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Soyasaponin I improved neuroprotection and regeneration in memory deficient model rats.

Hong SW, Heo H, Yang JH, Han M, Kim DH, Kwon YK - PLoS ONE (2013)

Bottom Line: Oral administration of soya-I exhibited significant memory-enhancing effects in the passive avoidance, Y-maze, and Morris water maze tests.Addition of soya-Ι in the cultured NPCs significantly elevated the markers for cell proliferation (Ki-67) and neuronal differentiation (NeuN, TUJ1, and MAP2).Soya-Ι may improve hippocampal learning and memory impairment by promoting proliferation and differentiation of NPCs in the hippocampus through facilitation of neuronal regeneration and minimization of neuro-inflammation.

View Article: PubMed Central - PubMed

Affiliation: Department of Life and Nanopharmaceutical Sciences, Kyung Hee University, Hoegi-dong, Dongdaemoon-gu, Seoul, Republic of Korea.

ABSTRACT
Soy (Glycine Max Merr, family Leguminosae) has been reported to possess anti-cancer, anti-lipidemic, estrogen-like, and memory-enhancing effects. We investigated the memory-enhancing effects and the underlying mechanisms of soyasaponin I (soya-I), a major constituent of soy. Impaired learning and memory were induced by injecting ibotenic acid into the entorhinal cortex of adult rat brains. The effects of soya-I were evaluated by measuring behavioral tasks and neuronal regeneration of memory-deficient rats. Oral administration of soya-I exhibited significant memory-enhancing effects in the passive avoidance, Y-maze, and Morris water maze tests. Soya-Ι also increased BrdU incorporation into the dentate gyrus and the number of cell types (GAD67, ChAT, and VGluT1) in the hippocampal region of memory-deficient rats, whereas the number of reactive microglia (OX42) decreased. The mechanism underlying memory improvement was assessed by detecting the differentiation and proliferation of neural precursor cells (NPCs) prepared from the embryonic hippocampus (E16) of timed-pregnant Sprague-Dawley rats using immunocytochemical staining and immunoblotting analysis. Addition of soya-Ι in the cultured NPCs significantly elevated the markers for cell proliferation (Ki-67) and neuronal differentiation (NeuN, TUJ1, and MAP2). Finally, soya-I increased neurite lengthening and the number of neurites during the differentiation of NPCs. Soya-Ι may improve hippocampal learning and memory impairment by promoting proliferation and differentiation of NPCs in the hippocampus through facilitation of neuronal regeneration and minimization of neuro-inflammation.

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Proliferation and maturation of NPCs and neurite outgrowth of MAP2-positive neurons increased by soya-Ι among cultured hippocampal cells.A. Confocal laser scanning microscopic images of Ki67-positive NPCs (green: Ki67-positive cells, red: PI). B. Confocal laser scanning microscopic images of NeuN-positive cells cultured from the rat embryonic hippocampus (green: NeuN-positive cells, red: PI). C. Confocal laser scanning microscopic images of MAP2 and TUJ1-positive NPCs (green: TUJ1-positive cells and MAP2-positive cells, red: PI). D. Average percentages of Ki67-positive cells, E. Average numbers of NeuN-positive cells, F. Average numbers of TUJ1-positive cells, G. Average numbers of MAP2-positive cells compared with PI-positive cells per microscopic field. H. Confocal laser scanning microscopic images of MAP2-positive cells cultured from the rat embryonic hippocampus (green: MAP2-positive cells, red: PI). I. Average neurite length of MAP2-positive cells with neurites longer than double the cell body width. J. The average number of branches per MAP2-positive cell, where the length of the primary branches is two-fold longer than the width of the cell bodies. All assays and counting were carried out as described in Materials and Methods. Over ten positions per cover slip (three times of Ki67, NeuN, TUJ1, and MAP2 immunostaining assay per one trial hippocampal primary culture carried out) were selected and counted. Data represent means ± SEM. (*p <0.05, **p <0.01, ***p <0.001, compared with the vehicle group by the Newman-Keuls Multiple Comparison Test (D,E) or unpaired t test (F, G, I, J)).
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pone-0081556-g006: Proliferation and maturation of NPCs and neurite outgrowth of MAP2-positive neurons increased by soya-Ι among cultured hippocampal cells.A. Confocal laser scanning microscopic images of Ki67-positive NPCs (green: Ki67-positive cells, red: PI). B. Confocal laser scanning microscopic images of NeuN-positive cells cultured from the rat embryonic hippocampus (green: NeuN-positive cells, red: PI). C. Confocal laser scanning microscopic images of MAP2 and TUJ1-positive NPCs (green: TUJ1-positive cells and MAP2-positive cells, red: PI). D. Average percentages of Ki67-positive cells, E. Average numbers of NeuN-positive cells, F. Average numbers of TUJ1-positive cells, G. Average numbers of MAP2-positive cells compared with PI-positive cells per microscopic field. H. Confocal laser scanning microscopic images of MAP2-positive cells cultured from the rat embryonic hippocampus (green: MAP2-positive cells, red: PI). I. Average neurite length of MAP2-positive cells with neurites longer than double the cell body width. J. The average number of branches per MAP2-positive cell, where the length of the primary branches is two-fold longer than the width of the cell bodies. All assays and counting were carried out as described in Materials and Methods. Over ten positions per cover slip (three times of Ki67, NeuN, TUJ1, and MAP2 immunostaining assay per one trial hippocampal primary culture carried out) were selected and counted. Data represent means ± SEM. (*p <0.05, **p <0.01, ***p <0.001, compared with the vehicle group by the Newman-Keuls Multiple Comparison Test (D,E) or unpaired t test (F, G, I, J)).

Mentions: Finally, we investigated whether soya-Ι could directly induce the proliferation or differentiation of NPCs. We conducted immunocytochemical assays in primary NPCs isolated from the embryonic day 16 (E16) rat hippocampus, where pyramidal cell progenitors begin to proliferate. We subcultured the hippocampal cells once to differentiate and immunostain with markers for cell proliferation (Ki67), neuronal differentiation (NeuN, TUJ1), and neurite outgrowth (MAP2). For immunobloting analyses, cell extracts were prepared to detect VGluT1, GAD65/67, and ChAT after 12 days of differentiation. The primary cells in culture were treated with soya-I for the last 1 day for proliferation or 6 days for differentiation. In this analysis, the numbers of cells stained with propidium iodide (PI-positive cells) were usually 80~110 cells in a microscopic field. An increase in Ki67-positive NPCs was observed in all groups treated with soya-Ι (0.5, 1, and 2 μM) for 1 day. Soya-Ι at 2 μM increased the number of proliferating cells (2.5-fold compared with vehicle treatment; Vehicle n = 3, Soya-I 0.5 μM n = 3, Soya-I 1 μM n = 3, Soya-I 2 μM n = 3; F3,8 = 9.426, p = 0.0053 by One-way ANOVA; Figure 6A, D). We also observed that addition of soya-Ι at 0.5 and 1 μM into NPCs cultured from the hippocampus promoted differentiation into late-stage NPCs (TUJ1-positive cells; Vehicle n = 8, Soya-I 0.5 μM n = 8; p < 0.0001 by unpaired t test; Figure 6C, F) and mature neurons (NeuN-positive cells; Vehicle n = 12, Soya-I 0.5 μM n = 12, Soya-I 1 μM n = 14; F2,35 = 14.21, p < 0.0001 by One-way ANOVA; Figure 6B, E) as well as neurite-growing cells (MAP2-positive cells; Vehicle n = 5, Soya-I 0.5 μM n = 5; p = 0.0073 by unpaired t test; Figure 6C, G). In this hippocampal NPC cultures, GFAP-positive astrocytes were hardly detected (data not shown).


Soyasaponin I improved neuroprotection and regeneration in memory deficient model rats.

Hong SW, Heo H, Yang JH, Han M, Kim DH, Kwon YK - PLoS ONE (2013)

Proliferation and maturation of NPCs and neurite outgrowth of MAP2-positive neurons increased by soya-Ι among cultured hippocampal cells.A. Confocal laser scanning microscopic images of Ki67-positive NPCs (green: Ki67-positive cells, red: PI). B. Confocal laser scanning microscopic images of NeuN-positive cells cultured from the rat embryonic hippocampus (green: NeuN-positive cells, red: PI). C. Confocal laser scanning microscopic images of MAP2 and TUJ1-positive NPCs (green: TUJ1-positive cells and MAP2-positive cells, red: PI). D. Average percentages of Ki67-positive cells, E. Average numbers of NeuN-positive cells, F. Average numbers of TUJ1-positive cells, G. Average numbers of MAP2-positive cells compared with PI-positive cells per microscopic field. H. Confocal laser scanning microscopic images of MAP2-positive cells cultured from the rat embryonic hippocampus (green: MAP2-positive cells, red: PI). I. Average neurite length of MAP2-positive cells with neurites longer than double the cell body width. J. The average number of branches per MAP2-positive cell, where the length of the primary branches is two-fold longer than the width of the cell bodies. All assays and counting were carried out as described in Materials and Methods. Over ten positions per cover slip (three times of Ki67, NeuN, TUJ1, and MAP2 immunostaining assay per one trial hippocampal primary culture carried out) were selected and counted. Data represent means ± SEM. (*p <0.05, **p <0.01, ***p <0.001, compared with the vehicle group by the Newman-Keuls Multiple Comparison Test (D,E) or unpaired t test (F, G, I, J)).
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC3852400&req=5

pone-0081556-g006: Proliferation and maturation of NPCs and neurite outgrowth of MAP2-positive neurons increased by soya-Ι among cultured hippocampal cells.A. Confocal laser scanning microscopic images of Ki67-positive NPCs (green: Ki67-positive cells, red: PI). B. Confocal laser scanning microscopic images of NeuN-positive cells cultured from the rat embryonic hippocampus (green: NeuN-positive cells, red: PI). C. Confocal laser scanning microscopic images of MAP2 and TUJ1-positive NPCs (green: TUJ1-positive cells and MAP2-positive cells, red: PI). D. Average percentages of Ki67-positive cells, E. Average numbers of NeuN-positive cells, F. Average numbers of TUJ1-positive cells, G. Average numbers of MAP2-positive cells compared with PI-positive cells per microscopic field. H. Confocal laser scanning microscopic images of MAP2-positive cells cultured from the rat embryonic hippocampus (green: MAP2-positive cells, red: PI). I. Average neurite length of MAP2-positive cells with neurites longer than double the cell body width. J. The average number of branches per MAP2-positive cell, where the length of the primary branches is two-fold longer than the width of the cell bodies. All assays and counting were carried out as described in Materials and Methods. Over ten positions per cover slip (three times of Ki67, NeuN, TUJ1, and MAP2 immunostaining assay per one trial hippocampal primary culture carried out) were selected and counted. Data represent means ± SEM. (*p <0.05, **p <0.01, ***p <0.001, compared with the vehicle group by the Newman-Keuls Multiple Comparison Test (D,E) or unpaired t test (F, G, I, J)).
Mentions: Finally, we investigated whether soya-Ι could directly induce the proliferation or differentiation of NPCs. We conducted immunocytochemical assays in primary NPCs isolated from the embryonic day 16 (E16) rat hippocampus, where pyramidal cell progenitors begin to proliferate. We subcultured the hippocampal cells once to differentiate and immunostain with markers for cell proliferation (Ki67), neuronal differentiation (NeuN, TUJ1), and neurite outgrowth (MAP2). For immunobloting analyses, cell extracts were prepared to detect VGluT1, GAD65/67, and ChAT after 12 days of differentiation. The primary cells in culture were treated with soya-I for the last 1 day for proliferation or 6 days for differentiation. In this analysis, the numbers of cells stained with propidium iodide (PI-positive cells) were usually 80~110 cells in a microscopic field. An increase in Ki67-positive NPCs was observed in all groups treated with soya-Ι (0.5, 1, and 2 μM) for 1 day. Soya-Ι at 2 μM increased the number of proliferating cells (2.5-fold compared with vehicle treatment; Vehicle n = 3, Soya-I 0.5 μM n = 3, Soya-I 1 μM n = 3, Soya-I 2 μM n = 3; F3,8 = 9.426, p = 0.0053 by One-way ANOVA; Figure 6A, D). We also observed that addition of soya-Ι at 0.5 and 1 μM into NPCs cultured from the hippocampus promoted differentiation into late-stage NPCs (TUJ1-positive cells; Vehicle n = 8, Soya-I 0.5 μM n = 8; p < 0.0001 by unpaired t test; Figure 6C, F) and mature neurons (NeuN-positive cells; Vehicle n = 12, Soya-I 0.5 μM n = 12, Soya-I 1 μM n = 14; F2,35 = 14.21, p < 0.0001 by One-way ANOVA; Figure 6B, E) as well as neurite-growing cells (MAP2-positive cells; Vehicle n = 5, Soya-I 0.5 μM n = 5; p = 0.0073 by unpaired t test; Figure 6C, G). In this hippocampal NPC cultures, GFAP-positive astrocytes were hardly detected (data not shown).

Bottom Line: Oral administration of soya-I exhibited significant memory-enhancing effects in the passive avoidance, Y-maze, and Morris water maze tests.Addition of soya-Ι in the cultured NPCs significantly elevated the markers for cell proliferation (Ki-67) and neuronal differentiation (NeuN, TUJ1, and MAP2).Soya-Ι may improve hippocampal learning and memory impairment by promoting proliferation and differentiation of NPCs in the hippocampus through facilitation of neuronal regeneration and minimization of neuro-inflammation.

View Article: PubMed Central - PubMed

Affiliation: Department of Life and Nanopharmaceutical Sciences, Kyung Hee University, Hoegi-dong, Dongdaemoon-gu, Seoul, Republic of Korea.

ABSTRACT
Soy (Glycine Max Merr, family Leguminosae) has been reported to possess anti-cancer, anti-lipidemic, estrogen-like, and memory-enhancing effects. We investigated the memory-enhancing effects and the underlying mechanisms of soyasaponin I (soya-I), a major constituent of soy. Impaired learning and memory were induced by injecting ibotenic acid into the entorhinal cortex of adult rat brains. The effects of soya-I were evaluated by measuring behavioral tasks and neuronal regeneration of memory-deficient rats. Oral administration of soya-I exhibited significant memory-enhancing effects in the passive avoidance, Y-maze, and Morris water maze tests. Soya-Ι also increased BrdU incorporation into the dentate gyrus and the number of cell types (GAD67, ChAT, and VGluT1) in the hippocampal region of memory-deficient rats, whereas the number of reactive microglia (OX42) decreased. The mechanism underlying memory improvement was assessed by detecting the differentiation and proliferation of neural precursor cells (NPCs) prepared from the embryonic hippocampus (E16) of timed-pregnant Sprague-Dawley rats using immunocytochemical staining and immunoblotting analysis. Addition of soya-Ι in the cultured NPCs significantly elevated the markers for cell proliferation (Ki-67) and neuronal differentiation (NeuN, TUJ1, and MAP2). Finally, soya-I increased neurite lengthening and the number of neurites during the differentiation of NPCs. Soya-Ι may improve hippocampal learning and memory impairment by promoting proliferation and differentiation of NPCs in the hippocampus through facilitation of neuronal regeneration and minimization of neuro-inflammation.

Show MeSH
Related in: MedlinePlus