The Exposure to Osteoarthritic Synovial Fluid Enhances the Immunomodulatory Profile of Adipose Mesenchymal Stem Cell Secretome.

OBJECTIVE: Several clinical studies have proposed the infusion of adipose mesenchymal stem cells (AMSCs) as an alternative therapy for joint diseases with inflammatory components, such as osteoarthritis. Indeed, AMSCs are able to stimulate tissue repair through a paracrine activity and the interaction with the inflammatory microenvironment seems to have a critical role. DESIGN: To reproduce the inflammatory microenvironment, AMSCs were exposed to osteoarthritic synovial fluid (SF) for 48 h and the effect of their secretome on differentiation of monocytes (M0) into macrophages M1-like and mature dendritic cells (mDCs) was evaluated. Furthermore, the effect of the secretome of AMSCs exposed to SF was evaluated on the T cell population in terms of T cell proliferation and expansion of T regulatory cells (T reg). RESULTS: Our data show that the exposure of AMSCs to SF activates cells and promotes the release of immunosuppressive factors, which induce macrophage polarization of M0 into the M2-like phenotype and inhibit differentiation of monocytes into mature dendritic cells (mDCs). Only the secretome of exposed AMSCs was able to inhibit T cell proliferation and promote T reg expansion. CONCLUSIONS: Our results suggest that the microenvironment plays a fundamental role for the development of anti-inflammatory and immunomodulatory properties of AMSCs.

Pro inflammatory stimuli enhance the immunosuppressive functions of adipose mesenchymal stem cells-derived exosomes.

The predominant mechanism by which adipose mesenchymal stem cells (AMSCs) participate to tissue repair is through a paracrine activity and their communication with the inflammatory microenvironment is essential part of this process. This hypothesis has been strengthened by the recent discovery that stem cells release not only soluble factors but also extracellular vesicles, which elicit similar biological activity to the stem cells themselves. We demonstrated that the treatment with inflammatory cytokines increases the immunosuppressive and anti-inflammatory potential of AMSCs-derived exosomes, which acquire the ability to shift macrophages from M1 to M2 phenotype by shuttling miRNA regulating macrophages polarization. This suggests that the immunomodulatory properties of AMSCs-derived exosomes may be not constitutive, but are instead induced by the inflammatory microenvironment.

Exploring the Possible Prognostic Role of B-Lymphocyte Stimulator (BLyS) in a Large Series of Patients with Neuroendocrine Tumors.

BACKGROUND AND OBJECTIVE: BLyS (B-Lymphocyte stimulator) is over-expressed in several tumoral settings, with direct or indirect effects on neoplastic proliferation and possibly representing a therapeutic target. In this study, we explored the role of BLyS in a large population of patients with neuroendocrine tumors (NETs). METHODS: The study analyzed the stored sera of 124 consecutive unselected patients with NETs: 36 lung carcinoids (24 typical, 12 atypical), 47 gastroenteric tract and 41 pancreatic (30 non-functioning and 11 functioning: 9 insulinomas, 2 glucagonomas). In 23 cases, BLyS was repeatedly assessed during the follow-up and the disease was monitored (progression, stabilization or remission) according to the RECIST criteria. Patients were compared to 92 age and sex-matched blood donors (BDs). Serum levels of BLyS and Chromogranin A (CgA) were analyzed by ELISA. RESULTS: NET patients showed significantly higher BLyS levels than BDs (1274±809 pg/ml vs. 587±173 pg/ml; p<0.0001). BLyS correlated weakly with CgA (r=0.19 and p=0.035) but did not correlate with Ki67, grading, metastasis, histological type and site. In patients with sustained remission after surgery, BLyS and CgA both showed a gradual reduction over time. Patients with progressing disease showed higher BLyS levels compared to stable patients (1524±694 pg/ml vs. 1168± 373 pg/ml; p= 0.033). BLyS serum levels remained stable in remission and therapy-controlled patients, while increased in the follow-up of progressing cases. CONCLUSION: Higher BLyS levels identify patients with a more severe disease, characterized by progression despite treatments, possibly representing a factor implicated in the proliferation of the neoplastic cells or in sustaining the neoplastic environment.

Serum Interleukin-10 Levels Correlate with Cerebrospinal Fluid Amyloid Beta Deposition in Alzheimer Disease Patients.

BACKGROUND AND OBJECTIVE: In Alzheimer disease (AD) inflammation becomes evident throughout the course of the disease. However, the association between inflammation, cognitive impairment, and cerebrospinal biomarkers (Aß42, t-tau, p-tau181, and Aß42/p-tau181 ratio) is poorly understood. METHODS: A large panel of inflammatory cytokines (interleukin [IL]-1ß, IL-1ra, IL-2, IL-4, IL-6, IL-10, IL-17, interferon-γ, tumor necrosis factor-α, and vascular endothelial growth factor) was analyzed using a multiplex immunoassay in 27 patients with a diagnosis of AD dementia and in 18 control subjects. In a subgroup with available cerebrospinal fluid (CSF) samples, cytokines in serum were correlated with the levels of neurodegenerative CSF biomarkers (Aß42, t-tau, p-tau181, and Aß42/p-tau181 ratio). RESULTS: Compared to control subjects, AD patients showed a significant upregulation of IL-10, IL-1ß, and IL-17 serum levels. Several cytokines appeared intercorrelated, and IL-10 in particular presented a significant inverse correlation with CFS levels of Aß42 and the Aß42/p-tau ratio. CONCLUSION: Our findings indicate that serum levels of IL-10 may represent a possible peripheral expression of amyloid beta deposition in AD patients.

Characterization of the Proinflammatory Profile of Synovial Fluid-Derived Exosomes of Patients with Osteoarthritis.

The purpose of this study is to characterize synovial fluid- (SF-) derived exosomes of patients with gonarthrosis comparing two methods of isolation and to investigate their immune regulatory properties. Extracellular vesicles (EVs) have been isolated from inflamed SF by polymer precipitation method and quantified by Exocet kit and by nanoparticle tracking analysis. Vesicles expressed all the specific exosomal markers by immunoblot and FACS. After isolation with Exoquick, a relevant contamination by immune complexes was detected, which required further magnetic bead-based purification to remove. SF-derived exosomes significantly stimulated the release of several inflammatory cytokines and chemokines and metalloproteinases by M1 macrophages but did not influence the expression of CD80 and CD86 costimulatory molecules. In conclusion, we characterized purified exosomes isolated from inflamed SF and demonstrate that purified exosomes are functionally active in their ability to stimulate the release of proinflammatory factors from M1 macrophages. Our data indicate that SF-derived exosomes from gonarthrosis patients play a role in disease progression.

Exploring the plasmatic platelet-activating factor acetylhydrolase activity in patients with anti-phospholipid antibodies.

PURPOSE: To explore the role of plasmatic platelet-activating factor acetylhydrolase (PAF-AH), a marker of cardiovascular risk, in patients with anti-phospholipid antibodies (aPL). METHODS: PAF-AH activity was assessed in a series of 167 unselected patients screened for aPL in a context of thrombotic events, risk of thrombosis or obstetric complications and in 77 blood donors. RESULTS: 116/167 patients showed positive results for at least one aPL among IgG/IgM anti-prothrombin/phosphatidylserine (aPS/PT), anti-cardiolipin (aCL), anti-beta2-glycoprotein I (aß2GPI) or lupus anticoagulant (LAC), while 51/167 patients resulted aPL-negative. LAC+ patients disclosed higher PAF-AH than LAC-negative (22.1 ± 6.4 nmol/min/ml vs. 19.5 ± 4.1 nmol/min/ml; p = 0.0032), and aPL-negative patients (p = 0.03). Patients presenting positive IgG aß2GPI disclosed higher PAF-AH than patients with only IgM aß2GPI-positive antibodies (23.1 ± 7.2 nmol/min/ml vs. 20.1 ± 5.3 nmol/min/ml; p = 0.035), as well as than patients showing only isolated LAC, aCL or aPS/PT (16.9 ± 3.8 nmol/min/ml; p = 0.003). CONCLUSIONS: PAF-AH plasmatic activity is particularly up-regulated in LAC+ and in aß2GPI IgG+ patients, possibly representing an alternative prognostic biomarker for the therapeutic management of APS patients.

The Detection of Anti-adalimumab Antibodies in a Series of Inflammatory Polyarthritis: three ELISA Methods Compared.

BACKGROUND: A reliable identification of ADAb represents a fundamental tool in the followup of patients under treatment with anti-TNF drugs. OBJECTIVES: To compare three immunoenzymatic assays for anti-adalimumab antibodies (AAA) detection. METHODS: The study was performed in 40 patients with chronic inflammatory polyarthritis, comprising both patients showing a good response to adalimumab (ADL) and patients who lost response or did not respond to ADL, recently or in the past (retrospective study). Thus, sera were collected before ADL administration or well after ADL discontinuation. AAA were analysed by three different bridging ELISAs, following manufacturers' instructions. RESULTS: All methods disclosed negative results in responder patients and univocally recognized 11/31 (35.5%) AAA highpositive samples in non-responder patients, including several cases that had discontinued ADL for a long time (range 3-48 months). Among the overall non-responder patients, 10/31 (32.3%) disclosed concordant clear-cut AAA-negative results, while negative versus low-positive or borderline results were found in another ten non-responder patients, indicating slight differences in sensitivity between the methods, especially in patients who were analysed retrospectively. Methotrexate in combination therapy with ADL tended to be more frequent in AAA-negative, than in AAA-positive patients (52.6% vs. 25%; p=ns). CONCLUSIONS: The three bridging ELISA methods under study showed a good agreement and were able to identify uniquely the presence of high positive AAA, even after a long time since ADL discontinuation.