Ambra Girardi

MicroRNA expression profiling of oral carcinoma identifies new markers of tumor progression.

Oral squamous cell carcinoma, the most frequently occurring malignant head and neck tumour, generally exhibits poor prognosis and metastases are the main cause of death. The discovery of reliable prognostic indicators of tumour progression could greatly improve clinical practice. MicroRNAs are involved in the regulation of basic cellular processes such as cell proliferation, differentiation, and apoptosis. Since miRNAs have been shown to be abnormally expressed in different tumours their importance as potential cancer prognostic indicators is increasing. To define the role of miRNA in OSCC tumours we investigated the expression profile of 15 OSCC (8 without metastasis and 7 with lymph node metastasis) using microarray analysis. Thirteen miRNA were significantly overexpressed (miR-489, miR-129, miR-23a, miR-214, miR-23b, miR-92, miR-25, miR-210, miR-212, miR-515, miR-146b, miR-21, miR-338) and 6 miRNA were underexpressed (miR-520h, miR-197, miR-378, miR-135b, miR-224, miR-34a) in oral tumours. Underexpression of mir-155, let-7i, mir-146a was found to characterize progression to metastastatic tumours. Further investigations will elucidate whether differentially expressed miRNAs will help to better classify OSCCs, thus improving diagnoses and patient care.

Biological effect of resorbable plates on normal osteoblasts and osteoblasts derived from Pfeiffer syndrome.

Biodegradable fixation devices made of the polymers polylactide, polyglycolide and their copolymers are used routinely during maxillofacial, craniofacial, and orthopedic reconstructive surgical procedures, thanks to their property of biodegradation that avoid the need for implant removal. In particular, they are used in the treatment of craniosynostosis in pediatric patients affected by Pfeiffer syndrome, where the resorption time of 1 year or less does not interfere with the normal growth of the skull. To study the mechanism how polylactide-polyglycolide (PLPG) acid plates can induce osteoblast differentiation and proliferation in normal osteoblasts and in osteoblasts derived from a patient with Pfeiffer syndrome, the expression levels of bone-related genes were analyzed using real-time reverse transcription-polymerase chain reaction. Osteoblasts grown on the PLPG acid plates resulted in significant upregulation of mRNA expression of many genes related to osteodifferentiation during the treatment, indicating that polylactide, polyglycolide biopolymers enhance proliferation, differentiation, and deposition of matrix in osteoblasts. This study also revealed some differences in gene expression between normal osteoblasts and osteoblasts derived from patients with Pfeiffer syndrome, cultivated on PLPG acid plates.

Expression and association data strongly support JARID2 involvement in nonsyndromic cleft lip with or without cleft palate.

Nonsyndromic cleft lip with or without cleft palate (CL/P) affects approximately 1 in 1,000 births. Genetic studies have provided evidence for the role of several genes and candidate loci in clefting; however, conflicting results have frequently been obtained and much have to be done to unravel the complex genetics of CL/P. In the present investigation we have focused on the candidate region in 6p23, a region that have been found linked to CL/P in several investigations, in the attempt to find out the susceptibility gene provisionally named OFC1. Gene expression experiments in mice embryo of positional candidate genes revealed that JARID2 was highly and specifically expressed in epithelial cells in merging palatal shelves. A family‐based linkage disequilibrium study confirmed the pivotal role of JARID2 in orofacial development and strongly supports a role for this gene in CL/P etiology (multiallelic haplotype test P=6×10−5). Understanding the molecular role of JARID2 within facial development may offer additional information to further unravel the complex genetics of CL/P. Hum Mutat 31:1–7, 2010.

IL6 and IL10 are genetic susceptibility factors of periodontal disease.

Background: Periodontitis is a disease mainly caused by a chronic infection of tissues that support the teeth. Several factors, such as diabetes, smoking and oral care, as well as genetic susceptibility can influence both the risk to develop periodontitis and its progression. The aim of the investigation was to test whether alleles of candidate genes were associated with periodontitis. Materials and Methods: A case control study was performed with a cohort of 184 patients with chronic periodontitis and 231 healthy controls from the Italian population. A total of six single nucleotide polymorphisms from five candidate genes, i.e., IL1A, IL1B, IL6, IL10 and vitamin D receptor, were investigated. Results: Evidence of association were obtained for rs1800795 mapping in IL6 (P value = 0.01) as well as for the rs1800872 mapping in IL10 (P = 0.04). The rarer variant allele lowered the risk to develop periodontitis at IL6 (Odds Ratio [OR] = 0.69 [95% confidence interval {CI} 0.51-0.93]) and increased the risk at IL10 (OR = 1.38 [95% CI 1.01-1.86]). Conclusions: The present investigation indicated that polymorphisms of IL6 and IL10 constitute risk factors for chronic periodontitis, while there was no evidence implicating a specific IL1A or IL1B genotype.

Trabecular Titanium Induces Osteoblastic Bone Marrow Stem Cells Differentiation

Pure titanium and titanium alloys are materials widely used in orthopedics and dental surgery because of their mechanical properties, chemical stability and biocompatibility. Although excellent clinical results have been shown, traditional porous metals have several inherent limitations (low volumetric porosity, relatively high modulus of elasticity and availability as a coating only). With the aim of going over these limits, improving the potentiality of osteointegration, a new highly porous titanium biomaterial (Trabecular Titaniumâ„¢, TT) has been developed. Because the molecular events due to TT and able to alter osteoblast activity to promote bone formation are poorly understood, expression of osteoblastic related genes in mesenchymal stem cells exposed to TT was investigated. The expression levels of bone related genes like RUNX2, SPP1, COL1A1, COL3A1, BGLAP, ALPL, and FOSL1) and mesenchymal stem cells marker (CD105) were analyzed, using real time Reverse Transcription-Polymerase Chain Reaction.TT causes induction of bone related genes osteopontin (SPP1), osteocalcin (BGLAP) alkaline phosphatase (ALPL) and indicating the differentiation effect of this biomaterial on mesenchymal stem cells.The obtained results can be relevant to better understand the molecular mechanism of bone regeneration and as a model for comparing other materials with similar clinical effects.

MIF Expression in Induced Peripheral Blood Mononuclear Cells by Vitamin D3 and its Potential Correlation with Resting Metabolic Rate in Obesity

Macrophage migration inhibitory factor (MIF) plays a pivotal role in systemic and local inflammatory and immune responses. The aim of this study is to assess MIF gene expression in PBMCs harvested from obese individuals and to compare it with that of lean subjects to analyze any potential relationship between resting metabolic rate as well as several different parameters and MIF expression in PBMC. We also aim to assess anti-inflammatory characteristics of vitamin D3 on MIF expression in vitro. Participants were 30 obese and 18 non-obese subjects who were assessed following an overnight fasting for RMR by means of indirect calorimetry. Body composition was measured using a Bodystat device. The PBMCs were separated from whole blood by the Ficoll-hypaque technique. The mRNA was extracted and the cDNA was synthesized. This process was followed by real-time PCR using primer pairs specific for MIF mRNA and beta actin as internal control. Our findings clearly demonstrate that there were significant differences in terms of BMI, BMR predict, fat proportion, fat mass, free fat mass, TBW, visceral fat, fasting serum glucose, TG, HDL, Hs-CRP and RMR between the two groups. Moreover, the level of MIF expression in the obese group was approximately 2.5 times higher compared to the lean group. An increased level of MIF expression in the obese group and a decreased expression of that non-obese was observed after inducing PBMCs with vitamin D3. One of the intriguing results of this study was the observed reverse correlation between MIF expression and fat mass as well as fat proportion after PBMCs were cultured in the presence of vitamin D3. Therefore, it could be concluded that MIF expression, which is in turn influenced by vitamin D3, has a role in the hyperactive immune profile and the pro-inflammatory state observed in obese individuals which is suggested to have a causal relationship with obesity.

Bio-Oss®acts on Stem cells derived from Peripheral Blood

OBJECTIVES This study aims to study how Bio-Oss® can induce osteoblast differentiation in mesenchymal stem cells, the expression levels of bone related genes and mesenchymal stem cells markers using real time Reverse Transcription-Polymerase Chain Reaction. METHODS PB-hMSCs stem preparations were obtained for gradient centrifugation from peripheral blood of healthy anonymous volunteers, using the Acuspin System-Histopaque 1077. The samples were then cultured for 7 days for RNA processing, and the expression was quantified using real time PCR. RESULTS Bio-Oss® caused an induction of osteoblast transcriptional factor like RUNX2 and of bone related genes; SPP1 and FOSL1. In contrast, the expression of ENG was significantly decreased in stem cells treated with Bio-Oss® with respect to untreated cells, indicating the differentiation effect of this biomaterial on stem cells. CONCLUSION The results obtained can be relevant to enhance the understanding of the molecular mechanism of bone regeneration and can act as a model for comparing other materials with similar clinical effects.

Role of the MIR146A polymorphism in the origin and progression of oral squamous cell carcinoma

Gene expression and cell behavior are regulated by several factors, including small non-coding RNAs. MicroRNAs affecting cell growth, differentiation, and apoptosis are thought to play an important role in tumorigenesis. The levels of miR-146 appear to be associated with cancer development and progression, including that of oral squamous cell carcinoma. The aim of this investigation was to ascertain whether the single nucleotide polymorphism, rs2910164, mapping in the MIR146A gene, has a role in oral squamous cell carcinoma progression. A genetic association study was performed with a sample set of 346 oral squamous cell carcinomas collected in Italy. Our data indicate that the rs2910164 polymorphism is not associated with tumor development. However, a slight increase in the frequency of the variant allele was observed in Stage II tumors. Further investigations are needed to verify a possible role of the variant allele or rs2910164 in oral squamous cell carcinoma progression.

No evidence for a role of CRISPLD2 in non‐syndromic cleft lip with or without cleft palate in an Italian population

Non-syndromic cleft lip with or without cleft palate (NSCLP) is a malformation with variable phenotypes, resulting from a mixture of genetic and environmental factors. Some studies have supported a role for the 16q24 region and its candidate gene, CRISPLD2, in clefting. A replication study is necessary to confirm these findings. The aim of the present study was to test, by genetic linkage and association analyses, whether the candidate gene, CRISPLD2, represents a risk factor for NSCLP. The analysis of 39 multigenerational families provided formal exclusion of a linkage between NSCLP and the CRISPLD2 locus under different genetic models and non-parametric analyses. The family-based study of 239 unrelated probands and their parents revealed no association between any particular allele or haplotype and NSCLP. Therefore, the present investigation did not support the hypothesis of the involvement of CRISPLD2 in NSCLP malformation, at least with regard to the Italian population.

Microflora and periodontal disease.

Background: Periodontitis is a disease that affects and destroys the tissues that support teeth. Tissue damage results from a prolonged inflammatory response to an ecological shift in the composition of subgingival biofilms. Three bacterial species that constitute the red complex group, Porphyromonas gingivalis, Tannerella forsythia, and Treponema denticola, are considered the main pathogens involved in periodontitis. Materials and Methods: In the present study, a real-time polymerase chain reaction bases assay was designed to detect and quantify red complex species, then used to investigate 307 periodontal pocket samples from 127 periodontitis patients and 180 controls. Results: Significant higher prevalence of red complex species and increased amount of P. gingivalis and T. denticola were detected in periodontal pocket of periodontitis patients. Conclusions: Results demonstrated that the test is a valuable tool to improve diagnosis of periodontal disease.