Experiments have previously demonstrated the therapeutic potential of mobilized dental pulp stem cells (MDPSCs) for complete pulp regeneration. The aim of the present pilot clinical study is to ...assess the safety, potential efficacy, and feasibility of autologous transplantation of MDPSCs in pulpectomized teeth.
Five patients with irreversible pulpitis were enrolled and monitored for up to 24 weeks following MDPSC transplantation. The MDPSCs were isolated from discarded teeth and expanded based on good manufacturing practice (GMP). The quality of the MDPSCs at passages 9 or 10 was ascertained by karyotype analyses. The MDPSCs were transplanted with granulocyte colony-stimulating factor (G-CSF) in atelocollagen into pulpectomized teeth.
The clinical and laboratory evaluations demonstrated no adverse events or toxicity. The electric pulp test (EPT) of the pulp at 4 weeks demonstrated a robust positive response. The signal intensity of magnetic resonance imaging (MRI) of the regenerated tissue in the root canal after 24 weeks was similar to that of normal dental pulp in the untreated control. Finally, cone beam computed tomography demonstrated functional dentin formation in three of the five patients.
Human MDPSCs are safe and efficacious for complete pulp regeneration in humans in this pilot clinical study.
► CXCR7 agonists suppress tube formation in HUVEC. ► Stimulation of CXCR7 regulates down-stream signal from CXCR4. ► The expression of CXCR4 protein is regulated by CXCR7.
The CXCL12/CXCR4 axis is ...involved in many cellular responses for host homeostasis, and malfunction of this signaling pathway is associated with a variety of diseases. It is now known that CXCL12 also binds to another newly identified chemokine receptor, CXCR7, which does not couple with a G-protein. CXCR7 can form homodimers, or heterodimers with CXCR4, and is believed to sequester the chemokine CXCL12, although the CXCL12/CXCR7 axis activates MAP kinases through β-arrestin. Therefore, it has not been well defined how CXCR7 activation affects CXCL12-induced cellular events. To elucidate the function of CXCR7, we prepared CXCR7 agonist Compound 1. Compound 1 is a selective and potent CXCR7 agonist that clearly has the activity to recruit β-arrestin toward CXCR7. It also activates MAP kinases Akt and ERK. Using this compound, we confirmed that the CXCR7 agonist, but not an antagonistic antibody, did inhibit CXCL12 induced HUVEC tube formation, suggesting that activation of CXCR7 ameliorates CXCL12 induced cellular events, probably by affecting on CXCR4 function. We show that β-arrestin recruitment to CXCR4 is reduced by over-expression of CXCR7 and activation of CXCR7 by agonist treatment reduces the protein level of CXCR4. Based on our results, together with reported information, we propose that CXCR7, when up-regulated upon inflammation, can act as a negative regulator of CXCR4 by heterodimerizing with CXCR4, inducing its internalization and degradation. This mechanism suggests that CXCR7 agonists can have a therapeutic effect on CXCL12 causing diseases by countering the effects of CXCL12.
Background:Dietary intake of ω3 polyunsaturated fatty acids (ω3-PUFAs) reduces progression of atherosclerosis and prevents future cardiovascular events. Macrophages are key players in the ...pathogenesis of aortic aneurysm. The effects of ω3-PUFAs on abdominal aortic aneurysm (AAA) formation and macrophage-mediated inflammation remain unclear.Methods and Results:The AAA model was developed by angiotensin II infusion in apolipoprotein E-deficient mice. Mice were supplemented with eicosapentaenoic acid (EPA) or docosahexaenoic acid (DHA). The development of AAA lesions and macrophage infiltration in the aorta were analyzed. Gene expression of inflammatory markers in aortic tissues and peritoneal macrophages were measured by using quantitative polymerase chain reaction. AAA formation and macrophage infiltration were significantly suppressed after EPA and DHA administration. EPA administration and DHA administration significantly decreased the expression of tumor necrosis factor-α, monocyte chemoattractant protein-1, transforming growth factor-β, matrix metalloproteinases (MMP)-2, MMP-9, and vascular cell adhesion molecule-1 in the aortas. The expression of arginase 2, which is a marker of pro-inflammatory macrophages, was significantly lower and that of Ym1, which is a marker of anti-inflammatory macrophages, and was significantly higher after EPA and DHA administration. The same trends were observed in peritoneal macrophages after EPA and DHA administration.Conclusions:Dietary intake of EPA and DHA prevented AAA development through the inhibition of aortic and macrophage-mediated inflammation. (Circ J 2015; 79: 1470–1478)
Plasminogen activator inhibitor-1 (PAI-1), an endogenous inhibitor of a major fibrinolytic factor, tissue-type plasminogen activator, can both promote and inhibit angiogenesis. However, the ...physiologic role and the precise mechanisms underlying the angiogenic effects of PAI-1 remain unclear. In the present study, we report that pharmacologic inhibition of PAI-1 promoted angiogenesis and prevented tissue necrosis in a mouse model of hind-limb ischemia. Improved tissue regeneration was due to an expansion of circulating and tissue-resident granulocyte-1 marker (Gr-1+) neutrophils and to increased release of the angiogenic factor VEGF-A, the hematopoietic growth factor kit ligand, and G-CSF. Immunohistochemical analysis indicated increased amounts of fibroblast growth factor-2 (FGF-2) in ischemic gastrocnemius muscle tissues of PAI-1 inhibitor-treated animals. Ab neutralization and genetic knockout studies indicated that both the improved tissue regeneration and the increase in circulating and ischemic tissue-resident Gr-1+ neutrophils depended on the activation of tissue-type plasminogen activator and matrix metalloproteinase-9 and on VEGF-A and FGF-2. These results suggest that pharmacologic PAI-1 inhibition activates the proangiogenic FGF-2 and VEGF-A pathways, which orchestrates neutrophil-driven angiogenesis and induces cell-driven revascularization and is therefore a potential therapy for ischemic diseases.
IL-17 is known to be a cytokine mainly secreted from Th17 cells, which well associate with autoimmune inflammatory responses. In the generation of Th17 cells, RORc and RORa have pivotal roles in ...controlling the transcription of Il17. We speculated additional regulation in Il17a transcription and randomly screened a 6344 clone cDNA library to identify specific modulators for Il17a promoter activity. After the screen, the E3 ubiquitin ligases SIAH1 and SIAH2 were investigated further and confirmed to increase Il17a promoter activity in a T-cell line and to promote Th17 development ex vivo. This enhancement was a consequence of enhanced expression of hypoxia-inducible factor-1α (HIF-1α) protein, which is reported to directly regulate expression of Il17a and Rorgt at the transcriptional level. In the absence of HIF-1α, both ubiquitin ligases had little effect on Th17 cell differentiation. These results suggest that the SIAH1 and SIAH2 play a pivotal role to promote Th17 cell differentiation through maintaining the stability of HIF-1α protein.
Abstract
Cholera toxin B (CTB) is a subunit of cholera toxin, a bacterial enterotoxin secreted by Vibrio cholerae and also functions as an immune adjuvant. However, it remains unclear how CTB ...activates immune cells. We here evaluated whether or how CTB induces production of a pro-inflammatory cytokine, interleukin-1β (IL-1β). CTB induced IL-1β production not only from bone marrow-derived macrophages (BMMs) but also from resident peritoneal macrophages in synergy with O111:B4-derived lipopolysaccharide (LPS O111:B4) that can bind to CTB. Meanwhile, when prestimulated with O55:B5-derived LPS (LPS O55:B5) that fails to bind to CTB, resident peritoneal macrophages, but not BMMs, produced IL-1β in response to CTB. The CTB-induced IL-1β production in synergy with LPS in both peritoneal macrophages and BMMs was dependent on ganglioside GM1, which is required for internalization of CTB. Notably, not only the NLRP3 inflammasome but also the pyrin inflammasome were involved in CTB-induced IL-1β production from resident peritoneal macrophages, while only the NLRP3 inflammasome was involved in that from BMMs. In response to CTB, a Rho family small GTPase, RhoA, which activates pyrin inflammasome upon various kinds of biochemical modification, increased its phosphorylation at serine-188 in a GM1-dependent manner. This phosphorylation as well as CTB-induced IL-1β productions were dependent on protein kinase A (PKA), indicating critical involvement of PKA-dependent RhoA phosphorylation in CTB-induced IL-1β production. Taken together, these results suggest that CTB, incorporated through GM1, can activate resident peritoneal macrophages to produce IL-1β in synergy with LPS through novel mechanisms in which pyrin as well as NLRP3 inflammasomes are involved.
CTB activates peritoneal macrophages via a novel inflammasome pathway
ABSTRACTThe granulocyte colony‐stimulating factor (G‐CSF) promotes angiogenesis. However, the exact mechanism is not known. We demonstrate that vascular endothelial growth factor (VEGF) was released ...by Gr‐1+CD11b− neutrophils but not Gr‐1−CD11b+ monocytes prestimulated with G‐CSF in vitro and in vivo. Similarly, in vivo, concomitant with an increase in neutrophil numbers in circulation, G‐CSF augmented plasma VEGF level in vivo. Local G‐CSF administration into ischemic tissue increased capillary density and provided a functional vasculature and contributed to neovascularization of ischemic tissue. Blockade of the VEGF pathway abrogated G‐CSF‐induced angiogenesis. On the other hand, as we had shown previously, VEGF can induce endothelial progenitor cell (EPC) mobilization. Here, we show that G‐CSF also augmented the number of circulating VEGF receptor‐2 (VEGFR2) EPCs as compared with untreated controls. Blocking the VEGF/VEGFR1, but to a much lesser extent, the VEGF/VEGFR2 pathway in G‐CSF‐treated animals delayed tissue revascularization in a hindlimb model. These data clearly show that G‐CSF modulates angiogenesis by increasing myelomonocytic cells (VEGFR1+ neutrophils) and their release of VEGF. Our results indicated that administration of G‐CSF into ischemic tissue provides a novel and safe therapeutic strategy to improve neovascularization.
Mast cells accumulate in tissues undergoing angiogenesis during tumor growth, wound healing, and tissue repair. Mast cells can secrete angiogenic factors such as vascular endothelial growth factor ...(VEGF). Ionizing irradiation has also been shown to have angiogenic potential in malignant and nonmalignant diseases. We observed that low-dose irradiation fosters mast cell-dependent vascular regeneration in a limb ischemia model. Irradiation promoted VEGF production by mast cells in a matrix metalloproteinase-9 (MMP-9)-dependent manner. Irradiation, through MMP-9 up-regulated by VEGF in stromal and endothelial cells, induced the release of Kit-ligand (KitL). Irradiation-induced VEGF promoted migration of mast cells from the bone marrow to the ischemic site. Irradiation-mediated release of KitL and VEGF was impaired in MMP-9-deficient mice, resulting in a reduced number of tissue mast cells and delayed vessel formation in the ischemic limb. Irradiation-induced vasculogenesis was abrogated in mice deficient in mast cells (steel mutant, Sl/Sl(d) mice) and in mice in which the VEGF pathway was blocked. Irradiation did not induce progenitor mobilization in Sl/Sl(d) mice. We conclude that increased recruitment and activation of mast cells following irradiation alters the ischemic microenvironment and promotes vascular regeneration in an ischemia model. These data show a novel mechanism of neovascularization and suggest that low-dose irradiation may be used for therapeutic angiogenesis to augment vasculogenesis in ischemic tissues.
Clinical hypertension (HT) is associated with renal inflammation and elevated circulating levels of proinflammatory cytokines. Interleukin (IL)-1 receptor antagonist (IL-1Ra) is one of the most ...important anti-inflammatory cytokines and plays a crucial role in inflammation. Inhibition of IL-1 may contribute to modulation of the Angiotensin II (Ang II)-induced HT response. The present study aimed to elucidate the effects of IL-1Ra and anti-IL-1β antibody (01BSUR) on Ang II-induced renal injury. To determine the contribution of IL-1Ra to Ang II-induced renal inflammation, male wildtype (WT) and IL-1Ra-deficient (IL-1Ra-/-) mice were infused with Ang II (1000 ng/kg/min) using subcutaneous osmotic pump for 14 days. We checked renal function, histological change, and several mRNA expressions 14 days after infusion. Fourteen days after infusion, systolic blood pressure (197 ± 5 vs 169 ± 9 mmHg, P<0.05) in IL-1Ra-/- mice significantly increased compared with WT mice. Furthermore, on day 14 of Ang II infusion, plasma IL-6 was 5.9-fold higher in IL-1Ra-/- versus WT mice (P<0.001); renal preproendothelin-1 mRNA expression was also significantly higher in IL-1Ra-/- mice (P<0.05). In addition, renal histology revealed greater damage in IL-1Ra-/- mice compared with WT mice 14 days after infusion. Finally, we administrated 01BSUR to both IL-1Ra-/- and WT mice, and 01BSUR treatment decreased Ang II-induced HT and renal damage (glomerular injury and fibrosis of the tubulointerstitial area) in both IL-1Ra-/- and WT mice compared with IgG2a treatment. Inhibition of IL-1 decreased Ang II-induced HT and renal damage in both IL-1Ra-/- and WT mice, suggesting suppression of IL-1 may provide an additional strategy to protect against renal damage in hypertensive patients.
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