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The role of brain cholesterol in Alzheimer's disease (AD) is currently a matter of debate. Experimental evidence suggests that reducing circulating and brain cholesterol protects against AD, however recent data indicate that low membrane cholesterol results in neurodegeneration and that the cholesterol synthesis catalyst seladin-1 is down-regulated in AD-affected brain regions. We previously reported a significant correlation between resistance to amyloid toxicity and content of membrane cholesterol in differing cultured cell types. Here we provide evidence that Aβ42 pre-fibrillar aggregates accumulate more slowly and in reduced amount at the plasma membrane of human SH-SY5Y neuroblastoma cells overexpressing seladin-1 or treated with PEG-cholesterol than at the membrane of control cells. The accumulation was significantly increased in cholesterol-depleted cells following treatment with the specific seladin-1 inhibitor 5,22Echolestadien- 3-ol or with methyl-β-cyclodextrin. The resistance to amyloid toxicity and the early cytosolic Ca2+ rise following exposure to Aβ42 aggregates were increased and prevented, respectively, by increasing membrane cholesterol whereas the opposite effects were found in cholesterol-depleted cells. These results suggest that seladin-1-dependent cholesterol synthesis reduces membrane-aggregate interaction and cell damage associated to amyloid-induced imbalance of cytosolic Ca2+. Our findings extend recently reported data indicating that seladin-1 overexpression directly enhances the resistance to Aβ toxicity featuring seladin-1/DHCR 24 as a possible new susceptibility gene for sporadic AD. [ABSTRACT FROM AUTHOR]

In consideration of the close relationship between radicular cysts and alveolar bone, it is important to evaluate the potential involvement of the neighbouring bone tissue in such lesions. In the present study, using cytochemical, immunocytochemical and morphological analyses, presumptive osteoclast precursors were revealed in the connective tissue of radicular cyst capsules. The osteoclastic nature of these cells was postulated by their positive staining for the enzyme tartrate-resistant acid phosphatase (TRAP) and by the expression of vitronectin receptor (VnR) on their cell surface. However, these cells did not express the vacuolar-type proton pump, suggesting that they may represent early osteoclast precursors infiltrating the cyst capsule. Cysts also contained activated small blood vessels whose endothelial cells expressed the VnR. This integrin receptor is important in the adhesion of preosteoclasts to the endothelial lining, a necessary step for their emigration out of the vasculature. Therefore, the intracystic vessels could represent a substrate for preosteoclast recruitment. These precursor cells may then reach the perialveolar bone surface and contribute to bone demolition together with those recruited by the resorbing surfaces. The bone-destroying potential of radicular cysts was confirmed by the presence of numerous osteoclasts with large resorption areas on the perialveolar bone surfaces exposed to the cyst capsules. The resorbed surfaces were usually located around the vascular canals of the Haversian systems. [ABSTRACT FROM AUTHOR]

In an attempt to verify the nature of bone resorption processes on the bone surrounding radicular cysts, fragments of cysts with the adjacent bone tissue were studied by morphological, cytochemical and ultrastructural methods. Cyst fluid was analysed for its content of cytokines with osteolytic activity. The cyst wall exhibited several connective tissue extensions which penetrated the adjacent bone. Numerous multinucleated tartrate-resistant acid phosphatase (TRAP)-positive cells were seen at the tip of the intraosseous extensions of the cyst capsule and in direct contact with the bone tissue. Typical resorption lacunae were identified on the bone surface by scanning electron microscopy. Moreover, mononuclear TRAP-positive cells were seen within the cyst capsule. High levels of prostaglandin E2 (PGE2) and interleukin-6 (IL-6) were detected in the cyst fluids. In conclusion, active bone resorption may contribute significantly to the growth of these lesions within the jaws. [ABSTRACT FROM AUTHOR]

New Findings: What is the central question of this study? Fibroblast-to-myofibroblast transition is a key mechanism in the reparative response to tissue damage, but myofibroblast persistence in the wound leads to fibrosis and organ failure. The role of relaxin as an antifibrotic agent capable of counteracting the acquisition of biophysical features of differentiated myofibroblasts deserves further investigation. What is the main finding and its importance? Electrophysiological analysis showed that relaxin, administered during profibrotic treatment, hyperpolarizes the membrane potential and attenuates delayed rectifier and inwardly rectifying K(+) currents, which usually increase in the transition to myofibroblasts. These findings provide further clues to the therapeutic potential of relaxin in fibrosis. The hormone relaxin (RLX) is produced by the heart and may be involved in endogenous mechanisms of cardiac protection against ischaemic injury and fibrosis. Recent findings in cultured cardiac stromal cells suggest that RLX can inhibit fibroblast-to-myofibroblast transition, thereby counteracting fibrosis. In order to explore its efficiency as an antifibrotic agent further, we designed the present study to investigate whether RLX may influence the electrophysiological events associated with differentiation of cardiac stromal cells to myofibroblasts. Primary cardiac proto-myofibroblasts and NIH/3T3 fibroblasts were induced to myofibroblasts by transforming growth factor-β1, and the electrophysiological features of both cell populations were investigated by whole-cell patch clamp. We demonstrated that proto-myofibroblasts and myofibroblasts express different membrane passive properties and K(+) currents. Here, we have shown, for the first time, that RLX (100 ng ml(-1) ) significantly reduced both voltage- and Ca(2+) -dependent delayed-rectifier and inward-rectifying K(+) currents that are typically increased in myofibroblasts compared with proto-myofibroblasts, suggesting that this hormone can antagonize the biophysical effects of transforming growth factor-β1 in inducing myofibroblast differentiation. These newly recognized effects of RLX on the electrical properties of cardiac stromal cell membrane correlate well with its well-known ability to suppress myofibroblast differentiation, further supporting the possibility that RLX may be used for the treatment of cardiac fibrosis., (© 2015 The Authors. Experimental Physiology © 2015 The Physiological Society.)

Mitochondria play a crucial role in pathways of stress conditions. They can be transported from one cell to another, bringing their features to the cell where they are transported. It has been shown in cancer cells overexpressing multidrug resistance (MDR) that mitochondria express proteins involved in drug resistance such as P-glycoprotein (P-gp), breast cancer resistant protein and multiple resistance protein-1. The MDR phenotype is associated with the constitutive expression of COX-2 and iNOS, whereas celecoxib, a specific inhibitor of COX-2 activity, reverses drug resistance of MDR cells by releasing cytochrome c from mitochondria. It is possible that COX-2 and iNOS are also expressed in mitochondria of cancer cells overexpressing the MDR phenotype. This study involved experiments using the human HCC PLC/PRF/5 cell line with and without MDR phenotype and melanoma A375 cells that do not express the MDR1 phenotype but they do iNOS. Western blot analysis, confocal immunofluorescence and immune electron microscopy showed that iNOS is localized in mitochondria of MDR1-positive cells, whereas COX-2 is not. Low and moderate concentrations of celecoxib modulate the expression of iNOS and P-gp in mitochondria of MDR cancer cells independently from inhibition of COX-2 activity. However, A375 cells that express iNOS also in mitochondria, were not MDR1 positive. In conclusion, iNOS can be localized in mitochondria of HCC cells overexpressing MDR1 phenotype, however this phenomenon appears independent from the MDR1 phenotype occurrence. The presence of iNOS in mitochondria of human HCC cells phenotype probably concurs to a more aggressive behaviour of cancer cells., (© 2015 The Authors. Journal of Cellular and Molecular Medicine published by John Wiley & Sons Ltd and Foundation for Cellular and Molecular Medicine.)

Aim: Comparing the efficacy of photoablative and photodynamic diode laser in adjunct to scaling -root planing (SRP) and SRP alone for the treatment of chronic periodontitis., Materials and Methods: Twenty-six patients were studied. Maxillary left or right quadrants were randomly assigned to sham-laser treatment + SRP or laser + SRP. This consisted of photoablative intra/extra-pocket de-epithelization with diode laser (λ = 810 nm), followed by single SRP and multiple photodynamic treatments (once weekly, 4-10 applications, mean ± SD: 3.7 ± 2.4) using diode laser (λ = 635 nm) and 0.3% methylene blue as photosensitizer. The patients were monitored at days 0 and 365 by clinical assessment (probing depth, PD; clinical attachment level, CAL; bleeding on probing, BOP) and at days 0, 15, 30, 45, 60, 75, 90, 365 by cytofluorescence analysis of gingival exfoliative samples taken in proximity of the teeth to be treated (polymorphonuclear leukocytes, PMN; red blood cells, RBC; damaged epithelial cells, DEC; bacteria)., Results: At day 365, compared with the control quadrants, the laser + SRP therapy yielded a significant (p < 0.001) reduction in PD (-1.9 mm), CAL (-1.7 mm) and BOP (-33.2% bleeding sites), as well as in bacterial contamination - especially spirochetes - and PMN and RBC shedding in the gingival samples (p < 0.001)., Conclusions: Diode laser treatment (photoablation followed by multiple photodynamic cycles) adjunctive to conventional SRP improves healing in chronic periodontitis patients., (© 2012 John Wiley & Sons A/S.)

The demonstration that the adult heart contains myocardial progenitor cells which can be recruited in an attempt to replace the injured myocardium has sparkled interest towards novel molecules capable of improving the differentiation of these cells. In this context, the peptide hormone relaxin (RLX), recently validated as a cardiovascular hormone, is a promising candidate. This study was designed to test the hypothesis that RLX may promote the growth and maturation of mouse neonatal immature cardiomyocytes in primary culture. The cultures were studied at 2, 12, 24 and 48 hrs after the addition of human recombinant H2 RLX (100 ng/ml), the main circulating form of the hormone, or plain medium by combining molecular biology, morphology and electrophysiology. RLX modulated cell proliferation, promoting it at 2 and 12 hrs and inhibiting it at 24 hrs; RLX also induced the expression of both cardiac-specific transcription factors (GATA-4 and Nkx2-5) and cardiac-specific structural genes (connexin 43, troponin T and HCN4 ion channel) at both the mRNA and protein level. Consistently, RLX induced the appearance of ultrastructural and electrophysiological signs of functionally competent, mature cardiomyocytes. In conclusion, this study provides novel circumstantial evidence that RLX specifically acts on immature cardiomyocytes by promoting their proliferation and maturation. This notion suggests that RLX, for which the heart is both a source and target organ, may be an endogenous regulator of cardiac morphogenesis during pre-natal life and could participate in heart regeneration and repair, both as endogenous myocardium-derived factor and exogenous cardiotropic drug, during adult life., (© 2012 Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.)

Skeletal muscle regeneration is severely compromised in the case of extended damage. The current challenge is to find factors capable of limiting muscle degeneration and/or potentiating the inherent regenerative program mediated by a specific type of myoblastic cells, the satellite cells. Recent studies from our groups and others have shown that the bioactive lipid, sphingosine 1-phosphate (S1P), promotes myoblast differentiation and exerts a trophic action on denervated skeletal muscle fibres. In the present study, we examined the effects of S1P on eccentric contraction (EC)-injured extensor digitorum longus muscle fibres and resident satellite cells. After EC, skeletal muscle showed evidence of structural and biochemical damage along with significant electrophysiological changes, i.e. reduced plasma membrane resistance and resting membrane potential and altered Na(+) and Ca(2+) current amplitude and kinetics. Treatment with exogenous S1P attenuated the EC-induced tissue damage, protecting skeletal muscle fibre from apoptosis, preserving satellite cell viability and affecting extracellular matrix remodelling, through the up-regulation of matrix metalloproteinase 9 (MMP-9) expression. S1P also promoted satellite cell renewal and differentiation in the damaged muscle. Notably, EC was associated with the activation of sphingosine kinase 1 (SphK1) and with increased endogenous S1P synthesis, further stressing the relevance of S1P in skeletal muscle protection and repair/regeneration. In line with this, the treatment with a selective SphK1 inhibitor during EC, caused an exacerbation of the muscle damage and attenuated MMP-9 expression. Together, these findings are in favour for a role of S1P in skeletal muscle healing and offer new clues for the identification of novel therapeutic approaches to counteract skeletal muscle damage and disease., (© 2011 The Authors Journal of Cellular and Molecular Medicine © 2011 Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.)

Recent evidence indicates that the adult heart contains sub-epicardial cardiogenic niches where cardiac stem cells and stromal supporting cells reside together. Such stromal cells include a special population, previously identified as interstitial Cajal-like cells and recently termed telocytes because of their long, slender processes (telopodes) embracing the myocardial precursors. Specific stromal cells, presumptively originated from the epicardium, have been postulated to populate the developing heart where they are thought to play a role in its morphogenesis. This study is designed to investigate the occurrence of telocytes in the developing heart and provide clues to better understand their role as supporting cells involved in the architectural organization of the myocardium during heart development. Our results showed that stromal cells with the immunophenotypical (vimentin, CD34) and ultrastructural features of telocytes were present in the mouse heart since early embryonic to adult life, as well as in primary cultures of neonatal mouse cardiac cells. These cells formed an extended network of telopodes which closely embraced the growing cardiomyocytes and appeared to contribute to the aggregation of cardiomyocyte clusters in vitro. In conclusion, the present findings strongly suggest that, during heart development, stromal cells identifiable as telocytes could play a nursing and guiding role for myocardial precursors to form the correct three-dimensional tissue pattern and contribute to compaction of the embryonic myocardial trabeculae. It is tempting to speculate that telocytes could be a novel, possible target for therapeutic strategies aimed at potentiating cardiac repair and regeneration after ischemic injury., (© 2010 The Authors Journal compilation © 2010 Foundation for Cellular and Molecular Medicine/Blackwell Publishing Ltd.)

In the post-infarcted heart, grafting of precursor cells may partially restore heart function but the improvement is modest and the mechanisms involved remain to be elucidated. Here, we explored this issue by transplanting C2C12 myoblasts, genetically engineered to express enhanced green fluorescent protein (eGFP) or eGFP and the cardiotropic hormone relaxin (RLX) through coronary venous route to swine with experimental chronic myocardial infarction. The rationale was to deliver constant, biologically effective levels of RLX at the site of cell engraftment. One month after engraftment, histological analysis showed that C2C12 myoblasts selectively settled in the ischaemic scar and were located around blood vessels showing an activated endothelium (ICAM-1-,VCAM-positive). C2C12 myoblasts did not trans-differentiate towards a cardiac phenotype, but did induce extracellular matrix remodelling by the secretion of matrix metalloproteases (MMP) and increase microvessel density through the expression of vascular endothelial growth factor (VEGF). Relaxin-producing C2C12 myoblasts displayed greater efficacy to engraft the post-ischaemic scar and to induce extracellular matrix re-modelling and angiogenesis as compared with the control cells. By echocardiography, C2C12-engrafted swine showed improved heart contractility compared with the ungrafted controls, especially those producing RLX. We suggest that the beneficial effects of myoblast grafting on cardiac function are primarily dependent on the paracrine effects of transplanted cells on extracellular matrix remodelling and vascularization. The combined treatment with myoblast transplantation and local RLX production may be helpful in preventing deleterious cardiac remodelling and may hold therapeutic possibility for post-infarcted patients.

Cannabinoids exert complex actions on neurotransmitter systems involved in cognition, locomotion, appetite, but no information was available so far on the interactions between the endocannabinoid system and histaminergic neurons that command several, similar behavioural states and memory. In this study, we investigated the effect of cannabimimetic compounds on histamine release using the microdialysis technique in the brain of freely moving rats. We found that systemic administration of the cannabinoid receptors 1 (CB1-r) agonist arachidonyl-2'chloroethylamide/N-(2chloroethyl)-5Z,8Z,11Z,14Z-eicosatetraenamide (ACEA; 3 mg/kg) increased histamine release from the posterior hypothalamus, where the histaminergic tuberomamillary nuclei (TMN) are located. Local infusions of ACEA (150 nm) or R(+)-methanandamide (mAEA; 1 microm), another CB1-r agonist, in the TMN augmented histamine release from the TMN, as well as from two histaminergic projection areas, the nucleus basalis magnocellularis and the dorsal striatum. When the endocannabinoid uptake inhibitor AM404 was infused into the TMN, however, increased histamine release was observed only in the TMN. The cannabinoid-induced effects on histamine release were blocked by co-administrations with the CB1-r antagonist AM251. Using double-immunofluorescence labelling and confocal laser-scanning microscopy, CB1-r immunostaining was found in the hypothalamus, but was not localized onto histaminergic cells. The modulatory effect of cannabimimetic compounds on histamine release apparently did not involve inhibition of gamma-aminobutyric acid (GABA)ergic neurotransmission, which provides the main inhibitory input to the histaminergic neurons in the hypothalamus, as local infusions of ACEA did not modify GABA release from the TMN. These profound effects of cannabinoids on histaminergic neurotransmission may partially underlie some of the behavioural changes observed following exposure to cannabinoid-based drugs.