Unraveling the Mechanisms of Hypertrophy-Induced Matrix Mineralization and Modifications in Articular Chondrocytes.

Chondrocyte hypertrophic differentiation is a main event leading to articular cartilage degradation in osteoarthritis. It is associated with matrix remodeling and mineralization, the dynamics of which is not well characterized during chondrocyte hypertrophic differentiation in articular cartilage. Based on an in vitro model of progressive differentiation of immature murine articular chondrocytes (iMACs) into prehypertrophic (Prehyp) and hypertrophic (Hyp) chondrocytes, we performed kinetics of chondrocyte differentiation from Prehyp to Hyp to follow matrix mineralization and remodeling by immunofluorescence, biochemical, molecular, and physicochemical approaches, including atomic force microscopy, scanning electron microscopy associated with energy-dispersive X-ray spectroscopy (SEM-EDS), attenuated total reflection infrared analyses, and X-ray diffraction. Chondrocyte apoptosis was determined by TUNEL assay. The results show the formation of a mineral phase 7 days after Hyp induction, which spreads within the matrices to form poorly crystalline carbonate-substituted hydroxyapatite after 14 days, then the proportions of crystalline relative to amorphous content increases over time. Hyp differentiation also induced a matrix turnover that occurs over the first 7 days, characterized by a decrease in type II collagen and aggrecan and the concomitant appearance of type X collagen. This is accompanied by an increase in the enzymatic activity of MMP-13, the main collagenase in cartilage. The number of apoptotic chondrocytes slightly increased with Hyp differentiation and SEM-EDS analyses detected phosphorus-rich structures that could correspond to apoptotic bodies. Our findings highlight the mechanisms of matrix remodeling events leading to the mineralization of articular cartilage that may occur in osteoarthritis.

Ion intercalation dynamics of electrosynthesized mesoporous WO3 thin films studied by multi-scale coupled electrogravimetric methods.

Mesoporous WO3 thin films were prepared electrochemically by using an ionic surfactant during the synthesis, and the electrochemical properties are investigated in comparison with their dense analogues. This report specifically highlights the suitability of a time resolved coupled electrogravimetric method to follow meticulously the ion intercalation/extraction phenomena which revealed the enhanced ion intercalation/extraction behavior of electrodeposited mesoporous WO3 thin films for diverse applications in energy storage and electrochromism. This methodology (electrochemical impedance spectroscopy (EIS) and its coupling with a fast quartz crystal microbalance (QCM)) has the ability to detect the contribution of the charged or uncharged species during the electrochemical processes, and to deconvolute the global EQCM responses into the anionic, cationic, and the free solvent contributions. Our study identifies the involvement of several charged species (Li(+), Li(+)·H2O) in the compensation of charge, and H2O molecules indirectly contribute to the process in both dense and mesoporous WO3 thin films. Even a slight contribution of ClO4(-) ions was detected in the case of mesoporous analogues. The results of the study indicate that the transfer resistances of Li(+) and Li(+)·H2O are decreased when the WO3 films are mesoporous. A more significant difference is observed for the larger and partially dehydrated Li(+)·H2O ions, suggesting that increased surface area and pore volume created by mesoporous morphology facilitate the transfer of larger charged species. The relative concentration changes of cations are also magnified in the mesoporous films. The final concentration variations are higher in mesoporous films than that in the dense analogues; ∼4 times and ∼10 times higher for Li(+) and for Li(+)·H2O, respectively. To the best of our knowledge, an unambiguous identification of species other than desolvated cations (e.g. Li(+) ions), the information on their transfer dynamics and quantification of the transferred species have never been reported in the literature to describe the charge compensation process in WO3 based electrodes.

Characterization of counterion and surface influence on micelle formation using tapping mode atomic force microscopy in air.

Cylindrical micelles prepared in aqueous solutions from cationic surfactants octadecyl trimethylammonium (OTA+) or cetyltrimethylammonium (CTA+) and parachlorobenzoate (PCB) counterion were successfully imaged after evaporation of water using tapping mode atomic force microscopy (TM-AFM) onto very smooth gold and glass substrates. With the help of the obtained topography AFM images, it was shown that the micellar structures are preserved on gold substrates after evaporation of the solvent despite the new set of stresses due mainly to capillary forces and dehydration. The influence of the substrate on the resulting micellar morphology observed in air was investigated for these two materials: cylindrical micelles were evidenced as loosely adherent on gold surface in the presence of parachlorobenzoate (PCB) and identical, geometrically speaking, to those known to exist in aqueous solutions. In this situation, topographic AFM images allowed us to determine accurately their geometrical characteristics such as diameter and length in the nanometer range. On the other hand, AFM images obtained in air on glass surfaces revealed micellar structures that are different from those existing in the bulk of the solution. Indeed, bilayer-type micelles with a thickness close to twice the surfactant monomer expected length were observed, indicating that the well-established and strong influence of glass on micelle geometry at the glass/solution interface is maintained after evaporation of water. These results have been analyzed on the basis of positive charge of gold deduced from electrochemical impedance spectroscopy (EIS) and Raman spectroscopy measurements on one hand and of the negative charge of glass on the other hand. Although these results appeal to new theoretical considerations dealing with dynamics of evaporation of micellar solution drops and/or with counterion contributions to macromolecular interactions in aqueous solutions and in air, this new AFM imaging method appears to be the more adequate one to image and measure the micelles formed in the presence of water.