We are sorry, but NCBI web applications do not support your browser and may not function properly. More information

Results: 3

1.
Figure 1

Figure 1. From: Minocycline corrects early, pre-plaque neuroinflammation and inhibits BACE-1 in a transgenic model of Alzheimer's disease-like amyloid pathology.

Minocycline corrects neuroinflammation in young, pre-plaque Tg mice. (A-C) Representative western blots for iNOS, COX-2 and IL-1β (typical markers of microglial and neuronal activation) in control, Non Tg Placebo mice (Non Tg), Tg Placebo mice (Tg Pl) and Tg mice treated with minocycline (Tg Mino). Note the significant up-regulation of iNOS and COX-2 in Tg Pl compared to Non Tg. Minocycline restored iNOS and COX-2 to levels similar to those of Non Tg, and significantly reduced the levels of IL-1β (* P < 0.05, **P < 0.01, one-way ANOVA with Tukey post-hoc test). See main text for the values of Non Tg treated with minocycline. (D) Schematic illustrating the sampling of images in the CA1 area of the hippocampus and from the lateral posterior thalamic nucleus as utilized for the morphological study represented in E-G. (E) Representative micrographs illustrating the ir of Iba-1 in microglial cells in the hippocampus of the Non Tg, Tg Pl and Tg Mino mice. Note the altered morphology of the microglial cells in Tg Pl compared to Non Tg. In Tg Pl mice, we observed an enlargement of the soma size, polarization and thickening of the microglial processes, which are classical indicators of microglial activation. Minocycline treatment resulted in correction of microglial soma size (note the small, roundish morphology), with some residual thickening of processes and increase in spiny processes. (F) Representative micrographs illustrating the ir of Iba-1 in microglial cells in the lateral posterior thalamic nucleus of the Non Tg, Tg Pl and Tg Mino. The cells were notably smaller and less ramified than in the hippocampus, but no differences in the morphology could be observed between experimental groups. Scale bar for E and F: 20 μm. (G,H) Quantification of cell soma size and density of microglial cells from hippocampus (G) and thalamus (H) in Non Tg, Tg Pl and Tg Mino. Note that minocycline treatment resulted in significant reduction of microglial cells soma size compared to Tg Pl in the hippocampus (G). No significant differences were observed in the soma size of cells in the thalamus (H). As previously reported [23], no significant changes in microglial cell density were observed between Non Tg and Tg. This pattern was not altered across experimental groups in any area. (* P < 0.05, one-way ANOVA followed by Tukey post-hoc test). Non Tg: Non Tg Placebo mice; Tg Mino: Tg mice treated with minocycline; Tg Pl: Tg Placebo.

Maria Teresa Ferretti, et al. J Neuroinflammation. 2012;9:62-62.
2.
Figure 3

Figure 3. From: Minocycline corrects early, pre-plaque neuroinflammation and inhibits BACE-1 in a transgenic model of Alzheimer's disease-like amyloid pathology.

Minocycline inhibits β-site APP cleaving enzyme 1 in young, pre-plaque Tg mice. (A) BACE-1 activity was quantified from hippocampal homogenates using a well-characterized fluorometric assay. Fluorescent units (FU) data were normalized on Non Tg Placebo values. Note the significant up-regulation of BACE-1 activity in Tg Placebo compared to Non Tg Placebo (***P < 0.001), which was corrected following minocycline treatment (*P < 0.05 compared to Tg Placebo). (B) Specificity of the BACE-1 fluorometric assay: note how the fluorescent units detected from brain homogenates (of a Non Tg Placebo animal) are seven times higher than the background (bg). The signal was completely abolished by co-incubation with a specific BACE inhibitor (BACEi), and was in the range of activity of a recombinant human BACE protein (rhBACE, 2.5 μg). All the data were analyzed with one-way ANOVA followed by Tukey post-hoc test. (C) Specificity of the polyclonal BACE-1 antibody used for western blotting: the specific band (close to 75 kDa) appeared in brain extracts from post natal day 7 (pnd 7) and from adult cortex (ctx), while it was hardly detectable in cerebellum (cereb) sample and was not detectable in a glial preparation from pups. (D) Representative western blots of BACE-1, NFkB and IkB from the same hippocampal samples used for the enzymatic assay (ns = non-specific band recognized by the polyclonal antibody directed against NFkB). (E) Quantification data of BACE-1, NFkB and IkB from hippocampal homogenates. Band intensity was quantified via densitometry and the values were normalized on the neuronal specific-β-tubulin content. Note the significant up-regulation of BACE-1 levels in Tg Placebo, which was corrected by the minocycline treatment (P < 0.05). The levels of NFkB and BACE-1 in each sample strongly correlated (P < 0.001, Spearman's correlation analysis). Given the not-normal distribution of the data, the values were analyzed with Kruskal-Wallis test and Spearman's correlation analysis. BACEi: BACE inhibitor; bg; background; cereb: cerebellum; ctx: cortex; pnd: postnatal day; rhBACE: recombinant human BACE.

Maria Teresa Ferretti, et al. J Neuroinflammation. 2012;9:62-62.
3.
Figure 2

Figure 2. From: Minocycline corrects early, pre-plaque neuroinflammation and inhibits BACE-1 in a transgenic model of Alzheimer's disease-like amyloid pathology.

Minocycline effects on amyloid precursor protein metabolism. Cortical brain homogenates from Tg animals treated with vehicle (Tg Pl) or minocycline (Tg Mino) were subjected to western blotting and ELISA to determine the levels of Aβ species, full-length APP (flAPP) and CTF. (A) Representative western blot of cortical homogenates from Tg animals treated with vehicle or with minocycline, using 6E10 antibody. Note the strong down-regulation of the approximately 12-kDa ir band (indicated with an asterisk) which likely represents a mixture of β-CTF of APP and Aβ trimers. (B) Densitometric analysis of the 12-kDa band detected with 6E10. Values were normalized on neuronal specific β-tubulin. Minocycline-treated animals showed significantly lower levels compared with placebo (**P < 0.01, Student's t-test). (C) Correlation analysis of the levels of the 12-kDa band ir to 6E10 and the levels of iNOS per sample. The correlation was found to be highly significant (r = 0.81; P = 0.004, Spearman's correlation), where Tg Placebo animals (black dots) had the highest levels of 6E10-ir and iNOS, while Tg Mino (yellow dots) mice displayed reduced levels of ir with 6E10 and lower levels of iNOS. (D) Representative western blots of cortical (ctx) homogenates from placebo and minocycline-treated animals using the pab27576 antibody. This antibody recognizes an epitope on the C-terminus of APP and therefore detects two prominent bands in the blots: a high molecular weight band (about 100 kDa), corresponding to flAPP and a faster band (approximately 12 kDa) corresponded to the CTF fragments. A calibration curve using known amounts of recombinant C100 was run in the same gel for quantification purposes (the recombinant peptide ran slightly slower due to the presence of a C-terminal hexa-His tag). (E) Quantification of flAPP and CTF levels in cortical homogenates from Tg Placebo and Tg-treated animals. FlAPP relative optical density values were normalized on neuronal specific β-tubulin. Note the significant down-regulation of flAPP (**P < 0.01, Student's t-test) in Tg Mino compared with Tg Placebo. CTF absolute levels were extrapolated from the calibration curve of recombinant CTF. There was a non-significant trend towards reduction following minocycline treatment. Normalization of the relative optical density of CTF over the relative optical density of flAPP confirmed a non-significant trend towards a reduction (ratio CTF/flAPP). All data were analyzed with Student's t-test. (F) Quantification of human Aβ levels in cortical homogenates from Tg Placebo and Tg Mino using ELISA. The treatment reduced both Aβ40 and Aβ42, but the effect did not reach significance (P = 0.09 and 0.08, respectively, Student's t-test; see main text for exact values). (G) The ratios between Aβ (pg/mg total protein) and CTF (ng/mg total protein) were calculated per sample and compared across groups. We observed 24.29 ± 6.43 pg of Aβ42 and 15.14 ± 3.08 pg of Aβ40 per each nanogram of CTF. Minocycline treatment resulted in a reduction of the ratio (which did not reach significance), suggesting that less Aβ was produced per CTF molecule after the treatment with minocycline.

Maria Teresa Ferretti, et al. J Neuroinflammation. 2012;9:62-62.

Supplemental Content

Recent activity

Your browsing activity is empty.

Activity recording is turned off.

Turn recording back on

See more...
Write to the Help Desk