Migration Of BMSCs Research Articles

Tryptophan and Kynurenine Enhances the Stemness and Osteogenic Differentiation of Bone Marrow-Derived Mesenchymal Stromal Cells In Vitro and In Vivo.

Aging tissues present a progressive decline in homeostasis and regenerative capacities, which has been associated with degenerative changes in tissue-specific stem cells and stem cell niches. We hypothesized that amino acids could regulate the stem cell phenotype and differentiation ability of human bone marrow-derived mesenchymal stromal cells (hBMSCs). Thus, we performed a screening of 22 standard amino acids and found that D-tryptophan (10 μM) increased the number of cells positive for the early stem cell marker SSEA-4, and the gene expression levels of OCT-4, NANOG, and SOX-2 in hBMSCs. Comparison between D- and L-tryptophan isomers showed that the latter presents a stronger effect in inducing the mRNA levels of Oct-4 and Nanog, and in increasing the osteogenic differentiation of hBMSCs. On the other hand, L-tryptophan suppressed adipogenesis. The migration and colony-forming ability of hBMSCs were also enhanced by L-tryptophan treatment. In vivo experiments delivering L-tryptophan (50 mg/kg/day) by intraperitoneal injections for three weeks confirmed that L-tryptophan significantly increased the percentage of cells positive for SSEA-4, mRNA levels of Nanog and Oct-4, and the migration and colony-forming ability of mouse BMSCs. L-kynurenine, a major metabolite of L-tryptophan, also induced similar effects of L-tryptophan in enhancing stemness and osteogenic differentiation of BMSCs in vitro and in vivo, possibly indicating the involvement of the kynurenine pathway as the downstream signaling of L-tryptophan. Finally, since BMSCs migrate to the wound healing site to promote bone healing, surgical defects of 1 mm in diameter were created in mouse femur to evaluate bone formation after two weeks of L-tryptophan or L-kynurenine injection. Both L-tryptophan and L-kynurenine accelerated bone healing compared to the PBS-injected control group. In summary, L-tryptophan enhanced the stemness and osteoblastic differentiation of BMSCs and may be used as an essential factor to maintain the stem cell properties and accelerate bone healing and/or prevent bone loss.

Asprin-loaded strontium-containing α-calcium sulphate hemihydrate/nano-hydroxyapatite composite promotes regeneration of critical bone defects.

Our laboratory originally synthesized strontium(Sr)‐containing α‐calcium sulphate hemihydrate/nano‐hydroxyapatite composite (Sr‐α‐CSH/n‐HA) and demonstrated its ability to repair critical bone defects. This study attempted to incorporate aspirin into it to produce a better bone graft material for critical bone defects. After 5% Sr‐α‐CSH was prepared by coprecipitation and hydrothermal methods, it was mixed with aspirin solution of different concentrations (50 μg/ml, 200 μg/ml, 800 μg/ml and 3200 μg/ml) at a fixed liquid‐solid ratio (0.54 v/w) to obtain aspirin‐loaded Sr‐α‐CSH/n‐HA composite. In vitro experiments were performed on the composite extracts. The tibial defects (3 mm*5 mm) in SD rat model were filled with the composite for 4 weeks and 12 weeks to evaluate its osteogenic capacity in vivo. Our results showed its capability of proliferation, migration and osteogenesis of BMSCs in vitro got improved. In vivo treatment with 800 μg/ml aspirin–loaded Sr‐α‐CSH/n‐HA composite led to significantly more new bone formation in the defects compared with Sr‐α‐CSH/n‐HA composite and significantly promoted the expression of osteogenic‐related genes and inhibited osteoclast activity. In general, our research suggests that aspirin‐loaded Sr‐α‐CSH/n‐HA composite may have a greater capacity of repairing tibial defects in SD rats than simple Sr‐α‐CSH/n‐HA composite.

HIF/Ca2+/NO/ROS is critical in roxadustat treating bone fracture by stimulating the proliferation and migration of BMSCs

AimsFracture site is regionally hypoxic resulting from vasculature disruption. HIF-1αplays an essential role in fracture repair. This study aims to investigate the influence of FG4592 on the femur fracture of SD rats and the proliferation, migration of BMSCs. Materials and methodsAfter the femoral fracture model was established, computed tomography imaging and histological analyses were used to quantify bone healing and the expression of CD90, HIF-1α, VEGF were observed by means of immunohistochemistry method on Day 10 and Day 20. In addition, CCK-8 assay, transwell, flow cytometric analysis, laser confocal microscopy assay, western blot and rT-PCR were performed to text the proliferation and migration of BMSCs using FG4592. Key findingsIn vivo, FG4592 facilitated the repair of bone fracture by increasing the number of BMSCs and cartilage formation. In vitro, FG4592 markedly improved the proliferation, migration of BMSCs via upregulation of intracellular Ca2+, NO and concomitant decrease of ROS. Gene silencing of HIF-1α resulted in the opposite phenomenon in BMSCs with the treatment of FG4592. SignificanceThe transplantation of BMSCs is the most promising candidate for the treatment of fracture non-union. We illustrated that FG4592 promoted the proliferation, migration of BMSCs via the HIF/Ca2+/NO/ROS pathway and further accelerated fracture healing. These results provide a deeper understanding for the mechanism of HIF in promoting fracture healing.

CREB activation affects mesenchymal stem cell migration and differentiation in periodontal tissues due to orthodontic force.

During the orthodontic tooth movement, cells in periodontal ligament could differentiate into osteoblasts to synthesize alveolar bone as well as affect the proliferation, migration and differentiation of mesenchymal stem cells, which also contribute to bone remodeling. However, the mechanism is still largely elusive. Here, we evaluated the expression of CREB at the tension site of mouse periodontal ligament under orthodontic mechanical strain and in the cyclic tension strain treated human periodontal ligament cells. Then, through gain and loss of function analysis, we revealed that CREB in PDLCs promotes SDF-1 and FGF2 secretion, which enhance the migration and osteoblastic differentiation of BMSCs. We further discovered that CREB transcriptionally activates FGF2 and SDF-1 expressions by binding to the promoter regions.In conclusion, this study confirms that CREB is an upregulated gene in periodontal ligament under orthodontic tension strain stimulation and plays an important role in regulating BMSCs' physiological activity in orthodontic tension strain-induced bone formation.

Bone mesenchymal stem cells are recruited via CXCL8-CXCR2 and promote EMT through TGF-β signal pathways in oral squamous carcinoma.

ObjectivesBone mesenchymal stem cells (BMSCs) play critical roles in tumour microenvironment. However, molecular mechanisms of how BMSCs to be recruited and effect subsequent tumour progression are poorly understood in oral squamous cell carcinoma (OSCC).Materials and MethodsThe distribution of CXCL8 was detected by immunohistochemical staining in OSCC tissues. The chemotaxis of conditioned media from different epithelial cells to BMSCs was examined by trans‐well assay. Real‐time quantitative PCR (qPCR) and ELISA were used to detect the expression of related cytokines and chemokine receptors. The migration of BMSCs was observed in BALB/c nude mice. The roles of BMSCs in proliferation, migration and invasion of OSCC were detected by CCK‐8, flow cytometry and trans‐well assay. Epithelial‐mesenchymal transition (EMT)–related markers were analysed by qPCR and Western blot in vitro, and growth was evaluated in BALB/c nude mice using subcutaneously implanted OSCC in nude mouse model in vivo.ResultsUsing OSCC, we show CXCL8, secreted by OSCC, binds to exclusively CXCR2 in BMSCs to facilitate migration of BMSCs to OSCC. TGF‐β secreted by BMSCs subsequently induces EMT of OSCC to promote their proliferation, migration and infiltration. We also showed that the Ras/Raf/Erk axis plays a critical role in tumour progression.ConclusionsOur results provide the molecular basis for BMSC recruitment into tumours, and how this process leads to tumour progression and leads us to develop a novel OSCC treatment target.

M2 macrophages promote vasculogenesis during retinal neovascularization by regulating bone marrow-derived cells via SDF-1/VEGF.

Macrophages promote vasculogenesis during retinal neovascularization (RNV) by increasing the recruitment and differentiation of bone marrow-derived cells (BMCs). Different subtypes of macrophages (M1 and M2 macrophages) are associated with RNV. However, the mechanism underlying the regulation of BMCs by different macrophage subtypes during RNV remains unclear. In the present study, we investigated the role and mechanism of action of different macrophage subtypes that regulate BMCs during the development of RNV. The retinal avascular area and neovascularization (NV) tuft area in M2 macrophage group in vivo were the largest compared to those in the control phosphate buffer saline (PBS), unpolarized-M0, and M1 macrophage groups. The number of recruited green fluorescent protein (GFP)-positive BMCs and the degree of differentiation of BMCs into CD31-positive endothelial cells (ECs) and alpha-smooth muscle actin (α-SMA)-positive smooth muscle cells (SMCs) were higher in the M2 macrophage group than in the other groups. M2-conditional medium (M2-CM) affected the in vitro migration and activation of bone marrow mesenchymal stem cells (BMSCs, a subset of BMCs) more than M1-CM. The expression of stromal cell-derived factor-1 (SDF-1) and vascular endothelial growth factor (VEGF) in M2 macrophages and BMSCs cultured with M2-CM was also higher than that in M1 macrophages and BMSCs cultured with M1-CM. Migration of BMSCs was reduced after inhibiting the SDF-1 signaling pathway. Our results indicate that M2 macrophages may express significantly higher levels of SDF-1 and VEGF than M1 macrophages, thus regulating the recruitment and differentiation of BMCs and further aggravating vasculogenesis during RNV.

Recombinant human BMP-2 accelerates the migration of bone marrow mesenchymal stem cells via the CDC42/PAK1/LIMK1 pathway in vitro and in vivo.

Biomaterials are widely used for bone regeneration and fracture repair. The migration of bone marrow mesenchymal stem cells (BMSCs) into bone defect sites or material implantation sites, and their differentiation into osteoblasts, is central to the fracture healing process, and the directional migration of BMSCs depends on cytokines or chemokines at the defect site. BMP-2 can stimulate the migration of a variety of cells, but it remains unclear whether BMSC migration can be induced. To provide evidence for BMP-2-induced BMSC migration, we tested the cytoskeletal changes and migration ability of BMSCs after treatment with recombinant human BMP-2 (rhBMP-2). We also explored the recruitment of BMSCs from the circulatory system using a collagen sponge incorporating rhBMP-2 that was implanted in vivo. Furthermore, to understand the mechanism underlying this migration, we investigated the effect of rhBMP-2 on migration-related signal pathways. Here, we found that, rhBMP-2 treatment significantly increased the migration of BMSCs in vitro via activation of the CDC42/PAK1/LIMK1 pathway, and that this migration could be blocked by silencing CDC42. In vivo, collagen sponge material loaded with rhBMP-2 could recruit BMSCs injected into the circulatory system. Moreover, inhibition using the small interfering RNA for CDC42 led to a significant decrease in the number of BMSCs within the material. In conclusion, our data prove that rhBMP-2 can accelerate BMSC migration via the CDC42/PAK1/LIMK1 pathway both in vivo and in vitro, and therefore provides a foundation for further understanding and application of rhBMP-2-incorporated materials by enhancing BMSC recruitment.

3D printed scaffolds of calcium silicate-doped \u03b2-TCP synergize with co-cultured endothelial and stromal cells to promote vascularization and bone formation

Synthetic bone scaffolds have potential application in repairing large bone defects, however, inefficient vascularization after implantation remains the major issue of graft failure. Herein, porous β-tricalcium phosphate (β-TCP) scaffolds with calcium silicate (CS) were 3D printed, and pre-seeded with co-cultured human umbilical cord vein endothelial cells (HUVECs) and human bone marrow stromal cells (hBMSCs) to construct tissue engineering scaffolds with accelerated vascularization and better bone formation. Results showed that in vitro β-TCP scaffolds doped with 5% CS (5%CS/β-TCP) were biocompatible, and stimulated angiogenesis and osteogenesis. The results also showed that 5%CS/β-TCP scaffolds not only stimulated co-cultured cells angiogenesis on Matrigel, but also stimulated co-cultured cells to form microcapillary-like structures on scaffolds, and promoted migration of BMSCs by stimulating co-cultured cells to secrete PDGF-BB and CXCL12 into the surrounding environment. Moreover, 5%CS/β-TCP scaffolds enhanced vascularization and osteoinduction in comparison with β-TCP, and synergized with co-cultured cells to further increase early vessel formation, which was accompanied by earlier and better ectopic bone formation when implanted subcutaneously in nude mice. Thus, our findings suggest that porous 5%CS/β-TCP scaffolds seeded with co-cultured cells provide new strategy for accelerating tissue engineering scaffolds vascularization and osteogenesis, and show potential as treatment for large bone defects.

Dynamics of early stem cell recruitment in skin flaps subjected to ischemia reperfusion injury

ObjectiveBone marrow-derived stromal cell (BMSCs) therapy improves survival of skin flaps subject to ischemia/reperfusion (I/R) injury. However, very little is known about the trafficking and distribution of BMSCs in post-ischemic skin tissue following intravenous administration. The aim of this study was to assess the behavior of BMSCs in post-ischemic skin flaps and to compare the magnitude and kinetics of accumulation of BMSCs and leukocytes following I/R. MethodsCutaneous flaps perfused by the inferior epigastric vessels were created in C57Bl6 mice. The flaps were subjected to 3.5h of ischemia followed by reperfusion. Wound healing and vascular perfusion were assessed in 3 groups of mice (sham, I/R, and I/R+BMSCs treatment) on days 3, 5, 7 and 14 post-reperfusion. The kinetics and magnitude of BMSCs and leukocyte recruitment were quantified in additional 2 groups (Sham and I/R) after I/R using intravital fluorescence microscopy at 2 and 4h after the intravenous injection of fluorescently labeled BMSCs. ResultsWound healing after I/R was significantly enhanced in skin flaps of mice treated with BMSCs, compared to controls. The rolling velocity of BMSCs was higher compared to leukocytes both in control mice (32.4±3.7μm/s vs 24.0±2.2μm/s, p<0.05) and in I/R mice (34.6±3.8μm/s vs 20.2±2.3μm/s, p<0.005). However, the rolling velocity of both cell populations was not altered by I/R. The firm adhesion and transendothelial migration of BMSCs did not differ from the values detected for leukocytes for both control and I/R mice. ConclusionsThe magnitude and kinetics of BMSCs recruitment in skin flaps subjected to I/R are not significantly different from the responses noted for leukocytes, suggesting that similar mechanisms may be involved in the recruitment of both cell populations following I/R.

Functional Self-Assembled Peptide Nanofibers for Bone Marrow Mesenchymal Stem Cell Encapsulation and Regeneration in Nucleus Pulposus.

Low back pain (LBP) is mainly caused by intervertebral disc degeneration (IDD). Recent studies have demonstrated that the transplantation of mesenchymal stem cells (MSCs) can regenerate regions that have undergone degeneration, and the regenerative effect can be enhanced by using a hydrogel carrier. This article describes an injectable functional hydrogel system manufactured by combining RADA16-I and RADA-KPSS (RADA-KPSS was manufactured by conjugating a bioactive motif derived from BMP-7 [KPSS] onto the C terminal of RADA16-I) at a volume ratio of 1:1. This hydrogel system can enhance the proliferation, differentiation, and chemotactic migration of BMSCs. In addition, the encapsulation of BMSCs with this system maintains cell viability for a long period after transplantation into an ex vivo cultured disc model. In conclusion, KPSS-conjugated RADKPS is an ideal encapsulation system for BMSCs in intervertebral disc (IVD) regeneration.

Cyclic mechanical stretching promotes migration but inhibits invasion of rat bone marrow stromal cells

Bone marrow stromal cells (BMSCs, also broadly known as bone marrow-derived mesenchymal stem cells) are multipotent stem cells that have a self-renewal capacity and multilineage differentiation potential. Mechanical stretching plays a vital role in regulating the proliferation and differentiation of BMSCs. However, little is known about the effects of cyclic stretching on BMSC migration and invasion. In this study, using a custom-made cell-stretching device, we studied the effects of cyclic mechanical stretching on rat BMSC migration and invasion using a Transwell Boyden Chamber. The protein secretion of matrix metalloproteinase-2 (MMP-2) and matrix metalloproteinase-9 (MMP-9) was detected by gelatin zymography, and the activation of focal adhesion kinase (FAK) and extracellular signal regulated kinase1/2 (ERK1/2) was measured by western blot. We found that cyclic mechanical stretching with 10% amplitude at 1Hz frequency for 8h promotes BMSC migration, but reduces BMSC invasion. FAK and ERK1/2 signals were activated in BMSCs after exposure to cyclic stretching. In the presence of the FAK phosphorylation blocker PF573228 or the ERK1/2 phosphorylation blocker PD98059, the cyclic-stretch-promoted migration of BMSCs was completely suppressed. On the other hand, cyclic mechanical stretching reduced the secretion of MMP-2 and MMP-9 in BMSCs, and PF573228 suppressed the cyclic-stretch-reduced secretion of MMP-2 and MMP-9. The decrease of BMSC invasion induced by mechanical stretching is partially restored by PF573228 but remained unaffected by PD98059. Taken together, these data show that cyclic mechanical stretching promotes BMSC migration via the FAK-ERK1/2 signalling pathway, but reduces BMSC invasion by decreasing secretion of MMP-2 and MMP-9 via FAK, independent of the ERK1/2 signal.

Radially oriented collagen scaffold with SDF-1 promotes osteochondral repair by facilitating cell homing

The migration of cells from the side and the bottom of the defect is important in osteochondral defect healing. Here, we designed a novel collagen scaffold that possessed channels in both the horizontal and the vertical directions, along with stromal cell-derived factor-1 (SDF-1) to enhance osteochondral regeneration by facilitating cell homing. Firstly we fabricated the radially oriented and random collagen scaffolds, then tested their properties. The radially oriented collagen scaffold had better mechanical properties than the random scaffold, but both supported cell proliferation well. Then we measured the migration of BMSCs in the scaffolds in vitro. The radially oriented collagen scaffold effectively promoted their migration, and this effect was further facilitated by addition of SDF-1. Moreover, we created osteochondral defects in rabbits, and implanted them with random or oriented collagen scaffolds with or without SDF-1, and evaluated cartilage and subchondral bone regeneration at 6 and 12 weeks after surgery. Cartilage regeneration with both the radially oriented scaffold and SDF-1 effectively promoted repair of the cartilage defect. Our results confirmed that the implantation of the radially oriented channel collagen scaffold with SDF-1 could be a promising strategy for osteochondral repair.

Matrix metalloproteinase-9 is up-regulated by CCL19/CCR7 interaction via PI3K/Akt pathway and is involved in CCL19-driven BMSCs migration

C–C chemokine receptor 7 (CCR7) and its ligands CCL19 contributes to the directional migration of certain cancer cell lines, but its role in the migration of BMSCs remains vague. The aim of this study was to determine the possible interaction between CCL19-induced conditions and matrix metalloproteinases-9 (MMP9) expression in BMSCs. Cell migration using Transwell assay indicated that activation of CCR7 by its specific ligand, exogenous chemokine ligand 19 (CCL19), was associated with a significant linear increase. Western blot and real-time PCR indicated that CCL19/CCR7 significantly upregulated expression of MMP9, which is related to metastasis-associated genes. The CCL19/CCR7 interaction significantly enhanced phosphorylation of Akt, as measured by Western blot. P-Akt and MMP9 protein expression exhibited a time-dependent pattern, and the peak was at 48h. LY294002 significantly abolished the effects of exogenous CCL19. These results suggest that CCL19/CCR7 contributes to the migration of BMSCs by upregulating MMP9 potentially via the PI3K/Akt pathway.

Decreased bone marrow stromal cells activity involves in unilateral anterior crossbite-induced early subchondral bone loss of temporomandibular joints

ObjectiveSubchondral bone loss in mandibular condyles was reported to be induced by experimentally created unilateral anterior crossbite (UAC) which altered the occlusal load distribution and hereafter the temporomandibular joint (TMJ) remodelling process. However, the initial cellular responses are poorly understood. In the present study, changes in osteoblast and osteoclast activities in TMJ subchondral bone were investigated using the rats treated with UAC. DesignForty rats were randomly divided into UAC and control groups, and sampled at 2 weeks after the operation. Subchondral bone loss was evaluated by micro-CT. Osteoclast and osteoblast activities were analyzed by real-time PCR. The osteoblast differentiation of the locally isolated BMSCs from TMJ subchondral bone was assessed by Alizarin red staining. The migration of BMSCs was detected by transwell assays. ResultsCompared with the age-matched controls, TMJ subchondral bone loss was observed in the UAC-treated rats (p<0.05). The osteoblast activity evaluated by real-time PCR and osteoblast number revealed by immunohistochemical staining were reduced in the TMJ subchondral bone of UAC rats (p<0.05), and the capability of proliferation, migration and osteoblast differentiation were all decreased in the locally isolated BMSCs from the UAC group (p<0.05). ConclusionsThe present data demonstrated an involvement of reduced BMSCs activity in the initiation of the mandibular subchondral bone loss at the early stage of installation of the aberrant prostheses.

SAT0169 Impaired migration and proliferation of bone marrow-derived mesenchymal stem cells from patients with systemic lupus erythematosus were mediated by IKK-β

BackgroundSystemic lupus erythematosus (SLE) is an autoimmune disease characterized by multi-organ involvements and a wide array of clinical manifestations. There is increasing evidence that dysregulation of stem cells is involved...

Oxidized phosphatidylcholine induces migration of bone marrow-derived mesenchymal stem cells through Krüppel-like factor 4-dependent mechanism

Platelet-activating factor (PAF; 1-O-alkyl-2-acetyl-sn-glycero-3-phosphorylcholine) and PAF-like oxidized phospholipids including 1-palmitoyl-2-oxovaleroyl-sn-glycero-3-phosphorylcholine (POVPC) are generated upon LDL oxidation. The aim of this study was to evaluate the question of whether POVPC can regulate migration of human bone marrow-derived stem cells (hBMSCs) and to characterize signaling mechanisms involved in the POVPC-induced cell migration. POVPC treatment resulted in dose- and time-dependent increase of hBMSCs migration. Treatment of cells with BN52021, a specific antagonist of PAF receptor, completely blocked cell migration induced by not only PAF but also POVPC. Silencing of endogenous PAF receptor expression using PAF receptor-specific small interfering RNA resulted in significant attenuation of cell migration induced by PAF or POVPC. Both PAF and POVPC induced expression of Krüppel-like factor 4 (KLF4) in hBMSCs. POVPC- or PAF-induced cell migration was abrogated by small interfering RNA-mediated depletion of endogenous KLF4. These results suggest that PAF receptor plays a pivotal role in POVPC-induced migration of human BMSCs through PAF receptor-mediated expression of KLF4.

The change of cytokines in tear and blood after different pterygium operation

Pterygium is an invasion of altered ocular tissue into the cornea. Bone marrow-derived stem cells have been reported to be involved in wound healing under chemotactic factors after pterygium removal and pain may act as a trigger signal. We evaluated the change of systemic and local chemotactic factors that could affect the mobilization and migration of BMSCs to the wound bed after conventional bare sclera pterygium excision. We also applied temporary amniotic membrane patch after pterygium removal, and compared the changes of cytokines with those of conventional bare sclera excision group. Substance-P (SP), vascular endothelial growth factor (VEGF), and stem cell factor (SCF) were measured in plasma and tear using ELISA and migrating CD34 + cells by flow cytometry. The results showed that post-operative pain was much reduced ( p < 0.05), and SP, VEGF and SCF kept consistently lower levels in plasma after temporary amniotic membrane application. Circulating CD34 + cells increased slightly in the temporary amniotic membrane patch group compared with marked increase in the bare sclera group. Thus, the application of a temporary amniotic membrane after pterygium removal might be an effective therapeutic means by controlling pain and excessive infiltration of bone marrow-derived stem cells.

Survival, migration and differentiation of allogenic BMSCs in injured spinal cord in rats

Objective To observe survival, migration and differentiation of marrow derived mes-enehymal stem cells (BMSCs) after transplantation into the injured spinal cord in rats. Methods Allo-genie BMSCs stained with Hoechst were transplanted into the spinal cord that had been semi-transected in rats. Two months later, the animals were killed to observe the survival, migration and differentiation of the BMSCs.Results Two months after transplantation, a dense mass of the transplanted Hoechst stained BMSCs was observed surrounding the injection spot and transferred into the center of injury site. lees than 10% of the transplanted BMSCs expressed the mark proteins of neurons and astroeytes. Conclusion Allogenic BMSCs can well survive, proliferate and transfer in the injured spinal cord in rats and a few of them can differentiate into neural-like cells. Key words: Bone marrow cells; Spinal cord injury; Transplantaion; Survival; Differenti-ation