Liposomes embedded with differentiating factors as a new strategy for enhancing DPSC osteogenic commitment.

Human dental pulp stem cell (DPSC) differentiation toward the osteoblastic phenotype is enhanced when culture media are supplemented with differentiating factors, i.e. ascorbic acid, ß-glycerophosphate and dexamethasone. Liposomes, spherical vesicles formed by a phospholipid bilayer, are frequently used as carriers for drugs, growth factors and hydrophobic molecules. The aim of this work was to speed up DPSC commitment to the osteogenic lineage by embedding differentiating factors within liposomes. Firstly, liposomes were prepared by rehydrating a phospholipidic thin film and characterised in terms of dimensions. Secondly, liposome-exposed DPSCs were characterised by their immunophenotypic profile. Levels of CD90 were significantly decreased in the presence of liposomes filled with ascorbic acid, ß-glycerophosphate and dexamethasone (Lipo-Mix) with respect to normal differentiation medium (DM), while CD73 and CD29 expression were enhanced, suggesting osteogenic commitment. Additionally, an appreciable extracellular matrix deposition is detected. Thirdly, the Lipo-Mix formulation better increases alkaline phosphatase activity and levels of Collagen I secretion with respect to DM. In parallel, the new liposome formulation is capable of decreasing the release of H2O2 and of triggering a precocious antioxidant cell response, redressing the redox balance required upon mesenchymal stem cell commitment to osteogenesis. It can be therefore hypothesised that Lipo-Mix could represent a suitable tool for clinical regenerative purposes in the field of tissue engineering by speeding up DPSC osteogenic commitment, mineralised matrix deposition and remodelling.

Submandibular approach and use of the harmonic instrument in lateral oral cavity and oropharyngeal oncologic surgery.

The approach to the surgical treatment of lateral oral cavity and oro-pharyngeal cancer has undergone many changes over the years, passing from a demolitive approach involving segmental resectioning of the mandible to a conservative approach with temporary mandibulotomy. The complications resulting from these two different approaches invited further review of the various indications related to them. The present report considers, through a retrospective analysis, whether the sub-mandibular approach, which envisages the use of the harmonic instrument, is an oncologically correct alternative and whether it actually reduces the number of early and late complications. From 2003 to 2009, 259 patients underwent surgery for lateral oral cavity and oro-pharyngeal cancer. A sub-mandibular approach was used in 163 patients, and a conservative trans-mandibular approach in 12. The results showed that the number of late complications and T recurrences was significantly lower in those patients in whom the sub-mandibular approach was carried out. There was no difference in the microscopic examination of the resection margins. The use of the harmonic instrument made it possible to perform "en bloc" (T+N) operations working in narrow spaces with less intra-operative bleeding, which contributed to adequate oncological radicality.

Respiratory epithelial adenomatoid hamartoma of the maxillary sinus: case report.

The case is described of a male patient with respiratory epithelial adenomatoid hamartoma of the left maxillary sinus that initially presented as a chronic sinus inflammation. This benign lesion is characterized by glandular proliferation originating from the surface of the respiratory epithelium. Maxillary sinus localisation is very rare but is very important to be able to distinguish hamartomas from schneiderian papillomas of the inverted type and adenocarcinomas, potentially requiring aggressive surgical treatment. Moreover, misinterpretation of the respiratory epithelial adenomatoid hamartoma as chronic sinus inflammation may result in inadequate treatment. The clinical and pathological features of this lesion are discussed.

Effect on prolactin secretion of Echinacea purpurea, hypericum perforatum and Eleutherococcus senticosus.

It has been recently reported that prolactin (PRL) plays an important role in immune system regulation. In this study we investigated the activity of three natural drugs with immunomodulatory activity: Echinacea purpurea (EP), Hypericum perforatum (HP) and Eleutherococcus senticosus (ES) on PRL production. Male rats were orally treated with two different doses (30 and 100 mg/kg) of extract of these drugs for 3 or 15 days. A 3-day treatment was not able to modify PRL serum levels, whereas a 15-day treatment with EP and HP at the higher dose significantly inhibits PRL production. A treatment with ES was always ineffective. A possible mechanism for this effect could be that both HP and EP extracts display a direct dopaminergic activity, although an involvement of the GABA-ergic system cannot be excluded.

Th1-Th2 response in hyperprolactinemic mice infected with Salmonella enterica serovar Typhimurium.

Prolactin (PRL) is a pituitary hormone and a cytokine known to regulate several physiological functions. It plays a role in modulating the immune system of rodents and humans. A hormonal protection against listeria and salmonella infections has been previously ascribed to effects of PRL on immunocompetent cells. Here, the role of PRL in the Th1-Th2 response was evaluated based on the pattern of cytokines release by splenocytes from hyperprolactinemic mice infected with Salmonella enterica serovar Typhimurium. Hyperprolactinemia by pituitary graft reduced the number of bacteria in spleens of in vivo infected mice. Modulation of Th1 (IFN-gamma, IL-12) and Th2 (IL-4, IL-10) cytokine production by splenic cells was found. Our results indicate that PRL can up-regulate IFN-c and IL-12 secretion in response to salmonella infection, confirming its in vivo immunostimulatory effect and suggesting hormonal participation in the genesis and sustenance of the Th1 response.

Elevated bioactive prolactin levels in systemic lupus erythematosus--association with disease activity.

OBJECTIVE: To assess the possibility that prolactin (PRL) is involved in the pathogenesis of systemic lupus erythematosus (SLE). METHODS: We determined serum PRL levels in 122 serum samples from 78 unselected patients with SLE (73 women, 5 men, age range 16-71 yrs). Disease activity was defined according to Lupus Activity Criteria Count (LACC) and scored by Systemic Lupus Disease Activity Index (SLEDAI). Serum PRL concentrations were determined by immunoradiometric assay (IRMA) and by biological assay (BA) that evaluates Nb2 lymphoma cell proliferation. RESULTS: Hyperprolactinemia (> 20 ng/ml) was found in 21 patients (26.9%) by IRMA and in 31 (39.7%) by BA. A significant correlation between IRMA and BA PRL levels was found (rs 0.46, p < 0.001). According to LACC, SLE was active in 29 patients and inactive in 49. In those with active disease median PRL levels were higher both by IRMA (18.5 ng/ml, range 2.2-51.2 vs 10.6 ng/ml, range 3.9-29.6; p < 0.001) and BA (21.0 ng/ml, range 12.4-84 vs 14.9 ng/ml, range 4.2-46.1; p < 0.001). Hyperprolactinemia was associated with active disease in 13/21 patients (61.9%) by IRMA and in 18/31 (58.1%) by BA (p < 0.01). SLEDAI scores correlated with PRL levels both by IRMA (rs 0.5, p < 0.001) and BA (rs 0.41, p < 0.02). A followup analysis on serum samples from 44 patients seen again after 6-8 mo confirmed the above results. There was no difference in the rate of different clinical manifestations in hyperprolactinemic and normoprolactinemic subjects, apart from the increased prevalence of malar rash and central nervous system manifestations in the patients with hyperprolactinemia (p < 0.03 and p < 0.01, respectively). CONCLUSION: Hyperprolactinemia was frequently detected in patients with SLE by IRMA and by BA and was associated with disease activity. Our findings suggest that PRL may play a role in the pathogenesis of SLE.

Inhibition of inducible nitric oxide synthase and cyclooxygenase-2 expression by flavonoids in macrophage J774A.1.

The present study focuses on the effect of various naturally occurring flavonoids (apigenin, galangin, morin, naringenin, quercetin, and silymarin) on nitric oxide (NO) and prostaglandin E2 (PGE2) production induced by lipopolysaccharide (LPS) in the macrophage cell line J774A.1. Moreover, we evaluated flavonoid modulation of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) enzyme expression by western blot analysis. Apigenin and quercetin (0.5-50 microM) were the most potent inhibitors of NO production and this effect was concentration-dependent and significant at 5 and 50 microM. These data were consistent with the modulation of iNOS enzyme expression. A similar pattern was observed considering the inhibitory effect of flavonoids on LPS-induced PGE2 release and COX-2 expression. Quercetin, galangin, apigenin, and naringenin markedly decreased PGE2 release and COX-2 expression in a concentration-dependent manner. This study suggests that inhibition of iNOS and COX-2 expression by flavonoids may be one of the mechanisms responsible for their anti-inflammatory effects.

[Rhinoscleroma with extramaxillary extension: report of a clinical case]. / Rhinoscleroma con estensione extramascellare: a proposito di un caso clinico.

Rhinoscleroma is a chronic, granulomatous infection most frequently affecting the respiratory tract mucosa. The disease is endemic in some geographic areas and is sporadic in Italy, where medio-facial granulomatoses must be differentiated from malignant naso-sinus neoplasms. The present work describes a case of rhino-sinus scleroma with extramaxillary extension, defining the most important clinical characteristics and the treatment performed.

[Eutectic mixture of local anesthetics (EMLA): evaluation of the analgesic effectiveness during ND: YAG laser turbinoplasty]. / Eutectic Mixture of Local Anaesthetics (EMLA): valutazione dell'efficacia analgesica nella turbinoplastica al laser Neodymio: ittrio-alluminio-granato (Nd: Yag).

The analgesic effect of EMLA anesthetic cream is evaluated during contact turbinoplasty with a Neodymium: yttrium-aluminum-garnet (Nd: Yag) laser. A total of 48 patients were selected and randomized in two groups. EMLA was applied on the nasal mucosa, respectively, for 15 and 30 minutes after using vasoconstrictor cottonoids. Both groups underwent turbinate reduction by contact with Nd: Yag laser using three discharge incisions traced parallel on the medial surface of the lower turbinate working from the tail to the head. The pain induced by the surgical procedure was evaluated with a visual analogic scale. The score obtained was significantly lower in the 1st group than in the 2nd group and this difference was statistically significant (p < 0.01). The EMLA anesthetic cream applied on the mucosa for 15 minutes allows to obtain a good analgesia during the functional contact turbinate surgery with a Nd: Yag laser and thus can be considered a valid alternative to anesthesia by local infiltration.

Effect of fumonisin B1 on inducible nitric oxide synthase and cyclooxygenase-2 in LPS-stimulated J774A.1 cells.

Fumonisin B1 (FB1) is a water-soluble fungal metabolite that elicits a wide spectrum of toxicological effects. Cellular targets of FB1 include immune cells and in particular macrophages. In the present study the cytotoxic effect of FB1 (1-100 microM) was evaluated using a murine macrophage cell line (J774A.1) as model system. The effect of FB1 on nitric oxide (NO) and prostaglandin E2 (PGE2) production induced by lipopolysaccharide (LPS, 10 and 100 ng/ml) was also investigated. Macrophages were pretreated with FB1 for 72 h and then stimulated with LPS for 24 h. The increase of LPS-induced production of these inflammatory mediators was observed at increasing concentrations of FB1 (0.1-10 microM) and was found to be concentration dependent. By western blot analysis we demonstrated that the observed increase of NO and PGE2 production by FB1 was related to an enhancement of iNOS and COX-2 expression.

Prolactin induction of nitric oxide synthase in rat C6 glioma cells.

We have examined the neuroimmunoregulatory function of prolactin (PRL) on astrocytic inducible nitric oxide synthase (iNOS) expression in the C6 glioma cell line. After 24 h of PRL (5-100 nM) stimulation, a concentration-dependent increase of NO release, evaluated as nitrite, was observed in C6 culture medium. Moreover, both NO release and iNOS expression induced by interferon-gamma (250 U/ml) were enhanced by PRL (18-100 nM). PRL-induced NO release was inhibited by dexamethasone, an inhibitor of de novo iNOS synthesis. We used erbstatin (5 microg/ml), a potent inhibitor of protein tyrosine kinases, to test whether these proteins were required for signaling events evoked by PRL in these cells. This inhibitor was able to inhibit completely the PRL-induced NO production and iNOS expression. In conclusion, we provide evidence that NO production in glial cells can be regulated not only by cytokines but also by neuroimmunoregulatory hormones such as PRL, which is present in normal brain but may be elevated in several pathological states.

Simple method of detecting enteroviruses in contaminated molluscs and sewage by using polymerase chain reaction coupled with a colorimetric microwell detection assay.

The methods normally used for the detection of enteroviruses in environmental samples involve the use of cell cultures, which are expensive and time consuming. The Polymerase Chain Reaction (PCR) is a useful tool for the detection of enteroviruses in several matrixes because primary cell culture is not needed and the increased sensitivity of PCR allows detection of the low numbers of target nucleic acids usually found in environmental samples. A 5-h, user-friendly PCR assay was used to detect enteroviruses in bivalves molluscs (clams) and sewage. Reverse transcription and amplification were performed in a one-step reaction using rTth polymerase. Carryover contamination was prevented with dUTP and uracil N-glycosylase. Detection was performed colorimetrically in a microwell titer plate. This method has greater advantages over conventional methodologies for routinely screening a large number of samples, namely, the rapid acquisition of results and cost effectiveness.

Preparation and local anaesthetic activity of N-[2-(tert-amino)ethyl]- and N-(lupinyl)-benzotriazol-1/2-ylacetamides.

Two sets of N-[2-(tert-amino)ethyl]- and N-[(quinolizidin-1 alpha-yl) methyl]-benzotriazol-2-ylacetamides, bearing substituents on position 5 or 5 and 6, were prepared and tested for local anaesthetic activity in comparison with lidocaine. Most of the prepared compounds exhibited a fairly good activity comparable or superior to that of lidocaine. The introduction of substituents on the benzene ring and the replacement of the usual tert-amino alkyl chains with the quinolizidin-1 alpha-ylmethyl (lupinyl) moiety were quite profitable for both the intensity and duration of activity. One selected compound was subjected to a large pharmacological screening and found endowed with a good level of the purported antiarrhythmic activity without any other disturbing activity.

Prolactin modulation of nitric oxide and TNF-alpha production by peripheral neutrophils in rats.

It has been demonstrated that prolactin (PRL) is a potent immunomodulator that exerts stimulatory effects on physiological responses of immune cells. In the present research we have investigated whether PRL may influence nitric oxide (NO) and/or tumor necrosis factor-alpha (TNF-alpha) production in neutrophils obtained from inflammatory exudate of carrageenin-induced experimental pleurisy in the rat. In this acute model of inflammation the role of endogenous NO was evaluated using an inhibitor of NO-synthase, NG-nitro-L-arginine methyl ester (L-NAME). A treatment of animals with L-NAME (10 mg/kg s.c.) induced a reduction of volume and cell number of pleural exudate and a decrease of nitrite production (measured by the Griees reaction) by polymorphonuclear cells after 24 h of incubation, while D-NAME, the inactive isomer, was without effect. Neutrophils from ovine prolactin (oPRL) treated rats (5 mg/kg for 5 times s.c.) or from rats with a hyperprolactinaemia induced by pituitary gland graft produced higher amounts of NO both after 24 and 48 h of incubation. On the contrary, a clear reduction in the production of NO was found in neutrophils from rats treated with bromocriptine (BRC) (2 mg/kg s.c.), a dopamine D2-receptor agonist. TNF-alpha production (measured by MTT/cytotoxic assay) by neutrophils was markedly increased in PRL-treated or pituitary-grafted rats in comparison to controls, whereas BRC treatment reduced TNF-alpha production.

Recombinant human prolactin induces protection against Salmonella typhimurium infection in the mouse: role of nitric oxide.

In the present study, we demonstrated that repeated treatment with recombinant human prolactin (rhPRL) protected mice against Salmonella typhimurium infection. The protective activity was statistically significant, dose-dependent and present only when rhPRL treatments were performed before the infection. This activity was probably related to the observed increases in phagocytosis and intracellular killing of peritoneal macrophages induced by the hormonal treatment. The number of peripheral leukocytes was not modified, excluding a mobilization of cells from other compartments. A decrease in the mortality rate after challenge was also observed in mice treated with the monoclonal antibody anti-PRL receptor U5, confirming that the protective activity was associated with receptor activation. Our studies also suggest that nitric oxide (NO) production was involved in the protective effect of rhPRL since pre-treatment of the animals with L-NAME, an inhibitor of NO-synthase, was able to completely revert the protective activity, whereas D-NAME, the inactive D-isomer, was without effect.

Effect of L-alpha glycerylphosphorylcholine on muscarinic receptors and membrane microviscosity of aged rat brain.

1. Old rats showed a significant decrease in the number of muscarinic M(1) receptors and a significant increase in membrane microviscosity in the striatum and hippocampus as compared to young animals. In contrast, no significant changes in the density of muscarinic M(2) receptors were observed with aging. 2. Chronic treatment of aged rats with L-alpha-glycerylphosphorylcholine (L-alpha-GPC) restored the number of M(1) receptors to levels found in the striatum and hippocampus from young animals. The same treatment to aged rats partially restored membrane microviscosity in both regions studied and hence increased membrane fluidity. 3. None of the major metabolites of L-alpha-GPC (choline, glycerophosphate or phosphorylcholine) was able to restore the number of striatal and hippocampal M(1) sites and membrane microviscosity of aged rats, neither did any of these treatments (including treatment with L-alpha-GPC) modify the level of M(1) receptors and microviscosity values in young rats.

The effect of age and sex on the expression of prolactin binding activity in the chicken bursa of Fabricius.

The binding of 125I-labeled prolactin (PRL) to membranes from the bursa of Fabricius of male and female chicks of different ages (15-30-45 and 60 days) was studied. In male chicks the binding was very low in 15 day-old animals and slightly increased in more aged animals. In female chicks the binding was more evident in young animals and decreased in 60 day-old animals. The binding showed a hormonal specificity and Scatchard analysis of the binding revealed the presence of binding sites with low capacity and high affinity. The presence of PRL receptors in the bursa of the chick, a structure that confers immunological competence to birds, gives further support to the involvement of the hormone in the immune processes.

Regulation of prolactin receptor mRNA expression in peripheral lymphocytes in rats in response to changes in serum concentrations of prolactin.

In the present study we have evaluated the absolute number of the two forms of prolactin (PRL) receptor mRNA in rat peripheral blood lymphocytes and the modulation of receptor mRNA induced by changes in serum levels of endogenous PRL or by administration of ovine PRL. Lymphocytes expressed low levels of both forms of PRL receptor transcripts. Repeated treatments with ovine PRL significantly reduced levels of mRNA encoding the long form PRL receptor, whereas expression was markedly increased by repeated doses of bromocriptine. In contrast, the mRNA level of short form PRL receptor was unchanged by both treatments. The expression of long form transcripts was also markedly decreased in lymphocytes from pituitary-grafted rats. Therefore it appears that in rat peripheral lymphocytes PRL has a negative effect on the expression of its own receptor.