Recruitment Of Progenitor Cells Research Articles

Hyaluronic acid hydrogels incorporating platelet lysate enhance human pulp cell proliferation and differentiation.

The restoration of dentine-pulp complex remains a challenge for dentists; nonetheless, it has been poorly addressed. An ideal system should modulate the host response, as well as enable the recruitment, proliferation and differentiation of relevant progenitor cells. Herein was proposed a photocrosslinkable hydrogel system based on hyaluronic acid (HA) and platelet lysate (PL). PL is a cocktail of growth factors (GFs) and cytokines involved in wound healing orchestration, obtained by the cryogenic processing of platelet concentrates, and was expected to provide the HA hydrogels specific biochemical cues to enhance pulp cells' recruitment, proliferation and differentiation. Stable HA hydrogels incorporating PL (HAPL) were prepared after photocrosslinking of methacrylated HA (Met-HA) previously dissolved in PL, triggered by the Ultra Violet activated photoinitiator Irgacure 2959. Both the HAPL and plain HA hydrogels were shown to be able to recruit cells from a cell monolayer of human dental pulp stem cells (hDPSCs) isolated from permanent teeth. The hDPCs were also seeded directly over the hydrogels (5 × 104 cells/hydrogel) and cultured in osteogenic conditions. Cell metabolism and DNA quantification were higher, in all time-points, for PL supplemented hydrogels (p < 0,05). Alkaline phosphatase (ALPL) activity and calcium quantification peaks were observed for the HAPL group at 21 days (p < 0,05). The gene expression for ALPL and COLIA1 was up-regulated at 21 days to HAPL, compared with HA group (p < 0,05). Within the same time point, the gene expression for RUNX2 did not differ between the groups. Overall, data demonstrated that the HA hydrogels incorporating PL increased the cellular metabolism and stimulate the mineralized matrix deposition by hDPSCs, providing clear evidence of the potential of the proposed system for the repair of damaged pulp/dentin tissue and endodontics regeneration.

Porcine small intestine submucosal grafts improve remucosalization and progenitor cell recruitment to sites of upper airway tissue remodeling.

To better understand upper airway tissue regeneration, the exposed cartilage and bone at donor sites of tissue flaps may serve as in vivo "Petri dishes" for active wound healing. The pedicled nasoseptal flap (NSF) for skull-base reconstruction creates an exposed donor site within the nasal airway. The objective of this study is to evaluate whether grafting the donor site with a sinonasal repair cover graft is effective in promoting wound healing. In this multicenter, prospective trial, subjects were randomized to intervention (graft) or control (no graft) intraoperatively after NSF elevation. Individuals were evaluated at 2, 6, and 12 weeks postintervention with endoscopic recordings. Videos were graded (Likert scale) by 3 otolaryngologists blinded to intervention on remucosalization, crusting, and edema. Scores were analyzed for interrater reliability and cohorts compared. Biopsy and immunohistochemistry at the leading edge of wound healing was performed in select cases. Twenty-one patients were randomized to intervention and 26 to control. Subjects receiving the graft had significantly greater overall remucosalization (p = 0.01) than controls over 12 weeks. Although crusting was less in the small intestine submucosa (SIS) group, this was not statistically significant (p = 0.08). There was no overall effect on nasal edema (p = 0.2). Immunohistochemistry demonstrated abundant upper airway basal cell progenitors in 2 intervention samples, suggesting that covering grafts may facilitate tissue proliferation via progenitor cell expansion. This prospective, randomized, controlled trial indicates that a porcine SIS graft placed on exposed cartilage and bone within the upper airway confers improved remucosalization compared to current practice standards.

The homing of bone marrow stem cells is differentially activated in ischemic and valvular heart\xa0diseases and influenced by beta-blockers

BackgroundCell homing is the mechanism by which an injury releases signaling molecules that cause recruitment, proliferation, migration and differentiation of progenitor cells. Stromal derived factor-1 (SDF-1) and its receptor CXCR4 are key molecules involved in homing and little is known about their activation in cardiopathies. Here, we assessed the homing activation status of bone marrow cells (BMC) concerning the SDF-1 and CXCR4 expression in ischemic (IHD) and valvular (VHD) heart diseases.MethodsThe SDF-1 and inflammatory profile were analyzed by ELISA from plasma obtained bone marrow of ischemic heart patients (IHD, n = 41), valvular heart patients (VHD, n = 30) and healthy controls (C, n = 9). Flow cytometry was used to evaluate CXCR4 (CD184) expression on the surface of bone marrow cells, and the CXCR4 expression was estimated by real-time quantitative PCR.ResultsThe SDF-1 levels in the groups IHD, VHD and control were, respectively, 230, 530 and 620 pg/mL (P = 0.483), and was decreased in VHD patients using beta-blockers (263 pg/mL) when compared with other (844 pg/mL) (P = 0.023). Compared with IHD, the VHD group showed higher CXCR4 (P = 0.071) and CXCR7 (P = 0.082) mRNA expression although no difference in the level of CXCR4+ bone marrow cells was found between groups (P = 0.360).ConclusionIn conclusion, pathophysiological differences between IHD and VHD can affect the molecules involved in the activation of homing. In addition, the use of beta-blockers appears to interfere in this mechanism, a fact that should be considered in protocols that use BMC.

Research progress of dipeptidyl peptidase 4 inhibitors on healing of chronic diabetic foot ulcers

To review the effect of dipeptidyl peptidase 4 (DPP-4) inhibitors on the wound healing and its mechanisms in chronic diabetic foot ulcers. The latest literature concerning DPP-4 inhibitors for chronic diabetic foot ulcers was extensively reviewed, as well as the potential benefit and mechanism of DPP-4 inhibitors on wound healing of diabetic foot ulcers was analyzed thoroughly. DPP-4 inhibitors can accelerated the ulcer healing. The mechanisms probably include inhibiting the expression of the matrix metalloproteinase (MMP) and restoring the balance of the wound MMP and the tissue inhibitors of MMP; promoting recruitment of endothelial progenitor cells and augmenting angiogenesis; optimizing extracellular matrix construction and the immune response to persistent hypoxia in chronic diabetes wounds, and so on. At present, clinical researches show that DPP-4 inhibitors may be considered as an adjuvant treatment for chronic diabetic foot ulcers. DPP-4 inhibitors show promise in the local wound healing of chronic diabetic foot ulcers. However, more strictly designed, adequately powered, long-term follow-up, and high-quality randomized control trials are needed to further verify their efficacy and safety for chronic diabetic foot ulcers.

Cell membrane coating for reducing nanoparticle-induced inflammatory responses to scaffold constructs.

The controlled release of therapeutics from micro or nanoparticles has been well-studied. Incorporation of these particles inside biomaterial scaffolds is promising for tissue regeneration and immune modulation. However, these particles may induce inflammatory and foreign body responses to scaffold constructs, limiting their applications. Here we show that widely used poly(lactic-co-glycolic acid) nanoparticles (PLGA NPs) formed by double emulsion dramatically increased neutrophil infiltration and pro-inflammatory cytokines in alginate scaffolds 1 day after the subcutaneous injection of the scaffolds into mice. The coating of red blood cell (RBC) membranes on PLGA NPs completely eliminated these short-term inflammatory responses. For a longer term of 10 days, neither PLGA NPs nor RBC membrane-coated nanoparticles exerted a significant effect on the infiltration of neutrophils or macrophages in alginate scaffolds possibly due to the degradation and/or clearance of nanoparticles by infiltrating cells by that time. Despite the extensive exploration of cell membrane-coated nanoparticles, our study is the first to investigate the effects of cell membrane coating on foreign body reaction to nanoparticles. Harnessing the natural biocompatibility of cell membranes, our strategy of anti-inflammatory protection for scaffolds may be pivotal for many applications, such as those relying on the recruitment of stem cells and/or progenitor cells to scaffolds.

Circulating endothelial and endothelial progenitor cells in microvascular repairs in pancreatic hyperglycaemic-induced damage: a review

The body vasculature contains an inner lining of endothelium, which consists of highly metabolic active endothelial cells (ECs). In microvascular damage resulting from the detrimental physiological effects of hyperglycaemia, ECs are unable to change their glucose transport rate. This results in the mobilisation and recruitment of endothelial progenitor cells (EPCs) to the site of injury for neovascularisation and vascular repair. This review assessed the implications of circulating endothelial and endothelial progenitor cells (CEPCs) on pancreatic microvascular repair and highlighted the impact of EPCs in neovascularisation. This insight information may open up new avenues of research on novel approaches to maintaining and regenerating microvasculature in diabetes mellitus.

Tumor necrosis factor receptor 2 promotes neuroprotection during chronic autoimmune neuroinflammation

Abstract Multiple sclerosis (MS) is an inflammatory autoimmune disorder affecting the central nervous system (CNS) which affects over 400,000 Americans and over 2.5 million people worldwide. Although most patients are initially diagnosed with relapsing-remitting MS, the majority of these patients later develop a chronic-progressive form of MS, for which there is no well-established mouse model. The most common genetic factor associated with MS susceptibility is the Human Leukocyte Antigen (HLA)-DR2b haplotype. Additionally, studies in the experimental autoimmune encephalomyelitis (EAE) mouse model of MS, showed that lack of TNF signaling through its receptor TNFR2 leads to disease exacerbation and severe demyelination. Here, we developed a mouse model which expresses HLA-DR2b and lacks TNFR2, designated DR2bΔR2. Strikingly, DR2bΔR2 mice develop progressive EAE with pathology and clinical features observed in progressive MS patients. Adoptive transfer studies revealed that the clinical phenotype of EAE in DR2bΔR2 mice are largely dependent on TNFR2 expression in the CNS. Subsequently, we showed that DR2bΔR2 mice have a significant increase of lesions in the cerebellum, associated with reduced of oligodendrocyte progenitor cells (OPC) recruitment or function. Moreover, we showed that DR2bΔR2 mice are presented with chronic astrogliosis in demyelinating lesions. Our studies provide key insights into CNS repair and regulatory mechanisms controlled by TNFR2 during neuroinflammation and provide novel therapeutic strategies for treating progressive MS.

Integrin-Linked Kinase Controls Choroidal Neovascularization by Recruitment of Endothelial Progenitor Cells.

Vasculogenesis has been shown to contribute to the formation of choroidal neovascularization (CNV). However, the mechanism behind the recruitment of endothelial progenitor cells (EPC) to CNV is not well understood. Therefore, we were interested to know whether integrin-linked kinase (ILK) plays a role in recruiting EPC to CNV, and its possible mechanism. We investigated the effect of hypoxia on retinal pigment epithelium (RPE) cells expressing ILK, hypoxia-inducible factor 1α (HIF-1α), stromal-derived factor-1 (SDF-1), and vascular endothelial growth factor (VEGF), and we further examined the effect of ILK small interfering RNA (siRNA) on their expression. The function of ILK expressed by RPE on EPC in vitro with regard to angiogenic effect was also studied. In vivo, we determined the expression levels of the above factors in CNV. We also examined the role of ILK on their expression, on EPC recruiting, and on the growth of CNV. We found that hypoxia strongly induced the expression of ILK, HIF-1α, SDF-1, and VEGF. Moreover, the silencing of ILK attenuated their expression. It also decreased the phosphorylation of protein kinase B (PKB/AKT) and extracellular regulated protein kinases (ERK) and nearly abolished the proliferation, migration, and adhesion of EPC to RPE cells. In vivo, we showed that these factors were upregulated in CNV. Inhibiting the expression of ILK prohibited the "homing" of EPC to CNV lesions and attenuated the growth of CNV. We demonstrate that ILK controls the development of CNV by regulating the recruitment of EPC to CNV lesions, possibly through ILK-dependent expression of SDF-1 and VEGF in RPE.

Do platelets promote cardiac recovery after myocardial infarction: roles beyond occlusive ischemic damage.

Our understanding of platelet function has traditionally focused on their roles in physiological hemostasis and pathological thrombosis, with the latter being causative of vessel occlusion and subsequent ischemic damage to various tissues. In particular, numerous in vivo studies have implicated causative roles for platelets in the pathogenesis of ischemia-reperfusion (I/R) injury to the myocardium. However, platelets clearly have more complex pathophysiological roles, particularly as a result of the heterogeneous nature of biologically active cargo secreted from their granules or contained within released microparticles or exosomes. While some of these released mediators amplify platelet activation and thrombosis through autocrine or paracrine amplification pathways, they can also regulate diverse cellular functions within the localized microenvironment and recruit progenitor cells to the damage site to facilitate repair processes. Notably, there is evidence to support cardioprotective roles for platelet mediators during I/R injury. As such, it is becoming more widely appreciated that platelets fulfill a host of physiological and pathological roles beyond our basic understanding. Therefore, the purpose of this perspective is to consider whether platelets, through their released mediators, can assume a paradoxically beneficial role to promote cardiac recovery after I/R injury.

Treatment of stress urinary incontinence with low-intensity extracorporeal shock wave therapy in a vaginal balloon dilation induced rat model.

BackgroundTo investigate the outcomes and mechanisms of low-intensity extracorporeal shock wave therapy (Li-ESWT) on stress urinary incontinence (SUI) in a vaginal balloon dilation (VBD) rat model.MethodsThirty Sprague-Dawley rats were randomly grouped into normal controls, VBD only, and VBD with Li-ESWT. Li-ESWT was administered twice per week for 3 weeks. Afterward, all 30 rats were assessed with functional and histological studies. To explore the acute effect of Li-ESWT, another 25 rats, given intraperitoneal 5-ethynyl-2-deoxyuridine (EdU) at birth, were treated with Li-ESWT followed by assessment of vascular endothelial growth factor (VEGF) expression and endogenous progenitor cells distribution at 24 hours or 1 week after the last Li-ESWT therapy. Additionally, rat myoblast L6 cells were used for myotube formation assay in vitro.ResultsFunctional analysis with leak-point pressure (LPP) testing showed that rats treated with Li-ESWT following VBD had significantly higher LPP relative to those receiving VBD only (44.8±3.2 versus 27.0±2.9 cmH2O, P<0.01). Histological examinations showed increased urethral sphincter regeneration in Li-ESWT group. The rats treated with Li-ESWT also had increased vascularity, which was confirmed by immunohistochemistry of rat endothelial cell antigen, while reverse-transcriptase polymerase chain reaction (RT-PCR) showed VEGF expression was significantly enhanced. Additionally, there were significantly increased EdU+ cells in Li-ESWT treated rats at 24 hours. In vitro, Li-ESWT promoted myotube formation from L6 cells.ConclusionsLi-ESWT ameliorated SUI by promoting angiogenesis, progenitor cell recruitment, and urethral sphincter regeneration in a rat model induced by VBD. Li-ESWT represents a potential novel non-invasive therapy for SUI.

19. Anti-Angiogenic Therapy for Endometriosis

Endometriosis is a gynecological disease affecting 10 to 15% women. The disease is characterized by the growth of endometrium (lining of the uterus) outside of the uterine cavity. Women affected by this condition can experience symptoms that include pelvic pain, irregular bleeding, and infertility. One of the key requirements for endometriotic lesions to survive is to develop a blood supply to support their growth. Our laboratory is investigating mechanisms of how endometriotic lesions establish their blood supply and how neo-angiogenesis is regulated by endothelial and hematopoietic progenitor cells. Results from our laboratory showed that stromal cell derived factor-1 plays an important role in the recruitment of endothelial progenitor cells. Blocking of SDF-1 in a mouse model of endometriosis resulted in reduced lesion growth and survival. Studies are in progress to evaluate safety and efficacy of anti-angiogenic peptide, ABT-898, in an immunodeficient mouse model of endometriosis.

High-Fat Diet-Induced Obesity Promotes Expansion of Bone Marrow Adipose Tissue and Impairs Skeletal Stem Cell Functions in Mice.

Obesity represents a risk factor for development of insulin resistance and type 2 diabetes. In addition, it has been associated with increased adipocyte formation in the bone marrow (BM) along with increased risk for bone fragility fractures. However, little is known on the cellular mechanisms that link obesity, BM adiposity, and bone fragility. Thus, in an obesity intervention study in C57BL/6J mice fed with a high-fat diet (HFD) for 12 weeks, we investigated the molecular and cellular phenotype of bone marrow adipose tissue (BMAT), BM progenitor cells, and BM microenvironment in comparison to peripheral adipose tissue (AT). HFD decreased trabecular bone mass by 29%, cortical thickness by 5%, and increased BM adiposity by 184%. In contrast to peripheral AT, BMAT did not exhibit pro-inflammatory phenotype. BM progenitor cells isolated from HFD mice exhibited decreased mRNA levels of inflammatory genes (Tnfα, IL1β, Lcn2) and did not manifest an insulin resistant phenotype evidenced by normal levels of pAKT after insulin stimulation as well as normal levels of insulin signaling genes. In addition, BM progenitor cells manifested enhanced adipocyte differentiation in HFD condition. Thus, our data demonstrate that BMAT expansion in response to HFD exerts a deleterious effect on the skeleton. Continuous recruitment of progenitor cells to adipogenesis leads to progenitor cell exhaustion, decreased recruitment to osteoblastic cells, and decreased bone formation. In addition, the absence of insulin resistance and inflammation in the BM suggest that BMAT buffers extra energy in the form of triglycerides and thus plays a role in whole-body energy homeostasis. © 2018 The Authors. Journal of Bone and Mineral Research Published by Wiley Periodicals, Inc.

Delivery of VEGFA in bone marrow stromal cells seeded in copolymer scaffold enhances angiogenesis, but is inadequate for osteogenesis as compared with the dual delivery of VEGFA and BMP2 in a subcutaneous mouse model

BackgroundIn bone tissue engineering (BTE), extensive research into vascular endothelial growth factor A (VEGFA)-mediated angiogenesis has yielded inconsistent results. The aim of this study was to investigate the influence on angio- and osteogenesis of adenoviral-mediated delivery of VEGFA alone or in combination with bone morphogenetic protein 2 (BMP2) in bone marrow stromal cells (BMSC) seeded onto a recently developed poly(LLA-co-CL) scaffold.MethodsHuman BMSC were engineered to express VEGFA alone or in combination with BMP2 and seeded onto poly(LLA-co-CL) scaffolds. Changes in angiogenic and osteogenic gene and protein levels were examined by quantitative reverse-transcription polymerase chain reaction (RT-PCR), PCR array, and alkaline phosphatase assay. An in vivo subcutaneous mouse model was used to investigate the effect on angio- and osteogenesis of VEGFA alone or in combination with BMP2, using microcomputed tomography (μCT), histology, immunohistochemistry, and immunofluorescence.ResultsCombined delivery of a lower ratio (1:3) of VEGFA and BMP2 (ad-BMP2 + VEGFA) led to upregulation of osteogenic and angiogenic genes in vitro at 3 and 14 days, compared with mono-delivery of VEGFA (ad-VEGFA) and other controls. In vivo, in a subcutaneous mouse model, both ad-VEGFA and ad-BMP2 + VEGFA scaffold explants exhibited increased angiogenesis at 2 weeks. Enhanced angiogenesis was largely related to the recruitment and differentiation of mouse progenitor cells to the endothelial lineage and, to a lesser extent, to endothelial differentiation of the implanted BMSC. μCT and histological analyses revealed enhanced de novo bone formation only in the ad-BMP2 + VEGFA group, corresponding at the molecular level to the upregulation of genes related to osteogenesis, such as ALPL, RUNX2, and SPP1.ConclusionsAlthough BMSC expressing VEGFA alone or in combination with BMP2 significantly induced angiogenesis, VEGFA alone failed to demonstrate osteogenic activity both in vitro and in vivo. These results not only call into question the use of VEGFA alone in bone regeneration, but also highlight the importance in BTE of appropriately formulated combined delivery of VEGFA and BMP2.

Plasticity of human adipose-derived stem cells - relevance to tissue repair.

In contrast to cold blooded vertebrates, the ability to regenerate morphologically and functionally complex structures is limited in adult mammals. Recruitment of progenitor cells is a key step in the regenerative process. The possibility of repairing missing or diseased tissues in humans has been potentiated by the increasing understanding of somatic stem cells, their plasticity and the possibility of modulating it, that could be harnessed either to stimulate endogenous repair or to engineer the required tissue. Here, we focus on human mesenchymal stem cells (MSCs), important players in tissue homeostasis in healthy organisms, with a particular emphasis on those derived from the adipose tissue (ADSCs). While a mark of MSC identity is the ability to differentiate into osteoblasts, chondrocytes and adipocytes, there is evidence that their potential goes beyond these three mesenchymal lineages. We discuss some differentiation and modulatory properties of MSCs and provide an overview of our recent work on ADSCs from paediatric patients (pADSCs) that has shown their ability to give raise to non-mesenchymal cells, consistent with a significant plasticity. Finally, we present novel data indicating that both mesenchymal lineages (adipogenic, chondrogenic and osteogenic) and neural and epithelial lineages can originate from clonal lines that like the parental line express markers of pluripotency as well as the stromal cell marker, GREM1. Together these data support the existence of pADSC multipotent stem cells.

Cardiosphere-Derived Cells and Ischemic Heart Failure.

After a myocardial infarction, heart tissue becomes irreversibly damaged, leading to scar formation and inevitably ischemic heart failure. Of the many available interventions after a myocardial infarction, such as percutaneous intervention or pharmacological optimization, none can reverse the ischemic insult on the heart and restore cardiac function. Thus, the only available cure for patients with scarred myocardium is allogeneic heart transplantation, which comes with extensive costs, risks, and complications. However, multiple studies have shown that the heart is, in fact, not an end-stage organ and that there are endogenous mechanisms in place that have the potential to spark regeneration. Stem cell therapy has emerged as a potential tool to tap into and activate this endogenous framework. Particularly promising are stem cells derived from cardiac tissue itself, referred to as cardiosphere-derived cells (CDCs). CDCs can be extracted and isolated from the patient's myocardium and then administered by intramyocardial injection or intracoronary infusion. After early success in the animal model, multiple clinical trials have demonstrated the safety and efficacy of autologous CDC therapy in humans. Clinical trials with allogeneic CDCs showed early promising results and pose a potential "off-the-shelf" therapy for patients in the acute setting after a myocardial infarction. The mechanism responsible for CDC-induced cardiac regeneration seems to be a combination of triggering native cardiomyocyte proliferation and recruitment of endogenous progenitor cells, which most prominently occurs via paracrine effects. A further understanding of the mediators involved in paracrine signaling can help with the development of a stem cell-free therapy, with all the benefits and none of the associated complications.

Sitagliptin Accelerates Endothelial Regeneration after Vascular Injury Independent from GLP1 Receptor Signaling.

Introduction DPP4 inhibitors (gliptins) are commonly used antidiabetic drugs for the treatment of type 2 diabetes. Gliptins also act in a glucose-independent manner and show vasoregenerative effects. We have shown that gliptins can remarkably accelerate vascular healing after vascular injury. However, the underlying mechanisms remain unclear. Here, we examined potential signaling pathways linking gliptins to enhanced endothelial regeneration. Methods and Results We used wild-type and GLP1 receptor knockout (Glp1r−/−) mice to investigate the underlying mechanisms of gliptin-induced reendothelialization. The prototype DPP4 inhibitor sitagliptin accelerated endothelial healing in both animal models. Improved endothelial growth was associated with gliptin-mediated progenitor cell recruitment into the diseased vascular wall via the SDF1-CXCR4 axis independent of GLP1R-dependent signaling pathways. Furthermore, SDF1 showed direct proproliferative effects on endothelial cells. Excessive neointimal formation was not observed in gliptin- or placebo-treated Glp1r−/− mice. Conclusion We identified the SDF1-CXCR4 axis as a crucial signaling pathway for endothelial regeneration after acute vascular injury. Furthermore, SDF1 can directly increase endothelial cell proliferation. Gliptin-mediated potentiation of endothelial regeneration was preserved in Glp1r−/− animals. Thus, gliptin-mediated endothelial regeneration proceeds through SDF-1/CXCR4 in a GLP1R-independent manner after acute vascular injury.

Calcium-containing scaffolds induce bone regeneration by regulating mesenchymal stem cell differentiation and migration

BackgroundOsteoinduction and subsequent bone formation rely on efficient mesenchymal stem cell (MSC) recruitment. It is also known that migration is induced by gradients of growth factors and cytokines. Degradation of Ca2+-containing biomaterials mimics the bone remodeling compartment producing a localized calcium-rich osteoinductive microenvironment. The aim of our study was to determine the effect of calcium sulfate (CaSO4) on MSC migration. In addition, to evaluate the influence of CaSO4 on MSC differentiation and the potential molecular mechanisms involved.MethodsA circular calvarial bone defect (5 mm diameter) was created in the parietal bone of 35 Balb-C mice. We prepared and implanted a cell-free agarose/gelatin scaffold alone or in combination with different CaSO4 concentrations into the bone defects. After 7 weeks, we determined the new bone regenerated by micro-CT and histological analysis. In vitro, we evaluated the CaSO4 effects on MSC migration by both wound healing and agarose spot assays. Osteoblastic gene expression after BMP-2 and CaSO4 treatment was also evaluated by qPCR.ResultsCaSO4 increased MSC migration and bone formation in a concentration-dependent manner. Micro-CT analysis showed that the addition of CaSO4 significantly enhanced bone regeneration compared to the scaffold alone. The histological evaluation confirmed an increased number of endogenous cells recruited into the cell-free CaSO4-containing scaffolds. Furthermore, MSC migration in vitro and active AKT levels were attenuated when CaSO4 and BMP-2 were in combination. Addition of LY294002 and Wortmannin abrogated the CaSO4 effects on MSC migration.ConclusionsSpecific CaSO4 concentrations induce bone regeneration of calvarial defects in part by acting on the host’s undifferentiated MSCs and promoting their migration. Progenitor cell recruitment is followed by a gradual increment in osteoblast gene expression. Moreover, CaSO4 regulates BMP-2-induced MSC migration by differentially activating the PI3K/AKT pathway. Altogether, these results suggest that CaSO4 scaffolds could have potential applications for bone regeneration.

Myocardial reparative functions of exosomes from mesenchymal stem cells are enhanced by hypoxia treatment of the cells via transferring microRNA-210 in an nSMase2-dependent way

Hypoxia treatment enhances paracrine effect of mesenchymal stem cells (MSCs). The aim of this study was to investigate whether exosomes from hypoxia-treated MSCs (ExoH) are superior to those from normoxia-treated MSCs (ExoN) for myocardial repair. Mouse bone marrow-derived MSCs were cultured under hypoxia or normoxia for 24 h, and exosomes from conditioned media were intramyocardially injected into infarcted heart of C57BL/6 mouse. ExoH resulted in significantly higher survival, smaller scar size and better cardiac functions recovery. ExoH conferred increased vascular density, lower cardiomyocytes (CMs) apoptosis, reduced fibrosis and increased recruitment of cardiac progenitor cells in the infarcted heart relative to ExoN. MicroRNA analysis revealed significantly higher levels of microRNA-210 (miR-210) in ExoH compared with ExoN. Transfection of a miR-210 mimic into endothelial cells (ECs) and CMs conferred similar biological effects as ExoH. Hypoxia treatment of MSCs increased the expression of neutral sphingomyelinase 2 (nSMase2) which is crucial for exosome secretion. Blocking the activity of nSMase2 resulted in reduced miR-210 secretion and abrogated the beneficial effects of ExoH. In conclusion, hypoxic culture augments miR-210 and nSMase2 activities in MSCs and their secreted exosomes, and this is responsible at least in part for the enhanced cardioprotective actions of exosomes derived from hypoxia-treated cells.

Inhibition of Cyclooxygenase-2 Suppresses the Recruitment of Endothelial Progenitor Cells in the Microvasculature of Endometriotic Lesions

The incorporation of endothelial progenitor cells (EPCs) into newly developing blood vessels contributes to the vascularization of endometriotic lesions. We analyzed whether cyclooxygenase (COX)-2 signaling regulates this vasculogenic process. Endometriotic lesions were surgically induced in irradiated FVB/N mice, which were reconstituted with bone marrow from FVB/N-TgN [Tie2/green fluorescent protein (GFP)] 287 Sato mice. The animals received β-estradiol 17-valerate once a week and were treated daily with the selective COX-2 inhibitor parecoxib (25 mg/kg) or vehicle (control) for 7 and 28 days. Analyses involved the determination of lesion growth, cyst formation, homing of GFP+/Tie2+ EPCs, numbers of circulating EPCs, vascularization, cell proliferation, apoptosis, and immune cell infiltration by means of high-resolution ultrasonography, caliper measurements, flow cytometry, histologic analysis, and immunohistochemical analysis. In parecoxib-treated mice, blood circulating EPCs were higher, but numbers of recruited EPCs in endometriotic lesions were significantly lower when compared with controls. This finding was associated with an impaired early vascularization and stromal tissue growth as well as reduced glandular secretory activity of the lesions. Parecoxib-treated lesions further contained less proliferating and more apoptotic cells and exhibited lower numbers of infiltrating macrophages and neutrophilic granulocytes. These findings demonstrate that the inhibition of COX-2 suppresses vasculogenesis in endometriotic lesions, which may contribute to an impaired lesion vascularization and growth.

Distinct NG2 proteoglycan-dependent roles of resident microglia and bone marrow-derived macrophages during myelin damage and repair.

We used a bone marrow transplantation approach to distinguish the activities of bone marrow-derived macrophages from the activities of central nervous system-resident microglia in phenomena associated with axon demyelination and remyelination. We transplanted wild type or germline NG2 null beta-actin-EGFP expressing bone marrow into irradiated wild type or NG2 null recipient mice, followed by analysis of lysolecithin-induced spinal cord demyelination and remyelination and quantification of Iba-1+/ F4/80+/ EGFP+ macrophages and Iba-1+/ F4/80+/ EGFP- microglia. One week after microinjection of 1% lysolecithin into the spinal cord, wild type recipients receiving NG2 null bone marrow exhibit greatly reduced infiltration of macrophages into lesions, compared to wild type recipients receiving wild type bone marrow. Wild type bone marrow recipients also exhibit larger numbers of demyelinated axons than NG2 null recipients, indicative of macrophage participation in the initial myelin damage. However, wild type bone marrow recipients also exhibit superior myelin repair at 6 weeks post-injury, compared to NG2 null bone marrow recipients, demonstrating the additional importance of macrophages in remyelination. Incompletely repaired lesions in NG2 null bone marrow recipients at 6 weeks post-injury retain elevated numbers of macrophages, in contrast to lower numbers of macrophages in more completely repaired lesions in wild type bone marrow recipients. This suggests that NG2 expression renders macrophages more effective in myelin repair and less likely to promote chronic inflammation. Effective macrophage involvement in myelin repair is due in part to effects on the proliferation and/or recruitment of oligodendrocyte progenitor cells. Reduced numbers of oligodendrocyte progenitors are seen in lesions in NG2 null bone marrow recipients, likely due to deficits in macrophage production of oligodendrocyte progenitor-relevant mitogens and in phagocytosis of inhibitory myelin debris. Microglia also appear to be important for clearance of myelin debris, as indicated by reduced phagocytosis in NG2 null recipients receiving wild type bone marrow.