RESUMO
The differentiation of myelin-forming Schwann cells (SC) is completed with the appearance of myelin proteins MBP and P(0) and a concomitant downregulation of markers GFAP and p75NTR, which are expressed by immature and adult non-myelin-forming SC. We have previously demonstrated that holotransferrin (hTf) can prevent SC dedifferentiation in culture (Salis et al., 2002), while apotransferrin (aTf) cannot. As a consequence, we used pure cultured SC and submitted them to serum deprivation in order to promote dedifferentiation and evaluate the prodifferentiating ability of ferric ammonium citrate (FAC) through the expression of MBP, P(0), p75NTR and c-myc. The levels of cAMP, CREB and p-CREB were also measured. Results show that Fe(3+), either in its free form or as hTf, can prevent the dedifferentiation promoted by serum withdrawal. Both FAC and hTf were proven to promote differentiation, probably through the increase in cAMP levels and CREB phosphorylation, as well as levels of reactive oxygen species. This effect was inhibited by deferroxamine (Dfx, an iron chelator), H9 (a cAMP-PKA antagonist) and N-acetylcysteine (NAC, a powerful antioxidant).
Assuntos
Diferenciação Celular/fisiologia , AMP Cíclico/fisiologia , Compostos Férricos/farmacologia , Compostos de Amônio Quaternário/farmacologia , Células de Schwann/fisiologia , Transferrina/fisiologia , Animais , Animais Recém-Nascidos , Diferenciação Celular/efeitos dos fármacos , Células Cultivadas , Ferro/fisiologia , Ratos , Ratos Wistar , Células de Schwann/efeitos dos fármacosRESUMO
Transferrin, the iron carrier protein, has been shown to be involved in oligodendroglial cell differentiation in the central nervous system but little is known about its role in the peripheral nervous system. In the present work, we have studied the presence of transferrin and of its mRNA in rat sciatic nerves and in Schwann cells isolated at embryonic and adult ages as well as during the regeneration process that follows nerve crush. We have also studied the correlation between the expression of the mRNAs of transferrin and the expression of mature myelin markers in the PNS. We show that transferrin is present in whole sciatic nerves at late stages of embryonic life as well as at postnatal day 4 and in adult rats. We demonstrate for the first time, that in normal conditions, the transferrin mRNA is expressed in Schwann cells isolated from sciatic nerves between embryonic days 14 and 18, being absent at later stages of development and in adult animals. In adult rats, 3 days after sciatic nerve crushing, the mRNA of transferrin is expressed in the injured nerve, but 7 days after injury its expression disappears. Transferrin protein in the sciatic nerve closely follows the expression of its mRNA indicating that under these circumstances, it appears to be locally synthesized. Transferrin in the PNS could have a dual role. During late embryonic ages it could be locally synthesized by differentiating Schwann cells, acting as a pro-differentiating factor. A similar situation would occur during the regeneration that follows Wallerian degeneration. In the adult animals on the other hand, Schwann cells could pick up transferrin from the circulation or/and from the axons, sub serving possible trophic actions closely related to myelin maintenance.
Assuntos
RNA Mensageiro/metabolismo , Células de Schwann/metabolismo , Nervo Isquiático/lesões , Transferrina/genética , Envelhecimento/metabolismo , Animais , Animais Recém-Nascidos , Senescência Celular , Embrião de Mamíferos/metabolismo , Feminino , Compressão Nervosa , Gravidez , Ratos , Ratos Wistar , Células de Schwann/patologia , Nervo Isquiático/embriologia , Nervo Isquiático/metabolismo , Nervo Isquiático/patologia , Fatores de Tempo , Transferrina/metabolismo , Ferimentos e Lesões/metabolismo , Ferimentos e Lesões/patologia , Ferimentos e Lesões/fisiopatologiaRESUMO
We have previously reported that in the distal stump of ligated sciatic nerves, there is a change in the distribution of myelin basic protein (MBP) and P0 protein immunoreactivities. These results agreed with the studies of myelin isolated from the distal stump of animals submitted to ligation of the sciatic nerve, showing a gradual increase in a 14 kDa band with an electrophoretic mobility similar to that of an MBP isoform, among other changes. This band, which was resolved into two bands of 14 and 15 kDa using a 16% gel, was found to contain a mixture of MBP fragments and peptides with great homology with alpha- and beta-globins. In agreement with these results, we have demonstrated that the mRNA of alpha-globin is present in the proximal and distal stumps of the ligated nerve. It is also detected at very low levels in Schwann cells isolated from normal nerves. These results could be due to the presence of alpha- and/or beta-globin arising from immature cells of the erythroid series. Also, they could be present in macrophages, which spontaneously migrate to the injured nerve to promote the degradation of myelin proteins. Cells isolated from normal adult rat bone marrow which were injected intraortically were found to migrate to the injured area. These cells could contribute to the remyelination of the damaged area participating in the removal of myelin debris, through their transdifferentiation into Schwann cells or through their fusion with preexisting Schwann cells in the distal stump of the injured sciatic nerve.
Assuntos
Células da Medula Óssea/fisiologia , Globinas/biossíntese , Degeneração Neural/patologia , Regeneração Nervosa/fisiologia , RNA Mensageiro/biossíntese , Nervo Isquiático/lesões , Nervo Isquiático/metabolismo , Animais , Western Blotting , Movimento Celular/fisiologia , Eletroforese em Gel de Poliacrilamida , Feminino , Imuno-Histoquímica , Masculino , Proteína Básica da Mielina/metabolismo , Ensaios de Proteção de Nucleases , Peptídeos/química , Nervos Periféricos/metabolismo , Ratos , Ratos Wistar , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Células de Schwann/fisiologia , Nervo Isquiático/patologia , Espectrometria de Massas por Ionização e Dessorção a Laser Assistida por Matriz , TripsinaRESUMO
Schwann cells (SCs) in culture, without the presence of axons, become de-differentiated, reaching a condition similar to that of their precursor cells. The cytoplasmic accumulation of transferrin (Tf) in the myelinated peripheral nerve has been reported and data in the literature support a role for apoTf in myelination in the CNS. In the present report, we used SC cultures to evaluate the capacity of apoTf and holoTf to prevent cell de-differentiation promoted by fetal calf serum deprivation. SCs incubated in a serum-free medium showed a decrease in the expression of myelin basic protein (MBP) and P(0), markers of mature myelin-forming SCs, together with an increase in the levels of p75NTR and glial fibrillary acidic protein, markers of immature SCs. Treatment with holoTf prevented the decrease in expression of MBP and P(0) and the increase in p75NTR. ApoTf was unable to prevent these changes except when iron was added to the cultures. These results suggest a role for holoTf in the regulation of myelin formation by SCs.