Search

Abstract The endocannabinoid system (ECS) regulates energy metabolism, has been implicated in the pathogenesis of metabolic diseases and exerts its actions mainly through the type 1 cannabinoid receptor (CB1). Likewise, autophagy is involved in several cellular processes. It is required for the normal development of muscle mass and metabolism, and its deregulation is associated with diseases. It is known that the CB1 regulates signaling pathways that control autophagy, however, it is currently unknown whether the ECS could regulate autophagy in the skeletal muscle of obese mice. This study aimed to investigate the role of the CB1 in regulating autophagy in skeletal muscle. We found concomitant deregulation in the ECS and autophagy markers in high-fat diet-induced obesity. In obese CB1-KO mice, the autophagy-associated protein LC3 II does not accumulate when mTOR and AMPK phosphorylation levels do not change. Acute inhibition of the CB1 with JD-5037 decreased LC3 II protein accumulation and autophagic flux. Our results suggest that the CB1 regulates autophagy in the tibialis anterior skeletal muscle in both lean and obese mice.

Carbon dioxide (CO2)-laser processing of glasses is a versatile maskless writing technique to engrave micro-structures with flexible control on shape and size.In this study, we present the fabrication of hundreds of microns quartz micro-channels and micro-holes by pulsed CO2-laser ablation with a focus on the great potential of the technique in microfluidics and biomedical applications. After discussing the impact of the laser processing parameters on the design process, we illustrate specific applications. First, we demonstrate the use of a serpentine microfluidic reactor prepared by combining CO2-laser ablation and post-ablation wet etching to remove surface features stemming from laser-texturing that are undesirable for channel sealing. Then, cyclic olefin copolymer micro-pillars are fabricated using laser-processed micro-holes as molds with high detail replication. The hundreds of microns conical and square pyramidal shaped pillars are used as templates to drive 3D cell assembly. Human Umbilical Vein Endothelial Cells are found to assemble in a compact and wrapping way around the micro-pillars forming a tight junction network. These applications are interesting for both Lab-on-a-Chip and Organ-on-a-Chip devices.

Introduction: Only a minority of patients with platinum refractory head and neck squamous cell carcinoma (PR/HNSCC) gain some lasting benefit from immunotherapy. Methods: The combined role of the comprehensive genomic (through the FoundationOne Cdx test) and immune profiles of 10 PR/HNSCC patients treated with the anti-PD-1 nivolumab was evaluated. The immune profiles were studied both at baseline and at the second cycle of immunotherapy, weighing 20 circulating cytokines/chemokines, adhesion molecules, and 14 soluble immune checkpoints dosed through a multiplex assay. A connectivity map was obtained by calculating the Spearman correlation between the expression profiles of circulating molecules. Results: Early progression occurred in five patients, each of them showing TP53 alteration and three of them showing a mutation/loss/amplification of genes involved in the cyclin-dependent kinase pathway. In addition, ERB2 amplification (1 patient), BRCA1 mutation (1 patient), and NOTCH1 genes alteration (3 patients) occurred. Five patients achieved either stable disease or partial response. Four of them carried mutations in PI3K/AKT/PTEN pathways. In the only two patients, with a long response to immunotherapy, the tumor mutational burden (TMB) was high. Moreover, a distinct signature, in terms of network connectivity of the circulating soluble molecules, characterizing responder and non-responder patients, was evidenced. Moreover, a strong negative and statistically significant (p-value ≤ 0.05) correlation with alive status was evidenced for sE-selectin at T1. Conclusions: Our results highlighted the complexity and heterogeneity of HNSCCs, even though it was in a small cohort. Molecular and immune approaches, combined in a single profile, could represent a promising strategy, in the context of precision immunotherapy.

Osteonecrosis of the jaw (ONJ) is a chronic complication affecting long-term bisphosphonate-treated subjects, recognized by non-healing exposed bone in the maxillofacial region. The pathophysiological mechanism underlying ONJ has not been fully elucidated. The aim of the present study was to investigate the role of RANK/RANKL/OPG signaling pathway and, in parallel, to evaluate angiogenic and matrix mineralization processes in jaw bone necrotic samples obtained from bisphosphonate-treated subjects with established ONJ. Necrotic bone samples and native bone samples were processed for Light and Field Emission in Lens Scanning Electron Microscope (FEISEM) analyses, for Real-Time RT-PCR to evaluate the gene expression of TNFRSF11A (RANK), TNFSF11 (RANKL), and TNFSF11B (OPG) and for immunohistochemical analyses of VEGF and BSP expression. Morphological analyses performed by Light microscope and FEISEM show empty osteocytic lacunae and alteration of lamellar organization with degradation of the mineralized bone matrix in necrotic bone samples. A significant increase in TNFRSF11A, TNFSF11, TRAF6 and NFAT2 gene expression, and a reduction of TNFSF11B gene transcription level compared is also showed in necrotic bone compared to control samples. No significant difference of VEGF expression is evidenced, while lower BSP expression in necrotic bone compared to healthy samples is found. Even if the pathogenesis of bisphosphonate-associated ONJ remains unknown, a link between oral pathogens and its development seems to exist. We suppose lipopolysaccharide produced by bacteria colonizing and infecting necrotic bone and the surrounding viable area could trigger RANK/RANKL/OPG signaling pathway and, in this context, osteoclasts activation could be considered as a protective strategy carried out by the host bone tissue to delimitate the necrotic area and to counteract infection.

Autologous bone is considered the gold standard for bone regeneration, even if different heterologous bone substitutes have been proposed to overcome the limits related to its use. The aim of this study was to analyze and to compare the molecular events switched on by autologous or heterologous bone graft insertion, focusing on TGFβ1 expression and OPG/RANKL ratio, to analyze resorption process, and estimating graft vascularization, new bone tissue deposition and its mineralization, through VEGF, BSP and SPARC expression evaluation, respectively. Patients needing pre-prosthetic rehabilitation of the posterior maxilla were treated using an equine-derived biomaterial (Group 1) or calvaria autologous bone (Group 2), according to the morphology of the bone defect. Bone graft integration was evaluated on bone samples obtained from the treated areas at the moment of dental implant insertion, by morphological and immunohistochemical analyses for TGFβ1, OPG, RANKL, VEGF, BSP, and SPARC expression. Morphological analysis shows the presence of biomaterial residual granules in Group 1, in parallel to a good integration between graft and host tissue. Moderate TGFβ1 expression is seen in both Group 1 and Group 2. OPG/RANKL ratio appear higher in Group 1; VEGF expression appears very strong in Group 1 and strong in Group 2, while BSP and SPARC expression results weak in Group 1 and moderate in Group 2. Results reveal the good integration between both types of graft and the host tissue, even though autologous graft seems to produce a faster regenerative process, as evidenced by the different expression of the investigated molecules. According to these observations, the clinical use of heterologous particulate equine-derived biomaterial may ensure long-term predictability of implant-prosthetic rehabilitation, comparable to that obtained with autologous bone graft.