BackgroundThe Visceral Adiposity Index (VAI) is a sex-specific mathematical index, based on Waist Circumference (WC), Body Mass Index (BMI), triglycerides (TG) and HDL cholesterol (HDL) levels, indirectly expressing visceral adipose function and insulin sensitivity. Our aim was to find the optimal cut-off points of VAI identifying a visceral adipose dysfunction (VAD) associated with cardiometabolic risk in a Caucasian Sicilian population.MethodsMedical check-up data of 1,764 Primary Care patients (PC patients) were retrospectively and cross-sectionally examined using a receiver-operating characteristic (ROC) curve to determine appropriate stratified-for-age cut-off of VAI, for the identification of PC patients with Metabolic Syndrome (MetS) according to the NCEP-ATP III criteria. The PC patients with higher VAI scores were subdivided into three groups according to VAI tertiles (i.e. PC patients with mild VAD, moderate VAD or severe VAD). Finally, VAD classes were compared to classical cardio- and cerebrovascular risk factors as independent predictors of coronary heart disease and/or myocardial infarction, transient ischemic attack and/or ischemic stroke.ResultsModerate and severe VADs proved to be independently associated with cardiovascular events [(OR: 5.35; 95% CI: 1.92-14.87; p = 0.001) and (OR: 7.46; 95% CI: 2.64-21.05; p < 0.001) respectively]. Mild, moderate and severe VADs were found to be independently associated with cerebrovascular events [(OR: 2.73; 95% CI: 1.12-6.65; p = 0.027), (OR: 4.20; 95% CI: 1.86-9.45; p = 0.001) and (OR: 5.10; 95% CI: 2.14-12.17; p < 0.001) respectively].ConclusionsOur study suggests that among Caucasian Sicilian subjects there are clear cut-off points of VAI able to identify a VAD strongly associated with cardiometabolic risk.

BackgroundChronic periodontal disease is an infectious disease consisting of prolonged inflammation of the supporting tooth tissue and resulting in bone loss. Guided bone regeneration procedures have become common and safe treatments in dentistry, and in this context dental stem cells would represent the ideal solution as autologous cells. In this study, we verified the ability of dental pulp mesenchymal stem cells (DPSCs) and gingival mesenchymal stem cells (GMSCs) harvested from periodontally affected teeth to produce new mineralized bone tissue in vitro, and compared this to cells from healthy teeth.MethodsTo characterize DPSCs and GMSCs, we assessed colony-forming assay, immunophenotyping, mesenchymal/stem cell phenotyping, stem gene profiling by means of flow cytometry, and quantitative polymerase chain reaction (qPCR). The effects of proinflammatory cytokines on mesenchymal stem cell (MSC) proliferation and differentiation potential were investigated. We also observed participation of several heat shock proteins (HSPs) and actin-depolymerizing factors (ADFs) during osteogenic differentiation.ResultsDPSCs and GMSCs were successfully isolated both from periodontally affected dental tissue and controls. Periodontally affected dental MSCs proliferated faster, and the inflamed environment did not affect MSC marker expressions. The calcium deposition was higher in periodontally affected MSCs than in the control group.Proinflammatory cytokines activate a cytoskeleton remodeling, interacting with HSPs including HSP90 and HSPA9, thioredoxin-1, and ADFs such as as profilin-1, cofilin-1, and vinculin that probably mediate the increased acquisition in the inflamed environment.ConclusionsOur findings provide evidence that periodontally affected dental tissue (both pulp and gingiva) can be used as a source of MSCs with intact stem cell properties. Moreover, we demonstrated that the osteogenic capability of DPSCs and GMSCs in the test group was not only preserved but increased by the overexpression of several proinflammatory cytokine-dependent chaperones and stress response proteins.Electronic supplementary materialThe online version of this article (doi:10.1186/s13287-017-0633-z) contains supplementary material, which is available to authorized users.

Hutchinson-Gilford progeria syndrome (HGPS) is a rare human genetic disease that leads to a severe premature ageing phenotype, caused by mutations in the LMNA gene. The LMNA gene codes for lamin-A and lamin-C proteins, which are structural components of the nuclear lamina. HGPS is usually caused by a de novo C1824T mutation that leads to the accumulation of a dominant negative form of lamin-A called progerin. Progerin also accumulates physiologically in normal ageing cells as a rare splicing form of lamin-A transcripts. From this perspective, HGPS cells seem to be good candidates for the study of the physiological mechanisms of ageing. Progerin accumulation leads to faster cellular senescence, stem cell depletion and the progeroid phenotype. Tissues of mesodermic origin are especially affected by HGPS. HGPS patients usually have a bad quality of life and, with current treatments, their life expectancy does not exceed their second decade at best. Though progerin can be expressed in almost any tissue, when death occurs, it is usually due to cardiovascular complications. In HGPS, severe epigenetic alterations have been reported. Histone-covalent modifications are radically different from control specimens, with the tendency to lose the bipartition into euchromatin and heterochromatin. This is reflected in an altered spatial compartmentalization and conformation of chromatin within the nucleus. Moreover, it seems that microRNAs and microRNA biosynthesis might play a role in HGPS. Exemplary in this connection is the suggested protective effect of miR-9 on the central nervous system of affected individuals. This mini-review will report on the state of the art of HGPS epigenetics, and there will be a discussion of how epigenetic alterations in HGPS cells can alter the cellular metabolism and lead to the systemic syndrome.

Background: Anaplastic thyroid carcinoma (ATC) is a rare and aggressive endocrine tumor with highly undifferentiated morphology. It has been suggested that cancer stem cells (CSCs) might play a central role in ATC. The objectives of this study were (i) to characterize CSCs from ex vivo ATC specimens by investigating the expression of several pluripotent stem cell markers, and (ii) to evaluate in vitro drug resistance modifications after specific CSC transcription factor switch-off. Methods: In ex vivo experiments, eight formalin-fixed, paraffin-embedded ATC specimens were analyzed by reverse-transcription and real-time quantitative PCR and immunohistochemistry. In in vitro experiments using ATC SW1736 cells, the expression levels of OCT-4, NANOG, and ABCG2 and the sensitivity to either cisplatin or doxorubicin were evaluated after silencing. Results: OCT-4, KLF4, and SOX2 transcription factors and C-KIT and THY-1 stem surface antigens showed variable up-regulation in all ATC cases. The SW1736 cell line was characterized by a high percentage of stem population (10.4 -2.1% of cells were aldehyde dehydrogenase positive) and high expression of several CSC markers (SOX2, OCT4, NANOG, C-MYC, and SSEA4). SOX2 silencing down-regulated OCT-4, NANOG, and ABCG2. SOX2 silencing sensitized SW1736 cells, causing a significant cell death increase (1.8-fold) in comparison to control cells with 10 lM cisplatin (93.9 -3.4% vs. 52.6 -9.4%, p < 0.01) and 2.7 fold with 0.5 lM doxorubicin (45.8 -9.9% vs. 17.1 -3.4% p < 0.01). ABCG2 silencing caused increased cell death with both cisplatin (74.9 -1.4%) and doxorubicin treatment (74.1 -0.1%) vs. no-target-treated cells (respectively, 45.8 -1.0% and 48.6 -1.0%, p < 0.001). Conclusions: The characterization of CSCs in ATC through the analysis of multiple pluripotent stem cell markers might be useful in identifying cells with a stem-like phenotype capable of resisting conventional chemotherapy. In addition, our data demonstrate that SOX2 switch-off through ABCG2 transporter down-regulation has a major role in overcoming CSC chemotherapy resistance.

Hypoxia-inducible factor-1a is found frequently overexpressed in solid tumors cells, exerting an important role in angiogenesis, glucose metabolism, cell proliferation, survival and invasion. In thyroid carcinomas, hypoxiainducible factor-1a expression was found increased in differentiated, poorly differentiated, medullary and anaplastic variants. Hypoxia represents the principal stimulus responsible for hypoxia-inducible factor-1a induction. Other nonhypoxic stimuli increase hypoxia-inducible factor-1a synthesis through the activation of phosphatidylinositol 3-kinase and mitogen-activated protein kinase pathways in a cell-type-specific manner. We have previously shown the role of BRAF V600E mutation in papillary thyroid cancer cells as a factor that facilitates tumor cell growth and progression. In this study, we tested the hypothesis that BRAF V600E mutation influences hypoxia-inducible factor-1a expression in papillary thyroid carcinoma cells. We analyzed 27 papillary thyroid carcinomas, 13 of which presented BRAF V600E mutation. In tumor tissues, immunoreactivity for hypoxiainducible factor-1a was detected in the majority of analyzed BRAF V600E mutated cases. Transcriptional analyses revealed elevated hypoxia-inducible factor-1a levels with significant differences between wild-type and mutated group. A BRAF wild-type papillary thyroid carcinoma cell line and a BRAF V600E mutated papillary thyroid carcinoma cell line were selected to study the effects of BRAF mutation on hypoxia-inducible factor-1a expression in vitro. Knockdown of mutant BRAF V600E or both the wild type and the BRAF V600E by RNA interference induced a significant reduction of hypoxia-inducible factor-1a expression at mRNA and protein levels. Pharmacological inhibition of BRAF significantly reduces hypoxia-inducible factor-1a expression levels in papillary thyroid carcinoma cell line harboring BRAF V600E mutation. Our results suggest that hypoxiainducible factor-1a is expressed in papillary thyroid carcinomas and is regulated not only by hypoxia but also by BRAF V600E -mediated signaling pathway.

BRAFV600E is the most common mutation found in papillary thyroid carcinoma (PTC). Tissue inhibitor of metalloproteinases (TIMP-1) and nuclear factor (NF)-kB have been shown to play an important role in thyroid cancer. In particular, TIMP-1 binds its receptor CD63 on cell surface membrane and activates Akt signaling pathway, which is eventually responsible for its antiapoptotic activity. The aim of our study was to evaluate whether interplay among these three factors exists and exerts a functional role in PTCs. To this purpose, 56 PTC specimens were analyzed for BRAF V600E mutation, TIMP-1 expression, and NF-kB activation. We found that BRAF V600E mutation occurs selectively in PTC nodules and is associated with hyperactivation of NF-kB and upregulation of both TIMP-1 and its receptor CD63. To assess the functional relationship among these factors, we first silenced BRAF gene in BCPAP cells, harboring BRAF V600E mutation. We found that silencing causes a marked decrease in TIMP-1 expression and NF-kB binding activity, as well as decreased invasiveness. After treatment with specific inhibitors of MAPK pathway, we found that only sorafenib was able to increase IkB-a and reduce both TIMP-1 expression and Akt phosphorylation in BCPAP cells, indicating that BRAF V600E activates NF-kB and this pathway is MEK-independent. Taken together, our findings demonstrate that BRAF V600E causes upregulation of TIMP-1 via NF-kB. TIMP-1 binds then its surface receptor CD63, leading eventually to Akt activation, which in turn confers antiapoptotic behavior and promotion of cell invasion. The recognition of this functional trilogy provides insight on how BRAF V600E determines cancer initiation, progression, and invasiveness in PTC, also identifying new therapeutic targets for the treatment of highly aggressive forms.

The stromal vascular cell fraction (SVF) of visceral and subcutaneous adipose tissue (VAT and SAT) has increasingly come into focus in stem cell research, since these compartments represent a rich source of multipotent adipose-derived stem cells (ASCs). ASCs exhibit a self-renewal potential and differentiation capacity. Our aim was to study the different expression of the embryonic stem cell markers NANOG (homeobox protein NANOG), SOX2 (SRY (sex determining region Y)-box 2) and OCT4 (octamer-binding transcription factor 4) and to evaluate if there exists a hierarchal role in this network in ASCs derived from both SAT and VAT. ASCs were isolated from SAT and VAT biopsies of 72 consenting patients (23 men, 47 women; age 45 ± 10; BMI between 25 ± 5 and 30 ± 5 range) undergoing elective open-abdominal surgery. Sphere-forming capability was evaluated by plating cells in low adhesion plastic. Stem cell markers CD90, CD105, CD29, CD31, CD45 and CD146 were analyzed by flow cytometry, and the stem cell transcription factors NANOG, SOX2 and OCT4 were detected by immunoblotting and real-time PCR. NANOG, SOX2 and OCT4 interplay was explored by gene silencing. ASCs from VAT and SAT confirmed their mesenchymal stem cell (MSC) phenotype expressing the specific MSC markers CD90, CD105, NANOG, SOX2 and OCT4. NANOG silencing induced a significant OCT4 (70 ± 0.05%) and SOX2 (75 ± 0.03%) downregulation, whereas SOX2 silencing did not affect NANOG gene expression. Adipose tissue is an important source of MSC, and siRNA experiments endorse a hierarchical role of NANOG in the complex transcription network that regulates pluripotency.

Background: In periodontal patients with jawbone resorption, the autologous bone graft is considered a "gold standard" procedure for the placing of dental prosthesis; however, this procedure is a costly intervention and poses the risk of clinical complications. Thanks to the use of adult mesenchymal stem cells, smart biomaterials, and active biomolecules, regenerative medicine and bone tissue engineering represent a valid alternative to the traditional procedures. Aims: In the past, mesenchymal stem cells isolated from periodontally compromised gingiva were considered a biological waste and discarded during surgical procedures. This study aims to test the osteoconductive activity of FISIOGRAFT Bone Granular R and Matriderm R collagen scaffolds on mesenchymal stem cells isolated from periodontally compromised gingiva as a low-cost and painless strategy of autologous bone tissue regeneration. Materials and Methods: We isolated human mesenchymal stem cells from 22 healthy and 26 periodontally compromised gingival biopsy tissues and confirmed the stem cell phenotype by doubling time assay, colony-forming unit assay, and expression of surface and nuclear mesenchymal stem cell markers, respectively by cytofluorimetry and realtime quantitative PCR. Healthy and periodontally compromised gingival mesenchymal stem cells were seeded on FISIOGRAFT Bone Granular R and Matriderm R scaffolds, and in vitro cell viability and bone differentiation were then evaluated. Results: Even though preliminary, the results demonstrate that FISIOGRAFT Bone Granular R is not suitable for in vitro growth and osteogenic differentiation of healthy and periodontally compromised mesenchymal stem cells, which, instead, are able to grow, homogeneously distribute, and bone differentiate in the Matriderm R collagen scaffold.