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The objective of this study is to evaluate whether a chronic obstructive pulmonary disease (COPD) self-management education program with coaching of a case manager improves patient-related outcomes and leads to practice changes in primary care. COPD patients from six family medicine clinics (FMCs) participated in a 1-year educational program offered by trained case managers who focused on treatment adherence, inhaler techniques, smoking cessation, and the use of an action plan for exacerbations. Health-care utilization, health-related quality of life (HRQL), treatment adherence, inhaler technique, and COPD knowledge were assessed at each visit with validated questionnaires. We also evaluated whether the use of spirometry and the assessment of individual patient needs led to a more COPD-targeted treatment by primary care physicians, based on changes in prescriptions for COPD (medication, immunization, and written action plan). Fifty-four patients completed the follow-up visits and were included in the analysis. The number of unscheduled physician visits went from 40 the year before intervention to 17 after 1 year of educational intervention ( p = 0.033). Emergency room visits went from five to two and hospitalizations from two to three (NS). Significant improvements were observed in HRQL ( p = 0.0001), treatment adherence ( p = 0.025), adequate inhaler technique ( p < 0.0001), and COPD knowledge ( p < 0.001). Primary care physicians increased their prescriptions for long-acting bronchodilators with/without inhaled corticosteroid, flu immunizations, and COPD action plans in the event patient had an exacerbation. The COPD self-management educational intervention in FMCs reduced unscheduled visits to the clinic and improved patients’ quality of life, self-management skills, and knowledge. The program had a positive impact on COPD-related practices by primary care physicians in the FMCs.

International audience; Background: The differential pathogenicity of anti-HLA donor-specific antibodies (DSAs) is not fully understood. The presence of complement-binding DSAs helps in better defining the prognosis of acute antibody-mediated rejection (ABMR). The evolution of these antibodies after the treatment of ABMR is unknown.Methods: We included patients from the French multicenter RITUX ERAH study diagnosed with acute ABMR within the first year of renal transplantation, with circulating anti-HLA DSAs and treated randomly by rituximab or placebo (and intravenous immunoglobulins, plasma exchange). We centrally analyzed serum samples at the time of ABMR, 3 and 6 months after ABMR, with anti-HLA DSAs specificities and C1q-binding capacity assessment.Results: Twenty-five patients were included: 68% had C1q-binding DSAs at the time of ABMR. The presence of C1q-binding DSAs was associated with a poorer evolution of chronic glomerulopathy at 6 months (P = 0.036). The persistence of C1q-binding DSAs at 3 and/or 6 months after ABMR was associated with more severe chronic glomerulopathy (P = 0.006), greater C4d score deposition score at 6 months after ABMR (P = 0.008), and graft loss 5 years after ABMR (P = 0.029). C1q-binding capacity was associated with the DSA MFI but 5 C1q-binding DSAs in 4 patients had low MFI values without a prozone effect.Conclusion: The presence and persistence of anti-HLA C1q-binding DSAs after ABMR is a detrimental marker, leading to transplant glomerulopathy and graft loss. Assessment of the complement-binding capacities of DSAs could help decide treatment intensification.

Although it is a key-recommendation of all recent asthma guidelines, self-management education is still insufficiently offered in primary care settings.To demonstrate the benefits of an educational program offered at the site of primary care (Family Medicine Clinics- FMC) by trained asthma educators on patient outcomes and healthcare use.This was a one-year pre-post intervention study. Patients with a diagnosis of mild to moderate asthma were enrolled from six FMC. After an initial encounter by the educator, an assessment of educational needs and a spirometry were done, followed by 3 follow-up visits at 4-6 weeks, 4-6 months and one year. Expiratory flows, asthma control criteria, knowledge about asthma, adherence to medication and healthcare and medication use were assessed at each visit.Data from 124 asthma patients (41M/83F), aged 55 ± 18 years, were analyzed. After initiating the intervention, there was a progressive increase in asthma knowledge and an improvement in medication adherence. The number of unscheduled visits for respiratory problems went from 137 to 33 (P 0.0001), the number of antibiotic treatments from 112 to 33 (P = 0.0002) and the number of oral corticosteroids treatments from 26 to 8 (NS). Marked improvements were observed in regard to inhaler technique and provision of a written action plan.This study shows that an educational intervention applied at the site of primary care can result in significant improvements in patient asthma outcomes and reduce unscheduled visits and inappropriate use of medications such as antibiotics.

Human hematopoietic stem/progenitor cells (HSPCs) can be mobilized into the circulation using granulocyte-colony stimulating factor (G-CSF), for graft collection in view of hematopoietic transplantation. This process has been related to bone marrow (BM) release of serine proteases and of the matrix metalloproteinase-9 (MMP-9). Yet, the role of these mediators in HSC egress from their niches remains questionable, because they are produced by nonstromal cells (mainly neutrophils and monocytes/macrophages) that are not a part of the niche. We show here that the G-CSF receptor (G-CSFR) is expressed by human BM mesenchymal stromal/stem cells (MSCs), and that G-CSF prestimulation of MSCs enhances the in vitro trans-stromal migration of CD34+ cells. Zymography analysis indicates that pro-MMP-2 (but not pro-MMP-9) is expressed in MSCs, and that G-CSF treatment increases its expression and induces its activation at the cell membrane. We further demonstrate that G-CSF-stimulated migration depends on G-CSFR expression and is mediated by a mechanism that involves MMPs. These results suggest a molecular model whereby G-CSF infusion may drive, by the direct action on MSCs, HSPC egress from BM niches via synthesis and activation of MMPs. In this model, MMP-2 instead of MMP-9 is implicated, which constitutes a major difference with mouse mobilization models.

Tissue Factor Pathway Inhibitor-2 (TFPI-2) is a potent inhibitor of plasmin which activates metalloproteinases (MMPs) involved in extracellular matrix (ECM) degradation. Its secretion in ECM makes TFPI-2 a potential inhibitor to regulate tumour invasion and metastasis. Moreover, TFPI-2 is frequently downregulated, particularly in aggressive cancers. In this study, we silenced TFPI-2 in the NCI-H460 non-small cell lung cancer cell line and evaluated the role of TFPI-2 in cell invasion and its impact on MMPs expression. As the effects of siRNA are transient, the consequences of both gene silencing and restoration to normal expression could be studied kinetically in the same cells. We showed that TFPI-2 expression by NCI-H460 cells was effectively downregulated using specific small interfering RNA and this silencing was associated with an increase in the invasive potential of tumour cells while migration was not affected. We also showed that mRNA levels and protein expression of MMP-2, -3, -9, -14 were not influenced by TFPI-2 silencing. Moreover, the gelatinase activity of MMP-2 and MMP-9 was unmodified. In contrast, MMP-1 mRNA levels and protein were significantly and similarly increased in cells transfected with TFPI-2 siRNA. In conclusion, this study confirms that TFPI-2 downregulation can contribute to tumour invasion of lung cancer cells.

The proinflammatory chemokine CC chemokine ligand 5 (CCL5) is a potent chemoattractant of immature dendritic cells (iDCs). It remains to be elucidated whether CCL5 may also enhance iDC migration through the basement membrane by affecting matrix metalloproteinase (MMP)-9 secretion. In this study, iDCs were differentiated in vitro from human monocytes of healthy donors. Zymographic analysis of cellular membranes of nontreated iDCs revealed a basal secretion of the pro- and active MMP-9, whereas only pro-MMP-9 was detected in conditioned media. Increasing concentrations of CCL5 significantly enhanced MMP-9 secretion by iDCs, peaking at 100 ng/ml, which optimally increased iDC migration through a reconstituted basement membrane (Matrigel™) in vitro. The CCL5-enhanced secretion of MMP-9 occurred early (2 h) and was maintained at least for 10 h. A significant increase in MMP-9 mRNA synthesis was detected by reverse transcriptase-polymerase chain reaction, only at 6 h of CCL5 treatment, which suggests that the early effect of CCL5 (0–4 h) on MMP-9 secretion was independent of mRNA synthesis, whereas the more delayed effect (6–10 h) could be mediated through an increase in MMP-9 gene expression. In a Matrigel migration assay, the CCL5-enhanced iDC migration was reduced significantly by specific inhibitors of MMP-9, such as tissue inhibitor of metalloproteinase-1 or an anti-MMP-9 antibody, which indicates that iDC migration through the basement membrane depends on MMP-9. These results suggest that under inflammatory conditions, the chemokine CCL5 may enhance iDC migration through the basement membrane by rapidly increasing their MMP-9 secretion.

Immature dendritic cells (DCs) can induce T-cell hyporesponsiveness, thus interfering with the process of DC maturation in a pro-inflammatory context, may therefore provide a novel approach to inducing allograft tolerance. We have studied the effects of mycophenolic acid (MPA), an immunosuppressive agent currently used in transplantation, using an in vitro model of a mixed human DC/alloreactive CD4(+) T lymphocyte culture. DCs differentiated from monocytes were exposed to MPA during maturation. MPA treatment affected the maturation of DCs, and this was reflected both in the impairment of the up-regulation of co-stimulatory molecule expression and the maintained endocytic capacity. However, MPA-DCs exhibited a distinctive microscopic morphology and secreted IL-10 and so could no longer be regarded as immature DC. Moreover, MPA-DCs had a mature phenotype for chemokine receptor expression, exhibiting down-regulation of CCR5 and up-regulation of CCR7. Interestingly, the abilities of the MPA-DCs to induce CD4(+) T-cell proliferation in response to alloantigens was impaired not only via direct but also via indirect pathways. The maintenance of endocytosis and the inhibition of syngeneic T-cell activation suggest that these cells could have a potential role to avoid chronic rejection. All these characteristics suggest that MPA-DCs may be used in cell therapy to induce allograft tolerance.

The mechanism of G-CSF-induced hematopoietic progenitor/stem cell (HPC/HSC) mobilization from bone marrow microenvironment to peripheral blood has been related to local production by neutrophils of proteases, including elastase, cathepsin G and metalloproteinases (MMPs). We previously showed that in vitro migration of hematopoietic cells across a layer of human marrow stromal cells (MSC) is induced when stimulated by G-CSF. In the present study, we investigated the role of MMPs produced by MSC used for the trans-stromal migration of MO7e line and marrow CD34+ cells. Normal human MSC were previously grown to confluence on Transwell® filters (pore diameter 5-μm) and then stimulated or not by IL-1 (15 U/mL/Day) or G-CSF (15 ng/mL/Day) for three consecutive days. In a second step, MO7e or marrow CD34+ cells, put in the upper chamber, were allowed to migrate through the layers (4 hrs, SDF-1 100 ng/mL in the bottom chamber). The contribution of MMPs was evaluated in this in vitro trans-stromal migration assay using specific blocking monoclonal antibodies (mAb), anti-MMP-1, anti-MMP-2 and anti-MMP-9 (each at 5 μg/mL). The amounts of antigenic MMP-2 and MMP-9 produced by MSC were evaluated in cell supernatants by enzyme-linked immunosorbent assay (ELISA) and gelatinolytic activity of MMPs by zymography. mRNA expression of MMPs in MSC was analyzed by reverse transcriptase-polymerase chain reaction (RT-PCR). None of the MSC layers used contained detectable hematopoietic cells since they were all CD45-. Migration of MO7e cells across unstimulated or G-CSF-stimulated stromal cells was significantly inhibited by specific mAb. Percentages of inhibition were 25+/−5 % and 54+/−9 % for anti-MMP-2 mAb, and 18+/−6 % and 53+/−9 % for anti-MMP-9 mAb, respectively. Using the combination of anti MMP-2 and anti-MMP-9, inhibition reached 71+/−24 % across stimulated layers. In contrast, anti-MMP-1 mAb did not show any significant inhibition. Migration of CD34+ across G-CSF-stimulated stromal cells was similarly inhibited with percentages of 38+/−18 % for anti-MMP- 2 mAb, and 34+/−11 % for anti-MMP-9 mAb, while migration was not inhibited again, using anti-MMP-1 mAb. In parallel, we demonstrated that MSC did express G-CSF receptor by RT-PCR. Production of MMPs by unstimulated and G-CSF-stimulated MSC was detected by ELISA, zymography and RT-PCR for MMP-2, and by zymography and RT-PCR for MMP-9. In conclusion, G-CSF-induction of HPC trans-stromal migration involves MMP-2 and MMP-9 but not MMP-1. These findings suggest that microenvironment, in the absence of neutrophils, could have a significant role in G-CSF mobilization of HPC/HSC. A local production of MMP-2 and MMP-9 by marrow stromal cells could be critical in the egress of HSC out of the hematopoietic niche.

International audience; Background: An efficient strategy for programing dendritic cells (DCs) for cancer immunotherapy is the optimiza‑ tion of their maturation so that they can efficiently stimulate cancer‑specific T cell responses. Interleukin (IL)‑4 has appeared as an essential cytokine, widely used in vitro with granulocyte macrophage‑colony stimulating factor (GM‑CSF) to differentiate monocytes into immature DCs (iDC) and to prevent macrophage formation. Conflicting data have been published regarding the effect of IL‑4 on functional DC maturation. To further understand IL‑4's effects on DC maturation and function in vitro, we choose the most commonly used maturation factor tumor necrosis factor (TNF)‑α. Methods: Human monocyte‑derived iDC were treated for 48 h with GM‑CSF and TNF‑α in the presence (IL‑4 + ‑DC) or absence (IL‑4 − ‑DC) of IL‑4 and functions of both DC populations were compared. Results: On mixed lymphocyte reaction assay, IL‑4 + ‑DC were less potent than IL‑4 − ‑DC at inducing the proliferation of allogeneic CD4 + T cells and the proportion of activated T cells expressing CD69 and/or CD25 was smaller. Interleu‑ kin‑4 reduced the cell‑surface expression of TNF‑α‑induced DC maturation markers CD83, CD86, HLA‑DR and CD25 and generated a heterogeneous population of DCs. IL‑4 + ‑DC secreted less IL‑12 and more IL‑10 than IL‑4 − ‑DC follow‑ ing activation by soluble CD40L, and IL‑4 + ‑DC‑activated T cells secreted lesser amounts of T helper (Th) 1 cytokines (IL‑2 and interferon‑γ). Importantly, IL‑4 impaired the in vitro migratory capacity of DCs in response to CCL21 and CCL19 chemokines. This effect was related to reduced expression of CCR7 at both mRNA and protein levels. Conclusion: Interleukin‑4 used with GM‑CSF and TNF‑α during the maturation of DCs in vitro impaired DC functions and disturbed the maturation effect of TNF‑α. Finally, our study reinforces the view that the quality of the DC matura‑ tion stimulus, which regulates DC migration and cytokine production, may be a decisive feature of the immunogenic‑ ity of DCs.