The CMS19 disease model specifies a pivotal role for collagen XIII in bone homeostasis.

Mutations in the COL13A1 gene result in congenital myasthenic syndrome type 19 (CMS19), a disease of neuromuscular synapses and including various skeletal manifestations, particularly facial dysmorphisms. The phenotypic consequences in Col13a1 null mice (Col13a1-/-) recapitulate the muscle findings of the CMS19 patients. Collagen XIII (ColXIII) is exists as two forms, a transmembrane protein and a soluble molecule. While the Col13a1-/- mice have poorly formed neuromuscular junctions, the prevention of shedding of the ColXIII ectodomain in the Col13a1tm/tm mice results in acetylcholine receptor clusters of increased size and complexity. In view of the bone abnormalities in CMS19, we here studied the tubular and calvarial bone morphology of the Col13a1-/- mice. We discovered several craniofacial malformations, albeit less pronounced ones than in the human disease, and a reduction of cortical bone mass in aged mice. In the Col13a1tm/tm mice, where ColXIII is synthesized but the ectodomain shedding is prevented due to a mutation in a protease recognition sequence, the cortical bone mass decreased as well with age and the cephalometric analyses revealed significant craniofacial abnormalities but no clear phenotypical pattern. To conclude, our data indicates an intrinsic role for ColXIII, particularly the soluble form, in the upkeep of bone with aging and suggests the possibility of previously undiscovered bone pathologies in patients with CMS19.

Curcumin induces apoptosis in breast cancer cell lines and delays the growth of mammary tumors in neu transgenic mice.

Breast cancer is a leading cancer in women and despite the benefits of the current therapies a significant number of patients with this tumor is at risk of relapse. Some of the alterations taking place in breast cancer cells are currently exploited by molecularly targeted drugs. Different drugs have been developed which target a single molecule but, given that the tumor originates from the dysregulation of many genes, there is the need to find new drugs that have more than one molecular target. Curcumin [1,7-bis-(4-hydroxy-3-methoxyphenyl)-1,6-heptadiene-3,5-dione] (CUR), a polyphenolic compound found in the spice turmeric, is a pleiotropic molecule able to interact with a variety of molecular targets and has antitumor, anti-inflammatory, antioxidant, immunomodulatory and antimicrobial activities. Here we demonstrate that CUR inhibits the growth of breast cancer cell lines in a dose dependent manner, with IC50 values in the micromolar range, and induces an increase in the percentage of cells in sub-G0 phase, representing the apoptotic cell population. The activation of apoptosis was confirmed by PARP-1 cleavage and by the increased ratio between the pro-apoptotic Bax and the anti-apoptotic Bcl-2 protein. In addition, in CUR-treated cells the activity of ERK1/ERK2 MAP kinases was down-regulated. The cytotoxic effects of CUR were observed in breast cancer cells expressing either high or low levels of ErbB2/neu. The in vivo antitumor activity of CUR was tested in BALB-neuT mice transgenic for the neu oncogene, which develop atypical hyperplasia of the mammary gland at 6 weeks of age and invasive carcinoma at 16 weeks of age. CUR, administered to mice both early and in an advanced stage of mammary carcinogenesis, induced a significant prolongation of tumor-free survival and a reduction of tumor multiplicity. In addition, CUR administration was safe, since no modification of hematological and clinical chemistry parameters could be observed in BALB-neuT and BALB/c mice treated with this compound for several weeks. These findings support further studies on the therapeutic potential of CUR in combination with standard therapies in breast cancer patients.

Brain Natriuretic Peptide (BNP) regulates the production of inflammatory mediators in human THP-1 macrophages.

Brain Natriuretic Peptide (BNP), besides retaining vasodilatory, diuretic and natriuretic properties, is a vasoactive hormone that it is also involved in several cardiac diseases as well as severe sepsis and septic shock. All these conditions are characterized by an ongoing inflammatory response consisting in a complex interaction of pleiotropic mediators derived from plasma or cells, including monocytes and macrophages. However, the relationship between this hormone and inflammation remains to be elucidated. Therefore, the aim of the present study was to evaluate a possible BNP immunomodulatory activity on macrophages. Our results demonstrate that BNP regulates the production of major inflammatory molecules, such as reactive oxygen- and nitrogen species (ROS and RNS), leukotriene B(4) (LTB(4)), prostaglandin E(2) (PGE(2)); modulates the cytokines (TNF-alpha, IL-12 and IL-10) profile, and affects cell motility. These results furnish novel and brand-new proofs on BNP ability of modulating the production of inflammatory mediators in macrophages whose role has broad implications in inflammatory states where increased BNP levels have been reported.

Endomorphin-1 inhibits the activation and the development of a hyporesponsive-like phenotype in lipopolysaccharide- stimulated THP-1 monocytes.

Endomorphin-1 (EM-1) is an endogenous opioid peptide selectively binding to micro opioid receptors (MORs). Besides its analgesic effects on the central nervous system (CNS), it has been recently reported that EM-1 can cross the blood-brain barrier (BBB) and diffuse into the blood, behaving as an analgesic/anti-inflammatory molecule on peripheral tissues, thus leading to the hypothesis that it could represent a soluble modulator of immune cell functions. Interestingly, nothing is known about its possible effects on monocytes, the main circulating cell-type involved in those systemic responses, such as fever and septic states, involving the release of high amounts of pyrogenic inflammatory factors. The aim of this work is to evaluate possible EM-1effects on lipopolisaccharide (LPS)-stimulated THP-1 monocytes in terms of the production of inflammatory mediators and the instauration of a hyporesponsive-like phenotype which is a main feature of systemic inflammatory responses, and on the development of peripheral monocytes to DC. Our data demonstrate for the first time that EM-1 is able to inhibit both LPS-stimulated monocyte activation, in terms of arachidonic acid, PGE2, ROI and NO2 production and instauration of a hyporesponsive phenotype without any macroscopic effect on DC development. These data support the hypothesis that EM-1 could be involved in modulating monocyte functions during systemic inflammatory reactions, also providing new evidence for its eventual clinical application in endotoxic states.

Activatory properties of lysophosphatidic acid on human THP-1 cells.

Excessive leukocyte proliferation and proinflammatory mediators release represent common phenomena in several chronic inflammatory diseases. Multiple evidences identify lysophosphatidic acid (LPA), a small lipid endowed with pleiotropic activities, as an important modulator of both proliferation and activation of different cell types involved in several inflammation-associated pathologies. However, its possible role on monocyte proinflammatory activation is not fully understood yet. Aim of the present study was to investigate LPA effects on THP-1 cells in terms of proliferation, reactive oxygen intermediates (ROI) production and release of arachidonic acid-derived inflammatory mediators. Actually, LPA significantly increased both DNA synthesis and ROI production as well as prostaglandin E(2) release and the upregulation of LPA(3) receptor expression. These findings identified LPA as both a growth factor and a triggering mediator of proinflammatory response in THP-1 cells.

Sphingosine 1-phosphate interferes on the differentiation of human monocytes into competent dendritic cells.

Sphingosine 1-phosphate (S1P) is a lipidic messenger known to exert several physiological functions within the cell. We tested here whether the stimulation of human monocytes with different doses of S1P might interfere with their differentiation into competent dendritic cells (DC). Monocytes cultured with granulocyte macrophage colony stimulating factor, interleukin-4 (IL-4) and S1P differentiated into a DC population lacking CD1a molecules on the surface and acquired some aspects of mature DC (mDC), though in the absence of maturation stimuli. When stimulated with lipopolisaccharide (LPS), CD1a(-) DC produce high amounts of tumour necrosis factor-alpha and IL-10, but not IL-12. Accordingly, these CD1a(-) DC were not capable of stimulating allogenic T lymphocytes so well as CD1a(+) DC generated from untreated monocytes and maturated with LPS. S1P monocyte-derived DC lost their polarizing capacity abrogating the production of gamma-interferon/IL-4 by co-cultured naïve CD4(+)CD45RA(+) T cells. Our findings suggest a mechanism through which S1P can favour the development of immune-related pathological states.

Activatory properties of lysophosphatidic acid on human THP-1 cells.

Excessive leukocyte proliferation and proinflammatory mediators release represent common phenomena in several chronic inflammatory diseases. Multiple evidences identify lysophosphatidic acid (LPA), a small lipid endowed with pleiotropic activities, as an important modulator of both proliferation and activation of different cell types involved in several inflammation-associated pathologies. However, its possible role on monocyte proinflammatory activation is not fully understood yet. Aim of the present study was to investigate LPA effects on THP-1 cells in terms of proliferation, reactive oxygen intermediates (ROI) production and release of arachidonic acid-derived inflammatory mediators. Actually, LPA significantly increased both DNA synthesis and ROI production as well as prostaglandin E(2) release and the upregulation of LPA(3) receptor expression. These findings identified LPA as both a growth factor and a triggering mediator of proinflammatory response in THP-1 cells.