Article pubs.acs.org/JAFC
Soyasapogenol B and Genistein Attenuate LipopolysaccharideInduced Memory Impairment in Mice by the Modulation of NF-κBMediated BDNF Expression Hae-Ji Lee,† Su-Min Lim,† Da-Bin Ko,† Jin-Ju Jeong,† Yun-Ha Hwang,‡ and Dong-Hyun Kim*,† †
Department of Life and Nanopharmaceutical Sciences, Kyung Hee University, 26, Kyungheedae-ro, Dongdaemun-gu, Seoul 02447, Korea ‡ DongWha Pharm Research Institute, 35-71, Topsil-ro, Giheung-gu, Yongin-Shi, Gyeonggi 446-902 Korea S Supporting Information *
ABSTRACT: Lactobacillus plantarum C29-fermented defatted soybean (FDS), which contains soyasaponins such as soyasaponin I (SI) and soyasapogenol B (SB) and isoflavones such as genistin (GE) and genistein (GT), attenuated memory impairment in mice. Moreover, in the preliminary study, FDS and its soyasaponins and isoflavones significantly inhibited NF-κB activation in LPS-stimulated microglial BV2 cells. Therefore, we examined the effects of FDS and its constituents SI, SB, GT, and GE on LPS-induced memory impairment in mice. Oral administration of FDS (80 mg/kg), which has higher concentrations of SB and GE than DS, recovered LPS-impaired cognitive function in Y-maze (55.1 ± 3.5%) and passive avoidance tasks (50.9 ± 19.2 s) to 129.2% (74.1 ± 3.5%) and 114.2% (290.0 ± 22.4 s) of normal mice, respectively (P < 0.05). SB and GE (10 μM) also more potently attenuated LPS-impaired cognitive behavior than SI and GT, respectively. SB (10 mg/kg) was the most effective: treatment recovered LPS-impaired spontaneous alternation and latency time to 105.7% and 126.8% of normal control mice, respectively (P < 0.05). SB and GE significantly increased BDNF expression and CREB phosphorylation in LPS-treated mice and corticosterone-stimulated SH-SY5Y cells. Furthermore, SB and GE (10 μM) also significantly inhibited NF-κB activation in LPStreated mice. These findings suggested that FDS and its constituent soyasaponins and isoflavones may attenuate memory impairment by the regulation of NF-κB-mediated BDNF expression. KEYWORDS: soybean, soyasapogenol B, genstein, memory impairment, Lactobacillus plantarum
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INTRODUCTION Dementia, including Alzheimer’s disease (AD), is a well-known neurodegenerative disorder characterized by the progressive deterioration of memory and cognitive function.1,2 The pathogenesis of AD includes neurofibrillary tangle formation, inflammatory processes, and neurotransmitter disturbances in the central nervous system, which result in the impairment of the cortical and hippocampal cholinergic and immune systems.3,4 Cholinesterase inhibitors such as rivastigmine and cholinergic agonists such as physostigmine are often used in the treatment of AD.3,5,6 Recently, the bidirectional networks between the brain and gut microbiota were reported to be maintained through the endocrine, neural, and immune pathways.7,8 The overexpression of gut microbiota lipopolysaccharide (LPS) disturbs gastrointestinal immune responses, which may result in fluctuations in the secretion of the neurotransmitters, such as serotonin and catecholamines; subsequently, systemic inflammatory diseases, such as ulcerative colitis, obesity, and AD, may result.9,10 LPS released from Bacteroides f ragilis, which is abundant in the gut, might represent a major contributory factor to systemic inflammation and diseases, such as Alzheimer’s disease.11 Additionally, Czerniawski and Guzowski reported that the peripheral injection of LPS resulted in hippocampus-dependent impairment of context discrimination memory.12 Hasegawa-Ishii et al. reported that a single peritoneal injection of LPS activated hippocampal astrocytes © XXXX American Chemical Society
through interaction between the cells of the brain-immune interface and cytokine signals; furthermore, after repeated injection, LPS activated microglia.13 Buttini et al. reported that a peritoneal injection of LPS activated microglia, produced proinflammatory cytokines.14 Goel et al. reported that LPS impaired memory by modulating NF-κB-mediated brainderived neurotrophic factor (BDNF) expression and cAMP response element binding protein (CREB) phosphorylation.15 These reports suggested that chronic endotoxemia may impair memory by the activation of microglia in the hypothalamus. Moreover, antioxidant and anti-inflammatory compounds, such as lipoic acid and resveratrol, have been reported to protect against memory impairment.16,17 These results suggest that inhibiting LPS-induced inflammation may be beneficial for the therapy of memory loss, including AD. Soybean (Glycine max, family Leguminosae), which constitutes protein, dietary fiber, and isoflavone and soyasaponin glycosides as main constituents, exhibits memory enhancing,18 antilipidemic,19 and phytoestrogenic effects.20 Its phytochemicals isoflavones and soyasaponins exhibit phytoestrogenic,20 anti-inflammatory,21 and memory-enhancing effects.22−24 When these constituents were orally administered, the Received: June 2, 2017 Revised: July 22, 2017 Accepted: July 26, 2017
A
DOI: 10.1021/acs.jafc.7b02569 J. Agric. Food Chem. XXXX, XXX, XXX−XXX
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Figure 1. Structures of soysaponin I, soyasapogenol B, genistin, and genistein. freeze-dried powder supplemented with C29 (1.25 × 1010 CFU/g) (DW2009) was used in the present study. Culture of C29. Lactobacillus plantarum C29 was isolated in the previous study. C29 (KCCM10885, Korea Culture Center of Microorganisms, Seoul, Korea) isolated from kimchi. C29 was cultured as previously reported.33 Briefly, C29 was grown in MRS broth (2 L) for 10 h (an optical density at 60 nm, 1−2), centrifuged at 10 000 × g for 20 min, and washed with saline. The collected cells were freezedried. These were suspended in 1% glucose (for in vivo) or PBS (for in vitro). Culture of BV-2 and SH-SY5Y Cells. BV-2 and SH-SY5Y cells were purchased from Korea Cell Line Bank (Seoul, Korea) and cultured at 37 °C in a 5% CO2-95% air atmosphere in DMEM containing 5% fetal bovine serum and 1% antibiotic-antimycotic. For the measurement of NF-κB activation, BV-2 cells were treated with LPS (100 ng/mL) in the absence or presence of test agents for 90 min (for NF-κB activation) For the measurement of BDNF expression, SH-SY5Y cells were treated with corticosterone (300 μM) in the presence or absence of test agents for 24 h according to the method of Zhang et al.34 The protein levels were measured by immunoblotting. Animals. Male ICR mice (weight, 25−28 g; age, 6-week old) were purchased from the Samtaco Animal Inc. (Seoul, Republic of Korea), fed with water and food ad libitum, and maintained in a ventilated room (temperature, 22 °C ± 1 °C; humidity, 50% ± 10% humidity); and a 12-h diurnal light cycle, 07:00−19:00) for 1 week before the animal experiment. All experiments were approved by the Committee for the Care and Use of Laboratory Animals in the Kyung Hee University (KHUASP(SE)-16-025) and were carried out according to the Kyung Hee University Guidelines for Laboratory Animals Care and Use. In the preliminary study, mice were intraperitoneally administered LPS solution (0, 0.8, 8, or 80 μg/kg, dissolved in saline) once a day for 10 days to cause LPS-induced memory impairment, according to the method of Buttini et al.,14 with slight modifications (Supporting Information, SI, Figure S1). Memory behaviors were measured 24 h after the final administration of LPS by using the Y-maze and passive avoidance tasks. The treatment doses of 8 or 80 μg/kg resulted in memory-impaired behaviors in the Y-maze and passive avoidance tasks. Therefore, we treated LPS at a dose of 8 μg/kg to prepare LPSinduced memory impairment. Additionally, when the mice were orally administered with SB and GE (1, 10, and 20 mg/kg) in order to decide the effective dose of SI and GE against memory impairment, these were effective at more than 10 mg/kg. Therefore, to evaluate the ameliorative effect of SI, SB, GT, and GE against LPS-induced
isoflavone glycosides genistin (GT) and daidzin were similarly metabolized to genistein (GE), daidzein, and equols by gut microbiota,25 and soyasaponins Ab and I (SI) were to their aglycones soyasapogenols A and B (SB), respectively.26,27 Moreover, these metabolites and their sulfate and glucuronate conjugates were detected in the blood.28,29 These transformations of isoflavone and soyasaponin glycosides to their aglycones also occurred through fermentation with Lactobacillus plantarum C29 (formerly Lactobacillus pentosus var. plantarum).30 Of these metabolites, GE exhibited the more potent anti-inflammatory effects than its glycosides GT both in vitro and in vivo.31,32 SB also exhibited a more potent phytoestrogenic effect than its glycoside SI.26 Nevertheless, the amelioration of memory impairment by these phytochemicals has not been studied thoroughly. In the preliminary study to evaluate the effect of C29fermented defatted soybean powder (FDS), which contained a higher content of GE and SB than defatted soybean powder (DS), against LPS-induced memory impairment in vivo, we found that FDS inhibited NF-κB activation in LPS-stimulated microglial BV-2 cells more potently than DS. Therefore, we examined the effects of FDS and its phytochemicals (SI, GT, and their aglycones SB and GE) (Figure 1) on LPS-induced memory impairment in mice.
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MATERIALS AND METHODS
Chemicals. Corticosterone, LPS purified from Escherichia coli O111:B4, Dulbecco’s modified Eagle’s medium (DMEM), fetal bovine serum (FBS), gentamicin solution, GE, GT, SI, and SB were purchased from Sigma-Aldrich (St.Louis, MO, U.S.A.). Antibodies for brainderived neurotrophic factor (BDNF), cAMP response element binding protein (CREB), p-CREB, p65, p-p65, and β-actin were purchased from Cell Signaling Technology (Beverly, MA, U.S.A.). Enzyme-linked immunosorbent assay (ELISA) kits were purchased from Ebioscience (San Diego, CA, U.S.A.). A Luminata Forte Western HRP Substrate was purchased from Millipore Corporation (Billerica, MA, U.S.A.). Other chemicals used were of the highest grade available. Preparation of FDS. FDS was prepared according to the modified method of Yu et al. Briefly, DS (6%) was inoculated with C29 (1 × 108 CFU/mL) and incubated at 37 °C for 24 h and freeze-dried. The B
DOI: 10.1021/acs.jafc.7b02569 J. Agric. Food Chem. XXXX, XXX, XXX−XXX
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Figure 2. Effect of DS, FDS, and their phytochemicals on TLR4-linked NF-κB activation in LPS-stimulated BV-2 cells. (A) Effects of FDS, DS, and C29. (B) Effects of SI and SB. (C) Effects of GT and GE. Cells (0.5 × 106) were incubated with LPS in the absence or presence of test agents (10 or 50 μg/mL [for FDS or DS], 5 or 10 μM [for SI, DB, GT, and GE], or 103 or 105 CFU/mL [for C29]) for 90 min. Proteins were measured by immunoblotting. FDS, C29-fermneted defatted soybean extract; SI, soyasaponin I; SB, soyasapogenol B; GT, genistin; and GE, genistein. memory impairment, we orally administered test agents (at doses of 5 or 10 mg/kg) with LPS once a day for 3 days from 24 h after the seventh intraperitoneal injection of LPS (8 μg/kg) (Figure S1). LPS was intraperitoneally treated 2 h after the oral administration of test agents. The normal group received saline instead of LPS and the test agents. Memory behaviors were measured at 18 h after the final administration of the test agents in the Y-maze and passive avoidance tasks. After a further 2 h, mice were killed, and the hippocampus and colon were removed. The specimens were stored at −80 °C until use in the ELISA assays and immunoblotting. Measurement of Memory Behaviors in Mice. The Y-maze task was performed in mice with LPS-induced memory impairment according to the method of Jeong et al.33 Briefly, the Y-maze consisted of black-colored acryl and positioned at equal angles. Mice were placed at the end of the arm and allowed to move freely through the maze. When the hind paws of mice were completely placed in the arm, arm entry sessions were recorded. Sequential entry into three arms in alternative order was decided as the successive entries on overlapping triplet sets. The alternation percentage was calculated as the ratio (%) of actual to possible alternations (counted as the total number of arm entries minus 2).
The passive avoidance task was performed in mice with LPSinduced memory impairment according to the method of Jeong et al.33 Acquisition and retention was performed using a two-compartment acrylic box consisted of a lighted compartment and a dark compartment), which is connected by an entrance door. Memory impairment was induced by the intraperitoneal injection of LPS. Test agents and vehicle were orally administered into mice once a day for 3 days. The passive avoidance maze task was performed 20 h after the final administration of test agents. ELISA and Immunoblotting. The hippocampus was removed 2 h after the passive avoidance task and homogenized with RIPA lysis buffer containing 50 mM Tris−HCl (pH 8.0), 1.0% Igepal CA-630 (NP-40), 150 mM sodium chloride, 0.1% sodium dodecyl sulfate (SDS), 0.5% sodium deoxycholate, 1% protease inhibitor cocktail, and a phosphatase inhibitor cocktail on the ice. The lysate was centrifuged at 10 000 × g for 10 min (4 °C) and the supernatant were electrophoresed by SDS-polyacrylamide gel electrophoresis and then transferred to a nylon membrane. Proteins were detected using the antibodies, as previously reported.33 Immunodetection was carried out using a Luminata Forte Western HRP Substrate. C
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Figure 3. Effects of FDS and C29 on LPS-induced memory impairment and hippocampal NF-kB activation and TNF-α expression in mice. (A) Effects on memory impairment in the Y-maze task. (B) Effects on memory impairment in the passive avoidance task. (C) Effects on NF-κB activation and BDNF expression. (D) Effects on TNF-α expression. Memory impairment was induced by the intraperitoneal injection of LPS (8 μg/ kg/day) for 10 days. NOR, vehicle alone; LP, LPS alone; LT, 10 mg/kg of tacrine (Tac) with LPS; LF, 80 mg/kg of FDS with LPS; LC, 1 × 109 CFU/mouse of C29 with LPS. NF-κB, BDNF, and β-actin were measured by immunoblotting. All values are expressed as mean ± SD (n = 8). # Significantly different from vehicle-treated mice (p < 0.05). *Significantly different from control treated with LPS alone (p < 0.05). Statistical Analysis. Experimental results are indicated as means ± standard deviation (SD), and were statistically analyzed using one-way ANOVA and Turkey’s multiple comparison tests (P < 0.05).
(Supplement Figure S2). These chemicals did not exhibit cytotoxic effects for 48 h in our experimental conditions (data not shown). Ameliorative Effect of FDS against Memory Impairment in Mice. Next, we examined the effects of FDS, a C29fermented DS, on LPS-induced memory impairment in mice. Treatment with LPS significantly reduced spontaneous alterations in the Y-maze task (Figure 3A). Treatment with FDS (80 mg/kg) and C29 (1 × 109 CFU/mouse) recovered the LPS-induced reduction of spontaneous alterations (55.1 ± 3.5%) to 129.2% (74.1 ± 3.5%) and 96.7% (63.4 ± 7.4%) of normal control mice, respectively. The effect of FDS was more effective than that of C29. However, these compounds did not affect spontaneous alterations in mice that were not treated with LPS (Figure S3). Treatment with LPS also significantly impaired the cognitive function in the passive avoidance task: treatment decreased the latency time (Figure 3B). However, treatment with FDS recovered the LPS-induced memory impairment (50.9 ± 19.2 s) to 114.2% (290.0 ± 22.4 s) of normal control mice. FDS attenuated LPS-induced memory impairment more potently than C29 alone. During the acquisition trial, no differences were observed in latency time among the test groups.
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RESULTS Effect of DS, FDS, and Their Constituents on LPSInduced NF-κB Activation in BV-2 Cells. LPS causes memory impairment by the activation of the NF-κB signaling pathway.11,14 FDS and its constituent SI improved scopolamine-induced memory impairment in mice.18 To understand the ameliorative effect of FDS and its constituents on LPSinduced memory impairment, we first investigated whether FDS, DS, and their ingredients C29, SI, GT, SI, and SB could inhibit the NF-κB signaling pathway in LPS-stimulated microglial BV-2 cells (Figure 2). Treatment with LPS significantly increased NF-κB activation, whereas FDS, DS, and their ingredients inhibited LPS-induced activation of NFκB, as well as the phosphorylation of NF-κB signaling molecules TAK1 and IκBα. SB showed the most potent inhibition of LPS-induced NF-κB activation, followed by GE. Moreover, the inhibitory effect of FDS was stronger than that of DS. However, these compounds did not affect NF-κB activation in BV2 cells treated that were not treated with LPS D
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Figure 4. Effects of soyasaponin I (SI) and soyasapogenol B (SB) on LPS-induced memory impairment and hippocampal NF-κB activation and TNF-α expression in mice. (A) Effects on memory impairment in the Y-maze task. (B) Effects on memory impairment in the passive avoidance task. (C) Effects on NF-κB activation. (D) Effects on TNF-α expression. Memory impairment was induced by the intraperitoneal injection of LPS (8 μg/ kg/day) for 10 days. NOR, vehicle alone; LP, LPS alone; LT, 10 mg/kg of tacrine (Tac) with LPS; LSI, 10 mg/kg of SI with LPS; LSL, 5 mg/kg of SB with LPS; and LSH, 10 mg/kg of SB with LPS. NF-κB, BDNF and β-actin were measured by immunoblotting. All values are expressed as mean ± SD (n = 8). #Significantly different from vehicle-treated mice (p < 0.05). *Significantly different from control treated with LPS alone (p < 0.05).
of normal control mice in the passive avoidance task, respectively. GE more potently but not significantly attenuated the cognitive function than GT. Furthermore, GE increased LPS-suppressed BDNF expression and inhibited LPS-induced NF-κB activation in the hippocampi of mice. Effects of FDS Phytochemicals on CorticosteroneImpaired BDNF Expression in SH-SY5Y Cells. To determine the components of FDS that were responsible for the induction of BDNF expression in vivo, we examined the effects of FDS, C29, SI, SB, GT, and GE on corticosteronesuppressed BDNF expression in SH-SY5Y cells (Figure 6). Treatment with corticosterone, which induced neuronal atrophy in the hippocampus because of stress,34 suppressed BDNF expression and ERK phosphorylation, whereas LPS treatment did not affect BDNF expression (data not shown). Treatment with FDS, its constituents, or C29 increased corticosterone-suppressed BDNF expression by corticosterone. SB and GE more potently increased the BDNF expression and ERK phosphorylation than their respective glycosides SI and GT. Of these phytochemicals, SB most potently increased BDNF expression. However, these compounds did not affect BDNF expression in SH-SY5Y cells stimulated without corticosterone (Figure S4). However, no cytotoxic effects of these chemicals were observed for 48 h of treatment in our experimental conditions (data not shown).
Furthermore, treatment with FDS or C29 inhibited LPSinduced NF-κB activation and TNF-α expression, and increased LPS-suppressed BDNF expression in the hippocampus (Figure 3C). Effects of SB, SI, GE, and GT on Memory Impairment in Mice. To understand the amelioration of memory impairment by FDS, we first examined the effects of SI and its aglycone SB on LPS-induced memory impairment in mice (Figure 4). Treatment with SI and SB (10 mg/kg) recovered the LPS-induced reduction of spontaneous alterations to 93.0% and 105.7% of normal control mice in the Y-maze, respectively. SB increased LPS-impaired spontaneous alterations more efficiently than SI. However, these did not affect spontaneous alterations in mice that were not treated with LPS (Figure S2). SI and SB also increased LPS-reduced latency time in the passive avoidance task. SB recovered LPS-reduced latency times to 126.8% of normal control mice. During the acquisition trial, no differences were observed in latency times among the test groups. Furthermore, SB increased LPS-suppressed BDNF expression and inhibited NF-κB activation and TNF-α expression in the mouse hippocampus. Next, we examined the effects of GT and GE on LPSinduced memory impairment in mice (Figure 5). Treatment with GT or GE recovered the LPS-induced reduction of spontaneous alterations to 96.8% and 101.0% of normal control mice in the Y-maze task and latency time to 98.8% and 116.0% E
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Figure 5. Effects of genistin (GT) and genistein (GE) on LPS-induced memory impairment and hippocampal NF-κB activation and TNF-α expression in mice. (A) Effects on memory impairment in the Y-maze task. (B) Effects on memory impairment in the passive avoidance task. (C) Effects on NF-kB activation. (D) Effects on TNF-α expression. Memory impairment was induced by the intraperitoneal injection of LPS (8 μg/kg/ day) for 10 days. NOR, vehicle alone; LP, LPS alone; LT, 10 mg/kg of tacrine (Tac) with LPS; LG, 10 mg/kg of GT with LPS; LGE, 10 mg/kg of GE with LPS. NF-κB, BDNF and β-actin were measured by immunoblotting. All values are expressed as mean ± SD (n = 8). #Significantly different from vehicle-treated mice (p < 0.05). *Significantly different from control treated with LPS alone (p < 0.05).
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DISCUSSION Isoflavone and soyasaponin glycosides, which are the main phytochemicals in soybean, have anti-inflammatory, estrogenic, anticancer, and antiosteoporotic effects.17−22 Soybean saponin and isoflavone also attenuated the scopolamine-impaired cognitive function in mice.18,35 FDS, a C29-fermented DS, showed more potent amelioration of the memory impairment than that of nonfermented soybean. Treatment with C29 alone mitigate scopolamine-induced memory impairment and decreased the number of Enterobacteriaceae in the gut microbiota of mice but increased the population of lactic acid bacteria.33 LPS, which is produced from gut microbiota including Enterobacteriaceae,36 caused topical and systemic inflammation, which resulted in memory loss by the regulation of NF-κB-mediated BDNF/CREB expression.15,37 These results suggested that C29 and FDS may attenuate memory impairment through the regulation of gut microbiota composition. In the present study, FDS inhibited LPS-induced NF-κB activation in BV-2 cells, increased corticosterone-suppressed BDNF expression in SH-SY5Y cells more potently than DS or C29, and attenuated LPS-induced memory loss in mice. Therefore, to understand which of the constituents in FDS
could ameliorate memory impairment, we here examined the effects of GT, GE, SI, and SB on LPS-impaired cognitive function in mice. These constituents attenuated LPS-induced memory impairment. Among these, SB and GE most potently attenuated LPS-impaired cognitive function. Additionally, Kamo et al. reported that, when the mixture of SI and its aglycone SB was orally administered, SI was not absorbed into the blood but SB could not be detectable.29 Yuan et al. reported that, when GT and GE were orally administered, GE and its sulfate and glucuronate conjugates were detected in the blood and total levels was higher in GT-treated women than in GEtreated ones.25 Thus, orally administered SI and GT may be absorbed into the blood via the transformation into their aglycones, respectively, as phytochemicals previously reported.38 However, orally administered SB and GE may be absorbed without the metabolic modification and then metabolized into glucuronate and sulfate conjugates in the tissues including the liver. Therefore, when these constituents are orally administered, aglycones and their sulfate and glucuronate conjugates are detected in the blood. Nevertheless, we found that when SI, SB, GT, or GE were orally treated in mice with LPS-induced memory impairment, treatment with SB and GE recovered LPS-induced memory impairment more F
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Figure 6. Effect of DS, FDS, and their phytochemicals on TLR4-linked NF-κB activation in corticosterone (CCS)-stimulated SH-SY5Y cells. Cells (0.5 × 106) were incubated with corticosterone (300 μM) in the absence or presence of test agents (10 or 50 μg/mL [for FDS or DS], 5 or 10 μM [for SI, DB, GT, and GE], or 103 or 105 CFU/mL [for C29]) for 90 min. Proteins were measured by immunoblotting. FDS, C29-fermneted defatted soybean extract; SI, soyasaponin I; SB, soyasapogenol B; GT, genistin; and GE, genistein.
hippocampal BDNF concentration is lower in AD patients than in healthy persons,42,43 which supports the importance of BDNF in memory and learning. CREB is also important in neuronal plasticity, as well as spatial and long-term memory formation in the brain through ERK phosphorylation.44 CREB phosphorylation is down-regulated in patients with AD.45 Therefore, the regulation of BDNF expression and CREB activation is considered beneficial for the therapy of dementia, particularly AD.15,43−45 Short- or long-term treatment with LPS in mice induced memory impairment, as the previously reported.13,14,46,47 Treatment with LPS for 5 days also impaired the cognitive function in mice, whereas treatment with FDS for 5 days from 24 h after LPS treatment attenuated memory impairment (data not shown). Scopolamine impaired cognitive function and increased TNF-α expression in rodents.48 Treatment with FDS, soyasaponin Ab, SI, or GE attenuated scopolamine-induced memory impairment in experimental animals by an increase in BDNF expression.18,48,49 Moreover, we found that SB potently alleviated LPS-induced memory impairment. These findings suggest that FDS and its constituents, particularly SB and GE, may attenuate memory impairment by the regulation of NF-κB-mediated BDNF expression.
potently than treatment with SI and GT, respectively. We also found that SB and GE more potently increased BDNF expression and CREB phosphorylation in the hippocampus of mice with LPS-induced memory impairment than their glycosides, respectively. SB and GE also increased corticosteroid-suppressed BDNF expression in SH-SY5Y cells. Furthermore, SB and GE more potently inhibited LPS-induced NF-κB activation in the hippocampus of mice and inhibited the LPS-stimulated NF-κB activation and TNF-α expression in microglial BV-2 cells. These results suggest that SB and GE may attenuate memory impairment by inhibiting NF-κB activation and increasing BDNF expression. Nevertheless, we could not clarify the active metabolite(s). LPS, a stressor produced by gut microbiota, impairs memory and induces inflammatory responses NF-κB activation, TNF-α expression, and corticosteroid levels in vivo.11,39 Furthermore, we found that BDNF expression was suppressed by the stimulation of LPS-induced NF-κB activation in mice, like the previously described.15 BDNF influences the neuronal synaptic plasticity of the central nervous system and the peripheral nervous system by depolarizing neurons,40 enhancing glutamatergic synaptic transmission,40,41 increasing the phosphorylation of the N-methyl-D-aspartate receptor subunit, and facilitating hippocampal long-term potentiation.41 Therefore, BDNF is vital to memory, learning, and thinking. Moreover, the G
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(8) Moloney, R. D.; Desbonnet, L.; Clarke, G.; Dinan, T. G.; Cryan, J. F. The microbiome: stress.; health and disease. Mamm. Genome 2014, 25, 49−74. (9) Thomas, L. V.; Suzuki, K.; Zhao, J. Probiotics: a proactive approach to health. A symposium report. Br. J. Nutr. 2015, 114 (S1), S1−S15. (10) Cani, P. D.; Everard, A. Talking microbes: When gut bacteria interact with diet and host organs. Mol. Nutr. Food Res. 2016, 60, 58− 66. (11) Lukiw, W. J. Bacteroides f ragilis Lipopolysaccharide and inflammatory signaling in Alzheimer’s disease. Front. Microbiol. 2016, 7, 1544. (12) Czerniawski, J.; Guzowski, J. F. Acute neuroinflammation impairs context discrimination memory and disrupts pattern separation processes in hippocampus. J. Neurosci. 2014, 34, 12470− 12480. (13) Hasegawa-Ishii, S.; Inaba, M.; Umegaki, H.; Unno, K.; Wakabayashi, K.; Shimada, A. Endotoxemia-induced cytokine-mediated responses of hippocampal astrocytes transmitted by cells of the brain-immune interface. Sci. Rep. 2016, 6, 25457. (14) Buttini, M.; Limonta, S.; Boddeke, H. W. Peripheral administration of lipopolysaccharide induces activation of microglial cells in rat brain. Neurochem. Int. 1996, 29, 25−35. (15) Goel, R.; Bhat, S. A.; Hanif, K.; Nath, C.; Shukla, R. Angiotensin II Receptor Blockers Attenuate Lipopolysaccharide-Induced Memory Impairment by Modulation of NF-κB-Mediated BDNF/CREB Expression and Apoptosis in Spontaneously Hypertensive Rats. Mol. Neurobiol. 2017, DOI: 10.1007/s12035-017-0450-5. (16) Holmquist, L.; Stuchbury, G.; Berbaum, K.; Muscat, S.; Young, S.; Hager, K.; Engel, J.; Münch, G. Lipoic acid as a novel treatment for Alzheimer’s disease and related dementias. Pharmacol. Ther. 2007, 113, 154−164. (17) Gocmez, S. S.; Gacar, N.; Utkan, T.; Gacar, G.; Scarpace, P. J.; Tumer, N. Protective effects of resveratrol on aging-induced cognitive impairment in rats. Neurobiol. Learn. Mem. 2016, 131, 131−136. (18) File, S. E.; Jarrett, N.; Fluck, E.; Duffy, R.; Casey, K.; Wiseman, H. Eating soya improves human memory. Psychopharmacology (Berl) 2001, 157, 430−436. (19) Murphy, P. A.; Hu, J.; Barua, K.; Hauck, C. C. Group B saponins in soy products in the U.S. Department of Agriculture–Iowa State University isoflavone database and their comparison with isoflavone contents. J. Agric. Food Chem. 2008, 56, 8534−8540. (20) Pan, M.; Li, Z.; Yeung, V.; Xu, R. J. Dietary supplementation of soy germ phytoestrogens or estradiol improves spatial memory performance and increases gene expression of BDNF, TrkB receptor and synaptic factors in ovariectomized rats. Nutr. Metab. 2010, 7, 75. (21) Lee, I. A.; Park, Y. J.; Yeo, H. K.; Han, M. J.; Kim, D. H. Soyasaponin I attenuates TNBS-Induced colitis in mice by inhibiting NF-κB pathway. J. Agric. Food Chem. 2010, 58, 10929−10934. (22) Ding, J.; Xi, Y. D.; Zhang, D. D.; Zhao, X.; Liu, J. M.; Li, C. Q.; Han, J.; Xiao, R. Soybean isoflavone ameliorates β-amyloid 1−42induced learning and memory deficit in rats by protecting synaptic structure and function. Synapse 2013, 67, 856−864. (23) Menze, E. T.; Esmat, A.; Tadros, M. G.; Abdel-Naim, A. B.; Khalifa, A. E. Genistein improves 3-NPA-induced memory impairment in ovariectomized rats: impact of its antioxidant, anti-inflammatory and acetylcholinesterase modulatory properties. PLoS One 2015, 10, e0117223. (24) Hong, S. W.; Yoo, D. H.; Woo, J. Y.; Jeong, J. J.; Yang, J. H.; Kim, D. H. Soyasaponins Ab and Bb prevent scopolamine-induced memory impairment in mice without the inhibition of acetylcholinesterase. J. Agric. Food Chem. 2014, 62, 2062−2068. (25) Yuan, B.; Zhen, H.; Jin, Y.; Xu, L.; Jiang, X.; Sun, S.; Li, C.; Xu, H. Absorption and plasma disposition of genistin differ from those of genistein in healthy women. J. Agric. Food Chem. 2012, 60, 1428− 1436. (26) Chang, S.-Y.; Han, M.-J.; Han, S.-J.; Kim, D.-H. Metabolism of soyasaponin I by human intestinal microflora and its estrogenic and cytotoxic effects. Biomol. Ther. 2009, 17, 430−437.
ASSOCIATED CONTENT
S Supporting Information *
The Supporting Information is available free of charge on the ACS Publications website at DOI: 10.1021/acs.jafc.7b02569. Figure S1. Experimental design. Figure S2. Effect of DS, FDS, and their phytochemicals on TLR4-linked NF-κB activation in BV-2 cells. Figure S3. Effects of FDS, C29, SI, SB, GT, and GE on learning and memory and hippocampal NF-κB activation in mice. Figure S4. Effect of DS, FDS, and their phytochemicals on TLR4-linked NF-κB activation in SH-SY5Y cells. (PDF)
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AUTHOR INFORMATION
Corresponding Author
*Tel: +82-2-961-0374. Fax: +82-2-957-5030. E-mail: dhkim@ khu.ac.kr (D.-H.K.). ORCID
Dong-Hyun Kim: 0000-0003-4783-7900 Notes
The authors declare no competing financial interest.
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ACKNOWLEDGMENTS This research was supported by the Senior-friendly Product R&D program through the Korea Health Industry Development Institute (KHIDI), funded by the Ministry of Health & Welfare, Republic of Korea (grant number: HIC0997).
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ABBREVIATIONS USED AD, Alzheimer’s disease; BDNF, brain-derived neurotrophic factor; CREB, cAMP response element binding protein; DMEM, Dulbecco’s modified Eagle’s medium; FBS, fetal bovine serum; DS, defatted soybean powder; FDS, C29fermented defatted soybean powder; GE, genistein; GT, genistin; IL, interleukin; LPS, lipopolysaccharide; NF-κB, nuclear factor-κB; SD, standard deviation; SB, soyasapogenol B; SI, soyasaponin I; TNF, tumor necrosis factor
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REFERENCES
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