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Lipopolysaccharide modulates astrocytic S100B secretion: a study in cerebrospinal fluid and astrocyte cultures from rats.

Guerra MC, Tortorelli LS, Galland F, Da Ré C, Negri E, Engelke DS, Rodrigues L, Leite MC, Gonçalves CA - J Neuroinflammation (2011)

Bottom Line: However, lower levels of LPS in astrocyte cultures were able to induce a decrease in S100B secretion after 24 h, without significant change in intracellular content of S100B.Together, these data contribute to the understanding of the effects of LPS on astrocytes, particularly on S100B secretion, and help us to interpret cerebrospinal fluid and serum changes for this protein in neuroinflammatory diseases.Moreover, non-brain S100B-expressing tissues may be differentially regulated, since LPS administration did not lead to increased serum levels of S100B.

View Article: PubMed Central - HTML - PubMed

Affiliation: Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal do Rio Grande do Sul, Ramiro Barcelos, Porto Alegre, Brazil.

ABSTRACT

Background: Inflammatory responses in brain are primarily mediated by microglia, but growing evidence suggests a crucial importance of astrocytes. S100B, a calcium-binding protein secreted by astrocytes, has properties of a neurotrophic or an inflammatory cytokine. However, it is not known whether primary signals occurring during induction of an inflammatory response (e.g. lipopolysaccharide, LPS) directly modulate S100B.

Methods: In this work, we evaluated whether S100B levels in cerebrospinal fluid (CSF) and serum of Wistar rats are affected by LPS administered by intraperitoneal (IP) or intracerebroventricular (ICV) injection, as well as whether primary astrocyte cultures respond directly to lipopolysaccharide.

Results: Our data suggest that S100B secretion in brain tissue is stimulated rapidly and persistently (for at least 24 h) by ICV LPS administration. This increase in CSF S100B was transient when LPS was IP administered. In contrast to these S100B results, we observed an increase in in TNFα levels in serum, but not in CSF, after IP administration of LPS. In isolated astrocytes and in acute hippocampal slices, we observed a direct stimulation of S100B secretion by LPS at a concentration of 10 μg/mL. An involvement of TLR4 was confirmed by use of specific inhibitors. However, lower levels of LPS in astrocyte cultures were able to induce a decrease in S100B secretion after 24 h, without significant change in intracellular content of S100B. In addition, after 24 h exposure to LPS, we observed a decrease in astrocytic glutathione and an increase in astrocytic glial fibrillary acidic protein.

Conclusions: Together, these data contribute to the understanding of the effects of LPS on astrocytes, particularly on S100B secretion, and help us to interpret cerebrospinal fluid and serum changes for this protein in neuroinflammatory diseases. Moreover, non-brain S100B-expressing tissues may be differentially regulated, since LPS administration did not lead to increased serum levels of S100B.

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GSH content is modified by LPS in astrocyte cultures and hippocampal slices. Rat cortical astrocytes were cultured in DMEM containing 10% FCS. After confluence, the medium was replaced by DMEM without serum in the presence or absence of LPS (from 0.01 to 30 μg/mL). Cells were lysed in 1 h (A) or 24 h (B) and intracellular GSH content was measured. Each value represents the mean (± standard error) of at least 5 independent experiments performed in triplicate. Means indicated by different letters are significantly different (one way ANOVA followed by Duncan's test, with a significance level of p < 0.05). (C) Adult Wistar rats were killed by decapitation and 0.3 mm hippocampal slices were obtained using a McIlwain chopper. After a metabolic recovery period, hippocampal slices were exposed to LPS (from 0.1 to 10 μg/mL) and intracellular content of S100B was measured by ELISA at 1 h. Each value is the mean (± standard error) of at least 5 independent experiments performed in triplicate. Means indicated by different letters are significantly different (one way ANOVA followed by Duncan's test, with a significance level of p < 0.05).
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Figure 5: GSH content is modified by LPS in astrocyte cultures and hippocampal slices. Rat cortical astrocytes were cultured in DMEM containing 10% FCS. After confluence, the medium was replaced by DMEM without serum in the presence or absence of LPS (from 0.01 to 30 μg/mL). Cells were lysed in 1 h (A) or 24 h (B) and intracellular GSH content was measured. Each value represents the mean (± standard error) of at least 5 independent experiments performed in triplicate. Means indicated by different letters are significantly different (one way ANOVA followed by Duncan's test, with a significance level of p < 0.05). (C) Adult Wistar rats were killed by decapitation and 0.3 mm hippocampal slices were obtained using a McIlwain chopper. After a metabolic recovery period, hippocampal slices were exposed to LPS (from 0.1 to 10 μg/mL) and intracellular content of S100B was measured by ELISA at 1 h. Each value is the mean (± standard error) of at least 5 independent experiments performed in triplicate. Means indicated by different letters are significantly different (one way ANOVA followed by Duncan's test, with a significance level of p < 0.05).

Mentions: Another parameter analyzed to evaluate astroglial activity was intracellular content of glutathione. After exposure of astrocytes to LPS (at concentrations from 0.01 to 30 μg/mL), we observed a decrease in intracellular content of glutathione after 24 h (p = 0.011), but not at 1 h (p = 0.49) (Figure 5A and 5B). Hippocampal slice preparations also exhibited a decrease in glutathione content after LPS exposure for 1 h (p = 0.015) (Figure 5C).


Lipopolysaccharide modulates astrocytic S100B secretion: a study in cerebrospinal fluid and astrocyte cultures from rats.

Guerra MC, Tortorelli LS, Galland F, Da Ré C, Negri E, Engelke DS, Rodrigues L, Leite MC, Gonçalves CA - J Neuroinflammation (2011)

GSH content is modified by LPS in astrocyte cultures and hippocampal slices. Rat cortical astrocytes were cultured in DMEM containing 10% FCS. After confluence, the medium was replaced by DMEM without serum in the presence or absence of LPS (from 0.01 to 30 μg/mL). Cells were lysed in 1 h (A) or 24 h (B) and intracellular GSH content was measured. Each value represents the mean (± standard error) of at least 5 independent experiments performed in triplicate. Means indicated by different letters are significantly different (one way ANOVA followed by Duncan's test, with a significance level of p < 0.05). (C) Adult Wistar rats were killed by decapitation and 0.3 mm hippocampal slices were obtained using a McIlwain chopper. After a metabolic recovery period, hippocampal slices were exposed to LPS (from 0.1 to 10 μg/mL) and intracellular content of S100B was measured by ELISA at 1 h. Each value is the mean (± standard error) of at least 5 independent experiments performed in triplicate. Means indicated by different letters are significantly different (one way ANOVA followed by Duncan's test, with a significance level of p < 0.05).
© Copyright Policy - open-access
Related In: Results  -  Collection

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Figure 5: GSH content is modified by LPS in astrocyte cultures and hippocampal slices. Rat cortical astrocytes were cultured in DMEM containing 10% FCS. After confluence, the medium was replaced by DMEM without serum in the presence or absence of LPS (from 0.01 to 30 μg/mL). Cells were lysed in 1 h (A) or 24 h (B) and intracellular GSH content was measured. Each value represents the mean (± standard error) of at least 5 independent experiments performed in triplicate. Means indicated by different letters are significantly different (one way ANOVA followed by Duncan's test, with a significance level of p < 0.05). (C) Adult Wistar rats were killed by decapitation and 0.3 mm hippocampal slices were obtained using a McIlwain chopper. After a metabolic recovery period, hippocampal slices were exposed to LPS (from 0.1 to 10 μg/mL) and intracellular content of S100B was measured by ELISA at 1 h. Each value is the mean (± standard error) of at least 5 independent experiments performed in triplicate. Means indicated by different letters are significantly different (one way ANOVA followed by Duncan's test, with a significance level of p < 0.05).
Mentions: Another parameter analyzed to evaluate astroglial activity was intracellular content of glutathione. After exposure of astrocytes to LPS (at concentrations from 0.01 to 30 μg/mL), we observed a decrease in intracellular content of glutathione after 24 h (p = 0.011), but not at 1 h (p = 0.49) (Figure 5A and 5B). Hippocampal slice preparations also exhibited a decrease in glutathione content after LPS exposure for 1 h (p = 0.015) (Figure 5C).

Bottom Line: However, lower levels of LPS in astrocyte cultures were able to induce a decrease in S100B secretion after 24 h, without significant change in intracellular content of S100B.Together, these data contribute to the understanding of the effects of LPS on astrocytes, particularly on S100B secretion, and help us to interpret cerebrospinal fluid and serum changes for this protein in neuroinflammatory diseases.Moreover, non-brain S100B-expressing tissues may be differentially regulated, since LPS administration did not lead to increased serum levels of S100B.

View Article: PubMed Central - HTML - PubMed

Affiliation: Departamento de Bioquímica, Instituto de Ciências Básicas da Saúde, Universidade Federal do Rio Grande do Sul, Ramiro Barcelos, Porto Alegre, Brazil.

ABSTRACT

Background: Inflammatory responses in brain are primarily mediated by microglia, but growing evidence suggests a crucial importance of astrocytes. S100B, a calcium-binding protein secreted by astrocytes, has properties of a neurotrophic or an inflammatory cytokine. However, it is not known whether primary signals occurring during induction of an inflammatory response (e.g. lipopolysaccharide, LPS) directly modulate S100B.

Methods: In this work, we evaluated whether S100B levels in cerebrospinal fluid (CSF) and serum of Wistar rats are affected by LPS administered by intraperitoneal (IP) or intracerebroventricular (ICV) injection, as well as whether primary astrocyte cultures respond directly to lipopolysaccharide.

Results: Our data suggest that S100B secretion in brain tissue is stimulated rapidly and persistently (for at least 24 h) by ICV LPS administration. This increase in CSF S100B was transient when LPS was IP administered. In contrast to these S100B results, we observed an increase in in TNFα levels in serum, but not in CSF, after IP administration of LPS. In isolated astrocytes and in acute hippocampal slices, we observed a direct stimulation of S100B secretion by LPS at a concentration of 10 μg/mL. An involvement of TLR4 was confirmed by use of specific inhibitors. However, lower levels of LPS in astrocyte cultures were able to induce a decrease in S100B secretion after 24 h, without significant change in intracellular content of S100B. In addition, after 24 h exposure to LPS, we observed a decrease in astrocytic glutathione and an increase in astrocytic glial fibrillary acidic protein.

Conclusions: Together, these data contribute to the understanding of the effects of LPS on astrocytes, particularly on S100B secretion, and help us to interpret cerebrospinal fluid and serum changes for this protein in neuroinflammatory diseases. Moreover, non-brain S100B-expressing tissues may be differentially regulated, since LPS administration did not lead to increased serum levels of S100B.

Show MeSH
Related in: MedlinePlus