ATP is an important mitogen in the developing retina and its proliferative response decreases as chick retinal cells differentiate in culture. Both non-stimulated or ATP-induced proliferative response was abolished if cycling cells were cocultured with cells from older embryos or cultured with conditioned medium (CM) from postmitotic cells. The effect of CM was dose-dependent and reversible, as removal of CM from the cultures restored both basal and ATP-induced incorporation of [3H]-thymidine. The effect of CM was also dependent on the developmental stage of the retina used to prepare the medium. As tissues from older embryos were used, inhibition of the basal and ATP-induced proliferative response of the cells increased. Similar inhibition of ATP-induced increase in [3H]-thymidine incorporation was observed using CM from purified glial cultures. Neither ARL 67156, an ecto-ATPase inhibitor, prevented nor TGF-beta1 and TGF-beta2 mimicked the inhibitory effect of conditioned medium. Incubation of cells with CM or ATP for 24 h completely abolished the formation of [3H]-phosphoinositides induced by ATP. These effects were blocked by the P2 receptor antagonist PPADS and were not observed with dialysed CM, suggesting that agonist-dependent desensitization of P2 receptors occurred in cultures incubated with CM. However, removal of small molecules such as nucleotides by dialysis did not affect the decline in the proliferative activity induced by CM, suggesting that desensitization is not responsible for the conditioned medium-dependent cell cycle arrest of early developing retinal cells in culture. These results suggest that factors released from postmitotic cells induce the arrest of retinal cells in the mitotic state, a phenomenon that is concomitant with agonist-dependent P2 receptor desensitization.

No abstract

Sustained chronic inflammation induces activation of genes involved in cellular proliferation and apoptosis, thereby causing skeletal muscle degeneration. To investigate in vitro effects of isolated pentacyclic triterpenes from Eugenia punicifolia (Ep-CM) upon signaling pathways involved in the regulation of skeletal muscle cell line proliferation, and in vivo muscular tissue remodeling. C2C12 cells were seeded on eight-well plates and [(3)H]-thymidine incorporation, TUNEL assays, mitochondria viability, zymography for matrix metalloproteases (MMPs), Western blot analysis for MAPKinase signaling pathway, NFκB activation and HMGB1 production subsequently determined under basal conditions and after Ep-CM treatment. A polymer containing Ep-CM was implanted on the volar surface of gastrocnemius muscles subjected to acute injury induced by bupivacaine for local slow and gradual release of bioactive compounds, and mice killed 4 days after surgery. Ep-CM inhibited proliferation of C2C12 myoblast cell line in a dose-dependent manner, confirmed by reduction of [(3)H]-thymidine uptake without affecting cell viability or inducing apoptosis. The cytostatic effect of Ep-CM occurred mainly via inhibition of phosphorylated extracellular signal-regulated kinase (pERK) activation and DNA synthesis, possibly inhibiting the G1 phase of the cell cycle, since Ep-CM increased pAkt and p27(kip1) but reduced Cyclin D1. Ep-CM in vitro treatment increased MMP-9 and MMP-2 activities of C2C12 myoblast cells, but reduced in vivo MMP-9 activity and acute muscular inflammation. Besides cytostatic and anti-inflammatory effects, Ep-CM pentacyclic triterpenes also contributed to degradation of basement membrane components by activating mechanisms of skeletal muscle remodeling in response to local injury.

When retinal cell cultures were mechanically scratched, cell growth over the empty area was observed. Only dividing and migrating, 2 M6-positive glial cells were detected. Incubation of cultures with apyrase (APY), suramin, or Reactive Blue 2 (RB-2), but not MRS 2179, significantly attenuated the growth of glial cells, suggesting that nucleotide receptors other than P2Y 1 are involved in the growth of glial cells. UTPγS but not ADPβS antagonized apyrase-induced growth inhibition in scratched cultures, suggesting the participation of UTP-sensitive receptors. No decrease in proliferating cell nuclear antigen (PCNA + ) cells was observed at the border of the scratch in apyrase-treated cultures, suggesting that glial proliferation was not affected. In apyrase-treated cultures, glial cytoplasm protrusions were smaller and unstable. Actin filaments were less organized and alfa-tubulinlabeled microtubules were mainly parallel to scratch. In contrast to control cultures, very few vinculin-labeled adhesion sites could be noticed in these cultures. Increased Akt and ERK phosphorylation was observed in UTP-treated cultures, effect that was inhibited by SRC inhibitor 1 and PI3K blocker LY294002. These inhibitors and the FAK inhibitor PF573228 also decreased glial growth over the scratch, suggesting participation of SRC, PI3K, and FAK in UTP-induced growth of glial cells in scratched cultures. RB-2 decreased dissociated glial cell attachment to fibronectin-coated dishes and migration through transwell membranes, suggesting that nucleotides regulated adhesion and migration of glial cells. In conclusion, mechanical scratch of retinal cell cultures induces growth of glial cells over the empty area through a mechanism that is dependent on activation of UTP-sensitive receptors, SRC, PI3K, and FAK.

O Diabetes Mellitus Tipo II é uma doença que cursa com uma resposta tecidual subnormal a determinadas concentrações de insulina, através de um mecanismo de resistência à insulina, representando cerca de 90 a 95% dos casos de diabetes mellitus diagnosticados. Há correlação direta entre a angiotensina II e resistência insulínica em portadores de hipertensão arterial e/ou diabetes mellitus tipo II. O objetivo deste artigo é explicar, através de aspectos bioquímicos (reações e mudanças que ocorrem a nível celular, de que forma o Diabetes Mellitus Tipo II e a Hipertensão Arterial Sistêmica se inter-relacionam, no sentido de uma doença poder gerar a outra. Para isso, foram levantados artigos sobre o tema nas bases de dados Scielo, PubMed-Medline, além de livros que abordam o tema. Conclui-se que o principal mecanismo bioquímico envolvido é a ativação da via JAK/STAT/SOCS3, mediado pela ativação do receptor AT1, que por sua vez, determina a ubiquitinação dos substratos responsivos de insulina. Dessa forma, a interface entre o Diabetes Mellitus Tipo II e a Hipertensão Arterial Sistêmica fica melhor compreendida através do entendimento das mudanças bioquímicas que ocorrem na gênese destas duas patologias.