Biological effects of resin monomers on oral cell populations: descriptive analysis of literature.

AIM: Recently, the application of restorative materials containing metacrilate monomers in the conservative and paediatric dentistry has focused on the possible negative effects due to the use of these composites. In particular the release of monomers from reconstructions as a result of an insufficient polymerisation, can spread along the mucosal and dental tissues with potential immunological ed cytotoxic effects. Regarding to the importance of this issue, the aim of this study is to provide a descriptive review of the literature on potential local and systemic interactions of metacrylic and acrylic monomers with the immune system, both in vitro and in vivo. RESULTS: The most highly used monomers in composite materials applied in conservative dentistry include: 2-hydroessietil- methacrylate (HEMA), triethylene glycol-dimethacrylate (TEGDMA), bisphenol A glycidyl-methacrylate (BisGMA) and urethane- dimethacrylate (UDMA). Different investigations have been performed for better understanding of the potential side effects of metacrylic monomers on immune system cells. Different factors such as cell population, exposure time and parameters more strictly connected to these materials, such as molecular weight, chemical composition and mechanical characteristics, seem to be directly involved in these reactions.

Long term effects of cigarette smoke extract or nicotine on nerve growth factor and its receptors in a bronchial epithelial cell line.

Long-term exposure to cigarette smoke induces severe injuries to respiratory system through several mechanisms, some of them are well defined, but many others are not yet elucidated. Beside its classical role in nervous system, we have previously shown that Nerve Growth Factor (NGF) and its receptors have a crucial role in airway inflammatory diseases, such as Chronic Obstructive Pulmonary Disease. To expand our knowledge about the relevance of NGF and its receptors in airway diseases induced by cigarette smoking, we exposed for 16 weeks the bronchial epithelial cell line BEAS-2B to sub-toxic concentrations of whole cigarette smoke extract or pure nicotine. Viability, cell cycle gene expression, cell morphology and migration ability were tested and compared to NGF release and gene expression. Modulation of its receptors TrKA and p75NTR was also analyzed. The present study shows that long term exposure of BEAS-2B cells to cigarette smoke extract or nicotine induces: (A) differences: in cell viability, in the expression of cell cycle-related genes, in NGF release and in gene expression of NGF and its receptors; (B) similarities: in morphology and migration ability. Taken together, our data provide new insights about the biological role of NGF and its receptors in airway diseases induced by long-term cigarette smoking and, finally, our data evidence the opportunity not to use nicotine lozenges or e-cigarettes as anti smoking replacement therapy in patients with a previous airway disease according to the ability of nicotine to increase the amount of the pro-inflammatory cytokine NGF into the bronchial environment.

In vitro human osteoblast and extracellular matrix changes after transforming growth factor beta 1 treatment.

AIMS: Normal bone tissue is characterised by a balancing of osteoblast and osteoclast activity. The activity and differentiation of these cells are regulated by vitamins, hormones and cytokines. The action of these factors on bone tissue cells depends on the composition and mineralisation of extracellular bone matrix. In particular, transforming growth factor beta 1 (TGFbeta1) acts on collagen fibres, glycosaminoglycan secretion and on the enzymes correlated to the turnover of glycosaminoglycans. The normal functions of bone tissue also depend on its mineralisation, which is highly altered in the process of uraemia. METHODS: In this study, we analysed in vitro the effect of transforming growth factor beta on osteoblast proliferation, collagen synthesis and glycosaminoglycan secretion with 3H-thymidine, 3H-proline or 3H-glucosamine incorporation, and on enzymes, such as beta-N-acetyl-D-glucosaminidase and beta-glucuronidase, involved in extracellular matrix turnover. Moreover, phosphatase alkaline activity and osteocalcin related to mineralisation of extracellular matrix were determined. RESULTS: Our data show that TGFbeta1 significantly decreases 3H-thymidine and 3H-proline incorporation and increases (p < or = 0.01) extracellular sulphated glycosaminoglycan synthesis. It also increases osteocalcin levels, phosphatase alkaline, beta-N-acetyl-D-glucosaminidase and beta-glucoronidase activities. CONCLUSION: TGFbeta1 changes the synthesis of extracellular matrix components by osteoblasts. These variations favour the action of cytokine and osteoclasts. Since the TGFbeta1 accumulates in bone tissue and increases during uraemia, with due limitations this action leads to an imbalance between synthesis and degradation and could explain bone alterations in uraemic patients.

TGFbeta and TGFalpha, antagonistic effect in vitro on extracellular matrix accumulation by chick skin fibroblasts at two distinct embryonic stages.

ECM macromolecules create a specific environment that participates in the control of cell proliferation and differentiation during embryogenesis. Quantitative and qualitative alterations in the ECM may depend on several growth factors that modify cell metabolism. Since transforming growth factor beta (TGFbeta) and alpha (TGFalpha) are abundantly expressed during embryonic development in organs in which epithelial-mesenchymal interactions occur, the aim of this study was to determine: a) the effect of TGFbeta on the phenotype of 7 and 14 day chick embryo back skin (CEBS) fibroblasts by evaluating the neosynthesis of GAG, collagen and fibronectin; b) whether TGFalpha and TGFbeta production, in particular TGFbeta3 and TGFbeta4, and the number of TGFbeta receptors change during these two stages of embryonic development. The results show that the neosynthesis of ECM macromolecules, tested using radiolabelled precursors, is increased by TGFbeta. The growth factor generally favours cellular accumulation more than secretion. As far as GAG is concerned, TGFbeta has a greater stimulatory effect on sulphated GAG than on HA. Specific bioassay shows that TGFbeta3 and TGFbeta4 activity is higher in 7 day than 14 day CEBS fibroblasts. Moreover, TGFbeta3 and TGFbeta4 mRNA expression is increased in the first stages of development. Instead, the level of TGFalpha increases in successive developmental stages. Since TGFalpha stimulates the synthesis and secretion of HA, and HA binds and inactivates TGFbeta, the greater quantity of HA in 14 day fibroblasts may contribute to reducing the TGFbeta effect. Overall our data suggest that the production of TGFbeta and TGFalpha are age-dependent and that the balance between the two growth factors may be a mechanism for controlling skin differentiation.

Synthesis and secretion of transforming growth factor-beta1 by human desmoid fibroblast cell line and its modulation by toremifene.

The present study provides evidence that the in vitro cultured fibroblast cell line from desmoid tumors differs from normal fibrobasts in its extracellular matrix (ECM) macromolecule composition and is modulated by treatment with toremifene, an antiestrogen that reduces tumor mass by an unknown mechanism. The results showed increased transforming growth factor-beta 1 (TGF-beta1) production, TGF-beta1 mRNA expression, and TGF-beta1 receptor number in desmoid fibroblasts compared with normal cells. As desmoid fibroblasts did not produce tumor necrosis factor-alpha (TNF-alpha) but were sensitive to it, which enhanced glycosaminoglycans (GAG) accumulation, we assessed the TGF-beta1 effects on TNF-alpha production by human monocytes. Our results showed TGF-beta1 significantly increased TNF-alpha secretion by monocytes. Toremifene mediated its effects in desmoid fibroblasts via an estrogen receptor-independent pathway. It inhibited GAG accumulation and the secretion of both latent and active forms of TGF-beta1 and had an inhibitory effect on TNF-alpha production by monocytes. Our results suggest that in reducing TGF-beta1 production by desmoid fibroblasts and TNF-alpha production by monocytes, toremifene may restore the balance between the two growth factors.

Coronary stenting for unstable angina: predictors of 30-day and long-term clinical outcome.

BACKGROUND: Unstable angina is usually caused by acute thrombosis superimposed on a fissured plaque. Coronary artery stenting has been shown to improve short- and long-term results of coronary angioplasty in mainly stable patients with one-vessel disease, but it is uncertain whether its use in an unstable clinical setting can be safe and useful. This study sought to evaluate the results of coronary stenting in unstable angina and to determine patient, lesion and procedure-related predictors of 30-day and long-term ischemic events. METHODS: We studied 266 consecutive patients (mean age 62 +/- 9 years) with unstable angina who underwent coronary artery stenting. The procedure was performed electively in 24%, in bailout situations in 11% and for a suboptimal result of conventional angioplasty in 65%. After stent implantation, patients were treated with anticoagulation (61) on combined antiplatelet therapy (200). Multivariate logistic regression analyses were performed to determine 30-day and long-term predictive factors of ischemic complications. RESULTS: Procedural success was obtained in 261 patients (98.1%). During the first 30 days after stenting, one patient died from cardiogenic shock (0.3%) and six (22%) suffered a non-fatal Q-wave myocardial infarction. Patients with combined antiplatelet therapy had a significantly lower stent thrombosis rate (1.5% versus 11.4%, P = 0.002) than those treated with anticoagulant regimen. At long-term follow-up (17.7 +/- 9.4 months) cardiac mortality myocardial infarction and target-vessel revascularization rates were 0.4%, 1.5% and 9.3%, respectively. In multivessel and diabetic patients, a worse long-term event-free survival was observed. Logistic multivariate analysis revealed bailout stenting, anticoagulant therapy, implantation of stents longer than 15 mm as predictors of 30-day ischemic events. In addition, multivessel coronary artery disease and stent application with balloon size of less than 3 mm were predictive of long-term ischemic events. CONCLUSIONS: This study demonstrates that, either electively or after failure of conventional angioplasty, coronary stenting represents an effective therapy for patients with unstable angina. In the same clinical setting, combined antiplatelet therapy is associated with a lower 30-day stent thrombosis rate than anticoagulant therapy. Bailout stenting, anticoagulant therapy, implantation of stents longer than 15 mm were shown to be predictors of 30-day ischemic events, whereas multivessel coronary artery disease and stent application with small balloon size were predictive of long-term ischemic events.

Ornithine decarboxylase, polyamines and CD11b expression in HL-60 cells during differentiation induced by retinoic acid.

Polyamines (PA) and retinoic acid affect mammalian cell growth, differentiation and apoptosis. Retinoic acid induces granulocytic differentiation of mieloid cell lines and, during this process, is responsible for the expression of CD11b, a surface antigen. In this study we investigate the effects of retinoic acid on HL-60 cells, monitoring ornithine decarboxylase (ODC) activity (enzyme rate of PA), putrescine (PUT), spermidine (SPD), spermine (SPM) levels, CD11b myeloid surface marker differentiation, cell cycle, and apoptosis. ODC activity and PUT levels are correlated with mieloid cell differentiation induced by retinoic acid treatment. Only the ODC/PUT ratio is connected with retinoic acid treated HL-60 cells. Treated cultures show a decrease of proliferation and a cell block in the G0/G1 phase, with consequent diminished S phase. The G0/G1 and S phases are significantly related to ODC activity and to PUT and SPD behavior, whereas in differentiating condition only the decrease of PUT is related to the S phase. CD11b expression, stimulated by retinoic acid treatment, is associated with the SPM trend. Total PA behavior agrees with apoptotic cell increase after 96 h of stimulation. Our data show that retinoic acid treatment modifies ODC activity and the turnover of PA. PUT, SPD and SPM, therefore, have a different role, and may be involved in the differentiative/apoptotic program of retinoic acid treated HL-60 cells.

In vitro comparison of bioabsorbable and non-resorbable membranes in bone regeneration.

BACKGROUND: Barrier membranes are used to prevent down-growth of the oral mucosa along the root surface and to allow alveolar bone regeneration in guided tissue regeneration. Several studies have demonstrated bone regenerates in the presence of bioabsorbable and non-resorbable membranes, but no studies have compared multiple bioabsorbable barriers to one another and to non-resorbable barriers. This study evaluated the in vitro influence of bioabsorbable and non-resorbable membranes on specific parameters of human osteoblast activity. METHODS: Human osteoblasts were cultured on bioabsorbable membranes made of collagen, hyaluronic acid, and poly DL-lactide, and the most common non-resorbable membrane which is made of expanded polytetrafluoroethylene (ePTFE). The osteoblasts were cultured in vitro for 24 hours on barrier membranes in the presence of 3H-thymidine and 3H-proline to study cell proliferation and collagen synthesis. Transforming growth factor-beta1 (TGF-beta1) secretion was evaluated in conditioned media using an ELISA kit. RESULTS: The results showed that collagen and poly DL-lactide stimulated DNA synthesis more than ePTFE and hyaluronic acid. All bioabsorbable membranes significantly increased collagen synthesis and alkaline phosphatase activity. Collagen and hyaluronic acid increased secretion of TGF-beta1, a growth factor involved in bone remodeling. CONCLUSIONS: These data suggest bioabsorbable membranes, particularly collagen and hyaluronic acid, may promote bone regeneration through their activity on osteoblasts.

Phenotype expression of gingival fibroblasts cultured on membranes used in guided tissue regeneration.

Human gingival fibroblasts were cultured in vitro using as substrates an extracellular matrix (matrix) and polytetrafluoride (PTFE) membranes, which are used in guided tissue regeneration. To test the degree of biocompatibility of these membranes, the cellular proliferation and the accumulation of extracellular matrix (ECM) macromolecules were considered as parameters. The fibroblasts were cultured in vitro for 24 and 48 hours without serum on plastic, matrix, and PTFE membranes in the presence of 3H-thymidine, 3H-glucosamine, and 3H-proline to study the neo-synthesis of DNA, glycosaminoglycans (GAG), and collagen proteins, respectively. Studies on cell proliferation showed that fibroblasts grown on matrix membrane significantly increased 3H-thymidine incorporation, while fibroblasts grown on PTFE membrane decreased 3H-thymidine incorporation, compared to plastic used as a control. Moreover, the PTFE membrane induced a marked decrease of collagen and GAG accumulation both in the cellular and extracellular pool, while the matrix membrane provoked a decrease of the two macromolecules in the cellular pool and an increase in the extracellular one, compared to the control. The data we obtained demonstrate that matrix membranes are the most suitable to stimulate both cellular proliferation and ECM macromolecule accumulation.

Glycosidases during chick embryo lung development and their colocalization with proteoglycans and growth factors.

During development, the epithelial component of the lung goes through a complex orderly process of branching, following strict patterns of space and time. Proteoglycans, glycosaminoglycans and growth factors are fundamental components of the extracellular matrix and perform a key role in differentiative processes. The embryonic chick lung shows a specific glycosaminoglycan composition at different levels of branching and at different embryonic stages. Proteoglycan and glycosaminoglycan accumulation is the result of secretion, absorption and degradation processes. In this pathway, enzymes, such as glycosidases, growth factors and cytokines are involved. We examined the behaviour of glycosidases, such as beta-hexosaminidases (beta-N-acetyl-D-glucosaminidase, beta-N-acetyl-D-galactosaminidase), beta-glucuronidase and beta-galactosidase, during the development of the lung bud. Our data show that the activity of the enzymes is closely linked to the processes of epithelial proliferation, bronchial tubule lengthening and infiltration of the surrounding mesenchyme. The glycosaminoglycans colocalize with transforming growth factor beta2 and interleukin-1 in the basement membrane and in the mesenchymal areas where the epithelium grows, and are complementary to the presence of the glycosidases. In conclusion, the activity of these glycosidases is spatially and temporally programmed and favors the release of the factors and the events which they influence.

Epithelial-mesenchymal interactions and lung branching morphogenesis. Role of polyamines and transforming growth factor beta1.

Lung branching morphogenesis is a result of epithelial-mesenchymal interactions, which are in turn dependent on extracellular matrix composition and cytokine regulation. Polyamines have recently been demonstrated as able to modify chick embryo skin differentiation. In this work we have examined the effects of putrescine and spermidine during chick embryo lung morphogenesis in organotypic cultures by morphological, histochemical and biochemical examination. To verify the role of polyamines, we used specific inhibitors, such as bis-cyclohexylammonium sulphate and alfa-difluoromethylornithine, and transforming growth factor beta1, an ornithine decarboxylase and polyamine stimulator. Our data show that lung morphogenesis is significantly altered following the induced mesenchymal glycosaminoglycan changes. The increase of mesenchymal glycosaminoglycans is correlated with a stimulation of lung development in the presence of polyamines, and with its inhibition when transforming growth factor beta1 is added to the culture medium. The morphometric data show a uniform increase of both the mesenchyme and epithelial branching with spermidine and putrescine stimulus, whereas the mesenchymal substance alone is significantly increased in apical-median lung sections with transforming growth factor beta1 and transforming growth factor beta1 + spermidine lung cultures. Transforming growth factor beta1 and transforming growth factor beta1 + spermidine confirm the blocking of epithelial branching formations and fibroblast activation, and show that polyamines are unable to prevent the blocking of epithelial cells due to the inhibitory effect of transforming growth factor beta1.

Increasing plasma homocysteine during follow-up in heart transplant recipients: effects of folate and renal function.

BACKGROUND: Hyperhomocysteinemia is a common finding in heart transplant recipients and may represent a risk factor for graft failure. However, the time-course, determinants and effects of medical therapy on total homocysteine plasma levels after heart transplantation remain undetermined. The aim of this study was to prospectively analyze 1) the time-course of total homocysteine in heart transplant recipients; 2) the effects of folate supplements and cyclosporine A on total homocysteine; 3) the relation among renal function, serum vitamin levels, and total homocysteine. METHODS: Fifty-two heart transplant recipients consecutively evaluated for routine follow-up during 1998 were included in the study (mean age 54 +/- 12 years; 28% female). Among the 52 patients, 10 patients were treated with folate for the entire period of the study (Group F), while 26 patients never received folate (Group NF). The remaining 16 patients who did not take folate on a regular basis were excluded from subgroup analysis. Total homocysteine and creatinine plasma levels were assayed at entry into the study (time 0) and at the end of the study, 12 months later (time 12). RESULTS: Homocysteinemia increased significantly from time 0 to time 12 (p < 0.001), regardless of creatinine plasma levels (p = 0.03) and folate intake (p < 0.01). However, total homocysteine levels were lower in Group F compared to Group NF at time 0 and time 12 (p < 0.02). On multivariate analysis, time of follow-up, serum creatinine and lack of folate intake were positive independent predictors of total homocysteine. CONCLUSIONS: Homocysteinemia increased over time in heart transplant recipients, regardless of renal function and folate administration. Lower total homocysteine levels were associated with folate intake, suggesting that folate supplements may play a role in the prevention of vascular allograft disease.

[Influence of age on the morbidity and mortality in coronary artery surgery]. / Influência da idade na morbidade e mortalidade da cirurgia arterial coronária.

To study the effects of age on the results of coronary artery bypass grafting (CABG), 250 patients operated on from 1986 to 1989 were divided into two groups: 1) less than 65 years of age and 2) older than 65. Pre, intra and postoperative data collected in all patients included: sex, type and class of angina, associated diseases, previous myocardial infarction, previous CABG, left ventricular aneurysm, bypass time, aortic cross-clamp time, number of grafts per patient, need for prolonged inotropic support, postoperative complications, and mortality. A large number of elderly patients had unstable angina (20.3% vs 6.2%), post-infarction angina (10.1% vs 7.8%), angina at rest (10.1% vs 3.6%), peripheral vascular disease (8.4% vs 2.6%), required prolonged inotropic support (18.6% vs 3.1%), had major neurological complications (8.4% vs 0.1%) and perioperative myocardial infarction (5.0% vs 0.5%). Overall mortality was 3.6% but mortality rates were significantly higher in elderly patients (11.8% vs 1.0%). These data suggest that elderly patients have an increased risk of cardiac and neurologic morbidity and mortality. It appear that the increased morbidity and mortality is related to an increased susceptibility of the elderly to serious postoperative complications.

Novel poly(L-lactide) PLLA/SWNTs nanocomposites for biomedical applications: material characterization and biocompatibility evaluation.

Poly(L-lactide) (PLLA)/single-walled carbon nanotubes (SWNTs) nanocomposite films were produced using the solvent casting method, and morphological, thermal and mechanical properties were investigated. Biocompatibility was evaluated by using human bone cells, performing adhesion and proliferation studies. The role of single-walled nanotube incorporation and functionalization on PLLA bio-polymers was investigated. Pristine (SWNTs) and carboxylated (SWNTs-COOH) carbon nanotubes were considered in order to control the interaction between PLLA and nanotubes. SWNTs and SWNTs-COOH showed a good dispersion in the polymer matrix and improved the PLLA crystallinity. Thermal, morphological and dynamic-mechanical analyses revealed that carboxylic groups on the tube sidewalls increased compatibility between PLLA and nanostructures. Mechanical properties demonstrated an enhancement related to introduction and functionalization of carbon nanotubes. Biological investigations showed osteoblasts cultured on PLLA/SWNTs-COOH nanocomposites has higher cell adhesion and proliferation than osteoblasts cultured on PLLA and PLLA/SWNTs nanocomposites. These studies suggest that combination of biodegradable polymers and SWNTs opens a new perspective in the self-assembly of nanomaterials and nanodevices for biomedical applications with tunable properties.

Dose comparison trial of sustained-release fampridine in multiple sclerosis.

OBJECTIVE: To examine the efficacy and safety of three different doses of sustained-release fampridine in people with multiple sclerosis (MS). METHOD: This multicenter, randomized, double-blind, placebo-controlled, parallel-group study recruited 206 participants at 24 centers in the United States and Canada. After a single-blind, 2-week placebo run-in, participants were randomly assigned to receive fampridine (10, 15, or 20 mg twice daily) or placebo for 15 weeks. The primary efficacy variable was percent change in walking speed based on the timed 25-foot walk. RESULTS: Trends for increased walking speed were consistent across dose groups vs placebo, but not significant, on the prospective analysis. An increase from baseline in lower extremity strength during the 12-week stable-dose period was seen in the groups receiving 10- and 15-mg doses, compared with placebo (p = 0.018 and 0.003). There were no significant changes in other secondary assessments. Post hoc analysis revealed subsets of participants in each dose group with walking speeds during the treatment period that were consistently faster than during the nontreatment period. There were significantly more "consistent responders" in the drug-treated groups than in the placebo group (36.7% compared with 8.5%). Consistent responders showed significantly greater improvement in self-assessed ambulation on the 12-Item MS Walking Scale than did nonresponders. Fampridine was generally well tolerated. Severe and serious adverse events were more frequent at the highest dose. CONCLUSIONS: This phase 2 study suggests that a subgroup of patients, when treated with fampridine, experiences a clinically relevant improvement in walking ability, which is sustained for at least 14 weeks.

Fampridine-SR in multiple sclerosis: a randomized, double-blind, placebo-controlled, dose-ranging study.

OBJECTIVE: To determine the safety of sustained-release 4-aminopyridine in subjects with mutiple sclerosis (MS) and to examine dose-related efficacy up to 40 mg twice daily. METHOD: Multicenter, randomized, double-blind, placebo-controlled, study. Following a 4-week baseline peroid, subjects were randomly assigned to receive Fampridine-SR (n=25, doses from 10 to 40 mg twice daily, increasing in 5 mg increments weekly) or placebo (n=11). A battery of assessments was performed weekly, including the MS Functional Composite (MSFC), fatigue questionnaires, and lower extremity manual muscle testing. RESULTS: The most common adverse events were dizziness, insomnia, paresthesia, asthenia, nausea, headache, and tremor. Five subjects were discontinued from Fampridine-SR because of adverse events at doses greater than 25 mg, and these included convulsions in two subjects at doses of 30 and 35 mg twice daily. Improvement were seen in lower extremity muscle strength (prospective analysis) and walking speed (post-hoc analysis) in the Fampridine-SR group compared to placebo (unadjusted p-values of 0.01 and 0.03, respectively). There were no significant differences in other MSFC measure or fatigue scores. CONCLUSIONS: Future studies should employ doses up to 20 mg twice daily with lower extremity strength and walking speed as potential outcome measures.