Interleukin-6 increases the survival of retinal ganglion cells in vitro.

Interleukin-6 is a pleiotropic cytokine that mediates cellular communication both in physiological and pathological states. In this work, we demonstrate that 50 ng/mL IL-6 increases the survival of retinal ganglion cells (RGCs) after 48 h in culture. This effect was blocked by an intracellular Ca(+2) chelator, by inhibition of ryanodinic receptors and by an inhibitor of L-type Ca(+2) channels. IL-6 effect is mediated by PKC, tyrosine kinase, PI3-kinase and MEK activity. The blockade of polypeptide release also abolished the effect of IL-6. These results suggest a role for this cytokine during the development of the central nervous system (CNS).

Interleukin-2 and interleukin-4 increase the survival of retinal ganglion cells in culture.

Natural cell death is a degenerative phenomenon observed during the normal development of the nervous system. The neuroprotective effects of cytokines produced by neuronal, glial or infiltrating cells on neurons have been extensively studied. In this work we studied the role of interleukin (IL)-2 and IL-4 on the survival of retinal ganglion cells (RGC) after 48 h in culture. Our results demonstrate that the effect of both ILs was dose-dependent and the treatment with either IL-2 (50 U/ml) or IL-4 (5 U/ml) induced a 2-fold increase in RGC survival. The effect of IL-4, but not of IL-2, was totally abolished by either 20 microM 5-fluoro-2'-deoxyuridine, an inhibitor of cell proliferation, or by 1 microM telenzepine, an inhibitor of M1 muscarinic receptor. Our results suggest that both cytokines could play an important role during the development of retinal tissue as well as during retina trauma.

Effect of spleen-cell-conditioned medium on [3H]-choline uptake by retinal cells in vitro is mediated by IL-2.

Cytokines are essential molecules throughout the development of the nervous system and also play an important role during the adult life span. In the present work, we analyzed in vitro the effect of spleen-cell-conditioned medium (SCM) on the survival and [3H]-choline uptake of neonatal rat retinal cells. SCM induced an increase in neuronal survival, glial cell proliferation and neurite outgrowth, as evaluated by biochemical and morphological criteria. These effects were time dependent; after 120 h, SCM induced a 6-fold increase in the total protein level. The effect of SCM was blocked both by the inhibition of protein tyrosine kinase activity (10 microM genistein) and by the inhibition of cell division (20 microM fluorodeoxyuridine). SCM also increased the uptake of [3H]-choline by retinal cells. The effect was time dependent. The maximum effect was obtained after 48 h and was maintained at a high level until 120 h. Treatment by 10 microM genistein and 15 microM bis(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid (BAPTA-AM) (an intracellular calcium chelator) completely blocked this effect. However, 20 microM fluorodeoxyuridine did not abolish it. Conditioned medium obtained from glial cells stimulated with SCM (S-GCM) induced an effect on [3H]-choline uptake earlier than that promoted by SCM. Anti-interleukin-2 (IL-2) antibodies blocked the effect of both SCM and S-GCM on [3H]-choline uptake after 48 and 72 h. IL-2 (50 U/ml) elicited the same effect as that observed when the cells were maintained in the presence of SCM. Taken together, our results suggest that IL-2 plays an important role in controlling the survival and differentiation of retinal cells in vitro.

The effect of PKC activation on the survival of rat retinal ganglion cells in culture.

Natural cell death is a degenerative phenomenon occurring during the development of the nervous system. Approximately half the neurons initially generated during this period die. The role of trophic molecules produced by target and afferent neurons as well as by glial cells controlling this regressive event has been extensively demonstrated. The aim of this work was to study the role of activated protein kinase C (PKC), an enzyme involved in apoptosis regulation, on the survival of retinal ganglion cells kept "in vitro" for 48 h. For this purpose, we used the phorbol 12-myristate 13-acetate (PMA), a tumor promoter agent that activates PKC. Our results showed that PMA increases the survival of ganglion cells. The effect was dose-dependent and PMA concentrations of 10 or 100 ng/ml produced the maximal effect (a two-fold increase on ganglion cells survival compared with 48 h control). This effect was totally abolished by 1.25 microM chelerythrine chloride (an inhibitor of PKC) and 30 microM genistein (an inhibitor of tyrosine kinase enzymes). Otherwise, PMA was effective only when it was chronically present in the cultures. On the other hand, treatment with 20 microM 5-fluoro-2'-deoxyuridine, an inhibitor of cell proliferation, or 25 microM BAPTA-AM, an intracellular calcium chelator, did not block PMA effect. Our results suggest that the survival of retinal ganglion cells "in vitro" may be mediated by a mechanism that involves PKC activation.

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Trophic factors produced by retinal cells increase the survival of retinal ganglion cells in vitro.

The naturally occurring neuron death of normal development has been shown to depend on trophic factors produced and released by target cells. It has also been shown that the afferent supply and local interactions play a role in the control of this degenerative phenomenon. We studied the effect of trophic factors produced by intrinsic retinal cells on the survival of retinal ganglion cells in vitro. Retinae of newborn hooded rats were retrogradely labelled with horseradish peroxidase injected into the superior colliculus to permit the identification of retinal ganglion cells in culture. We tested the effect of conditioned media either from aggregates or from explants of retinal cells from neonatal rats on the survival of ganglion cells in vitro. Our results showed that both conditioned media increased the survival of these cells. The trophic activity was dose-dependent, was maintained after dialysis against a 12 kDa membrane, was abolished by heating at 56 degrees C for 30 min, and was not found in conditioned medium from cerebral cortical explants. Conditioned medium obtained without fetal calf serum presented the same trophic effect. These results suggest that the local control of developmental neuron death by intrinsic retinal cells may be mediated by neurotrophic factors.

Survival of retinal ganglion cells in vitro: the effect of conditioned medium from retinal cell aggregates.

The time course of degeneration of retinal ganglion cells was studied in vitro. We used the retinae of newborn hooded rats retrogradely labeled with horseradish peroxidase injected bilaterally into the superior colliculus to permit the identification of retinal ganglion cells in culture. We tested the effect of conditioned medium either from aggregates or from explants of retinal cells on the survival of the ganglion cells. Both conditioned media approximately doubled the survival of ganglion cells after 48-72 h in culture. Our data are consistent with the hypothesis that retinal cells produce soluble trophic factors that influence ganglion cell survival.