Effects of nutrients, mainly from mediterranean dietary foods, on mesenchymal stem derived cells: growth or differentiation

During the last decade the interest for the mesenchymal cells is growing due to their possible uses in therapies to treat certain degenerative pathologies. Mesenchymal stem cells have been found in the bone marrow and they have been shown to be responsible for bone repair and fat cells production. Mesenchymal stromal cells can be obtained from a wide variety of tissues in addition to bone marrow and can differentiate into many other cell types. The study of cell differentiation and programming provides new models for drug discovery and cell therapy that now overcomes gene therapy. Senescence, cancer development and degenerative diseases depend on mesenchymal cells contribution to tissue homeostasis. On the other hand, diet and life style are included among risk factors, which can contribute to the success of pharmacological treatments. This review focuses on nutrients from Mediterranean diet and supplements, which have been shown to influence mesenchymal stem cells and cells derived from them. Dietary intake of nutrients impairs both in vitro and in vivo observations, this review aims to gather the results about the effects of food compounds on mesenchymal cells from which adipocytes and osteoblasts derive. Amino acids and proteins, carbohydrates, lipids, fatty acids and vegetable secondary metabolites, differently act on mesenchymal cells bearing on modulation of gene expression and controlling the fate of cell lineages. Remarkable, the analysis of literature shows that the main effect of nutrients on mesenchymal cells is the stimulation of transcription factors which address the cells toward proliferation or differentiation. For instance, carbohydrates, simple or complex, and lipids appear to stimulate the PPAR receptors, whereas proteins and amino acids result to act on the mTOR system and they can also stimulate the MyoD-1 transcription factor and cooperating proteins. In conclusion, nutrients can promote cell growth and differentiation of mesenchymal cells

A peptidyl-glucosamine derivative affects IKKα kinase activity in human chondrocytes

Nuclear factor-kappaB (NF-kappaB) transcription factor regulates several cell signaling pathways, such as differentiation and inflammation, which are both altered in osteoarthritis. Inhibitor kappaB kinase (IKK)alpha and IKKbeta are kinases involved in the activation of the NF-kappaB transcription factor. The aim of the present study was to determine the effects of glucosamine (GlcN), which is administered in the treatment of osteoarthritis, and of its 2-(N-Acetyl)-L-phenylalanylamido-2-deoxy-beta-D-glucose (NAPA) derivative on IKK kinases and, consequently, on NF-kappaB activation in human chondrocytes. The human chondrosarcoma cell line HTB-94 and human primary chondrocytes were stimulated with tumor necrosis factor (TNF)alpha after pre-treatment with GlcN or NAPA. Gene mRNA expression level was evaluated by real-time PCR. Inhibitor kappaB protein (IkappaB)alpha phosphorylation and p65 nuclear re-localization were analyzed by Western blotting; IKKalpha nuclear re-localization was also investigated by immunocytochemistry and Western blotting. IKK kinase activity was studied by in vitro kinase assay. After TNFalpha stimulation, the mRNA expression level of some of the genes under NF-kappaB control, such as interleukin (IL)-6 and IL-8, increased, while treatment with GlcN and NAPA reverted the effect. We investigated the possibility that GlcN and NAPA inhibit IKK kinase activity and found that NAPA inhibits the IKKalpha kinase activity, whereas GlcN does not. Interestingly, both GlcN and NAPA inhibit IKKalpha nuclear re-localization. Our results demonstrate that glucosamine and its peptidyl derivative can interfere with NF-kappaB signaling pathway by inhibiting IKKalpha activity in human chondrocytes. However, the mechanism of action of the two molecules is not completely overlapping. While NAPA can both specifically inhibit the IKKalpha kinase activity and IKKalpha nuclear re-localization, GlcN only acts on IKKalpha nuclear re-localization

The induction of Maspin expression by a glucosamine-derivative has an antiproliferative activity in prostate cancer cell lines

Mammary serine protease inhibitor or Maspin has been characterized as a class II tumor suppressor gene in several cancer types, among them prostate cancer (CaP). Androgen ablation is an effective therapy for CaP, but with short-term effectiveness, thus new therapeutic strategies are actively sought. The present study is aimed to explore the effects of a glucosamine derivative, 2-(N-Carbobenzyloxy)L-phenylalanylamido-2-deoxy-β-D-glucose (NCPA), on two CaP cell lines, PC3 and LNCaP. In particular we analyzed the impact of NCPA on Maspin production, cell viability and cell cycle progression and apoptosis/necrosis pathway activation has been determined in PC3 and LNCaP cell lines. NCPA is able to stimulate Maspin production in PC3 and not in LNCaP cell lines. NCPA blocks the PC3 cell cycle in G1 phase, by inhibiting Cyclin D1 production and induces the apoptosis, therefore interfering with aggressiveness of this androgen-insensitive cell line. Moreover, NCPA is able to induce the expression of Maspin in LNCaP cell line treated with androgen receptor inhibitor, Bicalutamide, and in turn to stimulate the apoptosis of these cells. These findings suggest that NCPA, stimulating the endogenous production of a tumor suppressor protein, could be useful in the design of new therapeutic strategies for treatment of CaP

Glucosamine affects intracellular signalling through inhibition of mitogen-activated protein kinase phosphorylation in human chondrocytes

The aim of this study was to determine the effects of glucosamine on matrix metalloprotease (MMP) production, on mitogen-activated protein kinase (MAPK) phosphorylation, and on activator protein (AP)-1 transcription factor activation in human chondrocytes. The human immortalized cell line lbpva55 and healthy human chondrocytes (obtained from healthy donors) were subjected to challenge with 10 ng/ml IL-1β after pretreatment with 2.5 or 10 mmol/l glucosamine. MMP mRNA expression levels were evaluated using quantitative real-time PCR, and MMP protein production levels were evaluated in the culture supernatant using ELISA. MAPK phosphorylation was evaluated using Western blotting. AP-1 transcription factor activation was evaluated by measuring AP-1 DNA-binding activity. After IL-1β stimulation, levels of MMP-1, MMP-3 and MMP-13 production were markedly increased. Treatment with 2.5 and 10 mmol/l glucosamine reduced expression of these metalloproteases. MMP expression is regulated by transcription factors such as the AP-1 complex, which is activated by phosphorylated MAPKs. IL-1β stimulated phosphorylation of c-jun amino-terminal kinase, p38 MAPK and extracellular signal-regulated kinase-1/2. Glucosamine inhibited c-jun amino-terminal kinase and p38 phosphorylation, and consequently c-jun binding activity. These findings demonstrate, for the first time, that glucosamine inhibits IL-1β-stimulated MMP production in human chondrocytes by affecting MAPK phosphorylation

In vitro antiviral and anti-inflammatory activities of N-Acetylglucosamine: development of an alternative and safe approach to fight viral respiratory infections

Viral respiratory tract infections (RTIs) are responsible for significant morbidity and mortality worldwide. A prominent feature of severe respiratory infections, such as severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection, is the cytokine release syndrome. Therefore, there is an urgent need to develop different approaches both against viral replication and against the consequent inflammation. N-acetylglucosamine (GlcNAc), a glucosamine (GlcN) derivative, has been developed as an immunomodulatory and anti-inflammatory inexpensive and non-toxic drug for non-communicable disease treatment and/or prevention. Recent studies have suggested that GlcN, due to its anti-inflammatory activity, could be potentially useful for the control of respiratory virus infections. Our present study aimed to evaluate in two different immortalized cell lines whether GlcNAc could inhibit or reduce both viral infectivity and the inflammatory response to viral infection. Two different viruses, frequent cause of upper and lower respiratory tract infections, were used: the H1N1 Influenza A virus (IAV) (as model of enveloped RNA virus) and the Human adenovirus type 2 (Adv) (as model of naked DNA virus). Two forms of GlcNAc have been considered, bulk GlcNAc and GlcNAc in nanoform to overcome the possible pharmacokinetic limitations of GlcNAc. Our study suggests that GlcNAc restricts IAV replication but not Adv infection, whereas nano-GlcNAc inhibits both viruses. Moreover, GlcNAc and mainly its nanoformulation were able to reduce the pro-inflammatory cytokine secretion stimulated by viral infection. The correlation between inflammatory and infection inhibition is discussed

Platelet rich fibrin (PRF) and its related products: biomolecular characterization of the liquid fibrinogen

Liquid fibrinogen is an injectable platelet concentrate rich in platelets, leukocytes, and fibrinogen obtained by blood centrifugation. The aim of this study was to analyze the release of different growth factors in the liquid fibrinogen at different times and to assess possible correlations between growth factors and cell counts. The concentration of transforming growth factor beta 1 (TGF-β1), platelet-derived growth factor-AB (PDGF-AB), platelet-derived growth factor-BB (PDGF-BB), bone morphogenetic protein 2 (BMP-2), fibroblast growth factor 2 (FGF-2) and vascular endothelial growth factor (VEGF) released by liquid fibrinogen were examined with ELISA at three time points (T0, time of collection; T7, 7 days; T14, 14 days). The cellular content of the liquid fibrinogen and whole blood was also calculated for each volunteer. A mean accumulation of platelets of almost 1.5-fold in liquid fibrinogen compared to whole blood samples was found. An increase of TGF-β1, PDGF-AB, FGF-2, and VEGF levels was detected at T7. At T14, the level of TGF-β1 returned to T0 level; PDGF-AB amount remained high; the levels of FGF-2 and VEGF decreased with respect to T7, but remained higher than the T0 levels; PDGF-BB was high at all time points; BMP-2 level was low and remained constant at all time points. TGF-β1, PDGF-AB, and PDGF-BB showed a correlation with platelet amount, whereas BMP-2, FGF-2, and VEGF showed a mild correlation with platelet amount. Due to the high concentration of platelets, liquid fibrinogen does contain important growth factors for the regeneration of both soft and hard tissue. The centrifugation protocol tested in this study provides a valid solution to stimulate wound healing in oral and periodontal surgery

A review of the effect of a nanostructured thin film formed by titanium carbide and titanium oxides clustered around carbon in graphitic form on osseointegration

Improving the biocompatibility of implants is an extremely important step towards improving their quality. In this review, we recount the technological and biological process for coating implants with thin films enriched in titanium carbide (TiC), which provide improved cell growth and osseointegration. At first, we discuss the use of a Pulsed Laser Ablation Deposition, which produced films with a good biocompatibility, cellular stimulation and osseointegration. We then describe how Ion Plating Plasma Assisted technology could be used to produce a nanostructured layer composed by graphitic carbon, whose biocompatibility is enhanced by titanium oxides and titanium carbide. In both cases, the nanostructured coating was compact and strongly bound to the bulk titanium, thus particularly useful to protect implants from the harsh oxidizing environment of biological tissues. The morphology and chemistry of the nanostructured coating were particularly desirable for osteoblasts, resulting in improved proliferation and differentiation. The cellular adhesion to the TiC-coated substrates was much stronger than to uncoated surfaces, and the number of philopodia and lamellipodia developed by the cells grown on the TiC-coated samples was higher. Finally, tests performed on rabbits confirmed in vivo that the osseointegration process of the TiC-coated implants is more efficient than that of uncoated titanium implants

Modulatory effects of a nutraceutical supplement on Saos-2 cells reveal its phlebotonic activity

Objective: Herbal extract compositions are largely used to manage vein diseases. We prepared a new composition of herbs, named FLEBO OKTM, that, when administered as a nutraceutical to patients affected by peripheral vascular diseases, was able to improve their health conditions. We analyzed the effects of this nutraceutical composition on in vitro cultured cells with the aim to obtain information about its mechanisms of action. Methods: A culture of human osteoblast cell line Saos-2 was stimulated with tumor necrosis factor (TNF)-a or interleukin (IL)-1b to induce the expression of some chemokines and matrix metalloproteases (MMPs). This cell culture was then exposed to the prepared composition and the amount of expression of the genes coding for the monocyte chemotactic protein (MCP)-1, IL-8, IL-1b, MMP-2, MMP-3, MMP-9 proteins was measured by real-time polymerase chain reaction (RT-PCR). The experiments were repeated exposing the cells to the same amount of the well-known micronized purified flavonoid fraction. Moreover, we describe the effects of the administration of nutraceutical composition to 20 patients affected by peripheral vascular diseases and 20 healthy individuals. Results: The RT-PCR analyses showed that the new composition induces the expression of MMP-3 and MMP-9 and downregulates MMP-2 in cell cultures stimulated with IL-1b, whereas it induces the expression of IL-8 and represses the expression of IL-1b and MCP-1 in cell cultures stimulated with TNF-a. The induction of the expression of MMP-3 and the downregulation of MCP-1 might result in an antiplatelet activity that was not observed for the micronized purified flavonoid fraction. Interviewed patients reported an improvement in their conditions after 1 month of FLEBO OK treatment. Conclusion: These findings could provide a hypothesis for the high efficiency of the identified nutraceutical composition to management of peripheral vascular diseases