Patrice Laquerriere

Importance of hydroxyapatite particles characteristics on cytokines production by human monocytes in vitro

Calcium phosphate bioceramics have been applied as bone substitutes for several decades. Aseptic loosening after total joint arthroplasty is a major problem in orthopaedic surgery. Hydroxyapatite particles from materials wear have been reported as the main cause of implant failure. For this reason, an investigation into possible wear particles from materials used in the implant may lead to longevity after arthroplasty. Monocytes are among the first cells to colonize the inflammatory site. In the present study, we have evaluated the inflammatory response after exposition to particles with different characteristics (size, sintering temperature and shape). Our data demonstrate that the most important characteristic was the shape and the size of the particles. The needle shaped particles induced the larger production of TNF-alpha, IL-6 and IL-10 by cells. To a less manner, the smallest particles induced an increase of the expression and production of the cytokines studied (TNF-alpha, IL-6 and IL-10). The sintering temperature appeared to be a less important characteristic even though it was involved in the dissolution/precipitation process.

Endotracheal Tubes Coated with Antiseptics Decrease Bacterial Colonization of the Ventilator Circuits, Lungs, and Endotracheal Tube

Background: Formation of a bacterial biofilm within the endotracheal tube (ETT) after tracheal intubation is rapid and represents a ready source of lung bacterial colonization. The authors investigated bacterial colonization of the ventilator circuit, the ETT, and the lungs when the ETT was coated with silver-sulfadiazine and chlorhexidine in polyurethane, using no bacterial/viral filter attached to the ETT. Methods: Sixteen sheep were randomized into two groups. Eight sheep were intubated with a standard ETT (control group), and eight were intubated with a coated ETT (study group). Animals were mechanically ventilated for 24 h. At autopsy, the authors sampled the trachea, bronchi, lobar parenchyma, and ETT for quantitative bacterial cultures. Qualitative bacterial cultures were obtained from the filter, humidifier, inspiratory and expiratory lines, and water trap. ETTs were analyzed with light microscopy, scanning electron microscopy, and laser scanning confocal microscopy. Results: In the control group, all eight ETTs were heavily colonized (105–108 colony-forming units [cfu]/g), forming a thick biofilm. The ventilator circuit was always colonized. Pathogenic bacteria colonized the trachea and the lungs in five of eight sheep (up to 109 cfu/g). In the study group, seven of eight ETTs and their ventilator circuits showed no growth, with absence of a biofilm; one ETT and the respective ventilator circuit showed low bacterial growth (103–104 cfu/g). The trachea was colonized in three sheep, although lungs and bronchi showed no bacterial growth, except for one bronchus in one sheep. Conclusions: Coated ETTs induced a nonsignificant reduction of the tracheal colonization, eliminated (seven of eight) or reduced (one of eight) bacterial colonization of the ETT and ventilator circuits, and prevented lung bacterial colonization.

A new insight into the dissociating effect of strontium on bone resorption and formation.

Calcium phosphates are widely used as biomaterials and strontium (Sr) is known to have the ability to modify the bone balance towards osteosynthesis. In the present study we investigated the capacity of Sr-substituted sol-gel calcium phosphate to modify the expression of genes and proteins involved in extracellular matrix synthesis by primary bone cells. We first determined the most effective concentration of strontium using human primary bone cells. Sol-gel biphasic calcium phosphate (BCP) powders were then synthesised to obtain release of the optimal concentration of strontium. Finally, human osteoblasts obtained from explant cultures were cultured in the presence of sol-gel BCP, Sr-substituted BCP (5% Sr-substituted BCP, corresponding to a release of 5×10(-5)M [Sr(2+)] under the culture conditions (BCP(5%))) and medium containing strontium chloride (SrCl(2)). Viability, proliferation, cell morphology, protein production and protein activity were studied. We demonstrated that 5×10(-5)M SrCl(2) and BCP(5%) increased the expression of type I collagen and SERPINH1 mRNA and reduced the production of matrix metalloproteinases (MMP-1 and MMP-2) without modifying the levels of the tissue inhibitors of MMPs (TIMPs). Thus strontium has a positive effect on bone formation.

A clinical assessment of the Mucus Shaver: A device to keep the endotracheal tube free from secretions

Objective:We evaluated a new device designed to clean the endotracheal tube in mechanically ventilated patients, the Mucus Shaver. Design:Prospective, randomized trial. Setting:University hospital intensive care unit. Patients:We enrolled 24 patients expected to remain ventilated for >72 hrs. Interventions:The Mucus Shaver is a concentric inflatable catheter for the removal of mucus and secretions from the interior surface of the endotracheal tube. The Mucus Shaver is advanced to the distal endotracheal tube tip, inflated, and subsequently withdrawn over a period of 3–5 secs. Patients were prospectively randomized within 2 hrs of intubation to receive standard endotracheal tube suctioning treatment or standard suctioning plus Mucus Shaver use until extubation. Measurements and Main Results:During the study period, demographic data, recent medical history, adverse events, and staff evaluation of the Mucus Shaver were recorded. At extubation, each endotracheal tube was removed, cultured, and analyzed by scanning electron microscopy. Twelve patients were assigned to the study group and 12 were assigned to the control group. No adverse events related to the use of the Mucus Shaver were observed. At extubation, only one endotracheal tube from the Mucus Shaver group was colonized, whereas in the control group ten endotracheal tubes were colonized (8% vs. 83%; p < .001). Scanning electron microscopy showed little secretions on the endotracheal tubes from the study group, whereas thick bacterial deposits were present on all the endotracheal tubes from the control group (p < .001 by Fisher exact test, using a maximum biofilm thickness of 30 &mgr;m as cut-off). The nursing staff was satisfied by the overall safety, feasibility, and efficacy of the Mucus Shaver. Conclusions:The Mucus Shaver is a safe, feasible, and efficient device for endotracheal tube cleaning in the clinical setting. The Mucus Shaver is helpful in preventing endotracheal tube colonization by potentially harmful microorganisms.

Inflammatory cell response to calcium phosphate biomaterial particles: an overview.

Bone is a metabolically active and highly organized tissue consisting of a mineral phase of hydroxyapatite (HA) and amorphous calcium phosphate (CaP) crystals deposited in an organic matrix. One objective of bone tissue engineering is to mimic the chemical and structural properties of this complex tissue. CaP ceramics, such as sintered HA and beta-tricalcium phosphate, are widely used as bone substitutes or prosthesis coatings because of their osteoconductive properties. These ceramic interactions with tissues induce a cell response that can be different according to the composition of the material. In this review, we discuss inflammatory cell responses to CaP materials to provide a comprehensive overview of mechanisms governing the integration or loosening of implants, which remains a major concern in tissue engineering. A focus on the effects of the functionalization of CaP biomaterials highlights potential ways to increase tissue integration and limit rejection processes.

Structural characterization of sol–gel derived Sr-substituted calcium phosphates with anti-osteoporotic and anti-inflammatory properties

Sol–gel chemistry has been successfully used to prepare un-doped and Sr-doped calcium phosphate ceramics exhibiting a porous structure. The samples composition is very close to the nominal one. All samples present phase mixtures of mainly hydroxyapatite (HAp) and tricalcium phosphate (β-TCP). Doping with Sr2+ ions has a clear effect on the proportions of the different phases, increasing the amount of β-TCP. An amorphous phase is also observed incorporating some 40% of the total amount of strontium. Strontium ions also substitute for calcium both in HAp and β-TCP in specific sites that have been determined from Rietveld refinement on synchrotron powder diffraction data. The soluble amorphous and TCP phases are responsible for a beneficial partial release of strontium ions in solution during interactions between the material and biological fluids. Preliminary in vitro study demonstrates anti-inflammatory effects of strontium for human monocytes cultured in contact with calcium phosphates.

The effect of zinc on hydroxyapatite-mediated activation of human polymorphonuclear neutrophils and bone implant-associated acute inflammation.

Hydroxyapatite (HA) is widely used as coating biomaterial for prosthesis metal parts and as bone substitute. The release of HA particles induces an inflammatory response and, if uncontrolled, could result in implant loss. At the inflamed site, the polymorphonuclear cells (PMNs) represent the earliest phagocytic cells that predominate the cellular infiltrate. We have recently proposed that HA wear debris activate polymorphonuclear cells (PMNs) initiating and/or amplifying thereby the acute inflammatory response. Previous studies have shown that activation of monocytes by HA could be modulated by supplementing this latter with the divalent cation, Zinc. The purpose of this work was to investigate the modulation of PMNs activation following exposure to zinc-substituted HA. Our study demonstrate that addition of zinc to HA particles resulted in decreased levels of the pro-inflammatory mediator interleukin-8 (IL-8) and the matrix metallo-proteinase-9. We also show that these changes involve IL-8 receptors (CXCR-1 and CXCR-2).

In Vitro Release of Vascular Endothelial Growth Factor From Gadolinium-Doped Biodegradable Microspheres

A drug delivery vehicle was constructed that could be visualized noninvasively with MRI. The biodegradable polymer poly(DL‐lactic‐co‐glycolic acid) (PLGA) was used to fabricate microspheres containing vascular endothelial growth factor (VEGF) and the MRI contrast agent gadolinium diethylenetriamine pentaacetic acid (Gd‐DTPA). The microspheres were characterized in terms of size, drug and contrast agent encapsulation, and degradation rate. The PLGA microspheres had a mean diameter of 48 ± 18 μm. The gadolinium loading was 17 ± 3 μg/mg polymer and the VEGF loading was 163 ± 22 ng/mg polymer. Electron microscopy revealed that the Gd was dispersed throughout the microspheres and it was confirmed that the Gd loading was sufficient to visualize the microspheres under MRI. VEGF and Gd‐DTPA were released from the microspheres in vitro over a period of ∼6 weeks in three phases: a burst, followed by a slow steady‐state, then a rapid steady‐state. Biodegradable Gd‐doped microspheres can be effectively used to deliver drugs in a sustained manner, while being monitored noninvasively with MRI. Magn Reson Med 51:1265–1271, 2004. Published 2004 Wiley‐Liss, Inc.

Integration of dense HA rods into cortical bone

HA ceramics are daily used in human surgery for bone healing partly due to their ability to integrate into bone. They are generally used under a macroporous form. The behaviour of dense HA after implantation is not so well known. We implanted within cortical sheep femurs dense pure HA-ceramics cylinders for periods from 2 weeks to 18 months. The samples were then sectioned and examined using back-scattered and secondary SEM and the interface was analysed using EDS. Histomorphometry measurement was also performed using an image analysis device coupled to a light microscope. It appeared that the cylinders were in direct contact with immature bone after three weeks. The bone maturated within three months. The implant surface showed moderate signs of resorption and some grains were released from the surface. The resorption zone was only a few microm thick after 18 months. The bulk ceramic contained default zones of increased porosity. They can constitute fragile zone when located close to the surface in which the resorption rate is increased. We conclude that dense pure HA is poorly degraded when implanted in cortical bone. Degradation depends on the defaults found on the ceramic structure and the remodelling of bone surrounding the material.

Effect of strontium-substituted biphasic calcium phosphate on inflammatory mediators production by human monocytes

Calcium phosphate materials are widely used as bone substitutes because of their properties close to those of the mineral phase of bones. Nevertheless, after several months, calcium phosphate-based materials release particles that may be phagocytosed by monocytes, leading to an inflammatory reaction. Strontium is well known to counteract the osteoporosis process, but little is known about its effect on inflammatory processes. The purpose of this work was to study the effect of biphasic calcium phosphate (BCP) particles substituted with strontium on the inflammatory reaction. Human primary monocytes stimulated or not by lipopolysaccharide (LPS) were exposed to BCP particles containing strontium for 6 and 24 h. Inflammatory mediators (cytokines and matrix metalloproteinases (MMPs)) production was then quantified by ELISA and zymography. We observed that the presence of strontium had few effects on unstimulated cells, but it decreased the production of pro-inflammatory cytokines and the chemokine interleukin 8 in LPS-stimulated cell-conditioned medium. This work suggests for the first time that strontium may be involved in the control of inflammatory processes following BCP phagocytosis by human monocytes.