Human adipose stem cells induced to osteogenic differentiation by an innovative collagen/hydroxylapatite hybrid scaffold.

Novel biomaterials are of paramount importance for bone regrowth. In this study, we investigated human adipose stem cells (hASCs) for osteogenic, osteoconductivity, and osteoinductivity effects of an innovative collagen/hydroxylapatite hybrid scaffold. In hASCs that were grown on this scaffold, osteogenic genes were analyzed for their expression profiles, together with adhesion and extracellular matrix genes. In hASC integrins, basement membrane constituents and collagens were up-regulated, together with cell proliferation. In addition, expression of osteopontin and activated focal adhesion kinase was studied at the protein level. Our in vitro data indicate that hASCs, together with hybrid biomaterial, is an important model of study to investigate in vitro bone induction.-Mazzoni, E., D'Agostino, A., Manfrini, M., Maniero, S., Puozzo, A., Bassi, E., Marsico, S., Fortini, C., Trevisiol, L., Patergnani, S., Tognon, M. Human adipose stem cells induced to osteogenic differentiation by an innovative collagen/hydroxylapatite hybrid scaffold.

Specific IgG Antibodies React to Mimotopes of BK Polyomavirus, a Small DNA Tumor Virus, in Healthy Adult Sera.

BK polyomavirus (BKPyV) was isolated in 1971 from the urine of a kidney transplant patient. Soon after its identification, BKPyV was characterized as a kidney-tropic virus, which is responsible of a significant fraction of the rejection of transplant kidney in the host. Moreover, in experimental conditions, BKPyV is able to transform different types of animal and human cells and to induce tumors of different histotypes in experimental animals. BKPyV DNA sequences have been detected in healthy individuals and cancer patients using polymerase chain reaction/Shouthern blot hybridization methods. Serum antibodies against this polyomavirus were revealed using immunological techniques, which, however, cross-react with other polyomaviruses such as JC (JCPyV) and Simian Virus 40. These non-specific data indicate the need of novel immunological methods and new investigations to check in a specific manner, BKPyV spread in humans. To this aim, mimotopes from BKPyV structural capsid protein 1 (VP1) were employed for specific immunological reactions to IgG antibodies of human serum samples. An indirect enzyme-linked immunosorbent assay with synthetic peptides mimicking immunogenic epitopes of BKPyV VP1 was set up and employed to test sera of healthy adult subjects. Data from this innovative immunological assay indicate that serum antibodies against BKPyV VP1 mimotopes are detectable in healthy subjects ranging from 18 to 90 years old. The overall prevalence of serum samples that reacted to BKPyV VP1 mimotopes was 72%. The strong points from this investigation are the novelty of the immunological method, its simplicity of the approach, and the specificity of BKPyV antibody reaction to VP1 mimotopes.

Merkel Cell Carcinomas Arising in Autoimmune Disease Affected Patients Treated with Biologic Drugs, Including Anti-TNF.

Purpose: The purpose of this investigation was to characterize Merkel cell carcinomas (MCC) arisen in patients affected by autoimmune diseases and treated with biologic drugs.Experimental Design: Serum samples from patients with MCC were analyzed for the presence and titer of antibodies against antigens of the oncogenic Merkel cell polyomavirus (MCPyV). IgG antibodies against the viral oncoproteins large T (LT) and small T (ST) antigens and the viral capsid protein-1 were analyzed by indirect ELISA. Viral antigens were recombinant LT/ST and virus-like particles (VLP), respectively. MCPyV DNA sequences were studied using PCR methods in MCC tissues and in peripheral blood mononuclear cells (PBMC). Immunohistochemical (IHC) analyses were carried out in MCC tissues to reveal MCPyV LT oncoprotein.Results: MCPyV DNA sequences identified in MCC tissues showed 100% homology with the European MKL-1 strain. PBMCs from patients tested MCPyV-negative. Viral DNA loads in the three MCC tissues were in the 0.1 to 30 copy/cell range. IgG antibodies against LT/ST were detected in patients 1 and 3, whereas patient 2 did not react to the MCPyV LT/ST antigen. Sera from the three patients with MCC contained IgG antibodies against MCPyV VP1. MCC tissues tested MCPyV LT-antigen-positive in IHC assays, with strong LT expression with diffuse nuclear localization. Normal tissues tested MCPyV LT-negative when employed as control.Conclusions: We investigated three new MCCs in patients affected by rheumatologic diseases treated with biologic drugs, including TNF. A possible cause-effect relationship between pharmacologic immunosuppressive treatment and MCC onset is suggested. Indeed, MCC is associated with MCPyV LT oncoprotein activity. Clin Cancer Res; 23(14); 3929-34. ©2017 AACR.

Antibodies Against Mimotopes of Simian Virus 40 Large T Antigen, the Oncoprotein, in Serum Samples From Elderly Healthy Subjects.

Simian Virus 40 (SV40), a monkey polyomavirus, was administered to human populations by early anti-poliomylitis vaccines contaminated by this small DNA tumor virus. Data on SV40 infection in humans remain controversial. Elderly subjects represent an interesting cohort to investigate, because they were not immunized with SV40-contaminated vaccines. Taking advantage of the Italian population, the second oldest worldwide, elderly subjects (n = 237) up to 100 years old were enrolled in this study. Their sera were analyzed, by ELISA tests with synthetic peptides mimicking the viral epitopes, for IgG antibodies reacting with SV40 large Tumor antigen (Tag), the viral oncoprotein. An overall seroprevalence of 22% was revealed in subjects aged 66-100 years, ranging from 19% in individuals 66-74 years old, to 24% in subjects 82-100 years old, with a lower SV40 titer detected in the oldest group. Our data show that: (i) SV40 infection is not frequent in old individuals; (ii) the infection rate increases in elderly with the age; (iii) the antibody titer of SV40 Tag decreases with the age. In conclusion, SV40 infection seems to spread in old subjects independently from SV40-contaminated vaccines. This study seems to confirm that SV40 is also a human virus. J. Cell. Physiol. 232: 176-181, 2017. © 2016 Wiley Periodicals, Inc.

Circulating microRNAs found dysregulated in ex-exposed asbestos workers and pleural mesothelioma patients as potential new biomarkers.

Malignant pleural mesothelioma (MPM), a fatal cancer, is an occupational disease mostly affecting workers ex-exposed to asbestos fibers. The asbestos, a cancerogenic mineral of different chemical composition, was widely employed in western Countries in industrial manufactures of different types. MPM may arise after a long latency period, up to five decades. MPM is resistant to conventional chemo- and radio-therapies. Altogether, these data indicate that the identification of new and specific markers are of a paramount importance for an early diagnosis and treatment of MPM. In recent years, microRNAs expression was found dysregulated in patients, both in cancer cells and sera, affected by tumors of different histotypes, including MPM. Cell and circulanting microRNAs, found to be dysregulated in this neoplasia, were proposed as new biomarkers. It has been reported that circulating microRNAs are stable in biological fluids and could be employed as potential MPM biomarkers. In this investigation, circulating microRNAs (miR) from serum samples of MPM patients and workers ex-exposed to asbestos fibers (WEA) and healthy subjects (HS) were comparatively analyzed by microarray and RT-qPCR technologies. Our results allowed (i) to select MiR-3665, an endogenous stable microRNA, as the internal control to quantify in our analyses circulating miRNAs; to detect (ii) miR-197-3p, miR-1281 and miR 32-3p up-regulated in MPM compared to HS; (iii) miR-197-3p and miR-32-3p up-regulated in MPM compared to WEA; (iv) miR-1281 up-regulated in both MPM and WEA compared to HS. In conclusion, three circulating up-regulated microRNAs, i.e. miR-197-3p, miR-1281 and miR-32-3p are proposed as potential new MPM biomarkers.

P2X7 targeting inhibits growth of human mesothelioma.

Malignant pleural mesothelioma (MPM) is an aggressive tumor refractory to anti-blastic therapy. MPM cells show several genetic and biochemical defects, e.g. overexpression of oncogenes, downregulation of onco-suppressor genes, dysregulation of microRNA, or alteration of intracellular Ca2+ homeostasis and of apoptosis. No information is as yet available on purinergic signalling in this tumor. Signalling via the P2X7 (P2RX7 or P2X7R) purinergic receptor is attracting increasing attention as a pathway involved in cancer cell death or proliferation. In this report we show that the P2X7R is expressed by three MPM cell lines established from MPM patients but not by mesothelial cells from healthy subjects (healthy mesothelial cells, HMCs). MPM cell proliferation was inhibited by in vitro incubation in the presence of selective P2X7R antagonists, as well as by stimulation with the P2X7R agonist BzATP. Systemic administration of the selective P2X7R blocker AZ10606120 inhibited in vivo growth of MPM tumors whether implanted subcutaneously (s.c.) or intraperitoneally (i.p.). Our findings suggest that the P2X7R might be a novel target for the therapy of mesothelioma.