Effects of a new nanocomposite system on Human Gingival Fibroblasts/Streptococcus mitis co-culture

In the broad field of biomaterials, Bisphenol A glycidylmethacrylate (BisGMA)/triethyleneglycol dimethacrylate (TEGDMA) thermosets are frequently used for dental restoration (Lehtinen et al 2008), but infections due to bacterial adhesion remain the main reason of dental devices failure. In order to avoid biofilm formation on the components used for restoration and to reduce cytotoxicity against eukaryotic cells, a new material with antimicrobial properties was developed. Indeed, silver nanoparticles (n-Ag), which have well-known antimicrobial properties, were stabilized with a polyelectrolyte solution-Chitlac (lactose-modified chitosan) and was used to coating methacrylic thermosets (Travan et al, 2011). This study was aimed at evaluating the in vitro biological response of human gingival fibroblasts (HGFs)/Streptococcus mitis co-colture to this nanocomposite system. HGFs were obtained from fragments of healthy marginal gingival tissue, co-cultured with the clinical strain of S. mitis and treated for 24 -48 h with thermosets (uncoated or coated with Chitlac or Chitlac n-Ag). Cytotoxicity was evaluated by LDH assay; cell morphology and adhesion were verified by means of SEM and optical microscopy; cell migration was studied by a modified Boyden chamber and finally IL-6 and PGE2 secretion were detected by ELISA assays. In vitro results showed that in our co-culture model, which mimics the microenvironment of the oral cavity, the nanocomposite material does not exert cytotoxic effect towards HGFs that are able to adhere and migrate. The secretion of IL-6 is significant, but PGE2 production is minimal suggesting that IL-6 production is not related to an inflammatory response. Basing on its good biocompatibility we suggest this new tool useful for the realization of dental devices

The Effect of a Silver Nanoparticle Polysaccharide System on Streptococcal and Saliva-Derived Biofilms

In this work, we studied the antimicrobial properties of a nanocomposite system based on a lactose-substituted chitosan and silver nanoparticles: Chitlac-nAg. Twofold serial dilutions of the colloidal Chitlac-nAg solution were both tested on Streptococcus mitis, Streptococcus mutans, and Streptococcus oralis planktonic phase and biofilm growth mode as well as on saliva samples. The minimum inhibitory and bactericidal concentrations of Chitlac-nAg were evaluated together with its effect on sessile cell viability, as well as both on biofilm formation and on preformed biofilm. In respect to the planktonic bacteria, Chitlac-nAg showed an inhibitory/bactericidal effect against all streptococcal strains at 0.1% (v/v), except for S. mitis ATCC 6249 that was inhibited at one step less. On preformed biofilm, Chitlac-nAg at a value of 0.2%, was able to inhibit the bacterial growth on the supernatant phase as well as on the mature biofilm. For S. mitis ATCC 6249, the biofilm inhibitory concentration of Chitlac-nAg was 0.1%. At sub-inhibitory concentrations, the Streptococcal strains adhesion capability on a polystyrene surface showed a general reduction following a concentration-dependent-way; a similar effect was obtained for the metabolic biofilm activity. From these results, Chitlac-nAg seems to be a promising antibacterial and antibiofilm agent able to hinder plaque formation

Transmission electron microscopy demonstrating the effects of carvacrol codrug 4 on <i>S</i>. <i>aureus</i> ATCC 29213 (B), <i>E</i>. <i>coli</i> ATCC 8739 (D), <i>C</i>. <i>albicans</i> ATCC 10231 (F), and untreated cultures (control), respectively (A, C, and E).

<p>Microorganisms incubated for 3 h in media containing 12.5 mg/mL [<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0120937#pone.0120937.ref045" target="_blank">45</a>] of carvacrol codrug <b>4</b> (B, D, and F). Irregular features of septa in <i>S</i>. <i>aureus</i> ATCC 29213 (B: arrows); numerous electron-dense bubbles protrude from the cell surface (higher magnification b 140000x and c 110000x) in <i>E</i>. <i>coli</i> ATCC 8739 treated (D) and electron-dense granules of the substance internalized into the cytoplasm (d 110000x); integrity of the membrane (a 110000x) in the controls (C); disintegration of membrane in <i>C</i>. <i>albicans</i> ATCC 10231 (F).</p

Physicochemical properties of carvacrol codrug 4.

<p>^aValues are means of three experiments, standard deviation is given in parentheses.</p><p>Physicochemical properties of carvacrol codrug 4.</p

Kinetic data for hydrolysis of carvacrol codrug 4 at 37°C.^a

<p>^aValues are means of three experiments, standard deviation is given in parentheses.</p><p>^bAbbreviations: SGF, simulated gastric fluid; SIF, simulated intestinal fluid.</p><p>Kinetic data for hydrolysis of carvacrol codrug 4 at 37°C.<a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0120937#t004fn001" target="_blank">^a</a></p

Synthesis of carvacrol codrugs 1–4 and 9.

<p>Reagents and conditions: a) Ac₂O, AcOH, 4 h, rt for compounds <b>11–14</b>; KOH, Propargyl bromide, in dry MeOH, 1 h, 60°C (reflux) for compound <b>15</b>; b) DCC in DMF/DCM, 1 h, rt, then carvacrol, DMAP, 15 h, rt; c) TIPS, TFA in DCM, 48 h, rt under nitrogen atmosphere.</p

PAMPA-GI permeability of carvacrol codrug 4 after 16 h of incubation.

<p>^a pH of both donor and acceptor compartment.</p><p>PAMPA-GI permeability of carvacrol codrug 4 after 16 h of incubation.</p

Hemolytic activity of carvacrol codrugs 1–10, CAR, and NAC (A) and in a wider range of values for carvacrol codrug 4 (B).

<p>Hemolytic activity of carvacrol codrugs 1–10, CAR, and NAC (A) and in a wider range of values for carvacrol codrug 4 (B).</p