EVs) in HF. Human AMSCs-EVs were isolated and identified. HF mice were constructed and treated with EVs. The fibrosis was observed by staining experiments and Western blot (WB) assay. Alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), and hepatic hydroxyproline (Hyp) were detected to confirm liver function. For the in vitro experiments, human hepatic stellate cells were induced with transforming growth factor-β and treated with EVs. To measure the degree of HF, the expression of alpha-smooth muscle actin (α-SMA) and Collagen I was detected by WB assay, and cell proliferation was detected by cell counting kit 8 assay. The levels of miR-200a, Zinc finger E-box binding homeobox 1 (ZEB1), and phosphoinositide-3-kinase regulatory subunit 3 (PIK3R3) were detected by WB and real-time quantitative polymerase chain reaction. The binding of ZEB1 to PIK3R3 and miR-200a to ZEB1 was analyzed by chromatin immunoprecipitation and dual luciferase assays to validate their relationships. Human AMSCs and AMSCs-EVs were obtained. Serum ALT, AST, TBIL, and hepatic Hyp were increased, implying the fibrosis degree was aggravated in HF mice, which was decreased again after EV treatment. EVs inhibited HF degree by reducing α-SMA and Collagen I and promoting cell proliferation. AMSCs-EVs delivered miR-200a into hepatocytes, which up-regulated miR-200a expression, inhibited ZEB1 expression, and reduced its enrichment on the PIK3R3 promoter, therefore inhibiting PIK3R3 expression and alleviating HF. Overexpression of ZEB1 or PIK3R3 attenuated the anti-fibrotic effect of AMSCs-EVs. Human AMSCs-derived EVs mediated miR-200a delivery and inhibition of intracellular ZEB1/PIK3R3 axis to exert anti-fibrosis effects.

Human amniotic mesenchymal stem cells inhibit immune rejection injury from allogeneic mouse heart transplantation: A preliminary study on the microRNA expression

BackgroundMesenchymal stem cell therapy is a new treatment for immune rejection in heart transplantation. The aim of this paper is to investigate the effect of human amniotic mesenchymal stem cells (hAMSCs) on alleviating immune rejection of allogeneic heart transplantation in mice and its possible underlying mechanism. MethodsWe injected hAMSCs into cervical ectopic heart transplantation model mice via tail vein to observe the survival time, the pathological changes of donor myocardium, and the fluorescent distribution of hAMSCs after the transplantation. MicroRNAs (miRs) with significantly differential expression were obtained by RNA sequencing and bioinformatic analysis, and a dual luciferase reporter gene assay together with real-time quantitative PCR (qRT-PCR) was performed to verify the relationship between miRs and their targeting genes. ResultsThe intervention of hAMSCs prolonged the graft survival time and alleviated the pathological damage of the donor heart. The injected hAMSCs were distributed mainly in the liver, spleen, and kidney, only a very small portion in the donor and recipient hearts. In the allogeneic transplantation models, the expression of miR-34b-5p significantly increased after hAMSC treatment. MiR-34b-5p showed a knockdown effect on gene Fc gamma receptor 2B (FCGR2B). ConclusionshAMSCs can reduce the immune rejection injury after allogeneic heart transplantation. This effect may be associated with the upregulation of miR-34b-5p expression to knock down its targeting gene FCGR2B.

Hypoxia preconditioning of human amniotic mesenchymal stem cells enhances proliferation and migration and promotes their homing via the HGF/C-MET signaling axis to augment the repair of acute liver failure

BackgroundTransplantation of mesenchymal stem cells (MSCs) is a newly developed strategy for treating acute liver failure (ALF). Nonetheless, the low survival rate of MSCs after transplantation and their poor homing to damaged tissues limit the clinical application of MSCs. The research assessed whether hypoxic preconditioning (HPC) can improve the biological activity of human amniotic mesenchymal stem cells (hA-MSCs), promote their homing ability to the liver of mice with ALF, and influence liver tissue repair. MethodsFlow cytometry, CCK8, Transwell, and Western blotting assays were conducted to assess the effects of hypoxic preconditioning on the phenotype, proliferation, and migration of hA-MSCs and the changes in the c-Met and CXCR4 gene expression levels were studied. To evaluate the effects of the transplantation of hypoxic preconditioning of hA-MSCs on the homing and repair of D-galactosamine (D-GalN)/LPS-induced ALF, the mechanism was elucidated by adding c-Met, CXCR4-specific blockers (SU11274 and AMD3100). ResultsAfter hypoxia pretreatment (1% oxygen volume fraction), hA-MSCs maintained the morphological characteristics of adherence and vortex colony growth and showed high CD44, CD90, and CD105 and low CD31, CD34, and CD45 expression levels. Hypoxic preconditioning of hA-MSCs significantly increased their proliferation and migration and highly expressed the c-Met and CXCR4 genes. In vivo and in vitro, this migration-promoting effect was suppressed by the c-Met specific blocker SU11274. In the acute liver failure mouse model, the HGF expression level was considerably elevated in the liver than that in the serum, lungs and kidneys. The transplantation of hypoxic preconditioned hA-MSCs introduced a remarkable improvement in the liver function and survival rate of mice with ALF and enhanced the anti-apoptosis ability of liver cells. The anti-apoptotic enhancing effect of hypoxic preconditioning was suppressed by the c-Met specific blocker SU11274. Hypoxic hA-MSCs administration was observed to have considerably increased the fluorescent cells in the liver than that recorded after administering normal oxygen-hA-MSCs. The number of hepatic fluorescent cells decreased remarkably after adding the c-Met inhibitor SU11274, compared to that recorded after hypoxic pretreatment, whereas the effect of c-Met inhibitor SU11274 on normal oxygen-hA-MSCs was not significant. ConclusionsHypoxic preconditioning depicted no impact on the morphology and phenotype features of the human amniotic mesenchymal stem cells, but it can promote their proliferation, migration, anti-apoptotic effect, and homing rate and improve the repair of acute liver failure, which might be mediated by the HGF/c-Met signaling axis.

Dual cross-linked COL1/HAp bionic gradient scaffolds containing human amniotic mesenchymal stem cells promote rotator cuff tendon–bone interface healing

The tendon-bone interface heals through scar tissue, while the lack of a natural interface gradient structure and collagen fibre alignment leads to the occurrence of retearing. Therefore, the promotion of tendon healing has become the focus of regenerative medicine. The purpose of this study was to develop a gradient COL1/ hydroxyapatite (HAp) biomaterial loaded with human amniotic mesenchymal stem cells (hAMSCs). The performance of common cross-linking agents, Genipin, 1-ethyl-3-(3-dimethylaminopropyl) carbodiimide/N-hydroxysuccinimide (EDC/NHS), and dual cross-linked materials were compared to select the best cross-linking mechanism to optimize the biological and mechanical properties of the scaffold. The optimal COL1/HAp-loaded with hAMSCs were implanted into the tendon-bone rotator cuff interfaces in rats and the effect on the tendon-bone healing was assessed by micro-CT, histological analysis, and biomechanical properties. The results showed that Genipin and EDC/NHS dual cross-linked COL1/HAp had good biological activity and mechanical properties and promoted the proliferation and differentiation of hAMSCs. Animal experiments showed that the group using a scaffold loaded with hAMSCs had excellent continuity and orientation of collagen fibers, increased fibrocartilage and bone formation, and significantly higher biomechanical functions than the control group at the interface at 12 weeks post operation. This study demonstrated that dual cross-linked gradient COL1/HAp-loaded hAMSCs could promote interface healing, thereby providing a feasible strategy for tendon-bone interface regeneration.

Severely Damaged Freeze-Injured Skeletal Muscle Reveals Functional Impairment, Inadequate Repair, and Opportunity for Human Stem Cell Application.

The regeneration of severe traumatic muscle injuries is an unsolved medical need that is relevant for civilian and military medicine. In this work, we produced a critically sized nonhealing muscle defect in a mouse model to investigate muscle degeneration/healing phases. We caused a freeze injury (FI) in the biceps femoris of C57BL/6N mice. From day 1 to day 25 post-injury, we conducted histological/morphometric examinations, an analysis of the expression of genes involved in inflammation/regeneration, and an in vivo functional evaluation. We found that FI activates cytosolic DNA sensing and inflammatory responses. Persistent macrophage infiltration, the prolonged expression of eMHC, the presence of centrally nucleated myofibers, and the presence of PAX7+ satellite cells at late time points and with chronic physical impairment indicated inadequate repair. By looking at stem-cell-based therapeutic protocols of muscle repair, we investigated the crosstalk between M1-biased macrophages and human amniotic mesenchymal stem cells (hAMSCs) in vitro. We demonstrated their reciprocal paracrine effects where hAMSCs induced a shift of M1 macrophages into an anti-inflammatory phenotype, and M1 macrophages promoted an increase in the expression of hAMSC immunomodulatory factors. Our findings support the rationale for the future use of our injury model to exploit the full potential of in vivo hAMSC transplantation following severe traumatic injuries.

Regulated intestinal microbiota and gut immunity to ameliorate type 1 diabetes mellitus: A novel mechanism for stem cell-based therapy

Although stem cell transplantation is an effective strategy in the treatment of type 1 diabetes mellitus (T1DM), the mechanisms underlying its therapeutic effects remain unclear. We hypothesized that stem cells target gut microbiota and intestinal mucosal immunity to promote therapeutic effects against T1DM. We investigated the effects of human amniotic mesenchymal stem cells (hAMSCs) on intestinal microbiota and mucosal immunity in streptozotocin-induced T1DM mice. hAMSCs promoted significant reductions in blood glucose levels and increased the number of insulin-secreting cells in the T1DM model. Compared with T1DM model mice, 16S rRNA sequencing revealed significant differences in the composition, diversity, and abundance of microbiota in the ileum of hAMSC-treated mice. Bifidobacterium, Prevotella, and Alcaligenes species were among the 15 most abundant differential bacterial species. LC-MS revealed significant changes in ileal metabolites, and among the top 100 differential metabolites identified, we found that a significant increase in taurine was closely associated with hAMSC therapy. Additionally, we detected significant differences between the two groups with respect to the frequency and phenotype of CD4+ T cell subsets in mesenteric lymph nodes, and hAMSCs promoted significant increases in Th2 and Treg cell frequencies and reduced the frequencies of Th1 and Th17 cells. Moreover, correlation analysis revealed pairwise correlations between differential microflora and differential metabolites and immune signatures. hAMSCs thus have positive effects on the microbiota and their metabolites in the ileum and intestinal mucosal immunity in T1DM. Our findings indicate that gut microbiota and intestinal mucosal immunity may play vital roles in the hAMSC-based treatment of T1DM.

Research on Runx2 gene induced differentiation of human amniotic mesenchymal stem cells into ligament fibroblasts in vitro and promotion of tendon-bone healing in rabbits

To investigate whether the Runx2 gene can induce the differentiation of human amniotic mesenchymal stem cells (hAMSCs) to ligament fibroblasts in vitro and promote the tendon-bone healing in rabbits. hAMSCs were isolated from the placentas voluntarily donated from healthy parturients and passaged, and then identified by flow cytometric identification. Adenoviral vectors carrying Runx2 gene (Ad-Runx2) and empty vector adenovirus (Ad-NC) were constructed and viral titer assay; then, the 3rd generation hAMSCs were transfected with Ad-Runx2 (Ad-Runx2 group) or Ad-NC (Ad-NC group). The real-time fluorescence quantitative PCR and Western blot were used to detect Runx2 gene and protein expression to verify the effectiveness of Ad-Runx2 transfection of hAMSCs; and at 3 and 7 days after transfection, real-time fluorescence quantitative PCR was further used to detect the expressions of ligament fibroblast-related genes [vascular endothelial growth factor (VEGF), collagen type Ⅰ, Fibronectin, and Tenascin-C]. The hAMSCs were used as a blank control group. The hAMSCs, hAMSCs transfected with Ad-NC, and hAMSCs were mixed with Matrigel according to the ratio of 1 : 1 and 1 : 2 to construct the cell-scaffold compound. Cell proliferation was detected by cell counting kit 8 (CCK-8) assay, and the corresponding cell-scaffold compound with better proliferation were taken for subsequent animal experiments. Twelve New Zealand white rabbits were randomly divided into 4 groups of sham operation group (Sham group), anterior cruciate ligament reconstruction group (ACLR group), anterior cruciate ligament reconstruction+hAMSCs transfected with Ad-NC-scaffold compound group (Ad-NC group), and anterior cruciate ligament reconstruction+hAMSCs transfected with Ad-Runx2-scaffold compound group (Ad-Runx2 group), with 3 rabbits in each group. After preparing the ACL reconstruction model, the Ad-NC group and the Ad-Runx2 group injected the optimal hAMSCs-Matrigel compunds into the bone channel correspondingly. The samples were taken for gross, histological (HE staining and sirius red staining), and immunofluorescence staining observation at 1 month after operation to evaluate the inflammatory cell infiltration as well as collagen and Tenascin-C content in the ligament tissues. Flow cytometric identification of the isolated cells conformed to the phenotypic characteristics of MSCs. The Runx2 gene was successfully transfected into hAMSCs. Compared with the Ad-NC group, the relative expressions of VEGF and collagen type Ⅰ genes in the Ad-Runx2 group significantly increased at 3 and 7 days after transfection ( P<0.05), Fibronectin significantly increased at 3 days ( P<0.05), and Tenascin-C significantly increased at 3 days and decreased at 7 days ( P<0.05). CCK-8 detection showed that there was no significant difference ( P>0.05) in the cell proliferation between groups and between different time points after mixed culture of two ratios. So the cell-scaffold compound constructed in the ratio of 1∶1 was selected for subsequent experiments. Animal experiments showed that at 1 month after operation, the continuity of the grafted tendon was complete in all groups; HE staining showed that the tissue repair in the Ad-Runx2 group was better and there were fewer inflammatory cells when compared with the ACLR group and the Ad-NC group; sirius red staining and immunofluorescence staining showed that the Ad-Runx2 group had more collagen typeⅠ and Ⅲ fibers, tending to form a normal ACL structure. However, the fluorescence intensity of Tenascin-C protein was weakening when compared to the ACLR and Ad-NC groups. Runx2 gene transfection of hAMSCs induces directed differentiation to ligament fibroblasts and promotes tendon-bone healing in reconstructed anterior cruciate ligament in rabbits.

Evaluation of Safety and Efficacy of Amniotic Mesenchymal Stem Cells for POI in Animals.

The efficacy of human amniotic mesenchymal stem cell (hAMSC) ovarian injection in improving ovarian function in primary ovarian insufficiency (POI) patients has been shown in some reports. However, the safety and efficacy of hAMSC vein injection remains unclear. In this study, we evaluated the safety and efficacy of hAMSC intravenous injection in cynomolgus macaques and SD rats and provided evidence for clinical trials. The hAMSCs were transplanted three times in SD rats at low, medium, and high doses. The animal behavior and biochemical and biophysical parameters were routinely monitored on a 2-month period posttransplantation, and histopathologic examinations were also performed. Experiments on the acute toxicity, allergy test, and hemolysis test showed that hAMSCs possess good biocompatibility. Our results showed that the maximum tolerated dose of hAMSCs in SD rats was 4.0 × 107 cells/kg. The maximum safe dose with three injections of hAMSCs in SD rats was 5.0 × 106 cells/kg. In addition, the results demonstrated that hAMSCs may restore POI rat ovarian function after two injections of 2.5 × 106 cells/kg or 5.0 × 106 cells/kg, which improved the disturbed estrous cycle, hormone levels, and ovarian lesions induced by pZP3. In conclusion, the preclinical results suggested that the transplantation of hAMSCs may be safe and efficacious for SD rats at doses of 5.0 × 106 cells/kg and lower.

Tumor Necrosis Factor-α-Induced Protein-8-like 2 Transfected Adipose-Derived Stem Cells Regulated the Dysfunction of Monocrotaline Pyrrole-Induced Pulmonary Arterial Smooth Muscle Cells and Pulmonary Arterial Endothelial Cells.

Pulmonary arterial hypertension (PAH) is characterized by pulmonary arterial endothelial cell (PAEC) dysfunction and pulmonary arterial smooth muscle cell (PASMC) activation. For decades, the therapies for PAH based on stem cells have been shown to be effective. Meanwhile, tumor necrosis factor-α-induced protein-8-like 2 (TIPE2) promote the viability of human amniotic mesenchymal stem cells. Therefore, we aimed to explore the role of TIPE2 in adipose-derived stem cells (ADSCs) and the function of TIPE2-transfected ADSCs in the regulation of PAH. We first explored the role and underlying molecular mechanism of TIPE2 in viability and migration of ADSCs. Moreover, the ADSCs transfected with TIPE2 were cocultured with monocrotaline pyrrole (MCTP)-stimulated PASMCs or PAECs. The effects and mechanisms of TIPE2-transfected ADSCs on MCTP-induced PASMCs and PAECs were further investigated. The results showed that TIPE2 overexpression promoted viability and migration of ADSCs by activating the TLR4-ERK1/2 pathway. In addition, TIPE2-transfected ADSCs inhibited the abnormal proliferation and the impaired apoptosis of PASMCs via NF-κB signaling and promoted the conversion of PASMCs from synthetic to contractile. Meanwhile, TIPE2-transfected ADSCs reduced the apoptosis, endothelial-to-mesenchymal transition, and migration of PAECs via PI3K/AKT signaling after MCTP treatment. MCTP-induced oxidative stress and inflammation of PAECs were significantly decreased by TIPE2-transfected ADSCs. In rat model, TIPE2-ADSCs administration further decreased the monocrotaline-induced increase in the right ventricular systolic pressure and ratio of right ventricle weight/left ventricle and septa weight (L + S) and right ventricle weight/body weight compared with the ADSCs group. In conclusion, TIPE2-transfected ADSCs dramatically attenuated the PAH via inhibiting the dysfunction of PASMCs and PAECs.

Phenolated alginate hydrogel induced osteogenic properties of mesenchymal stem cells via Wnt signaling pathway

Osteogenic properties of phenolated alginate (1.2 %) hydrogel containing collagen (0.5 %)/nano-hydroxyapatite (1 %) were studied on human mesenchymal stem cells in vitro. The phenolation rate and physical properties of the hydrogel were assessed using nuclear magnetic resonance (NMR), Fourier-transform infrared spectroscopy (FTIR), Scanning electron microscope (SEM), swelling ratio, gelation time, mechanical assay, and degradation rate. The viability of encapsulated cells was monitored on days 7, 14, and 21 using an MTT assay. Osteoblast differentiation was studied using western blotting, and real-time PCR. Using PCR array analysis, the role of the Wnt signaling pathway was also investigated. Data showed that the combination of alginate/collagen/nanohydroxyapatite yielded proper mechanical features. The addition of nanohydroxyapatite, and collagen reduced degradation, swelling rate coincided with increased stiffness. Elasticity and pore size were also diminished. NMR and FTIR revealed suitable incorporation of collagen and nanohydroxyapatite in the structure of alginate. Real-time PCR analysis and western blotting indicated the expression of osteoblast-related genes such as Runx2 and osteocalcin. PCR array revealed the induction of numerous genes related to Wnt signaling pathways during the maturation of human stem cells toward osteoblast-like cells. In vivo data indicated that transplantation of phenolated alginate/collagen/nanohydroxyapatite hydrogel led to enhanced de novo bone formation in rats with critical-sized calvarial defects. Phenolated alginate hydrogel can promote the osteogenic capacity of human amniotic membrane mesenchymal stem cells in the presence of nanohydroxyapatite and collagen via engaging the Wnt signaling pathway.

Immune and cytokine alterations and RNA-sequencing analysis in gestational tissues from pregnant women after recovery from COVID-19

BackgroundCOVID-19 is a global pandemic. Understanding the immune responses in pregnant women recovering from COVID-19 may suggest new therapeutic approaches.MethodsWe performed a cross-sectional study between March 1, 2020, and September 1, 2020. Participants were assigned into the convalescent COVID-19 group if they had a previous COVID-19 infection during pregnancy or the healthy control group. RNA-Seq was performed on human umbilical cord mesenchymal stem cells (hUMSCs) and human amniotic mesenchymal stem cells (hAMSCs). Immunohistochemical staining, cytokine testing, lymphocyte subset analysis, RNA-Seq, and functional analyses were performed on the placental and umbilical cord blood (UCB) and compared between the two groups.ResultsA total of 40 pregnant women were enrolled, with 13 in the convalescent group and 27 in the control group. There were 1024, 46, and 32 differentially expressed genes (DEGs) identified in the placental tissue, hUMSCs, and hAMSCs between the convalescent and control groups, respectively. Enrichment analysis showed those DEGs were associated with immune homeostasis, antiviral activity, cell proliferation, and tissue repair. Levels of IL-6, TNF-α, total lymphocyte counts, B lymphocytes, Tregs percentages, and IFN-γ expressing CD4+ and CD8+ T cells were statistically different between two groups (p ≤ 0.05). ACE2 and TMPRSS2 expressed on the placenta were not different between the two groups (p > 0.05).ConclusionMultiple changes in immune responses occurred in the placental tissue, hUMSCs, and hAMSCs after maternal recovery from COVID-19, which might imply their protective roles against COVID-19 infection.

Osteogenic differentiation of human amniotic mesenchymal stem cells by phycocyanin and phycoerythrin pigments isolated from Spirulina platensis and Gracilaria gracilis algae

Bone regeneration is a multistep and regular physiological process that occurs normally in fracture repair and bone defects. However, some factors such as aging, particular diseases and some drugs prevent or slowdown bone natural healing. Cell therapy using stem cells and differentiation activating factors is an effective treatment method for bone regeneration triggering in unusual conditions. Therefore, in the present study the effect of phycocyanin and phycoerythrin pigments which isolated from Spirulina platensis and Gracilaria gracilis algae was investigate on osteogenic differentiation potency of human Amniotic Mesenchymal Stem Cells (hAMSCs). For this purpose, hAMSCs were exposed to 300, 500, and 700 µg/ml concentrations of phycocyanin and phycoerythrin pigments and then the cells viability was measured with MTT assay in 48 and 72 h after treatment. The osteo-differentiation level of cells was studied by measuring ALP activity using calorimetric method and Alizarin red staining for calcium deposition in 7 and 21 days after treatment. Also, total RNA of cells was extracted in different time periods and then cDNA synthesized with specific primers, and relative expression of Runx2, β-catenin and Osteocalcin genes were investigated using SYBR Green RT-qPCR technique. Osteogenic differentiation of hAMSCs that treated with pigments was confirmed by mineral deposits staining and increased level of ALP activity. Furthermore, these pigments elevated significantly the expression of osteogenic marker genes compared to control samples and caused hAMSCs to differentiate into osteoblast cells. According to these results, phycocyanin and phycoerythrin may suggest as suitable osteogenic supplements with low toxicity, low cost and high efficiency, although the molecular mechanism of its efficacy is not available yet.

The application of MDT model for calciphylaxis management in patients with end-stage renal disease.

This study focuses on the application of nurse-led multidisciplinary collaborative therapy (MDT) management model for calciphylaxis prevention of patients with terminal renal disease. Through the establishment of a multidisciplinary management team spanning nephrology department, blood purification center, dermatology department, burn and plastic surgery department, infection department, stem cell platform, nutrition department, pain department, cardiology department, hydrotherapy group, dermatology group, and outpatient treatment room, the distribution of duties among team members were clarified to bring out the best advantages of a multidisciplinary teamwork during treatment and nursing. For patients with calciphylaxis symptoms in terminal renal disease, a case-by-case management model was carried out with the focus on personalised problem. We emphasised on personalised wound care, precise medication care, active pain management, psychological intervention and palliative care, the amelioration of calcium and phosphorus metabolism disorder, nutritional supplementation, and the therapeutic intervention based on human amniotic mesenchymal stem cell regeneration. The MDT model effectively compensates for traditional nursing mode and could serve as a novel clinical management modality for calciphylaxis prevention in patients with terminal renal disease.

Intraocular Pressure-Lowering and Retina-Protective Effects of Exosome-Rich Conditioned Media from Human Amniotic Membrane Stem Cells in a Rat Model of Glaucoma.

Glaucoma is one of the most devastating eye diseases, since the disease can develop into blindness and no effective therapeutics are available. Although the exact mechanisms and causes of glaucoma are unknown, increased intraocular pressure (IOP) has been demonstrated to be an important risk factor. Exosomes are lipid nanoparticles secreted from functional cells, including stem cells, and have been found to contain diverse functional molecules that control body function, inhibit inflammation, protect and regenerate cells, and restore damaged tissues. In the present study, exosome-rich conditioned media (ERCMs) were attained via hypoxic culture (2% O2) of human amniotic membrane mesenchymal stem cells (AMMSCs) and amniotic membrane epithelial stem cells (AMESCs) containing 50 times more exosome particles than normoxic culture (20% O2) medium (NCM). The exosome particles in ERCM were confirmed to be 77 nm in mean size and contain much greater amounts of growth factors (GFs) and neurotrophic factors (NFs) than those in NCM. The glaucoma-therapeutic effects of ERCMs were assessed in retinal cells and a hypertonic (1.8 M) saline-induced high-IOP animal model. CM-DiI-labeled AMMSC exosomes were found to readily penetrate the normal and H2O2-damaged retinal ganglion cells (RGCs), and AMMSC-ERCM not only facilitated retinal pigment epithelial cell (RPEC) proliferation but also protected against H2O2- and hypoxia-induced RPEC insults. The IOP of rats challenged with 1.8 M saline increased twice the normal IOP (12-17 mmHg) in a week. However, intravitreal injection of AMMSC-ERCM or AMESC-ERCM (3.9-4.5 × 108 exosomes in 10 μL/eye) markedly recovered the IOP to normal level in 2 weeks, similar to the effect achieved with platelet-derived growth factor-AB (PDGF-AB, 1.5 μg), a reference material. In addition, AMMSC-ERCM, AMESC-ERCM, and PDGF-AB significantly reversed the shrinkage of retinal layers, preserved RGCs, and prevented neural injury in the glaucoma eyes. It was confirmed that stem cell ERCMs containing large numbers of functional molecules such as GFs and NFs improved glaucoma by protecting retinal cells against oxidative and hypoxic injuries in vitro and by recovering IOP and retinal degeneration in vivo. Therefore, it is suggested that stem cell ERCMs could be a promising candidate for the therapy of glaucoma.

Human amniotic mesenchymal stem cell-conditioned medium inhibited LPS-induced cell injury and inflammation by modulating CD14/TLR4-mediated signaling pathway in monocytes

Human amniotic mesenchymal stem cells (hAMSCs) have attracted considerable attention as a promising regenerative therapy. Many studies reported that the conditioned medium of hAMSCs (AM-CM) exerted anti-inflammatory and immunomodulatory functions, while its underlying mechanism is poorly understood. In this study, we first confirmed that AM-CM (25%, 50%, 100%) was optimal for anti-inflammation at 24 h. Lipopolysaccharide (LPS)-induced alteration of cell morphology, the decrease of cell proliferation, and the upregulation of cell apoptosis were significantly reversed in AM-CM-treated THP-1 cells. 25% and 50% AM-CM significantly decreased LPS-induced intracellular reactive oxygen species (ROS) production and proinflammatory cytokines secretion. Mechanistically, we found that AM-CM treatment suppressed LPS-induced activation of MAPK and NF-κB pathways by inhibiting CD14/TLR4 in THP-1 cells. Meanwhile, activation of NLRP3 inflammasome was also dose-dependently attenuated by AM-CM treatment. Thus, AM-CM may exert positive influences on the inflammation microenvironment and provide a novel strategy for improving tissue regeneration.

Combined Application of Human Amniotic Membrane Mesenchymal Stem Cells and a Modified PGS-co-PCL Film in an Experimental Model of Myocardial Ischemia-Reperfusion Injury.

According to the World Health Organization (WHO), about 3.9 million people die annually of ischemic heart disease (IHD). Several clinical trials have shown that stem cell therapy is a promising therapeutic approach to IHD. Human amniotic membrane mesenchymal stem cells (hAMSCs) positively affect the repair of myocardial ischemia-reperfusion (MI/R) injury by stimulating endogenous repair mechanisms. The differentiated hAMSCs with and without modified PGS-co-PCL film were applied in the myocardium. MI/R injury was induced by ligating the left anterior descending artery in 48 male Wistar rats. The rats were divided into four groups, (n = 12) animals: heart failure (HF) as the control group, HF + MSCs, HF + MSCs + film, and HF + film. Echocardiography was performed 2 and 4weeks after MI/R injury moreover the expression of the VEGF protein was assessed in the rat heart tissue via immunohistochemistry. In vitro, our result shows fantastic cell survival when seeded on film. In vivo, the left ventricle ejection fraction (LEVD), fractional shortening (FS), end-diastolic (EDV), and stroke volume (SV) have been increased and systolic volumes decreased in all treatment groups in comparison with control. Although combination therapy has a more positive effect on hemodynamic parameters, there is no significant difference between HF + MSCs + film with other treatment groups. Also, In the IHC assay, expression of the VEGF protein significantly increased in all intervention groups. The implantation of MSCs and the modified film significantly enhanced the cardiac functional outcome; in this regard, enhancement in cell survival and VEGF expression are involved as underlying mechanisms in which cardiac film and MSCs exert a beneficial effect.

Human Amniotic Mesenchymal Stem Cells Alleviate aGVHD after allo-HSCT by Regulating Interactions between Gut Microbiota and Intestinal Immunity

Acute graft-versus-host disease (aGVHD) after allogeneic hematopoietic stem cell transplantation poses one of the most vexing challenges. Gut microbiota dysbiosis can proceed aGVHD and mesenchymal stem cells (MSCs) have promising therapeutic potential for aGVHD. However, whether hAMSCs affect the gut microbiota during aGVHD mitigation remains unknown. Accordingly, we sought to define the effects and underlying mechanisms of human amniotic membrane-derived MSCs (hAMSCs) regulating the gut microbiota and intestinal immunity in aGVHD. By establishing humanized aGVHD mouse models and hAMSCs treatment, we found that hAMSCs significantly ameliorated aGVHD symptoms, reversed the immune imbalance of T cell subsets and cytokines, and restored intestinal barrier. Moreover, the diversity and composition of gut microbiota were improved upon treatment with hAMSCs. Spearman’s correlation analysis showed that there was a correlation between the gut microbiota and tight junction proteins, immune cells as well as cytokines. Our research suggested that hAMSCs alleviated aGVHD by promoting gut microbiota normalization and regulating the interactions between the gut microbiota and intestinal barrier, immunity.Graphical