NLRP3 and AIM2 inflammasomes expression is modified by LPS and titanium ions increasing the release of active IL-1ß in alveolar bone-derived MSCs.

Periodontitis and peri-implantitis are inflammatory diseases of infectious etiology that lead to the destruction of the supporting tissues located around teeth or implants. Although both pathologies share several characteristics, it is also known that they show important differences which could be due to the release of particles and metal ions from the implant surface. The activation of the inflammasome pathway is one of the main triggers of the inflammatory process. The inflammatory process in patients who suffer periodontitis or peri-implantitis has been mainly studied on cells of the immune system; however, it is also important to consider other cell types with high relevance in the regulation of the inflammatory response. In that context, mesenchymal stromal cells (MSCs) play an essential role in the regulation of inflammation due to their ability to modulate the immune response. This study shows that the induction of NLRP3 and absent in melanoma 2 (AIM2) inflammasome pathways mediated by bacterial components increases the secretion of active IL-1ß and the pyroptotic process on human alveolar bone-derived mesenchymal stromal cells (hABSCs). Interestingly, when bacterial components are combined with titanium ions, NLRP3 expression is further increased while AIM2 expression is reduced. Furthermore, decrease of NLRP3 or AIM2 expression in hABSCs partially reverses the negative effect observed on the progression of the inflammatory process as well as on cell survival. In summary, our data suggest that the progression of the inflammatory process in peri-implantitis could be more acute due to the combined action of organic and inorganic components.

Inflammasomes NLRP3 and AIM2 in peri-implantitis: A cross-sectional study.

BACKGROUND: Inflammasome components NLRP3 and AIM2 contribute to inflammation development by the activation of caspase-1 and IL-1ß. They have not been yet evaluated in samples from patients with active peri-implantitis. Thus, the aim of the present study is to analyze the expression of inflammasomes NLRP3 and AIM2 and subsequent caspase 1 and IL-1ß assessing the microenvironment of leukocyte subsets in samples from patients with active peri-implantitis. METHODS: Biopsies were collected from 33 implants in 21 patients being treated for peri-implantitis. Biopsies from gingival tissues from 15 patients with healthy periodontium were also collected for control. These tissues were evaluated through conventional histological stainings. Then, immunohistochemical detection was performed to analyze NLRP3, AIM2, caspase-1, and IL-1ß and markers of different leukocyte subsets. PCR for inflammasomes and related genes was also done. RESULTS: This manuscript reveals a high immunohistochemical and mRNA expression of NLRP3 and AIM2 inflammasomes, caspase-1, and IL-1ß in biopsies collected from human peri-implantitis. The expression of the tested markers was significantly correlated with the increase in inflammatory infiltrate, probing depth, presence of biofilm, and bleeding on probing. In these peri-implantitis lesions, the area of biopsy tissue occupied by inflammatory infiltrate was intense while the area occupied by collagen was significantly lower. In comparison with periodontal healthy tissues, the inflammatory infiltrate was statistically significantly higher in the peri-implantitis biopsies and was mainly composed of plasma cells, followed by T and B lymphocytes. CONCLUSION: In human peri-implantitis, chronic inflammation can be explained in part by the action of IL-1ß/caspase 1 induced through NLRP3 and AIM2 inflammasome activation.

Physico-chemical and biological characterization of a new bovine bone mineral matrix available for human usage.

BACKGROUND: Anorganic bovine bone has been deeply studied for bone regeneration in the oral cavity. Different manufacturing processes can modify the final composition of the biomaterial and the responses that induce. AIM: To evaluate the physico-chemical characteristics of a bovine bone mineral matrix and the clinical, radiographical, histological, and mRNA results after using it for maxillary sinus floor augmentation in humans. MATERIALS AND METHODS: First, the physical-chemical characteristics of the biomaterial were evaluated by X-ray powder diffraction, X-ray fluorescence, and electron microscopy. A frequently used biomaterial with the same animal origin was used as comparator. Then, a clinical study was designed for evaluating clinical, radiographical, histological, and mRNA outcomes. Patients in need of two-stage maxillary sinus floor augmentation were included in the study. Six months after the grafting procedure, a bone biopsy was collected for evaluation. RESULTS: In terms of physico-chemical characteristics, no differences were found between both biomaterials. Clinically, 10 patients were included in the study. After 6 months, clinical and radiographical data showed adequate outcomes for allowing implant placement. Histological, immunohistochemical and mRNA analyses showed that the biomaterial in use provides biological support to induce responses similar to those of other commonly used biomaterials. CONCLUSION: Bovine bone mineral matrix (Creos™ Xenogain) used as a single material for maxillary sinus floor augmentation shows adequate biological, clinical, and radiological outcomes. In fact, the results from this study are similar to those reported in the literature for another bovine bone-derived biomaterial with whom it shares composition and micro- and nanoscale characteristics.

GARP promotes the proliferation and therapeutic resistance of bone sarcoma cancer cells through the activation of TGF-ß.

Sarcomas are mesenchymal cancers with poor prognosis, representing about 20% of all solid malignancies in children, adolescents, and young adults. Radio- and chemoresistance are common features of sarcomas warranting the search for novel prognostic and predictive markers. GARP/LRRC32 is a TGF-ß-activating protein that promotes immune escape and dissemination in various cancers. However, if GARP affects the tumorigenicity and treatment resistance of sarcomas is not known. We show that GARP is expressed by human osteo-, chondro-, and undifferentiated pleomorphic sarcomas and is associated with a significantly worse clinical prognosis. Silencing of GARP in bone sarcoma cell lines blocked their proliferation and induced apoptosis. In contrast, overexpression of GARP promoted their growth in vitro and in vivo and increased their resistance to DNA damage and cell death induced by etoposide, doxorubicin, and irradiation. Our data suggest that GARP could serve as a marker with therapeutic, prognostic, and predictive value in sarcoma. We propose that targeting GARP in bone sarcomas could reduce tumour burden while simultaneously improving the efficacy of chemo- and radiotherapy.

GARP is a key molecule for mesenchymal stromal cell responses to TGF-ß and fundamental to control mitochondrial ROS levels.

Multipotent mesenchymal stromal cells (MSCs) have emerged as a promising cell therapy in regenerative medicine and for autoimmune/inflammatory diseases. However, a main hurdle for MSCs-based therapies is the loss of their proliferative potential in vitro. Here we report that glycoprotein A repetitions predominant (GARP) is required for the proliferation and survival of adipose-derived MSCs (ASCs) via its regulation of transforming growth factor-ß (TGF-ß) activation. Silencing of GARP in human ASCs increased their activation of TGF-ß which augmented the levels of mitochondrial reactive oxygen species (mtROS), resulting in DNA damage, a block in proliferation and apoptosis. Inhibition of TGF-ß signaling reduced the levels of mtROS and DNA damage and restored the ability of GARP-/low ASCs to proliferate. In contrast, overexpression of GARP in ASCs increased their proliferative capacity and rendered them more resistant to etoposide-induced DNA damage and apoptosis, in a TGF-ß-dependent manner. In summary, our data show that the presence or absence of GARP on ASCs gives rise to distinct TGF-ß responses with diametrically opposing effects on ASC proliferation and survival.

TGF-ß and mesenchymal stromal cells in regenerative medicine, autoimmunity and cancer.

Multipotent mesenchymal stromal cells (MSCs) represent a promising cell-based therapy in regenerative medicine and for the treatment of inflammatory/autoimmune diseases. Importantly, MSCs have emerged as an important contributor to the tumor stroma with both pro- and anti-tumorigenic effects. However, the successful translation of MSCs to the clinic and the prevention of their tumorigenic and metastatic effect require a greater understanding of factors controlling their proliferation, differentiation, migration and immunomodulation in vitro and in vivo. The transforming growth factor(TGF)-ß1, 2 and 3 are involved in almost every aspect of MSC function. The aim of this review is to highlight the roles that TGF-ß play in the biology and therapeutic applications of MSCs. We will discuss the how TGF-ß modulate MSC function as well as the paracrine effects of MSC-derived TGF-ß on other cell types in the context of tissue regeneration, immune responses and cancer. Finally, taking all these aspects into consideration we discuss how modulation of TGF-ß signaling/production in MSCs could be of clinical interest.

Mesenchymal stromal cells express GARP/LRRC32 on their surface: effects on their biology and immunomodulatory capacity.

Mesenchymal stromal cells (MSCs) represent a promising tool for therapy in regenerative medicine, transplantation, and autoimmune disease due to their trophic and immunomodulatory activities. However, we are still far from understanding the mechanisms of action of MSCs in these processes. Transforming growth factor (TGF)-ß1 is a pleiotropic cytokine involved in MSC migration, differentiation, and immunomodulation. Recently, glycoprotein A repetitions predominant (GARP) was shown to bind latency-associated peptide (LAP)/TGF-ß1 to the cell surface of activated Foxp3(+) regulatory T cells (Tregs) and megakaryocytes/platelets. In this manuscript, we show that human and mouse MSCs express GARP which presents LAP/TGF-ß1 on their cell surface. Silencing GARP expression in MSCs increased their secretion and activation of TGF-ß1 and reduced their proliferative capacity in a TGF-ß1-independent manner. Importantly, we showed that GARP expression on MSCs contributed to their ability to inhibit T-cell responses in vitro. In summary, we have found that GARP is an essential molecule for MSC biology, regulating their immunomodulatory and proliferative activities. We envision GARP as a new target for improving the therapeutic efficacy of MSCs and also as a novel MSC marker.

CD105 (endoglin)-negative murine mesenchymal stromal cells define a new multipotent subpopulation with distinct differentiation and immunomodulatory capacities.

Administration of in vitro expanded mesenchymal stromal cells (MSCs) represents a promising therapy for regenerative medicine and autoimmunity. Both mouse and human MSCs ameliorate autoimmune disease in syn-, allo- and xenogeneic settings. However, MSC preparations are heterogeneous which impairs their therapeutic efficacy and endorses variability between experiments. This heterogeneity has also been a main hurdle in translating experimental MSC data from mouse models to human patients. The objective of the present manuscript has been to further characterize murine MSCs (mMSCs) with the aim of designing more efficient and specific MSC-based therapies. We have found that mMSCs are heterogeneous for endoglin (CD105) expression and that this heterogeneity is not due to different stages of MSC differentiation. CD105 is induced on a subpopulation of mMSCs early upon in vitro culture giving rise to CD105(+) and CD105(-) MSCs. CD105(+) and CD105(-) mMSCs represent independent subpopulations that maintain their properties upon several passages. CD105 expression on CD105(+) mMSCs was affected by passage number and cell confluency while CD105(-) mMSCs remained negative. The CD105(+) and CD105(-) mMSC subpopulations had similar growth potential and expressed almost identical mMSC markers (CD29(+)CD44(+)Sca1 (+) MHC-I(+) and CD45(-)CD11b(-)CD31(-)) but varied in their differentiation and immunoregulatory properties. Interestingly, CD105(-) mMSCs were more prone to differentiate into adipocytes and osteocytes and suppressed the proliferation of CD4(+) T cells more efficiently compared to CD105(+) mMSCs. Based on these studies we propose to redefine the phenotype of mMSCs based on CD105 expression.