Ascorbic acid enhances bone parameter expression in human gingival mesenchymal stem cells.

Ascorbic acid (AS), also known as vitamin C or ascorbate, is an essential dietary nutrient which plays a vital role in biological processes through various different mechanisms, in particular for the biosynthesis of collagen. The aim of the study was to establish the possibility of enhancing the osteogenic differentiation potential by manipulating the cellular micro-environment, through AS supplementation in human gingival mesenchymal stem cells (hGMSCs) at different concentrations, such as 60 and 90 µg/mL, for three weeks. Human GMSCs are considered a stem cell population, easily obtainable and displaying a remarkable immunotherapeutic potential and regenerative repair expression. Osteogenic differentiation level induced from AS was assayed by histochemical characterization, using light microscopy through Alizarin red S staining. The transcript levels of Collagen 1A1 (COL1A1), runtrelated transcription factor 2 (RUNX2), bone morphogenetic protein 2/4 (BMP2/4), osteopontin (OPN) and osteonectin (SPARC) were determined by quantitative RT-PCR. Protein expression of COL1A1, RUNX2, BMP2/4, OPN, SPARC were studied through Western blotting and confocal laser scanning microscopy (CLSM). Our results demonstrate that AS supports osteogenic differentiation in stem cells from gingiva niche as shown by osteogenic marker upregulation and by de novo production of calcium phosphate deposits as revealed by Alizarin red S staining. In summary, the results of the current study provide evidence that hGMSCs undergo osteogenic differentiation with AS treatment, for that reason AS could be a promising candidate for the prevention and healing of bone-related diseases.

ENOX2 (or tNOX): a new and old molecule with cancer activity involved in tumor prevention and therapy.

Cancer includes a number of related diseases due to abnormal cell proliferation that spreads to nearby tissues. Many compounds (physical, chemical and biological) have been used to try to halt this abnormal proliferation, but the therapeutic results are poor, due also to the side effects. It has been reported that ecto-nicotinamide adenine dinucleotide oxidase di-sulfide-thiol exchanger 2 (ENOX2), also known as tumor-associated nicotinamide adenine dinucleotide oxidase (tNOX), was found to be located on the cancer cell surface, essential for cancer cell growth. Capsaicin and other anti-oxidants are capable of inhibiting tNOX, causing apoptosis of cells, exerting anti-tumor activity. It is interesting that some authors reported that ENOX2 is present in the serum of cancer patients several years before the clinical symptoms of the tumor. However, this result has to be confirmed. In this article we discuss ENOX2 and its inhibition as a hope of improving cancer therapy.

Human periodontal ligament stem cells cultured onto cortico-cancellous scaffold drive bone regenerative process.

The purpose of this work was to test, in vitro and in vivo, a new tissue-engineered construct constituted by porcine cortico-cancellous scaffold (Osteobiol Dual Block) (DB) and xeno-free ex vivo culture of human Periodontal Ligament Stem Cells (hPDLSCs). hPDLSCs cultured in xeno-free media formulation preserved the stem cells' morphological features, the expression of stemness and pluripotency markers, and their ability to differentiate into mesenchymal lineage. Transmission electron microscopy analysis suggested that after one week of culture, both noninduced and osteogenic differentiation induced cells joined and grew on DB secreting extracellular matrix (ECM) that in osteogenic induced samples was hierarchically assembled in fibrils. Quantitative RT-PCR (qRT-PCR) showed the upregulation of key genes involved in the bone differentiation pathway in both differentiated and undifferentiated hPDLSCs cultured with DB (hPDLSCs/DB). Functional studies revealed a significant increased response of calcium transients in the presence of DB, both in undifferentiated and differentiated cells stimulated with calcitonin and parathormone, suggesting that the biomaterial could drive the osteogenic differentiation process of hPDLSCs. These data were confirmed by the increase of gene expression of L-type voltage-dependent Ca2+ (VDCCL), subunits α1C and α2D1 in undifferentiated cells in the presence of DB. In vivo implantation of the hPDLSCs/DB living construct in the mouse calvaria evidenced a precocious osteointegration and vascularisation process. Our results suggest consideration of DB as a biocompatible, osteoinductive and osteoconductive biomaterial, making it a promising tool to regulate cell activities in biological environments and for a potential use in the development of new custom-made tissue engineering.

Endocrinology of the skin: intradermal neuroimmune network, a new frontier.

Endocrinology systems exert an important effect on vascular function and have direct actions on blood vessels. Estrogens provoke an increase in skin elasticity, epidermal hydration, skin thickness, reduce skin wrinkles and augment the content of collagen and the level of vascularisation. Therefore, there is an intricate cross-talk between skin conditions and stress. In stress, ß2--adrenoreceptor (ß2AR) pathway, cortisol, epinephrine and norepinephrine increase DNA damage and interfere with the regulation of the cell cycle, contributing to aging and skin diseases. Substance P is a neuropeptide released in the skin from the peripheral nerve and is related to stress and inflammation. SP provokes infiltration of inflammatory cells in the skin and induces a variety of cytokines/chemokines. Corticotropin-releasing hormone (CRH), produced by mast cells, is a neuropeptide also expressed in skin and responds to stress. CRH initiates diverse intracellular signaling pathways, including cAMP, protein kinase C, and mitogen-activated protein kinases (MAPK). Under stress, CRH, glucocorticoids, epinephrine and cytokines are generated. Moreover, the release of ACTH binds the receptor MC2-R and stimulates the generation of glucocorticoids such as corticosterone and cortisol, which interact with the transcription factors AP-1 and NF-kB. In skin keratinocytes, ACTH promotes the generation of pro-inflammatory cytokines, which enhances T-cell activity. Cortisol is immunosuppressive by inhibiting Th1 and Th2 cell response, antigen presentation, antibody and cytokine/chemokine production. However, glucocorticoids are certainly helpful in Th1-mediated autoimmune disorders. On the other hand, cytokines, such as TNF, IL-1 and IL-6, stimulate the generation of CRH and activate HPA axis in inflammatory states. Here, we describe for the first time a cross-talk between endocrinology and skin, including pro-inflammatory cytokines and neurogenic inflammatory pathways.

HTK solution helps to preserve endothelial integrity of saphenous vein: an immunohistochemical and ultrastructural analysis.

The aim of the study is to demonstrate the ability of HTK (Histidine-tryptophan-ketoglutarate) solution to preserve endothelium. Ten saphenous veins (SVs) were prospectively collected from 10 patients who underwent coronary artery bypass grafting (CABG). The SVs were divided into two sets of segments, one of which preserved in HTK solution at 4°C (group A), and the other preserved at 4°C in saline solution NaCl 0.9% (group B); ten pieces from the SVs were processed as control. The control sample was fixed in 10% neutral buffered formalin immediately after harvesting. The observation lasted up to the 5th postoperative day. A morphological, ultrastructural, and immunohistochemical analysis (CD31) was performed on each piece. Immunohistochemical analysis demonstrated significant protection on endothelium in group A compared to group B starting from the 1st observational day. Ultrastructural data confirmed immunohistochemistry. These preliminary results represent a basis for further analysis. They suggest the protective role of HTK solution in preserving endothelial integrity and may imply some clinical benefits in organ protection.

Does optical coherence tomography identify arterial healing after stenting? An in vivo comparison with histology, in a rabbit carotid model.

OBJECTIVE: To verify whether optical coherence tomography (OCT) can accurately monitor the occurrence of arterial healing after stenting. SETTING: Delayed stent endothelialisation may predispose to stent thrombosis. OCT is a high-resolution intravascular imaging technique that accurately identifies stent struts and arterial tissues. DESIGN AND INTERVENTIONS: Eight New Zealand white rabbits underwent the implantation of single bare metal stents (diameter 2-2.5 mm, length 8-13 mm) in the right common carotid artery through the external carotid artery. After a median of 11 days (range 2-28), the stented arteries were visualised by OCT, with images acquired at a pull-back speed of 0.5 mm/sec. The rabbits were then euthanised, vessels were formalin-fixed and finally processed for histopathology. RESULTS: We analysed 32 cross-sections from eight stented carotid arteries, for a total of 384 stent struts. OCT detected all of the stent struts in 30 of 32 cross-sections (93.7%), and correctly identified the presence/absence of tissue for every strut. Histological and OCT measurements of mean neointima thickness (0.135 (SD 0.079) mm and 0.145 (SD 0.085) mm, respectively, p = NS) were similar and closely related (r = 0.85, p<0.001). Neointima area progressively increased with longer time intervals from stent deployment to sacrifice; histological and OCT measurements were similar for each time interval. The intra-observer and interobserver reproducibility of OCT neointima measurements were excellent (R2 = 0.90 and 0.88, respectively). CONCLUSIONS: OCT is a promising means for monitoring stent strut coverage and vessel wall healing in vivo, the relevance of which will become even more significant with the increasing use of drug-eluting stents.

Morphological and cytofluorimetric analysis of adult mesenchymal stem cells expanded ex vivo from periodontal ligament.

Many adult tissues contain a population of stem cells that have the ability of regeneration after trauma, disease or aging. Recently, there has been great interest in mesenchymal stem cells and their roles in maintaining the physiological structure of tissues, and their studies have been considered very important and intriguing, after having shown that this cell population can be expanded ex vivo to regenerate tissues not only of the mesenchymal lineage, such as intervertebral disc cartilage, bone, tooth-associated tissue, cardiomyocytes, but also to differentiate into cells derived from other embryonic layers, including neurons. Currently, different efforts have been focused on the identification of odontogenic progenitors from oral tissues. In this study we isolated and characterized a population of homogeneous human mesenchymal stem cells proliferating in culture with an attached well-spread morphology derived from periodontal ligament, a tissue of ectomesenchymal origin, with the ability to form a specialized joint between alveolar bone and tooth. The adherent cells were harvested and expanded ex vivo under specific conditions and analysed by FACScan flow cytometer and morphological analysis was carried out by light, scanning and transmission electron microscopy. Our results displayed highly evident cells with a fibroblast-like morphology and a secretory apparatus, probably indicating that the enhanced function of the secretory apparatus of the mesenchymal stem cells may be associated with the secretion of molecules that are required to survive and proliferate. Moreover, the presence in periodontal ligament of CD90, CD29, CD44,CD166, CD 105, CD13 positive cells, antigens that are also identified as stromal precursors of the bone marrow, indicate that the periodontal ligament may turn out to be a new efficient source of the cells with intrinsic capacity to self-renewal, high ability to proliferate and differentiate, that can be utilized for a new approach to regenerative medicine and tissue engineering.

[Sclerosing Sertoli cell tumor of the testis. Report of a case and review of the literature]. / Tumore a cellule del Sertoli sclerosante del testicolo. Descrizione di un caso e revisione della letteratura.

Sclerosing Sertoli cell tumor of the testis is a rare neoplasm; only 12 cases of patients aged from 18 to 80 years (mean age 34,6) are reported in the Literature. No malignant behavior has been observed in any of the repoted cases. The tumors were all unilateral and hormonally inactive. A case of sclerosing Sertoli cell tumor of the right testis identified in a 36-year-old man and treated by orchdectomy is reported. The patient had no evidence of disease after at 4 years from treatment.