Composites between hydroxyapatite and poly(epsilon-caprolactone) synthesized in open system at room temperature.

The hydroxyls present on the surface of hydroxyapatite (HA) granules, annealed at 700 composite function, 900 composite function and 1,100 composite function C, are able to initiate the polymerization of epsilon-caprolactone (CL), not only at 185 composite function C under vacuum, but also at room temperature in open system. A polymer layer ionically linked to the substrate is formed on HA surface, enhancing the compatibility between the organic phase and the inorganic one in composite biomaterials. We studied the characteristics of the polymer, produced by the reaction carried out at room temperature in open system, as well as the percentages of the poly(epsilon-caprolactone) (PCL) ionically bonded to the HA structure and of the "free" one. Both percentages appear very dependent on the annealing temperature; in particular, HA annealed for 1 h at 1,100 composite function C is the most efficient initiator of the reaction leading to ionically bonded PCL. The percentages of "free" polymer are much higher than at 185 composite function C under vacuum. Its formation is attributed to the role of water in opening the CL rings, and to the presence of CO(3) (2-) and HPO(4) (2-) ions in the HA annealed at lower temperatures. The presence of water appears to be the limiting factor for the production of PCL not bonded to the HA structure.

Release of 5-fluorouracil by biodegradable poly(ester-ether-ester)s. Part I: release by fused thin sheets.

The 5-fluorouracil release by biodegradable epsilon-caprolactone and L-lactide copoly(ester-ether-ester)s was tested. The drug-copolymer mixture was formed by fusion in thin sheets, which were dipped in Dulbecco's PBS for time intervals ranging from one hour to two months. Each experiment shows a fast initial release, which subsequently slows down and stops at a limiting value, depending on the copolymer composition. This behavior was attributed to an extraction of the drug present on the sheet surface, due only to its shape, and to hydrogen bonds between the drug and the copolymers. The results obtained lead to a possibility of using such copolymers as "time-delayed" drug-releasing systems, when formed in specimens with smaller surface-to-volume ratio, which could minimize the fast initial extraction.

Six novel MEN1 gene mutations in sporadic parathyroid tumors.

We report nine mutations of the multiple endocrine neoplasia type 1 (MEN1) gene in sporadic parathyroid adenomas. Six of them have not previously been described: E60X, P32R, 261delA, 934+2T-->G, S443P, and 1593insC. The tissue samples were initially submitted to LOH analysis at 11q13 followed by SSCP screening of LOH-positive samples. Mutations were identified by direct sequencing and subcloning. Three (E60X, P32R, and 261delA) were in exon 2, one (934+2bp) in the splice junction of exon 5, one (S443P) in exon 9, and one (1593insC) in exon 10. The 3 mutations in exon 2 were associated with loss and/or creation of a restriction site. The corresponding germline sequence of the MEN1 gene was normal. Most mutations would likely result in a nonfunctional menin protein, and therefore in the loss of a tumor suppressor protein.

Parathyroid expression of calcium-sensing receptor protein and in vivo parathyroid hormone-Ca(2+) set-point in patients with primary hyperparathyroidism.

A reduced expression of calcium-sensing receptor (CaR) messenger ribonucleic acid and protein accompanied by abnormalities in parathyroid cell proliferation and PTH secretion are present in primary hyperparathyroidism. We studied the expression of CaR protein by immunohistochemistry in 36 sporadic parathyroid adenomas and investigated the relationship between CaR expression and several preoperative clinical parameters, including the set-point of Ca(2+)-regulated PTH secretion (measured in vivo). The adenomas were classified in 4 categories according to the intensity of immunohistochemical staining: 5 (14%) showed a CaR staining intensity similar to that of normal parathyroid ( ), 10 (27%) showed moderate staining (++), 16 (45%) showed weak staining (+), and 5 (14%) were negative (-). The intensity of CaR staining was not related to preoperative serum Ca(2+), PTH levels or adenoma volume. Twenty-nine patients underwent preoperatively the calcium infusion test to evaluate the PTH-Ca(2+) set-point. Individual values of PTH-Ca(2+) set-point ranged from 1.38-1.93 mmol/L and were significantly correlated with basal Ca(2+) levels (r = 0.96; P: = 0. 0001) and adenoma volume (r = 0.5; P: = 0.01). The mean PTH-Ca(2+) set-point values were significantly different in the 4 groups of patients classified according to immunohistochemical staining intensity of their adenoma (P: = 0.025; F = 3.78); the mean PTH-Ca(2+) set-point was significantly higher in the groups classified as negative than in those classified as weak or moderate. No correlation was observed between the PTH-Ca(2+) set-point and basal PTH levels or between the percent maximal PTH inhibition and adenoma volume and basal PTH or Ca(2+) levels. In summary, our data suggest that there is a relationship between apparent CaR protein expression and PTH-Ca(2+) set-point abnormality, suggesting that a reduced receptor content might have an important role in the pathogenesis of primary hyperparathyroidism.

Fibers by bioresorbable poly(ester-ether-ester)s as potential suture threads: a preliminary investigation.

Fibers made by a bioresorbable poly(epsilon-caprolactone)-block-poly(oxyethylene)-block-poly(epsilon-caprolactone) copolymer, having a number average molecular mass of about 200,000 Da and an average molar composition of 66% oxycaproyl units and 34% oxyethylene units, were melt-spun, with the aim at using them as suture threads. Their properties were investigated by the stress-strain test and by differential scanning calorimetry (DSC). The results obtained show that the properties of this material depend very strongly on the alignment of its macromolecules. In particular, the only partial alignment, obtainable by a relatively moderate drawing just after the extrusion, leads to values of elongation at break too high for use of the fibers as suture threads. The DSC analysis reveals interesting properties of the material, but also confirms their strong dependence on the extrusion procedure and on the mechanical treatment. In conclusion, the results of this preliminary study show that the spinning technique must be improved, and that further investigations are necessary to ascertain the possibility of using these poly(ester-ether-ester)s for the fabrication of suture threads.

Periodontal membranes from composites of hydroxyapatite and bioresorbable block copolymers.

Biomembranes are frequently proposed as devices for "guided bone regeneration." Such membranes consist generally of a thin sheet of polymeric material, mostly textured from polymeric yarns or clots, which all have a diffuse very fine winding porosity. The cross-section size of the holes of such porosity is nanometric (diameter < 0.1 microm); thus these holes can be indicated as nanoholes. Whatever the method of production, the surface density of nanoholes (number per square centimeter) has to be as high as possible. It is important also that no variation of this density occurs. The fine dimension of these microholes allows the crossing of small molecules (O2, CO2, H2O, sugars, many nutritional organic compounds and even some simple proteins) but not other larger molecules and particulates, including cells of any kind. These biomembranes have, consequently, a semipermeable behavior, providing the functional role which is the interposition of a barrier for the cells, separating the bone from the surrounding soft tissues. The kinetic of proliferation of osteoblasts is lower than that of fibroblasts. Most membranes of this kind are not resorbable. The main problem for the resorbable ones is the speed of size increase of the holes during the time. Their diameter must not exceed a threshold value until the reconstruction of bone is complete, otherwise soft tissue cells will invade the growing bone tissue with formation of undesirable mixed tissue. The present paper deals with a resorbable membrane made with a composite polymer/ceramic. A poly(epsilon-caprolactone)-block-poly(oxyethylene)-block-poly(epsilon-caprolactone) copolymer is the polymeric matrix which contains dispersed ceramic hydroxyapatite microgranules, a stiff filling additive. The main possible use is that of periodontal membranes. The copolymer, obtained by thermal polymerization of epsilon-caprolactone onto poly(ethylene-glycol), presents good biological tolerance, is resorbable under physiological conditions and can promote cell growth. Histological tests, performed 6 months after implantation, showed that the polymeric matrix is almost totally resorbed. New-formed bone colonizes even the innermost parts of the membrane, with bone trabeculae closely surrounding the hydroxyapatite granules.

New composites of hydroxyapatite and bioresorbable macromolecular material.

Composite materials were prepared by mixing in different proportions of hydroxyapatite (HA) and poly(epsilon-caprolactone-oxyethylene-epsilon-caprolactone) block copolymer (PCL-POE-PCL) to produce a new resorbable material for biomedical applications. This material has proved to be very interesting for production of periodontal membranes. Mechanical properties are linearly proportional to the amount of HA introduced. Fourier transform infrared (FTIR) investigations have pointed out that HA is able to influence some close epsilon-caprolactone molecules to start its homopolymerization giving PCL with an end chain ionic bonding. HA grains are therefore surrounded by a film of PCL which grants close connection of HA grains within copolymeric matrix. This interface bond with PCL is, however, an interesting occurrence for preparations of HA/PCL composites.

In vitro cytotoxicity testing of chitosan-containing polyelectrolyte complexes.

Two chitosan-containing polyelectrolyte complexes, chitosan-poly(acrylic acid) and chitosan-poly(styrenesulphonate), were synthesized by polymerizing acrylic acid and sodium styrenesulphonate in the presence of chitosan and chitosan hydrochloride, respectively. The complexes were studied by optical microscopy and tested for cytotoxicity by the Neutral Red uptake, Kenacid Blue R-Binding and 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyl tetrazolium bromide assays. The optical microscopy confirmed the differences in crystallinity and structure already found for the two polycomplexes by other characterization techniques. The cytoxicity tests showed different influences on the cell activity by the extracts of the two polyelectrolyte complexes. Such results were discussed and correlated to the different structures of the two materials.

Cell cultures in the biocompatibility study of synthetic materials.

In vitro cytotoxicity (Neutral Red uptake, Kenacid Blue and MTT) and cytocompatibility (cell adhesion and proliferation) tests were applied to the biocompatibility study of a series of poly(ester-ether-ester) block copolymers of potential interest as biomaterials. Our results indicate that the copolymer extracts after 72 hours incubation with a 3T3 mouse fibroblast cell line do not induce significant toxic effects. Furthermore, human umbilical vein endothelial cells seeded on thin copolymer films show a normal pattern of growth. We conclude that the in vitro tests used are a valid instrument to evaluate the potential toxic action of synthetic materials on different cell compartments and that the tested materials seem to be promising for future applications in the field of biomedical devices.

Cell cultures in the biocompatibility study of synthetic materials.

In vitro cytotoxicity (Neutral Red uptake, Kenacid Blue and MTT) and cytocompatibility (cell adhesion and proliferation) tests were applied to the biocompatibility study of a series of poly(ester-ether-ester) block copolymers of potential interest as biomaterials. Our results indicate that the copolymer extracts after 72 hours incubation with a 3T3 mouse fibroblast cell line do not induce significant toxic effects. Furthermore, human umbilical vein endothelial cells seeded on thin copolymer films show a normal pattern of growth. We conclude that thein vitro tests used are a valid instrument to evaluate the potential toxic action of synthetic materials on different cell compartments and that the tested materials seem to be promising for future applications in the field of biomedical devices.

On a mathematical model for body tissue inflammation.

In the present paper I will try to prove the mathematical validity of a model on the localized bacterial infection for tissue inflammation. This model was proposed by Lauffenburger and Kennedy, and it describes the inflammatory response to bacterial invasion of body tissue. I prove the mathematical validity of the model by means of a positivity theorem, an existence theorem and a uniqueness theorem. In spite of the apparent simplicity of the problem, the solution requires a delicate set of techniques. It seems very difficult to extend these techniques to a model in more than one dimension.