ARHGAP30 is a Wrch-1-interacting protein involved in actin dynamics and cell adhesion.

The atypical Rho GTPase Wrch-1 has been proposed roles in cell migration, focal adhesion dissolution, stress fibre break down and tight junction heterogeneity. A screen for Wrch-1 binding-partners identified the novel RhoGAP protein, ARHGAP30, as a Wrch-1 interactor. ARHGAP30 is related to the Cdc42- and Rac1-specific RhoGAP CdGAP, which was likewise found to bind Wrch-1. In contrast to CdGAP, ARHGAP30 serves as a Rac1- and RhoA-specific RhoGAP. Ectopic expression of ARHGAP30 results in membrane blebbing and dissolution of stress-fibres and focal adhesions. Our data suggest roles for ARHGAP30 and CdGAP in regulation of cell adhesion downstream of Wrch-1.

RANKL/OPG in primary cultures of osteoblasts from post-menopausal women. Differences between osteoporotic hip fractures and osteoarthritis.

The OPG/RANKL/RANK system is important in the balance between bone formation and resorption. We used primary human osteoblasts (hOBs) cells to examine the impact of 17-beta-estradiol (E2) or/and 1,25-dihydroxyvitamin D (1,25D) in OPG/RANKL system in 28 post-menopausal (PM) women; (a) with hip fracture (OP) or (b) with osteoarthritis (OA). The hOB from OP patients proliferated slower during the first stage, than the OA women (31.5+/-2.6 and 21.4+/-1.3 days, respectively, p<0.05). The OP group secreted significantly higher OPG protein levels than the OA women (10.1+/-2.6 and 4.4+/-0.8pmol/L, respectively, p<0.05). The 1,25D and 1,25D+E2 induce an increase in RANKL and RANKL/OPG mRNA expression in OP patients above 200% (p<0.05). HOBs from the osteoporotic hip initially proliferate slower but after reaching the first cellular confluence, the proliferation rate is equal in both groups. Furthermore, hOBs from hips with OP present a higher protein secretion of OPG, and higher RANKL and RANKL/OPG expression ratio in response to 1,25D and 1,25D+E2, than hOBs from OA women. All this could suggest that the greater bone loss that characterizes OP patients can be mediated due to differences in the secretion and expression of the RANKL/OPG system in response to different stimuli.

Beneficial pleiotropic actions of melatonin in an experimental model of septic shock in mice: regulation of pro-/anti-inflammatory cytokine network, protection against oxidative damage and anti-apoptotic effects.

Septic shock, the most severe problem of sepsis, is a lethal condition caused by the interaction of a pathogen-induced long chain of sequential intracellular events in immune cells, epithelium, endothelium, and the neuroendocrine system. The lethal effects of septic shock are associated with the production and release of numerous pro-inflammatory biochemical mediators including cytokines, nitric oxide and toxic oxygen and nitrogen radicals, together with development of massive apoptosis. As melatonin has remarkable properties as a cytokine modulator, antioxidant and anti-apoptotic agent, the present study was designed to evaluate the possible protective effect of melatonin against LPS-induced septic shock in Swiss mice. We observed that intraperitoneally (i.p.) administered-melatonin (10 mg/kg) 30 min prior, and 1 hr after i.p. LPS injection (0.75 mg/animal) markedly protected mice from the LPS lethal effects with 90% survival rates for melatonin and 20% for LPS-injected mice after 72 hr. The melatonin effect was mediated by modulating the release of pro-/anti-inflammatory cytokine levels, protection from oxidative damage and counteracting apoptotic cell death. Melatonin was able to partially counteract the increase in LPS-induced pro-inflammatory cytokine levels such as tumor necrosis factor-alpha, IL-12 and interferon-gamma at the local site of injection, while it increased the production of the anti-inflammatory cytokine IL-10 both locally and systemically. Furthermore, melatonin inhibited the LPS-induced nitrite/nitrate and lipid peroxidation levels in brain and liver and counteracted the sepsis-associated apoptotic process in spleen. In conclusion, we have demonstrated that melatonin improves the survival of mice with septic shock via its pleiotropic functions as an immunomodulator, antioxidant and anti-apoptotic mediator.

Expression of membrane and nuclear melatonin receptors in mouse peripheral organs.

Previous studies have shown that melatonin acts through specific receptors, including MT(1) and MT(2) membrane receptors as well as a nuclear receptor belonging to the orphan nuclear receptor family. Therefore, the goal of this study was to determine whether melatonin receptors mRNA is expressed in mouse peripheral tissues. To study the different receptors subtype expression, we have used a reverse-transcription polymerase chain reaction (RT-PCR) procedure followed by Southern hybridization with specific digoxigenin-labeled probes. RT-PCR studies revealed the presence of both MT(1) membrane receptors and ROR(alpha)1 nuclear receptors in all the peripheral tissues examined (brain, heart, lung, liver and kidney). Moreover, the expression of ROR(alpha)1 nuclear receptors was also demonstrated by Western-blot. In contrast, expression of MT(2) membrane receptors was only observed in brain and lung. These results suggest that melatonin, acting through its different subtypes receptors, plays a role in the neuroendocrine regulation of peripheral tissues function.