The FiberTAG project: Tagging dietary fibre intake by measuring biomarkers related to the gut microbiota and their interest for health.

The scientific rationale for dietary fibre intake recommendations comes from the recognition of their benefits for health based on studies first published many years ago. It remains unclear which are the key physiological effects generated by dietary fibre in view of the diversity of the food components considered as dietary fibre, of the relevance of their classification (soluble and insoluble) and from the recent discoveries putting forward their interactions with the gut microbiota. The project FiberTAG (Joint Programming Initiative 'A Healthy Diet for a Healthy Life' 2017-2020 https://www.fibertag.eu/) aims to establish a set of biomarkers (markers of gut barrier function and bacterial co-metabolites including volatile compounds and lipid derivatives), measured in different biological compartments (faeces, blood or breath) linking dietary fibre intake and gut microbiota-related health effects. The FiberTAG consortium brings together academic and industrial partners from Belgium, France, Germany and Canada to share data and samples obtained from existing as well as new intervention studies in order to evaluate the relevance of such biomarkers. The FiberTAG consortium is currently working on five existing cohorts (prospective observational or nutritional interventions in healthy or obese patients), and a number of new intervention studies to analyse the effect of insoluble dietary fibre (wheat bran and chitin-glucan, provided by the industrial partners) in healthy individuals or in obese patients at high cardiometabolic risk.

Single-blind, placebo controlled randomised clinical study of chitosan for body weight reduction.

BACKGROUND: Chitosan is a dietary fibre which acts by reducing fat absorption and thus used as a means for controlling weight. Weight loss clinical trial outcomes, however, have contradictory results regarding its efficacy. The primary objective of the present study was to evaluate the efficacy and safety of a chitosan from fungal origin in treatment of excess weight in the absence of dietary restrictions. METHODS: A phase IV, randomised, multicentre, single-blind, placebo-controlled, clinical study was conducted by administering chitosan capsules (500 mg, five/day) and indistinguishable placebo capsules as daily supplements to 96 overweight and obese subjects for 90 days. The study participants were divided in 2:1 ratio to receive either chitosan (n = 64) or placebo (n = 32). Efficacy was assessed by measuring body weight, body composition parameters, anthropometric measurements, HbA1C level and lipid profile at day 45 and day 90. Also, short form-36 quality of life (QoL) questionnaire was assessed to evaluate improvement in life-style and dietary habits were recorded for calorie intake. Safety was assessed by evaluating safety parameters and monitoring adverse events. RESULTS: The mean changes in body weight were -1.78 ± 1.37 kg and -3.10 ± 1.95 kg at day 45 and day 90 respectively in chitosan group which were significantly different (p < 0.0001) as compared to placebo. BMI was decreased by10.91 fold compared to placebo after 90 day administration. In concert with this, there was also reduction in body composition and anthropometric parameters together with improvement in QoL score. Chitosan was also able to reduce HbA1C levels (below 6 %) in subjects who had initial higher values. The mean caloric intake shows that there was no change in dietary habits of subjects in both groups. Lipid levels were unaffected and all adverse events were mild in nature and unrelated to study treatment. CONCLUSION: Chitosan from fungal origin was able to reduce the mean body weight up to 3 kg during the 90 day study period. Together with this, there was also improvement in body composition, anthropometric parameters and HbA1C, reflecting overall benefits for the overweight individuals. Additionally, there was also improvement in QoL score. It was safe and well tolerated by all subjects. TRIAL REGISTRATION: CTRI/2014/08/004901.

Protective effect of a new biomaterial against the development of experimental osteoarthritis lesions in rabbit: a pilot study evaluating the intra-articular injection of alginate-chitosan beads dispersed in an hydrogel.

OBJECTIVE: This study aimed to evaluate the structural benefit of a new biomaterial composed of alginate-chitosan (AC) beads dispersed in a hydrogel (H) derived from chitosan on the development of osteoarthritis (OA) in rabbit. DESIGN: OA was induced by the surgical transection of the anterior cruciate ligament in rabbits. Animals received a single intra-articular injection (900 µl) of AC beads in H hydrogel, H hydrogel alone or saline a week after surgery. OA development was followed by X-rays. Blood samples were collected throughout the study to measure biological markers (Prostaglandins E2 - PGE2 and C reactive protein - CRP). Macroscopic observation and histological evaluation of articular cartilage and synovial membrane were performed 6 weeks after surgery. RESULTS: AC beads in H hydrogel prevented from the development of OA based on the reduction of the Kellgren & Lawrence (K&L) score. It also significantly reduced the histological score of cartilage lesion severity. This effect was homogenous on every joint compartment. It was due to a significant effect on cartilage structure and cellularity scores. The injection of AC beads in H hydrogel also tended to reduce the synovial membrane inflammation. No significant variation of biological markers was noted. CONCLUSIONS: The present pilot study provides interesting and promising results for the use of AC beads in H hydrogel in animal. It indeed prevented the development of OA cartilage lesions without inflammatory signs. The potencies of this biomaterial to protect OA joint should be further documented. It could then represent a new alternative for viscosupplementation in human OA management.

Chitin-glucan fiber effects on oxidized low-density lipoprotein: a randomized controlled trial.

BACKGROUND/OBJECTIVES: Elevated oxidized low-density lipoprotein (OxLDL) may promote inflammation, and is associated with increased risk of atherosclerotic coronary heart disease and worsening complications of diabetes mellitus. The primary objective of this study was to evaluate the efficacy of chitin-glucan (CG), alone and in combination with a potentially anti-inflammatory olive oil (OO) extract, for reducing OxLDL in subjects with borderline to high LDL cholesterol (LDL-C) levels. SUBJECTS/METHODS: This 6-week, randomized, double-blind, placebo-controlled study of a novel, insoluble fiber derived from the Aspergillus niger mycelium, CG, evaluated 130 subjects free of diabetes mellitus with fasting LDL-C 3.37-4.92 mmol/l and glucose ≤ 6.94 mmol/l. Participants were randomly assigned to receive CG (4.5 g/day; n=33), CG (1.5 g/day; n=32), CG (1.5 g/day) plus OO extract (135 mg/day; n=30), or matching placebo (n=35). RESULTS: Administration of 4.5 g/day CG for 6 weeks significantly reduced OxLDL compared with placebo (P=0.035). At the end of study, CG was associated with lower LDL-C levels relative to placebo, although this difference was statistically significant only for the CG 1.5 g/day group (P=0.019). CG did not significantly affect high-density lipoprotein cholesterol, triglycerides, glucose, insulin or F2-isoprostane levels. Adverse events did not substantively differ between treatments and placebo. CONCLUSIONS: In this 6-week study, CG (4.5 g/day) reduced OxLDL, an effect that might affect the risk for atherosclerosis.

Long-term in vitro degradation of PDLLA/bioglass bone scaffolds in acellular simulated body fluid.

The long-term (600days) in vitro degradation of highly porous poly(D,L-lactide) (PDLLA)/Bioglass-filled composite foams developed for bone tissue engineering scaffolds has been investigated in simulated body fluid (SBF). Foams of ∼93% porosity were produced by thermally induced phase separation (TIPS). The degradation profile for foams of neat PDLLA and the influence of Bioglass addition were comprehensively assessed in terms of changes in dimensional stability, pore morphology, weight loss, molecular weight and mechanical properties (dry and wet states). It is shown that the degradation process proceeded in several stages: (a) a quasi-stable stage, where water absorption and plasticization occurred together with weight loss due to Bioglass particle loss and dissolution, resulting in decreased wet mechanical properties; (b) a stage showing a slight increase in the wet mechanical properties and a moderate decrease in dimensions, with the properties remaining moderately constant until the onset of significant weight loss, whilst molecular weight continued to decrease; (c) an end stage of massive weight loss, disruption of the pore structure and the formation of blisters and embrittlement of the scaffold (evident on handling). The findings from this long-term in vitro degradation investigation underpin studies that have been and continue to be performed on highly porous poly(α-hydroxyesters) scaffolds filled with bioactive glasses for bone tissue engineering applications.

Subchronic (13-week) oral toxicity study in rats with fungal chitin-glucan from Aspergillus niger.

Chitin-glucan is an insoluble biopolymer, composed of chitin and beta-(1,3)-D-glucan, that is a component of the fungal cell wall. This study was conducted to assess the safety of chitin-glucan from the mycelium of Aspergillus niger (Artinia brand) for use as dietary supplement and food ingredient. Chitin-glucan was fed to Wistar rats (20/sex/group) at dietary levels of 0, 1, 5 and 10% for 13 weeks. Clinical and neurobehavioural observations, growth, feed and water consumption, ophthalmoscopy, haematology, clinical chemistry, urinalysis, organ weights, necropsy and histopathological examination revealed no adverse effects of chitin-glucan. Rats fed chitin-glucan at 10% consumed more feed than controls, probably due to lower energy density of their diet. Water intake was increased slightly at all dose levels. These changes were not toxicologically significant. Full and empty caecum weights were increased in mid-dose males and high-dose males and females. This caecal enlargement was a physiological response to the consumption of a high amount of poorly digestible carbohydrate and considered of no toxicological concern. In conclusion, feeding chitin-glucan at dietary levels up to 10% for 13 weeks was tolerated without any signs of toxicity. This level corresponded to 6.6 and 7.0 g chitin-glucan/kg body weight/day in male and female rats, respectively.

Controlled release of drugs from multi-component biomaterials.

In order to control their release, drugs are encapsulated into systems which are expected to provide a certain site with a predetermined amount of drug over a well-defined period of time. Here we report on a multi-component drug delivery biomaterial that consists of a hydrogel matrix in which drug-loaded biodegradable microcarriers are dispersed, and whose potential applications could be found in the design of implantable devices with long-term activity, as required by contraceptive and hormone replacement treatments. The release profile of the drug can actually be tuned by the complex interplay of several release mechanisms, including the permeability and eventually the degradation rate of the microcarriers and the diffusion through the hydrogel. The hydrogel consisted of 2-hydroxyethyl methacrylate cross-linked by ethylene glycol dimethacrylate. The microcarriers were biodegradable poly-epsilon-caprolactone (PCL) microspheres in which active molecules, such as levonorgestrel (LNG), were encapsulated. The hydrogels were characterized by water swelling, thermal properties, LNG diffusion through drug-free and drug-depleted hydrogel membranes and LNG release from devices with drug dispersed in the hydrogel. The PCL microspheres were observed by scanning electron microscopy; their size distribution, LNG loading and release were also investigated. The hydrogel-microsphere assemblies were characterized in terms of the distribution of the microspheres within the hydrogel, water swelling and the release of the encapsulated molecules. The developed device, due to its composite structure, has the ability to combine several release mechanisms, leading to drug release obeying zero-order kinetics for most of the time.

Vaterite deposition on biodegradable polymer foam scaffolds for inducing bone-like hydroxycarbonate apatite coatings.

Hydroxycarbonate apatite (HCA) coatings on the surface of bioresorbable materials for bone tissue engineering scaffolds were produced using macroporous poly(DL-lactide) (PDLLA) foams impregnated by calcium carbonate in vaterite crystalline form. Stable and homogeneous vaterite deposition on PDLLA foams was achieved using a slurry dipping technique. In vitro studies in simulated body fluid (SBF) were performed to induce formation of (HCA) on the surface of vaterite/PDLLA composite foams. HCA was detected after immersion of foams in SBF for 7 days. Hence, depositing vaterite on materials followed by immersion in SBF is confirmed to induce HCA coatings on the surface of the material. The HCA coated, bioactive and resorbable PDLLA foams are intended for use as bone tissue engineering scaffolds.

Prolonged maintenance of neointestine using subcutaneously implanted tubular scaffolds in a rat model.

Tissue-engineered small intestine offers a possible alternative to long-term parenteral nutrition or intestinal transplantation in patients with short bowel syndrome. The aim of this study was to investigate the prolonged development of neointestine grown on subcutaneously implanted scaffolds. Tubular polylactide-coglycolide (PLGA) scaffolds were implanted into adult Lewis rats. Four weeks after scaffold implantation, a suspension of organoid units was delivered to the lumen of each scaffold. Organoid units were manufactured from small intestine harvested from neonatal Lewis rats by partial digestion using collagenase and dispase. Scaffolds were removed at 4, 8, and 12 weeks after organoid unit implantation, processed to paraffin, and sectioned. Hematoxylin and eosin staining demonstrated well-developed and well-differentiated intestinal mucosa and a vascularised submucosa within the scaffolds at 4, 8, and 12 weeks. Appearances were similar to native small intestine. Immunohistochemistry performed using primary antibody against proliferating cell nuclear antigen, a marker for cellular proliferation, demonstrated positively staining cells within the mucosa and submucosa at all time points. In the mucosal layer these positively staining cells were found primarily in the crypts. These findings show that neointestinal mucosa can be maintained for at least 12 weeks on a subcutaneous PLGA scaffold, and the presence of actively proliferating cells at 12 weeks suggests potential for further development beyond this.

Olfactory ensheathing cells, olfactory nerve fibroblasts and biomatrices to promote long-distance axon regrowth and functional recovery in the dorsally hemisected adult rat spinal cord.

Cellular transplantation, including olfactory ensheathing cells (OEC) and olfactory nerve fibroblasts (ONF), after experimental spinal cord injury in the rat has previously resulted in regrowth of severed corticospinal (CS) axons across small lesion gaps and partial functional recovery. In order to stimulate CS axon regrowth across large lesion gaps, we used a multifactorial transplantation strategy to create an OEC/ONF continuum in spinal cords with a 2-mm-long dorsal hemisection lesion gap. This strategy involved the use of aligned OEC/ONF-poly(D,L)-lactide biomatrix bridges within the lesion gap and OEC/ONF injections at 1 mm rostral and caudal to the lesion gap. In order to test the effects of this complete strategy, control animals only received injections with culture medium rostral and caudal to the lesion gap. Anatomically, our multifactorial intervention resulted in an enhanced presence of injured CS axons directly rostral to the lesion gap (65.0 +/- 12.8% in transplanted animals versus 13.1 +/- 3.9% in control animals). No regrowth of these axons was observed through the lesion site, which may be related to a lack of OEC/ONF survival on the biomatrices. Furthermore, a 10-fold increase of neurofilament-positive axon ingrowth into the lesion site as compared to untreated control animals was observed. With the use of quantitative gait analysis, a modest recovery in stride length and swing speed of the hind limbs was observed. Although multifactorial strategies may be needed to stimulate repair of large spinal lesion gaps, we conclude that the combined use of OEC/ONF and poly(D,L)-lactide biomatrices is rather limited.

Chronically injured corticospinal axons do not cross large spinal lesion gaps after a multifactorial transplantation strategy using olfactory ensheathing cell/olfactory nerve fibroblast-biomatrix bridges.

Transplantation of mixed cultures containing olfactory ensheathing cell (OEC) and olfactory nerve fibroblasts (ONF) has been shown to stimulate regrowth of both acutely and chronically injured corticospinal (CS) axons across small spinal cord lesion gaps. Here, we used a multifactorial transplantation strategy to stimulate regrowth of chronically injured CS axons across large spinal cord lesion gaps. This strategy combined the transplantation of aligned OEC/ONF-biomatrix complexes, as described previously (Deumens et al. [2004] Neuroscience 125:591-604), within the lesion gap with additional OEC/ONF injections rostral and caudal to the lesion site. We show an enhanced presence of injured CS axons directly rostral to the lesion gap, with no effects on injured CS axons at or caudal to the lesion gap. Furthermore, injured CS axons did not penetrate the OEC/ONF-biomatrix complex within the lesion gap. The enhanced presence of CS axons rostral to the lesion gap was not accompanied by any recovery of behavioral parameters assessed with the BBB locomotor rating scale or CatWalk gait analysis. We conclude that our multifactorial transplantation strategy should be optimized to create an OEC/ONF continuum in the injured spinal cord and thereby stimulate regrowth of injured CS axons across large spinal lesion gaps.

A pilot study investigating a novel subcutaneously implanted pre-cellularised scaffold for tissue engineering of intestinal mucosa.

Tissue engineering of the small intestine offers an alternative to long-term intravenous nutrition and transplantation in patients with intestinal failure. Initial work, although encouraging, is limited by the volume of neonatal tissue required to produce a small neomucosal cyst. Our novel approach is to implant tubular poly-lactide-co-glycolide (PGLA) foam scaffolds subcutaneously. The aim of this study was to investigate whether these scaffolds would support growth of intestinal neomucosa. PGLA scaffolds were implanted subcutaneously into 8 Lewis rats; after 5 weeks, 'organoid units' were injected into the lumens. Tissue was assessed histologically after harvesting and quantitative immunohistochemistry was performed using antibodies against vascular endothelial growth factor (VEGF), vascular endothelial growth factor receptor 2 (VEGF-R2), fibroblast growth factor basic (bFGF) and fibroblast growth factor receptor 2 (FGF-R2). At 4 weeks post organoid unit implantation, clearly recognisable mucosa and submucosa was present on the luminal surface of the scaffold. Densities of VEGF and VEGF-R2 positive cells increased with time post organoid unit implantation. This pilot study demonstrates that it is possible to tissue engineer small intestinal neomucosa using subcutaneously implanted PLGA scaffolds. The yield of the process compares favourably to the published literature. Further work is required to optimise the technique.

Biocompatibility of poly-DL-lactic acid (PDLLA) for lung tissue engineering.

This study explores the possibility of growing lung cells on poly-DL-lactic acid (PDLLA) scaffolds, with a view to in future engineer pulmonary tissue for human implantation. As a first step in this process, the ability of PDLLA to maintain the growth of lung epithelium is tested using a robust cell line. Poly-DL-lactic acid has been investigated in two forms, as planar discs and as 3-D foams, and it has been demonstrated that PDLLA is not only nontoxic to pneumocytes but it also actively supports their growth. The initial findings suggest that the material is an appropriate matrix for engineering of distal lung tissue.

Study of the connectivity properties of Bioglass -filled polylactide foam scaffolds by image analysis and impedance spectroscopy.

The porous structure of two series of poly(D,L-lactide)/Bioglass composite foams prepared by thermal-induced phase separation was investigated by image analysis and impedance spectroscopy. Polymer solutions of either low or high molecular weight containing different concentrations (up to 50 wt.%) of Bioglass particles of mean particle size d < 5 microm were studied. The morphology of both macro- and micropores was studied by scanning electron microscopy and image analysis of both neat and composite foams (containing 10-50 wt.% Bioglass). The pore connectivity of both neat polymer and composite foams was characterized by impedance spectroscopy in relation with their transport properties. The influence of the foam composition (i.e., polymer molecular weight and concentration of Bioglass on pore microstructure was studied using these non-destructive methods. It was found that addition of Bioglass particles has a pronounced effect on pore orientation, leading to increasing loss of order of pore structure, especially for low-molecular weight PDLLA foams.

Mechanical properties of highly porous PDLLA/Bioglass composite foams as scaffolds for bone tissue engineering.

This study developed highly porous degradable composites as potential scaffolds for bone tissue engineering. These scaffolds consisted of poly-D,L-lactic acid filled with 2 and 15 vol.% of 45S5 Bioglass particles and were produced via thermally induced solid-liquid phase separation and subsequent solvent sublimation. The scaffolds had a bimodal and anisotropic pore structure, with tubular macro-pores of approximately 100 microm in diameter, and with interconnected micro-pores of approximately 10-50 microm in diameter. Quasi-static and thermal dynamic mechanical analysis carried out in compression along with thermogravimetric analysis was used to investigate the effect of Bioglass on the properties of the foams. Quasi-static compression testing demonstrated mechanical anisotropy concomitant with the direction of the macro-pores. An analytical modelling approach was applied, which demonstrated that the presence of Bioglass did not significantly alter the porous architecture of these foams and reflected the mechanical anisotropy which was congruent with the scanning electron microscopy investigation. This study found that the Ishai-Cohen and Gibson-Ashby models can be combined to predict the compressive modulus of the composite foams. The modulus and density of these complex foams are related by a power-law function with an exponent between 2 and 3.

Alignment of glial cells stimulates directional neurite growth of CNS neurons in vitro.

Olfactory ensheathing cells (OECs) together with olfactory nerve fibroblasts (ONFs) and neonatal astrocytes are potent stimulators of neurite growth in adulthood and during development, respectively. Since it is known that alignment of glial cells is important for the correct outgrowth of axon tracts, it was hypothesized that the alignment of glial cells stimulates directional and enhanced neurite outgrowth. Adult OEC/ONF and neonatal astrocytes were cultured either on biodegradable poly(d,l)-lactide matrices or in Petri dishes for 4 days. Thereafter neonatal cerebral cortical neurons were added. After a 2-days coculture period the cultures were fixed and processed for a combined MAP-2 and phosphorylated neurofilament (RT97) staining. The neurite growth (neurite elongation and neurite formation) and the neurite direction were assessed. We show that (1). OEC/ONF cultures are more potent in stimulating the length of the longest neurite of cocultured neurons, (2). alignment of glial is achieved in vitro on our biomatrices, (3). aligned glial/biomatrix complexes do not enhance neurite growth, and (4). aligned glial/biomatrix complexes direct neurite outgrowth. These data have significant implications for in vivo experiments focusing on glial transplantation. Transplanting glial/biomatrix complexes may stimulate the directional regrowth of severed axons across a lesion site.

Porous poly(alpha-hydroxyacid)/Bioglass composite scaffolds for bone tissue engineering. I: Preparation and in vitro characterisation.

Highly porous composites scaffolds of poly-D,L-lactide (PDLLA) and poly(lactide-co-glycolide) (PLGA) containing different amounts (10, 25 and 50 wt%) of bioactive glass (45S5 Bioglass)were prepared by thermally induced solid-liquid phase separation (TIPS) and subsequent solvent sublimation. The addition of increasing amounts of Bioglass into the polymer foams decreased the pore volume. Conversely, the mechanical properties of the polymer materials were improved. The composites were incubated in phosphate buffer saline at 37 degrees C to study the in vitro degradation of the polymer by measurement of water absorption, weight loss as well as changes in the average molecular weight of the polymer and in the pH of the incubation medium as a function of the incubation time. The addition of Bioglass to polymer foams increased the water absorption and weight loss compared to neat polymer foams. However, the polymer molecular weight, determined by size exclusion chromatography, was found to decrease more rapidly and to a larger extent in absence of Bioglass. The presence of the bioactive filler was therefore found to delay the degradation rate of the polymer as compared to the neat polymer foams. Formation of hydroxyapatite on the surface of composites, as an indication of their bioactivity, was recorded by EDXA, X-ray diffractometry and confirmed by Raman spectroscopy.

In vitro evaluation of novel bioactive composites based on Bioglass-filled polylactide foams for bone tissue engineering scaffolds.

Highly porous poly(DL-lactic acid) (PDLLA) foams and Bioglass-filled PDLLA composite foams were characterized and evaluated in vitro as bone tissue engineering scaffolds. The hypothesis was that the combination of PDLLA with Bioglass in a porous structure would result in a bioresorbable and bioactive composite, capable of supporting osteoblast adhesion, spreading and viability. Composite and unfilled foams were incubated in simulated body fluid (SBF) at 37 degrees C to study the in vitro degradation of the polymer and to detect hydroxyapatite (HA) formation, which is a measure of the materials' in vitro bioactivity. HA was detected on all the composite samples after incubation in SBF for just 3 days. After 28 days immersion the foams filled with 40 wt % Bioglass developed a continuous layer of HA. The formation of HA for the 5 wt % Bioglass-filled foams was localized to the Bioglass particles. Cell culture studies using a commercially available (ECACC) human osteosarcoma cell line (MG-63) were conducted to assess the biocompatibility of the foams and cell attachment to the porous substrates. The osteoblast cell infiltration study showed that the cells were able to migrate through the porous network and colonize the deeper regions within the foam, indicating that the composition of the foams and the pore structures are able to support osteoblast attachment, spreading, and viability. Rapid formation of HA on the composites and the attachment of MG-63 cells within the porous network of the composite foams confirms the high in vitro bioactivity and biocompatibility of these materials and their potential to be used as scaffolds in bone tissue engineering and repair.

Preparation of macroporous biodegradable poly(L-lactide-co-epsilon-caprolactone) foams and characterization by mercury intrusion porosimetry, image analysis, and impedance spectroscopy.

Two poly(L-lactide-co-epsilon-caprolactone) random copolymers containing 5 and 40 mol % of epsilon-CL, namely P(LA-co-CL(5)) and P(LA-co-CL(40)), respectively, have been made macroporous by freeze-drying solutions in dimethylcarbonate. Most of the freeze-dried foams, prepared by varying polymer concentration and cooling rate, exhibited two main pore populations: (1). longitudinally oriented tube-like macropores with diameters >or=100 microm, and (2). interconnected micropores (10-100 microm). Pore characteristics, including macropore density, mean diameter, and interdistance, as well as micropore density, area, and shape, were determined by image analysis of scanning electron micrographs in order to study the influence of processing and formulation parameters on foam structure and properties. The pore orientation and the 3-D texture also were studied by image analysis and impedance spectroscopy. In the case of the P(LA-co-CL(5)), the macropore diameter increased with the cooling rate while the micropore diameter decreased. The micropores also became more circular when the cooling rate was increased. The pore size and morphology of the P(LA-co-CL(40)) were quite unchanged by varying the cooling rate. All the other conditions being the same, the P(LA-co-CL(5)) foams were better organized than the P(LA-co-CL(40)) foams, and pore orientation was improved at the higher cooling rate. Pore size and morphology also can be controlled by changing the polymer concentration (Cp), as we showed by studying P(LA-co-CL(5)) foams prepared by freeze-drying solutions in the 1-10 w/v % Cp range. Macropore density, average diameter, and interdistance of P(LA-co-CL(5)) foams increased with Cp, but the micropore characteristics remained almost unchanged no matter the Cp. The reliability of the characterization methods has been discussed, with special attention to mercury intrusion porosimetry, which is used primarily for measurement of pore volume and pore size distribution. However, this technique is reported here as a destructive and unreliable method for the characterization of fragile P(LA-co-CL(40)) foams. This study shows that image analysis and impedance spectroscopy can give reliable information relative to the pore morphology and anisotropy of freeze-dried foams.

Preparation, characterization, and in vitro degradation of bioresorbable and bioactive composites based on Bioglass-filled polylactide foams.

Highly porous poly(D,L-lactide)/Bioglass composites scaffolds were prepared by thermally induced phase separation process of polymer solutions and subsequent solvent sublimation. A series of composite foams with different polymer/Bioglass weight ratios was prepared to study the influence of Bioglass content on the foam characteristics such as porous structure, density, and pore volume. The pore volume was decreased from 9.5 to 5.7 cm(3)/g when the Bioglass content was increased up to 40 wt %, but the overall pore morphology was not affected very much by changing the polymer/glass composition ratio. The composites foams were then incubated in phosphate-buffered saline at 37 degrees C to study the in vitro degradation of the polymer and to detect hydroxyapatite (HA) formation as an indication of their bioactivity. The addition of Bioglass to polymer foams increased the water absorption and weight loss as compared with pure polymer foams. However, the polymer molecular weight, determined by size exclusion chromatography, was found to decrease more rapidly and to a larger extent in absence of Bioglass. This delayed degradation rate in the composite foams was probably caused by the dissolution of alkaline ions from the Bioglass, resulting in a buffering effect of the incubation medium. After incubation for 7 days, HA was detected by X-ray diffractometry and Raman spectroscopy and confirmed by environmental scanning electron microscopy and energy-dispersive X-ray analysis. The porous composites developed here are promising materials for bone regeneration applications because the formation of HA on the surface of the pore walls should provide good environment for the adhesion and proliferation of osteoblasts and osteoprogenitor cells.