Aggregation of human mesenchymal stromal cells (MSCs) into 3D spheroids enhances their antiinflammatory properties

Previous reports suggested that culture as 3D aggregates or as spheroids can increase the therapeutic potential of the adult stem/progenitor cells referred to as mesenchymal stem cells or multipotent mesenchymal stromal cells (MSCs). Here we used a hanging drop protocol to prepare human MSCs (hMSCs) as spheroids that maximally expressed TNFα stimulated gene/protein 6 (TSG-6), the antiinflammatory protein that was expressed at high levels by hMSCs trapped in the lung after i.v. infusion and that largely explained the beneficial effects of hMSCs in mice with myocardial infarcts. The properties of spheroid hMSCs were found to depend critically on the culture conditions. Under optimal conditions for expression of TSG-6, the hMSCs also expressed high levels of stanniocalcin-1, a protein with both antiinflammatory and antiapoptotic properties. In addition, they expressed high levels of three anticancer proteins: IL-24, TNFα-related apoptosis inducing ligand, and CD82. The spheroid hMSCs were more effective than hMSCs from adherent monolayer cultures in suppressing inflammatory responses in a coculture system with LPS-activated macrophages and in a mouse model for peritonitis. In addition, the spheroid hMSCs were about one-fourth the volume of hMSCs from adherent cultures. Apparently as a result, larger numbers of the cells trafficked through the lung after i.v. infusion and were recovered in spleen, liver, kidney, and heart. The data suggest that spheroid hMSCs may be more effective than hMSCs from adherent cultures in therapies for diseases characterized by sterile tissue injury and unresolved inflammation and for some cancers that are sensitive to antiinflammatory agents.     

Anti-inflammatory protein TSG-6 secreted by activated MSCs attenuates zymosan-induced mouse peritonitis by decreasing TLR2/NF-κB signaling in resident macrophages

Human mesenchymal stem/progenitor cells (hMSCs) repair tissues and modulate immune systems but the mechanisms are not fully understood. We demonstrated that hMSCs are activated by inflammatory signals to secrete the anti-inflammatory protein, TNF-α-stimulated gene 6 protein (TSG-6) and thereby create a negative feedback loop that reduces inflammation in zymosan-induced peritonitis. The results demonstrate for the first time that TSG-6 interacts through the CD44 receptor on resident macrophages to decrease zymosan/TLR2-mediated nuclear translocation of the NF-κB. The negative feedback loop created by MSCs through TSG-6 attenuates the inflammatory cascade that is initiated by resident macrophages and then amplified by mesothelial cells and probably other cells of the peritoneum. Because inflammation underlies many pathologic processes, including immune responses, the results may explain the beneficial effects of MSCs and TSG-6 in several disease models.     

Mesenchymal stem/stromal cells precondition lung monocytes/macrophages to produce tolerance against allo- and autoimmunity in the eye

Intravenously administered mesenchymal stem/stromal cells (MSCs) engraft only transiently in recipients, but confer long-term therapeutic benefits in patients with immune disorders. This suggests that MSCs induce immune tolerance by long-lasting effects on the recipient immune regulatory system. Here, we demonstrate that i.v. infusion of MSCs preconditioned lung monocytes/macrophages toward an immune regulatory phenotype in a TNF-α–stimulated gene/protein (TSG)-6–dependent manner. As a result, mice were protected against subsequent immune challenge in two models of allo- and autoimmune ocular inflammation: corneal allotransplantation and experimental autoimmune uveitis (EAU). The monocytes/macrophages primed by MSCs expressed high levels of MHC class II, B220, CD11b, and IL-10, and exhibited T-cell–suppressive activities independently of FoxP3⁺ regulatory T cells. Adoptive transfer of MSC-induced B220⁺CD11b⁺ monocytes/macrophages prevented corneal allograft rejection and EAU. Deletion of monocytes/macrophages abrogated the MSC-induced tolerance. However, MSCs with TSG-6 knockdown did not induce MHC II⁺B220⁺CD11b⁺ cells, and failed to attenuate EAU. Therefore, the results demonstrate a mechanism of the MSC-mediated immune modulation through induction of innate immune tolerance that involves monocytes/macrophages.Intravenous administration of mesenchymal stem/stromal cells (MSCs) has emerged as a promising cell-based immunotherapy for autoimmune diseases, graft-vs.-host disease, and transplantation (1–3). A significant body of data from preclinical and clinical studies has demonstrated remarkable immunosuppressive capacities of MSCs in various diseases, but the mechanisms are still difficult to explain.One key observation is that therapeutic benefits of MSC administration in animal models are achieved without significant engraftment of the cells; after i.v. infusion, most MSCs reside transiently within the lung and disappear rapidly with a t_1/2 of approximately 24 h in mice (4, 5). Therefore, direct suppressive effects of MSCs on the immune system are short-lived, and do not explain the long-term therapeutic effects observed with MSCs in clinical and animal studies. In this context, it has been suggested that MSCs trigger a state of immune tolerance, for example, through the induction of regulatory T cells (Tregs) (6, 7).Classically, lymphoid cells such as Tregs have been known to play a major role in regulating immune responses and maintaining immune tolerance. Recently, however, myeloid cells, including monocytes/macrophages, have gained attention as important mediators of immune regulation and tolerance (8, 9). In line with this, a few studies demonstrated that MSCs induce the expansion of myeloid cells with immunosuppressive activity and modulate monocytes/macrophages to an antiinflammatory phenotype, thereby inhibiting excessive inflammatory responses (10–12).However, it is not clear whether the MSC-educated myeloid cells can induce significant immune tolerance to repress adaptive immune responses in a setting of allo- or autoimmunity. Moreover, little is known about the mechanism(s) whereby MSCs induce immune tolerance through myeloid cells.In this study, we demonstrate that i.v. injection of MSCs into naive mice before immune challenge induces a significant immune tolerance in TNF-α–stimulated gene/protein (TSG)-6–dependent manner, and prevents the development of immune responses in two models of allo- and autoimmune ocular inflammation: corneal allotransplantation and experimental autoimmune uveitis (EAU). The MSC-induced tolerance involves a distinct subset of suppressive monocytes/macrophages in the lung, and is transferable independently of FoxP3⁺ Tregs. These data suggest a mechanism of MSCs in regulating adaptive immunity through induction of nonspecific innate tolerance.     

TSG-6 as a biomarker to predict efficacy of human mesenchymal stem/progenitor cells (hMSCs) in modulating sterile inflammation in vivo

Human mesenchymal stem/progenitor cells (hMSCs) from bone marrow and other tissues are currently being administered to large numbers of patients even though there are no biomarkers that accurately predict their efficacy in vivo. Using a mouse model of chemical injury of the cornea, we found that bone-marrow–derived hMSCs isolated from different donors varied widely in their efficacy in modulating sterile inflammation. Importantly, RT-PCR assays of hMSCs for the inflammation-modulating protein TSG-6 expressed by the TNFα-stimulated gene 6 (TSG-6 or TNFAIP6) predicted their efficacy in sterile inflammation models for corneal injury, sterile peritonitis, and bleomycin-induced lung injury. In contrast, the levels of TSG-6 mRNA were negatively correlated with their potential for osteogenic differentiation in vitro and poorly correlated with other criteria for evaluating hMSCs. Also, a survey of a small cohort suggested that hMSCs from female donors compared with male donors more effectively suppressed sterile inflammation, expressed higher levels of TSG-6, and had slightly less osteogenic potential.Human mesenchymal stem/progenitor cells (hMSCs) from bone marrow, adipose tissues, placenta, umbilical cord, and other tissues are currently being administered to large numbers of patients. Over 80 clinical trials with hMSCs have been registered (clinicaltrials.gov), and 5 have reached the Phase II or III stage of development (1). The trials are proceeding even though cultures of the cells are heterogeneous, and there is large variability in quality of hMSCs dependent on conditions, such as differences among donors, conditions used to expand the cells in culture, and random sampling in harvesting the cells from bone marrow and other tissues (2–5). Accounting for the variability among preparations is confounded by the lack of definitive markers for the cells. In addition, there are no biomarkers to predict the efficacy of hMSC samples in vivo. Therefore, the value of the data obtained from different clinical trials may well be compromised by variations in the quality of the hMSCs used.Recent data suggest that the therapeutic effects of the cells were explained in part by their paracrine effects, such as expression of factors that modulate inflammatory and immune responses, that limit growth of cancers, or that enhance tissue repair (2, 6–8). We recently observed that i.v.-infused hMSCs modulated excessive sterile inflammation and thereby improved mouse models for myocardial infarction (9), corneal injury (10), or peritonitis (11), in part because the hMSCs were activated to secrete TNFα-stimulated gene 6 (TSG-6), a protein that is a natural modulator of inflammation (12–14). Of special interest was that a well-known model of chemical injury of the cornea made it possible to obtain quantitative dose–response data for the effectiveness of recombinant TSG-6 on both neutrophil infiltration and the functional integrity of the tissue (15). Using this model and two other models for sterile inflammation, here, we demonstrated that bone-marrow–derived hMSCs isolated from different donors showed wide variations in their efficacy in modulating inflammation. Also, we identified a biomarker of mRNA levels for TSG-6 that predicts the in vivo efficacy of different donor-derived hMSCs in suppressing inflammation. The biomarker should prove useful in selecting preparations of hMSCs for therapy of patients.     

Time course of myocardial stromal cell–derived factor 1 expression and beneficial effects of intravenously administered bone marrow stem cells in rats with experimental myocardial infarction

Objective The chemokine stromal cell–derived factor–1 (SDF–1) has been implicated in homing of bone marrow cells to sites of injury. We investigated the time course of myocardial SDF–1 expression and effects of intravenously administered bone marrow mesenchymal stem cells (MSC) in rats with myocardial infarction (MI). Methods SDF–1 expression was measured by RT–PCR and Western blot in sham operated or infarcted hearts at 1/2, 1, 2, 4, 8 and 16 days post operation. MSCs from donor rats were labeled with BrdU. A total of 5 × 10⁶ cells in 2.5 mL of PBS or equal volume PBS alone were injected through the tail vein at above mentioned time points. The number of the labeled MSCs in the infarcted hearts was counted 3 days post injection. Cardiac function and vessel numbers were assessed 28 days post injection. Results Myocardial SDF–1 expression increased and peaked at the first day and decreased thereafter post MI and remained unchanged in sham operated hearts. The MSCs enrichment and angiogenesis in the host hearts were more abundant in the 1 day transplantation group than in the other groups (P < 0.01). Cardiac function was only improved in rats received intravenous MSCs injection within 4 days post MI and not affected by PBS injection. Conclusions Myocardial SDF–1 expression was increased only in the early phase post MI. MSCs intravenous infused at the early phase of MI were recruited to injured heart, enhanced angiogenesis and improved cardiac function.Dr. Ma and Prof. Ge contributed equally to this work     

Secretory Profile of Adipose-Tissue-Derived Mesenchymal Stem Cells from Cats with Calicivirus-Positive Severe Chronic Gingivostomatitis

The feline calicivirus (FCV) causes infections in cats all over the world and seems to be related to a broad variety of clinical presentations, such as feline chronic gingivostomatitis (FCGS), a severe oral pathology in cats. Although its etiopathogeny is largely unknown, FCV infection is likely to be a main predisposing factor for developing this pathology. During recent years, new strategies for treating FCGS have been proposed, based on the use of mesenchymal stem cells (MSC) and their regenerative and immunomodulatory properties. The main mechanism of action of MSC seems to be paracrine, due to the secretion of many biomolecules with different biological functions (secretome). Currently, several pathologies in humans have been shown to be related to functional alterations of the patient’s MSCs. However, the possible roles that altered MSCs might have in different diseases, including virus-mediated diseases, remain unknown. We have recently demonstrated that the exosomes produced by the adipose-tissue-derived MSCs (fAd-MSCs) from cats suffering from FCV-positive severe and refractory FCGS showed altered protein contents. Based on these findings, the goal of this work was to analyze the proteomic profile of the secretome produced by feline adipose-tissue-derived MSCs (fAd-MSCs) from FCV-positive patients with FCGS, in order to identify differences between them and to increase our knowledge of the etiopathogenesis of this disease. We used high-resolution mass spectrometry and functional enrichment analysis with Gene Ontology to compare the secretomes produced by the fAd-MSCs of healthy and calicivirus-positive FCGS cats. We found that the fAd-MSCs from cats with FCGS had an increased expression of pro-inflammatory cytokines and an altered proteomic profile compared to the secretome produced by cells from healthy cats. These findings help us gain insight on the roles of MSCs and their possible relation to FCGS, and may be useful for selecting specific biomarkers and for identifying new therapeutic targets.     

Mesenchymal stem cells: biology, pathophysiology, translational findings, and therapeutic implications for cardiac disease

Mesenchymal stem cells (MSCs) are a prototypical adult stem cell with capacity for self-renewal and differentiation with a broad tissue distribution. Initially described in bone marrow, MSCs have the capacity to differentiate into mesoderm- and nonmesoderm-derived tissues. The endogenous role for MSCs is maintenance of stem cell niches (classically the hematopoietic), and as such, MSCs participate in organ homeostasis, wound healing, and successful aging. From a therapeutic perspective, and facilitated by the ease of preparation and immunologic privilege, MSCs are emerging as an extremely promising therapeutic agent for tissue regeneration. Studies in animal models of myocardial infarction have demonstrated the ability of transplanted MSCs to engraft and differentiate into cardiomyocytes and vasculature cells, recruit endogenous cardiac stem cells, and secrete a wide array of paracrine factors. Together, these properties can be harnessed to both prevent and reverse remodeling in the ischemically injured ventricle. In proof-of-concept and phase I clinical trials, MSC therapy improved left ventricular function, induced reverse remodeling, and decreased scar size. This article reviews the current understanding of MSC biology, mechanism of action in cardiac repair, translational findings, and early clinical trial data of MSC therapy for cardiac disease.     

Allogeneic human mesenchymal stem cells for treatment of E. coli endotoxin-induced acute lung injury in the ex vivo perfused human lung

Recent studies have suggested that bone marrow-derived multipotent mesenchymal stem cells (MSCs) may have therapeutic applications in multiple clinical disorders including myocardial infarction, diabetes, sepsis, and hepatic and acute renal failure. Here, we tested the therapeutic capacity of human MSCs to restore alveolar epithelial fluid transport and lung fluid balance from acute lung injury (ALI) in an ex vivo perfused human lung preparation injured by E. coli endotoxin. Intra-bronchial instillation of endotoxin into the distal airspaces resulted in pulmonary edema with the loss of alveolar epithelial fluid transport measured as alveolar fluid clearance. Treatment with allogeneic human MSCs or its conditioned medium given 1 h following endotoxin-induced lung injury reduced extravascular lung water, improved lung endothelial barrier permeability and restored alveolar fluid clearance. Using siRNA knockdown of potential paracrine soluble factors, secretion of keratinocyte growth factor was essential for the beneficial effect of MSCs on alveolar epithelial fluid transport, in part by restoring amiloride-dependent sodium transport. In summary, treatment with allogeneic human MSCs or the conditioned medium restores normal fluid balance in an ex vivo perfused human lung injured by E. coli endotoxin.     

Amelioration of collagen-induced arthritis in DBA/1J mice by recombinant TSG-6, a tumor necrosis factor/interleukin-1-inducible protein

OBJECTIVE: To examine the effect of recombinant TSG-6 on collagen-induced arthritis (CIA) in DBA/1J mice. TSG-6 is a tumor necrosis factor (TNF)/ interleukin-1 (IL-1)-inducible hyaluronan-binding protein produced by synovial cells and chondrocytes that is present in synovial fluids of patients with rheumatoid arthritis. METHODS: To determine the effect of TSG-6 on chronic inflammatory joint disease, we induced CIA in DBA/1J mice by immunization with bovine type II collagen. Animals were treated with 12 intraperitoneal doses of 200 microg of recombinant TSG-6, beginning 3 days before the expected onset of disease symptoms. Progression of arthritis was monitored by determining the disease incidence, arthritis index, and footpad swelling. Levels of IgG1, IgG2a, and IgG2b antibodies against bovine and murine type II collagen and serum concentrations of IL-6 were determined at various time points. Histologic examination of affected joints was performed approximately 20 days after the onset of arthritis. RESULTS: Treatment with recombinant TSG-6 protein had a potent ameliorative effect, manifested by decreases in the disease incidence, arthritis index, and footpad swelling. Histologic examination of affected joints in TSG-6-treated animals revealed little pannus formation and cartilage erosion, features which were conspicuous in control mice. Animals treated with recombinant TSG-6 developed significantly reduced levels of IgG1, IgG2a, and IgG2b antibodies against bovine and murine type II collagen. CONCLUSION: The antiinflammatory effect of the TNF/IL-1-inducible TSG-6 protein in murine CIA suggests a role for this protein as an endogenous regulator of the inflammatory process.     

Engraftment and migration of human bone marrow stromal cells implanted in the brains of albino rats—similarities to astrocyte grafts

Neurotransplantation has been used to explore the development of the central nervous system and for repair of diseased tissue in conditions such as Parkinson’s disease. Here, we examine the effects of direct injection into rat brain of human marrow stromal cells (MSCs), a subset of cells from bone marrow that include stem-like precursors for nonhematopoietic tissues. Human MSCs isolated by their adherence to plastic were infused into the corpus striatum. Five to 72 days later, brain sections were examined for the presence of the donor cells. About 20% of the infused cells had engrafted. There was no evidence of an inflammatory response or rejection. The cells had migrated from the injection site along known pathways for migration of neural stem cells to successive layers of the brain. After infusion into the brain, the human MSCs lost their immunoreactivity to antibodies for collagen I. Initially, the human cells continued to stain with antibodies to fibronectin but the region of staining with fibronectin was significantly decreased at 30 and 72 days. The results suggest that MSCs may be useful vehicles for autotransplantation in both cell and gene therapy for a variety of diseases of the central nervous system.     

Intravenous vs intraperitoneal mesenchymal stem cells administration: What is the best route for treating experimental colitis?

AIM: To investigate the therapeutic effects of mesenchymal stem cells (MSCs) transplanted intraperitoneally and intravenously in a murine model of colitis.METHODS: MSCs were isolated from C57BL/6 mouse adipose tissue. MSC cultures were analyzed according to morphology, cellular differentiation potential, and surface molecular markers. Experimental acute colitis was induced in C57BL/6 mice by oral administration of 2% dextran sulfate sodium (DSS) in drinking water ad libitum from days 0 to 7. Colitis mice were treated with 1 × 10⁶ MSCs via intraperitoneal or intravenous injection on days 2 and 5. The disease activity index was determined daily based on the following parameters: weight loss, stool consistency and presence of blood in the feces and anus. To compare morphological and functional differences in tissue regeneration between different MSC injection modalities, mice were euthanized on day 8, and their colons were examined for length, weight, and histopathological changes. Inflammatory responses were determined by measuring the levels of different serum cytokines using a CBA Th1/Th2/Th17 kit. Apoptotic rates were evaluated by terminal deoxynucleotidyl transferase-mediated dUDP-biotin nick end labeling assay.RESULTS: Intravenous infusion of MSCs was more effective than intraperitoneal treatment (P < 0.001) in reducing the clinical and histopathologic severity of colitis, which includes weight loss, diarrhea and inflammation. An histological evaluation demonstrated decreased colonic inflammation based on reduced crypt loss and reduced infiltration of inflammatory cells. This therapeutic effect was most likely mediated by the down-regulation of pro-inflammatory cytokines [interleukin (IL)-6 and tumor necrosis factor (TNF)]; and by the up-regulation of anti-inflammatory cytokines (IL-10 and IL-4). Intravenous transplantation also induced high levels of IFN that lead to activation of the immunosuppressive activity of the MSCs, which did not occur with intraperitoneal transplantation (P = 0.006). An increase in apoptotic T cells was observed after intravenous, but not intraperitoneal, MSC infusion, suggesting that MSCs can induce apoptosis in resistant T cells in colonic inflammation (P = 0.027).CONCLUSION: Our results demonstrate that intravenous treatment is a superior method for reducing colon inflammation compared with intraperitoneal therapy.     

结肠炎大鼠炎症黏膜提取液对骨髓间质干细胞增殖和表面分化抗原的影响

背景:动物实验和临床试验均表明间质干细胞(MSCs)对受损肠道组织有一定修复作用,然而目前尚不清楚肠道炎症微环境对MSCs移植治疗炎症性肠病(IBD)有何影响。目的:通过体外实验观察结肠炎大鼠模型肠道炎症黏膜提取液对骨髓MSCs增殖和表面分化抗原的影响。方法:以全骨髓贴壁法分离和原代培养大鼠MSCs并行传代扩增。取三硝基苯磺酸(TNBS)结肠炎大鼠模型肠道炎症黏膜提取液(0、1、2、3 ml)与MSCs共培养,倒置相差显微镜观察各组细胞贴壁和生长情况,绝对计数法绘制细胞生长曲线,流式细胞术检测细胞表面分化抗原表达。结果:全骨髓贴壁法可成功分离MSCs,所获细胞CD29、CD44表达阳性,CD105表达低度阳性,CD34、CD45表达阴性。3 ml炎症黏膜提取液处理组MSCs接种6 h后见细胞贴壁,36 h开始克隆性增生,此后细胞增殖速度较空白对照组明显加快,第6 d即达100%融合,但表面分化抗原表达与空白对照组相比无明显变化。结论:结肠炎大鼠炎症黏膜提取液可促进骨髓MSCs增殖,但对细胞表面分化抗原无明显影响。     

Mesenchymal stem cells participating in ex vivo endothelium repair and its effect on vascular smooth muscle cells growth

BACKGROUND: Previous studies have shown that mesenchymal stem cells (MSCs) transplantation can promote neovascularization and regenerate damaged myocardium. However, it remains unknown whether MSCs seeding can be used to repair injured cellular components in vascular diseases. In this study we explored the feasibility of applying MSCs to endothelium repair in endothelial damage and vasoproliferative disorders. METHODS: Ex vivo model of endothelium repair was developed in which rabbit vascular smooth muscle cells (SMCs) were inoculated into the upper chamber and rabbit endothelial cells (ECs)/human MSCs into the lower chamber of a co-culture system. 3H-TdR incorporation and PCNA protein expression were assayed and migrated number of SMCs was calculated to evaluate the effect of MSCs seeding on SMCs growth. Flk-1 and vWF protein expressions were observed to analyze the plasticity of the seeded MSCs along endothelial lineage. RESULTS: In this co-culture system, no vWF protein but Flk-1 protein was observed in the 25.71% of MSCs after having been co-cultured with mature rabbit ECs for 5 days. Compared with the control group, the proliferation and migration of SMCs was significantly increased by proliferative ECs but decreased by confluent ECs (n=6, P<0.01). MSCs seeding decreased the proliferation and migration of SMCs compatible with the effect of proliferative ECs (n=6, P<0.001). However, no inhibition on SMCs growth was observed with MSCs seeding in comparison to the effect of confluent ECs. CONCLUSIONS: MSCs seeding can inhibit the proliferation and migration of SMCs. MSCs co-cultured with mature ECs have the ability to undergo milieu-dependent differentiation toward ECs.     

Bone marrow-derived mesenchymal stem cells are capable of mediating hepatitis B virus infection in injured tissues

Summary.  We have previously showed that endothelial progenitor cells (EPCs) through uptake of hepatitis B virus (HBV) may play a critical role in mediating extrahepatic HBV diseases. However, it remains to be elucidated whether mesenchymal stem cells (MSCs) are capable of mediating HBV trans-infection into extrahepatic tissues. Methods and Results: In this study, we showed that HBV antigens, HBV DNA and the viral particles were detected in MSCs after 3 days virus challenge. Neither HBV covalently closed circular DNA nor pregenomic RNA were detected in MSCs. Intravenously transplantation of HBV-exposed MSCs into myocardial infarction mouse model resulted in incorporation of HBV into injured heart and other damaged tissues. Conclusion: These results indicate that MSCs could serve as an additional extrahepatic virus reservoir, which may play a role at least in part in mediating HBV trans-infection into the injured tissues through the process of MSCs recruitment.     

Biological mechanisms and applied prospects of mesenchymal stem cells in premature ovarian failure

Premature ovarian failure (POF), also known as primary ovarian insufficiency (POI), refers to the loss of ovarian function in women after puberty and before the age of 40 characterized by high serum gonadotropins and low estrogen, irregular menstruation, amenorrhea, and decreased fertility. However, the specific pathogenesis of POF is unexplained, and there is no effective therapy for its damaged ovarian tissue structure and reduced reserve function. Mesenchymal stem cells (MSCs), with multidirectional differentiation potential and self-renewal ability, as well as the cytokines and exosomes they secrete, have been studied and tested to play an active therapeutic role in a variety of degenerative pathologies, and MSCs are the most widely used stem cells in regenerative medicine. MSCs can reverse POI and enhance ovarian reserve function through differentiation into granulosa cells (GCs), immune regulation, secretion of cytokines and other nutritional factors, reduction of GCs apoptosis, and promotion of GCs regeneration. Many studies have proved that MSCs may have a restorative effect on the structure and fertility of injured ovarian tissues and turn to be a useful clinical approach to the treatment of patients with POF in recent years. We intend to use MSCs-based therapy to completely reverse POI in the future.     

同种异体骨髓间充质干细胞在急性坏死性胰腺炎大鼠中的迁移和分化

目的 观察同种异体来源的骨髓间充质干细胞(bone marrow mesenchymal stem cells,MSCs)在急性坏死性胰腺炎(ANP)大鼠中的迁移及分化状况.方法 分离、纯化雄性SD大鼠的骨髓MSCs.按数字表法将雌性SD大鼠分为正常移植组、ANP移植组和ANP组,每组10只.采用腹腔注射L-精氨酸方法制备ANP模型.24 h后2个移植组大鼠经尾静脉输注MSCs.移植后72 h处死大鼠,取胰腺及心脏、肝脏、肾脏组织标本.胰腺组织常规病理检查并评分,采用原位杂交方法检测各组织Y染色体雄性鉴别基因sry片段的存在.结果 分离的MSCs在培养3～5 d内快速分裂增殖,形成集落,传代培养至第3代,CD29+CD44+CD45-细胞群达到95%以上.ANP组大鼠胰腺组织大片坏死,大量炎细胞浸润,病理分值为(10.31±0.85)分,显著高于ANP移植组的(7.30±0.79)分(P＜0.05).移植组大鼠的心脏、肝脏、胰腺、肾脏均可检测到sry基因.正常移植组胰腺组织可见散在分布的sry阳性细胞;ANP移植组胰腺组织可见较多sry阳性细胞,且多聚集于损伤较严重的部位.结论 炎性损伤的胰腺组织可能具有招募MSCs的能力,并能减轻大鼠胰腺局部的炎症反应.     

NF-κB inhibits osteogenic differentiation of mesenchymal stem cells by promoting β-catenin degradation

Mesenchymal stem cell (MSC)-based transplantation is a promising therapeutic approach for bone regeneration and repair. In the realm of therapeutic bone regeneration, the defect or injured tissues are frequently inflamed with an abnormal expression of inflammatory mediators. Growing evidence suggests that proinflammatory cytokines inhibit osteogenic differentiation and bone formation. Thus, for successful MSC-mediated repair, it is important to overcome the inflammation-mediated inhibition of tissue regeneration. In this study, using genetic and chemical approaches, we found that proinflammatory cytokines TNF and IL-17 stimulated IκB kinase (IKK)–NF-κB and impaired osteogenic differentiation of MSCs. In contrast, the inhibition of IKK–NF-κB significantly enhanced MSC-mediated bone formation. Mechanistically, we found that IKK–NF-κB activation promoted β-catenin ubiquitination and degradation through induction of Smurf1 and Smurf2. To translate our basic findings to potential clinic applications, we showed that the IKK small molecule inhibitor, IKKVI, enhanced osteogenic differentiation of MSCs. More importantly, the delivery of IKKVI promoted MSC-mediated craniofacial bone regeneration and repair in vivo. Considering the well established role of NF-κB in inflammation and infection, our results suggest that targeting IKK–NF-κB may have dual benefits in enhancing bone regeneration and repair and inhibiting inflammation, and this concept may also have applicability in many other tissue regeneration situations.     

Therapeutic potential of mesenchymal stem cells in the treatment of acute liver failure

Acute liver failure (ALF) is a severe and life-threatening condition in which rapid deterioration of liver function develops in a patient who has no preexisting liver disease. Mesenchymal stem cells (MSCs) are immunoregulatory stem cells which are able to modulate phenotype and function of all immune cells that play pathogenic role in the development and progression of ALF. MSCs in juxtacrine and paracrine manner attenuate antigen-presenting properties of dendritic cells and macrophages, reduce production of inflammatory cytokines in T lymphocytes, suppress hepatotoxicity of natural killer T (NKT) cells and promote generation and expansion of immunosuppressive T, B and NKT regulatory cells in acutely inflamed liver. Due to their nano-sized dimension and lipid envelope, intravenously injected MSC-derived exosomes (MSC-Exos) may by-pass all biological barriers to deliver MSC-sourced immunoregulatoy factors directly into the liver-infiltrated immune cells and injured hepatocytes. Results obtained by us and others revealed that intravenous administration of MSCs and MSC-Exos efficiently attenuated detrimental immune response and acute inflammation in the liver, suggesting that MSCs and MSC-Exos could be considered as potentially new remedies in the immunotherapy of ALF. In this review, we emphasize the current knowledge about molecular and cellular mechanisms which are responsible for MSC-based modulation of liver-infiltrated immune cells and we discuss different insights regarding the therapeutic potential of MSCs in liver regeneration.