Transcriptomic and metabolomic changes might predict frailty in SAMP8 mice.

Frailty is a geriatric, multi-dimensional syndrome that reflects multisystem physiological change and is a transversal measure of reduced resilience to negative events. It is characterized by weakness, frequent falls, cognitive decline, increased hospitalization and dead and represents a risk factor for the development of Alzheimer's disease (AD). The fact that frailty is recognized as a reversible condition encourages the identification of earlier biomarkers to timely predict and prevent its occurrence. SAMP8 (Senescence-Accelerated Mouse Prone-8) mice represent the most appropriate preclinical model to this aim and were used in this study to carry transcriptional and metabolic analyses in the brain and plasma, respectively, upon a characterization at cognitive, motor, structural, and neuropathological level at 2.5, 6, and 9 months of age. At 2.5 months, SAMP8 mice started displaying memory deficits, muscle weakness, and motor impairment. Functional alterations were associated with a neurodevelopmental deficiency associated with reduced neuronal density and glial cell loss. Through transcriptomics, we identified specific genetic signatures well distinguishing SAMP8 mice at 6 months, whereas plasma metabolomics allowed to segregate SAMP8 mice from SAMR1 already at 2.5 months of age by detecting constitutively lower levels of acylcarnitines and lipids in SAMP8 at all ages investigated correlating with functional deficits and neuropathological signs. Our findings suggest that specific genetic alterations at central level, as well as metabolomic changes in plasma, might allow to early assess a frail condition leading to dementia development, which paves the foundation for future investigation in a clinical setting.

Exogenous Nucleotides Improve the Skin Aging of SAMP8 Mice by Modulating Autophagy through MAPKs and AMPK Pathways.

The skin, serving as the body's primary defense against external elements, plays a crucial role in protecting the body from infections and injuries, as well as maintaining overall homeostasis. Skin aging, a common manifestation of the aging process, involves the gradual deterioration of its normal structure and repair mechanisms. Addressing the issue of skin aging is increasingly imperative. Multiple pieces of evidence indicate the potential anti-aging effects of exogenous nucleotides (NTs) through their ability to inhibit oxidative stress and inflammation. This study aims to investigate whether exogenous NTs can slow down skin aging and elucidate the underlying mechanisms. To achieve this objective, senescence-accelerated mouse prone-8 (SAMP8) mice were utilized and randomly allocated into Aging, NTs-low, NTs-middle, and NTs-high groups, while senescence-accelerated mouse resistant 1 (SAMR1) mice were employed as the control group. After 9 months of NT intervention, dorsal skin samples were collected to analyze the pathology and assess the presence and expression of substances related to the aging process. The findings indicated that a high-dose NT treatment led to a significant increase in the thickness of the epithelium and dermal layers, as well as Hyp content (p < 0.05). Additionally, it was observed that low-dose NT intervention resulted in improved aging, as evidenced by a significant decrease in p16 expression (p < 0.05). Importantly, the administration of high doses of NTs could improve, in some ways, mitochondrial function, which is known to reduce oxidative stress and promote ATP and NAD+ production significantly. These observed effects may be linked to NT-induced autophagy, as evidenced by the decreased expression of p62 and increased expression of LC3BI/II in the intervention groups. Furthermore, NTs were found to upregulate pAMPK and PGC-1α expression while inhibiting the phosphorylation of p38MAPK, JNK, and ERK, suggesting that autophagy may be regulated through the AMPK and MAPK pathways. Therefore, the potential induction of autophagy by NTs may offer benefits in addressing skin aging through the activation of the AMPK pathway and the inhibition of the MAPK pathway.

Gonadotropin elevation is ootoxic to ovulatory oocytes and inhibits oocyte maturation, and activin decoy receptor ActRIIB:Fc therapeutically restores maturation.

BACKGROUND: Elevated FSH often occurs in women of advanced maternal age (AMA, age ≥ 35) and in infertility patients undergoing controlled ovarian stimulation (COS). There is controversy on whether high endogenous FSH contributes to infertility and whether high exogenous FSH adversely impacts patient pregnancy rates. METHODS: The senescence-accelerated mouse-prone-8 (SAMP8) model of female reproductive aging was employed to assess the separate impacts of age and high FSH activity on the percentages (%) of viable and mature ovulated oocytes recovered after gonadotropin treatment. Young and midlife mice were treated with the FSH analog equine chorionic gonadotropin (eCG) to model both endogenous FSH elevation and exogenous FSH elevation. Previously we showed the activin inhibitor ActRIIB:Fc increases oocyte quality by preventing chromosome and spindle misalignments. Therefore, ActRIIB:Fc treatment was performed in an effort to increase % oocyte viability and % oocyte maturation. RESULTS: The high FSH activity of eCG is ootoxic to ovulatory oocytes, with greater decreases in % viable oocytes in midlife than young mice. High FSH activity of eCG potently inhibits oocyte maturation, decreasing the % of mature oocytes to similar degrees in young and midlife mice. ActRIIB:Fc treatment does not prevent eCG ootoxicity, but it restores most oocyte maturation impeded by eCG. CONCLUSIONS: FSH ootoxicity to ovulatory oocytes and FSH maturation inhibition pose a paradox given the well-known pro-growth and pro-maturation activities of FSH in the earlier stages of oocyte growth. We propose the FOOT Hypothesis ("FSH OoToxicity Hypothesis), that FSH ootoxicity to ovulatory oocytes comprises a new driver of infertility and low pregnancy success rates in DOR women attempting spontaneous pregnancy and in COS/IUI patients, especially AMA women. We speculate that endogenous FSH elevation also contributes to reduced fecundity in these DOR and COS/IUI patients. Restoration of oocyte maturation by ActRIB:Fc suggests that activin suppresses oocyte maturation in vivo. This contrasts with prior studies showing activin A promotes oocyte maturation in vitro. Improved oocyte maturation with agents that decrease endogenous activin activity with high specificity may have therapeutic benefit for COS/IVF patients, COS/IUI patients, and DOR patients attempting spontaneous pregnancies.

PRG ameliorates cognitive impairment in Alzheimer's disease mice by regulating ß-amyloid and targeting the ERK pathway.

BACKGROUND: PRG is derived from Phellinus ribis and is a homogeneous polysaccharide with well-defined structural information. PRG was found to have significant in vitro neurotrophic and neuroprotective activities. Thus, PRG might be a potential treatment for Alzheimer's disease. However, the related mechanisms of action are still unclear, so deeper in vivo experimental validation and the potential mechanisms need to be investigated. PURPOSE: The effects of PRG on AD mice were investigated using Senescence-accelerated SAMP8 mice as an AD model to elucidate the crucial molecular mechanisms. METHODS: PRG was obtained from Phellinus ribis by water-alcohol precipitation, column chromatography, and ultrafiltration. The Morris water maze and novel object recognition behavioral assays were used to evaluate the effects of PRG in AD mice. Nissl staining, the TUNEL apoptosis assay, and Golgi staining were used to assess brain neuronal cell damage, apoptosis, and neuronal status. Enzyme-linked immunosorbent assays, Western blotting, and immunofluorescence were used to explore the impacts of correlated factors and protein pathways under relevant mechanisms. RESULTS: The findings suggest that PRG improved learning ability and spatial memory capacity in SAMP8 mice. PRG hastened the disintegration of ß-amyloid, reduced the content and abnormal accumulation of the toxic Aß1-42 protein, and decreased apoptosis. PRG activated the BDNF/ERK/CREB signaling pathway through a cascade, exerted neurotrophic effects, regulated cell proliferation and differentiation, increased neuronal dendritic branching and spine density, and improved synaptic plasticity. CONCLUSION: PRG promoted ß-amyloid degradation to reduce neuronal damage and apoptosis. It exerted neurotrophic effects by activating the BDNF/ERK/CREB pathway, promoting neuronal dendritic branching and dendritic spine growth, regulating cell proliferation and differentiation, and improving synaptic plasticity, which improved AD. Taken together, as a novel natural active polysaccharide with a well-defined structure, PRG affected AD symptoms in senescence-accelerated mice by interacting with multiple targets. The results indicate that PRG is a promising potential anti-AD drug candidate.

Mechanisms underlying LncRNA SNHG1 regulation of Alzheimer's disease involve DNA methylation.

Alzheimer's disease (AD) is a neurodegenerative disease associated with long non-coding RNAs and DNA methylation; however, the mechanisms underlying the role of lncRNA small nucleolar RNA host gene 1 (lncRNA SNHG1) and subsequent involvement of DNA methylation in AD development are not known. The aim of this study was to examine the regulatory mechanisms attributed to lncRNA SNHG1 gene utilizing 2 strains of senescence-accelerated mouse prone 8 (SAMP8) model of AD and compared to senescence-accelerated mouse resistant (SAMR) considered a control. Both strains of the mouse were transfected with either blank virus, psLenti-U6-SNHG1(low gene expression) virus, and psLenti-pA-SNHG1(gene overexpression) virus via a single injection into the brains for 2 weeks. At 2 weeks mice were subjected to a Morris water maze to determine any behavioral effects followed by sacrifice to extract hippocampal tissue for Western blotting to measure protein expression of p-tau, DNMT1, DNMT3A, DNMT3B, TET1, and p-Akt. No marked alterations were noted in any parameters following blank virus transfection. In SAMP8 mice, a significant decrease was noted in protein expression of DNMT1, DNMT3A, DNMT3B, and p-Akt associated with rise in p-tau and TET1. Transfection with ps-Lenti-U6-SNHG1 alone in SAMR1 mice resulted in a significant rise in DNMTs and p-Akt and a fall in p-tau and TET1. Transfection of SAMP8 with ps-Lenti-U6-SNHG1 blocked effects on overexpression noted in this mouse strain. However, knockdown of lncRNA SNHG1 yielded the opposite results as found in SAMR1 mice. In conclusion, the knockdown of lncRNA SNHG1 enhanced DNA methylation through the PI3K/Akt signaling pathway, thereby reducing the phosphorylation levels of tau in SAMP8 AD model mice with ameliorating brain damage attributed to p-tau accumulation with consequent neuroprotection.

Multi-strain probiotics ameliorate Alzheimer's-like cognitive impairment and pathological changes through the AKT/GSK-3ß pathway in senescence-accelerated mouse prone 8 mice.

BACKGROUND: Alzheimer's disease (AD), the most prevalent type of dementia, still lacks disease-modifying treatment strategies. Recent evidence indicates that maintaining gut microbiota homeostasis plays a crucial role in AD. Targeted regulation of gut microbiota, including probiotics, is anticipated to emerge as a potential approach for AD treatment. However, the efficacy and mechanism of multi-strain probiotics treatment in AD remain unclear. METHODS: In this study, 6-month-old senescence-accelerated-mouse-prone 8 (SAMP8) and senescence-accelerated-mouse-resistant 1 (SAMR1) were utilized. The SAMP8 mice were treated with probiotic-2 (P2, a probiotic mixture of Bifidobacterium lactis and Lactobacillus rhamnosus) and probiotic-3 (P3, a probiotic mixture of Bifidobacterium lactis, Lactobacillus acidophilus, and Lactobacillus rhamnosus) (1 × 109 colony-forming units) once daily for 8 weeks. Morris water maze (MWM) and novel object recognition (NOR) tests were employed to assess the memory ability. 16S sequencing was applied to determine the composition of gut microbiota, along with detecting serum short-chain fatty acids (SCFAs) concentrations. Neural injury, Aß and Tau pathology, and neuroinflammation level were assessed through western blot and immunofluorescence. Finally, potential molecular mechanisms was explored through transcriptomic analysis and western blotting. RESULTS: The MWM and NOR test results indicated a significant improvement in the cognitive level of SAMP8 mice treated with P2 and P3 probiotics compared to the SAMP8 control group. Fecal 16S sequencing revealed an evident difference in the α diversity index between SAMP8 and SAMR1 mice, while the α diversity of SAMP8 mice remained unchanged after P2 and P3 treatment. At the genus level, the relative abundance of ten bacteria differed significantly among the four groups. Multi-strain probiotics treatment could modulate serum SCFAs (valeric acid, isovaleric acid, and hexanoic acid) concentration. Neuropathological results demonstrated a substantial decrease in neural injury, Aß and Tau pathology and neuroinflammation in the brain of SAMP8 mice treated with P3 and P2. Transcriptomic analysis identified the chemokine signaling pathway as the most significantly enriched signaling pathway between SAMP8 and SAMR1 mice. Western blot test indicated a significant change in the phosphorylation level of downstream AKT/GSK-3ß between the SAMP8 and SAMR1 groups, which could be reversed through P2 and P3 treatment. CONCLUSIONS: Multi-strain probiotics treatment can ameliorate cognitive impairment and pathological change in SAMP8 mice, including neural damage, Aß and Tau pathology, and neuroinflammation. This effect is associated with the regulation of the phosphorylation of the AKT/GSK-3ß pathway.

Differential Epigenetic Changes in the Dorsal Hippocampus of Male and Female SAMP8 Mice: A Preliminary Study.

Alzheimer's disease (AD) is the most common age-related neurodegenerative disease characterized by memory loss and cognitive impairment. The causes of the disease are not well understood, as it involves a complex interaction between genetic, environmental, and epigenetic factors. SAMP8 mice have been proposed as a model for studying late-onset AD, since they show age-related learning and memory deficits as well as several features of AD pathogenesis. Epigenetic changes have been described in SAMP8 mice, although sex differences have never been evaluated. Here we used western blot and qPCR analyses to investigate whether epigenetic markers are differentially altered in the dorsal hippocampus, a region important for the regulation of learning and memory, of 9-month-old male and female SAMP8 mice. We found that H3Ac was selectively reduced in male SAMP8 mice compared to male SAMR1 control mice, but not in female mice, whereas H3K27me3 was reduced overall in SAMP8 mice. Moreover, the levels of HDAC2 and JmjD3 were increased, whereas the levels of HDAC4 and Dnmt3a were reduced in SAMP8 mice compared to SAMR1. In addition, levels of HDAC1 were reduced, whereas Utx and Jmjd3 were selectively increased in females compared to males. Although our results are preliminary, they suggest that epigenetic mechanisms in the dorsal hippocampus are differentially regulated in male and female SAMP8 mice.

Effects of exercise intensity on spatial memory performance and hippocampal synaptic function in SAMP8 mice.

Learning and memory impairment is commonly noted in Alzheimer's disease (AD), which is regarded as a progressive synaptic failure disease. Exercise is a nonpharmacological strategy that may help prevent cognitive decline and reduce the risk of AD, which is usually thought to be related to synaptic damage in the hippocampus. However, the effects of exercise intensity on hippocampal memory and synaptic function in AD remain unclear. In this study, senescence-accelerated mouse prone-8 (SAMP8) mice were randomly assigned to the control group (Con), low-intensity exercise group (Low), and moderate-intensity exercise group (Mid). Here, we showed that eight weeks of treadmill exercise beginning in four-month-old mice improved spatial memory and recognition memory in six-month-old SAMP8 mice, while the Con group exhibited impaired spatial memory and recognition memory. Treadmill exercise also improved hippocampal neuron morphology in SAMP8 mice. Furthermore, dendritic spine density and the levels of postsynaptic density protein-95 (PSD95) and Synaptophysin (SYN) increased significantly in the Low and Mid groups as compared with the Con group. We further showed that moderate-intensity exercise (60 % of maximum speed) was more efficacious in increasing dendritic spine density、PSD95 and SYN, than low-intensity exercise (40 % of maximum speed). In conclusion, the positive effect of treadmill exercise is closely related to exercise intensity, with moderate-intensity exercise showing the most optimal effects.

Yuanzhi Powder inhibits tau pathology in SAMP8 mice: Mechanism research of a traditional Chinese formula against Alzheimer's disease.

ETHNOPHARMACOLOGICAL RELEVANCE: Yuanzhi Powder (YZP), a classical Chinese medicine formula, is good at tonifying heart-Qi and improving cognitive ability. YZP has been reported to show therapeutic effect on alleviating the symptoms of Alzheimer's disease (AD). AIM OF THE STUDY: This study was conducted to observe the effects of YZP on improving the cognitive abilities of SAMP8 mice, and explore the involved mechanisms on inhibiting the excessive accumulation of phosphorylated tau. MATERIAL AND METHODS: Thirty SAMP8 mice were randomly divided into five groups: AD group, AD + DO group, AD + YZP group, AD + LAC group and AD + LAC + YZP group. Age-matched SAMR1 mice were served as CTL group. AD + LAC group and AD + LAC + YZP group received 1 µg Lactacystin solution via intra-cerebroventricular injection. All mice (except the CTL group and AD + LAC group) were intragastrically administrated for 8 consecutive weeks. Then, the Morris Water Maze (MWM) test was conducted for evaluation of learning and memory abilities. The pathological changes of hippocampal CA1 were observed by Hematoxylin & eosin (H&E) staining. The expression of 26S proteasome in the hippocampus was measured by Western Blot (WB) and immunohistochemistry (IHC). The expressions of total tau (Tau5) and hyperphosphorylated tau (pS199, pT231 and pS396) were detected by WB. The aggregation of hyperphosphorylated tau and the binding ability of tau protein to microtubules were evaluated respectively by immunostaining and Thioflavin-S staining and double-label immunofluorescence. RESULTS: SAMP8 mice showed serious cognitive impairment in behavioral tests. However, treatment of YZP significantly ameliorated the cognitive deficits of SAMP8 mice. The H&E staining suggested that YZP could protect against neuronal loss in SAMP8 mice. The IHC and WB results showed that YZP increases 26S proteasome expression in SAMP8 mice and 26S proteasome expression was effectively inhibited by Lactacystin. Meanwhile, The WB results demonstrated that YZP can inhibit the expression of hyperphosphorylated tau (pT231, pS396 and pS199). Furthermore, the immunostaining and Thioflavin-S staining and double-label immunofluorescence results indicated that YZP attenuates the excessive aggregation of hyperphosphorylated tau and enhances the binding ability of tau to stabilize microtubules in SAMP8 mice. CONCLUSIONS: YZP could enhance cognitive performance and learning of AD, ameliorate tau pathology and significantly improve the binding ability of tau to microtubules, based potentially on inhibiting the excessive aggregation of hyperphosphorylated tau via the 26Sproteasome pathway but not necessarily the only one.

Aging-Accelerated Mouse Prone 8 (SAMP8) Mice Experiment and Network Pharmacological Analysis of Aged Liupao Tea Aqueous Extract in Delaying the Decline Changes of the Body.

Aging and metabolic disorders feedback and promote each other and are closely related to the occurrence and development of cardiovascular disease, type 2 diabetes, neurodegeneration and other degenerative diseases. Liupao tea is a geographical indication product of Chinese dark tea, with a "red, concentrated, aged and mellow" flavor quality. In this study, the aqueous extract of aged Liupao tea (ALPT) administered by continuous gavage significantly inhibited the increase of visceral fat and damage to the intestinal-liver-microbial axis in high-fat modeling of SAMP8 (P8+HFD) mice. Its potential mechanism is that ALPT significantly inhibited the inflammation and aggregation formation pathway caused by P8+HFD, increased the abundance of short-chain fatty acid producing bacteria Alistipes, Alloprevotella and Bacteroides, and had a calorie restriction effect. The results of the whole target metabolome network pharmacological analysis showed that there were 139 potential active components in the ALPT aqueous extract, and the core targets of their actions were SRC, TP53, AKT1, MAPK3, VEGFA, EP300, EGFR, HSP90AA1, CASP3, etc. These target genes were mainly enriched in cancer, neurodegenerative diseases, glucose and lipid metabolism and other pathways of degenerative changes. Molecular docking further verified the reliability of network pharmacology. The above results indicate that Liupao tea can effectively delay the body's degenerative changes through various mechanisms and multi-target effects. This study revealed that dark tea such as Liupao tea has significant drinking value in a modern and aging society.

Probing the relevance of the accelerated aging mouse line SAMP8 as a model for certain types of neuropsychiatric symptoms in dementia.

Introduction: People with dementia (PwD) often present with neuropsychiatric symptoms (NPS). NPS are of substantial burden to the patients, and current treatment options are unsatisfactory. Investigators searching for novel medications need animal models that present disease-relevant phenotypes and can be used for drug screening. The Senescence Accelerated Mouse-Prone 8 (SAMP8) strain shows an accelerated aging phenotype associated with neurodegeneration and cognitive decline. Its behavioural phenotype in relation to NPS has not yet been thoroughly investigated. Physical and verbal aggression in reaction to the external environment (e.g., interaction with the caregiver) is one of the most prevalent and debilitating NPS occurring in PwD. Reactive aggression can be studied in male mice using the Resident-Intruder (R-I) test. SAMP8 mice are known to be more aggressive than the Senescence Accelerated Mouse-Resistant 1 (SAMR1) control strain at specific ages, but the development of the aggressive phenotype over time, is still unknown. Methods: In our study, we performed a longitudinal, within-subject, assessment of aggressive behaviour of male SAMP8 and SAMR1 mice at 4, 5, 6 and 7 months of age. Aggressive behaviour from video recordings of the R-I sessions was analysed using an in-house developed behaviour recognition software. Results: SAMP8 mice were more aggressive relative to SAMR1 mice starting at 5 months of age, and the phenotype was still present at 7 months of age. Treatment with risperidone (an antipsychotic frequently used to treat agitation in clinical practice) reduced aggression in both strains. In a three-chamber social interaction test, SAMP8 mice also interacted more fervently with male mice than SAMR1, possibly because of their aggression-seeking phenotype. They did not show any social withdrawal. Discussion: Our data support the notion that SAMP8 mice might be a useful preclinical tool to identify novel treatment options for CNS disorders associated with raised levels of reactive aggression such as dementia.

Effects of Scrophularia buergeriana Extract (Brainon®) on Aging-Induced Memory Impairment in SAMP8 Mice.

Alzheimer's disease (AD) is a worldwide problem. Currently, there are no effective drugs for AD treatment. Scrophularia buergeriana Miquel (SB) is a traditional herbal medicine used in Korea to treat various diseases. Our previous studies have shown that ethanol extract of SB roots (SBE, Brainon®) exhibits potent anti-amnesic effects in Aß1-42- or scopolamine-treated memory impairment mice model and neuroprotective effects in a glutamate-induced SH-SY5Y cell model. In this study, we evaluated the therapeutic effects of Brainon® and its mechanism of action in senescence-accelerated mouse prone 8 (SAMP8) mice. Brainon® (30 or 100 mg/kg/day) was orally treated to six-month-old SAMP8 mice for 12 weeks. Results revealed that Brainon® administration effectually ameliorated cognitive deficits in Y-maze and passive avoidance tests. Following the completion of behavioral testing, western blotting was performed using the cerebral cortex. Results revealed that Brainon® suppressed Aß1-42 accumulation, Tau hyperphosphorylation, oxidative stress, and inflammation and alleviated apoptosis in SAMP8 mice. Brainon® also promoted synaptic function by downregulating the expression of AChE and upregulating the expression of p-CREB/CREB and BDNF. Furthermore, Brainon® restored SAMP8-reduced expression of ChAT and -dephosphorylated of ERK and also decreased AChE expression in the hippocampus. Furthermore, Brainon® alleviated AD progression by promoting mitophagy/autophagy to maintain normal cellular function as a novel finding of this study. Our data suggest that Brainon® can remarkably improve cognitive deficiency with the potential to be utilized in functional food for improving brain health.

[Effect of electroacupuncture on complement C1q and microglia phagocytosis in hippocampus of SAMP8 mice].

OBJECTIVE: To observe the effect of electroacupuncture (EA) on the expression of Iba-1, complement C1q and CD68 in hippocampus of SAMP8 mice, so as to explore its mechanisms underlying improvement of Alzheimer's disease (AD). METHODS: Twenty-four male SAMP8 mice were randomly and equally divided into model and EA groups, and 12 SAMR1 mice were used as the control group. EA (2 Hz, 1.5-2.0 mA) was applied to "Baihui" (GV20), "Dazhui"(GV14) and "Shen-shu"(BL23) for 20 min once daily in the EA group, each course of treatment was 8 days, with an interval of 2 days between two courses, and the mice were treated for 3 courses. Morris water maze test was performed to assess the learning-memory ability of mice. The positive expression levels of Iba-1 and CD68 proteins in the hippocampus CA1 region were detected by immunohistochemistry. The mRNA and protein expression levels of Iba-1,C1q and CD68 in the hippocampus were detected by real-time PCR and Western blot, separately. RESULTS: Compared with the control group, the average escape latency of Morris water maze test was prolonged in the model group (P<0.01), duration of swimming in the original platform quadrant and the number of original platform crossing were significantly shorter and decreased respectively (P<0.01). Compared with the model group, the average escape latency in the EA group was shortened (P<0.05, P<0.01), the duration of swimming in the original platform quadrant and the number of original platform crossing were significantly prolonged and increased (P<0.01). The immunoactivity of Iba-1 and CD68 in hippocampal CA1 region, and mRNA and protein expression levels of hippocampal Iba-1,C1q and CD68 were significantly up-regulated in the model group in contrast to the control group (P<0.01, P<0.05), and obviously down-regulated except the mRNA expression level of hippocampal Iba-1 in the EA group relevant to the model group (P<0.01, P<0.05). CONCLUSION: EA can improve the learning and memory ability of SAMP8 mice, which may be associated with its effect in inhibiting of complement C1q-dependent microglial phagocytosis in the hippocampus.

Stress upregulates 2-arachidonoylglycerol levels in the hypothalamus, midbrain, and hindbrain, and it is sustained by green nut oil supplementation in SAMP8 mice revealed by DESI-MSI.

The endocannabinoid 2-arachidonoylglycerol (2AG) is an important modulator of stress responses. Its level in the brain increases in response to stress, but region-specific effects of stress on brain 2AG are not well known yet. Moreover, green nut oil (GNO), oil extracted from the seeds of Plukenetia volubilis has several health benefits, but its effects on brain 2AG levels are unknown. Therefore, we conducted this study to explore the effects of stress and GNO supplementation on 2AG levels in specific brain regions of senescence-accelerated mouse prone 8 (SAMP8). In this study, desorption electrospray ionization-mass spectrometry imaging (DESI-MSI) revealed that water-immersion stress for three days significantly increased 2AG levels in several brain regions of SAMP8 mice, including the hypothalamus, midbrain, and hindbrain. No significant change was observed in the relative abundance of brain 2AG in stress given SAMP8 mice after eighteen days of removing stress load compared to control SAMP8 mice. GNO supplementation also increased brain 2AG in SAMP8 mice without stress load. Additionally, GNO supplementation sustained the increased brain 2AG levels in stress given SAMP8 mice after eighteen days of removing stress load. Among all brain regions, a relatively higher accumulation of 2AG was noted in the hypothalamus, midbrain, and hindbrain of GNO-fed SAMP8. Our data explored the potentiality of GNO supplementation to improve brain 2AG levels which might be used to treat anxiety and depressive behaviors.

An Overlooked Prebiotic: Beneficial Effect of Dietary Nucleotide Supplementation on Gut Microbiota and Metabolites in Senescence-Accelerated Mouse Prone-8 Mice.

Nucleotides (NTs) are regulatory factors in many biological processes and play important roles in the growth, development, and metabolism of living organisms. We used senescence-accelerated mouse prone-8 (SAMP8) to investigate the effects of NTs on the gut microbiota and metabolites. And the promoting effect of NTs on the growth of a probiotic (Lactobacillus casei) was explored through in vitro experiments. The results showed that the sequencing depth of 16S rDNA covered all microbial species in the feces of SAMP8. Supplementation with exogenous NTs to the diet enhanced the diversity of the gut microbiota, reduced the abundance of bacteria with negative effects on the body (such as Verrucomicrobia, Ruminococcaceae, Akkermansia and Helicobacter), and increased the abundance of the microbiota, which had beneficial effects on the mice (such as Lactobacillus, Candidatus saccharimonas and Lachnospiraceae_NK4A136_group). Metabonomic analysis showed that NT deficiency in the diet significantly affected metabolites in the mouse feces. The metabolites in mice supplemented with NTs tended to be normal (SAMR1). The differentially expressed metabolites caused by NT addition are involved in various pathways in the body, including linoleic acid metabolism, vitamin B6 metabolism, and histidine metabolism. Correlation analysis revealed a significant correlation between the gut microbiota and differentially expressed metabolites caused by the addition of NTs. In vitro experiments showed that NTs significantly promoted the growth, secretion of biofilm and extracellular polymeric substance of L. casei. NTs also promoted the ability of the crude extract of L. casei to resist the secretion of Shigella biofilm. Thus, NTs can regulate the abundance of the gut microbiota and alter the metabolic expression of the intestinal microbiome.

Kampo formulas alleviate aging-related emotional disturbances and neuroinflammation in male senescence-accelerated mouse prone 8 mice.

Aging-induced neuroinflammation, also known as neuroinflammaging, plays a pivotal role in emotional disturbances, including depression and anxiety, in older individuals, thereby leading to cognitive dysfunction. Although numerous studies have focused on therapeutic strategies for cognitive impairment in older individuals, little research has been performed on treating its preceding emotional disturbances. Here, we examined whether Kampo formulas (kososan [KS], nobiletin-rich kososan [NKS], and hachimijiogan [HJG]) can ameliorate aging-induced emotional disturbances and neuroinflammation in mice. The depression-like behaviors observed in SAMP8 mice, relative to normally aging SAMR1 mice, were significantly prevented by treatment with Kampo formulas for 13 weeks. Western blot analysis revealed that hippocampal neuroinflammation was significantly abrogated by Kampo formulas. KS and NKS also significantly attenuated the hippocampal neuroinflammatory priming induced by lipopolysaccharide (LPS, 0.33 mg/kg, i.p.) challenge in SAMP8 mice. Hippocampal IL-1ß, IL-6, and MCP-1 levels were significantly decreased in NKS-treated SAMP8 mice. KS and NKS showed significantly reduced tau accumulation in the brains of SAMP8 mice. RNA-sequencing revealed that each Kampo formula led to unique dynamics of hippocampal gene expression and appeared to abrogate hippocampal inflammatory responses. HJG significantly blocked the LPS-induced increase in serum IL-6 and MCP-1. These results suggest that Kampo formulas would be useful for treating aging-induced depression, in part by regulating neuroinflammatory pathways. This finding may pave the way for the development of therapeutic strategies for aging-related emotional disturbances, which may contribute to the prevention of cognitive dysfunction in older individuals.

Effects and mechanisms of melatonin combined with enriched environment on learning and memory ability of SAMP8 mice / 中华行为医学与脑科学杂志

Objective:To explore the effects and possible mechanisms of melatonin combined with enriched environment on the learning and memory ability of senescence-accelerated mouse prone 8(SAMP8).Methods:Forty-eight SAMP8 male mice aged 4 months were randomly divided into model group, enriched environment group, melatonin group and melatonin combined with enriched environment group (combined intervention group) by random number table method, with 12 mice in each group. Mice in the melatonin group and combined intervention group were subcutaneously injected with melatonin at a dose of 8 mg·kg -1·d -1, and the mice in the model group and the enriched environment group were given the same amount of normal saline instead.The mice in model group and melatonin group were raised in a standard environment, and the mice in enriched environment group and combined intervention group were raised in an enriched environment.The intervention lasted 28 days. The aging degree of mice was scored before and 28 days after the intervention. Morris water maze test was used to detect the learning and memory ability of mice. Nissl staining and TUNEL staining were used to observe the Nissl staining positive cells and apoptotic cells in the CA1 area of hippocampus.ELISA was used to detect the levels of interleukin-1β (IL-1β), interleukin-6 (IL-6) and tumor necrosis factor-α (TNF-α) in the hippocampus of mice. Western blot was used to detect the levels of amyloid β-protein (Aβ) 1-42, microtubule-associated protein tau (tau) phosphorylated at threonine (Thr) 205 (Tau pT205), Toll-like receptor 4 (TLR4), and nuclear factor-κB (NF-κB) p65 protein in the hippocampus of mice. qRT-PCR was used to detect the levels of TLR4, NF-κB p65 mRNA in the hippocampus of mice. SPSS 22. 0 statistical software was used for repeated measure ANOVA, one-way ANOVA and LSD test. Results:(1) Aging score: after intervention, the aging scores of mice in the four groups were significantly different ( F=120.601, P<0.01). The aging scores of mice in the enriched environment group, melatonin group, and combined intervention group were lower than those in the model group (all P<0.05), while the aging score of mice in the combined intervention group was significantly lower than those in the enriched environment group and melatonin group (both P<0.05). (2) The results of the location navigation experiment showed that the time × group interaction effect of the escape latencies of mice in the four groups were significant ( F=30.524, P<0.001). From the 2nd to 4th day, the escape latencies of mice in the enriched environment group, melatonin group and combined intervention group were all lower than that in the model group (all P<0.05). The results of the space exploration experiment showed that the residence time in the target quadrant and the number of platform crossings of mice in the four groups were significantly different ( F=291.328, 113.482, both P<0.01). The residence time in the target quadrant ((29.45±1.70)s, (32.44±1.55)s, (37.48±0.84) s) and the number of platform crossings ((6.44±0.61) times, (7.16±0.70) times, (12.60±1.23) times) of mice in the enriched environment group, melatonin group and combined intervention group were higher than those in the model group ((15.07±1.28) s, (4.10±0.61) times), while the residence time in the target quadrant and the number of platform crossings of mice in the enriched environment group and the melatonin group were significantly lower than those in the combined intervention group (all P<0.05). (3) Nissl and TUNEL staining showed that the number of Nissl positive neurons in the hippocampal CA1 region of mice in the four groups were significantly different ( F=809.264, P<0.01), and the number of apoptotic cells in the hippocampal CA1 region were also significantly different ( F=1 060.583, P<0.01). The number of Nissl stained positive neurons in the hippocampal CA1 region of mice in the combined intervention group was more than those in the model group, enriched environment group, and melatonin group (all P<0.05), and the number of apoptotic cells were less than those in the model group, enriched environment group, and melatonin group (all P<0.05). (4) The results of ELISA assay showed that there were significantly different in the levels of IL-1β, IL-6 and TNF-α in the hippocampus of mice in the four groups ( F=152.887, 63.506, 432.026, all P<0.01). The contents of IL-1β, IL-6 and TNF-α in the hippocampus of mice in the enriched environment group, melatonin group, and combined intervention group were lower than those in the model group(all P<0.05). Among them, the contents of IL-1β, IL-6 and TNF-α in the hippocampus of mice in the enriched environment group and melatonin group were significantly higher than those in the combined intervention group (all P<0.05). (5) Western blot analysis showed that there were significantly different in the protein expression levels of Aβ1~42, tau pT205, TLR4, NF-κB p65 in the hippocampus of mice in the four groups ( F=122.349, 98.934, 201.635, 116.553, all P<0.01). The protein expression levels of Aβ1-42, tau pT205, TLR4, and NF-κB p65 in the hippocampus of mice in the enriched environment group, melatonin group, and combined intervention group were lower than those in the model group.Among them, the protein expression levels of Aβ1-42, tau pT205, TLR4, NF-κB p65 in the hippocampus of mice in the enriched environment group and melatonin group were significantly higher than those in the combined intervention group (all P<0.05). (6) qRT-PCR showed that the mRNA expression levels of TLR4 and NF-κB p65 in the hippocampus of mice in the four groups were significantly different ( F=42.913, 102.446, both P<0.01). The mRNA expression levels of TLR4 ((0.63±0.05), (0.55±0.04), (0.42±0.03)) and NF-κB p65 ((0.98±0.06), (0.82±0.04), (0.72±0.04)) in the hippocampus of mice in the enriched environment group, melatonin group and combined intervention group were lower than those in the model group ((0.74±0.07), (1.20±0.05)) (all P<0.05). Among them, the mRNA expression levels of TLR4 and NF-κB p65 in the hippocampus of mice in the enriched environment group and melatonin group were significantly higher than those in the combined intervention group (all P<0.05). Conclusion:Melatonin combined with enriched environment can improve the learning and memory ability and neuroinflammatory response of SAMP8 mice, and its mechanism may be related with the down-regulation of TLR4/NF-κB p65 signaling pathway.

Evaluation of the Anti-Aging Effects of a Probiotic Combination Isolated From Centenarians in a SAMP8 Mouse Model.

Population aging is a prominent global problem in today's society. However, there are currently no good methods to treat or prevent aging, so anti-aging research has crucial implications. In this research, we screened bacteria from centenarians, and finally selected four probiotics (Lactobacillus fermentum SX-0718, L. casei SX-1107, Bifidobacterium longum SX-1326, and B. animalis SX-0582) to form a probiotic combination. By using the senescence accelerated mouse prone 8 (SAMP8) model, the anti-aging effects of the probiotic combination were evaluated by using behavioural testing, neuroinflammation, intestinal inflammation, and intestinal microbiota. The results showed that probiotic combination improved the impaired spatial memory, motor dysfunction, and decreased exploratory behavior in aging mice. The probiotic combination inhibited Toll-like receptor 4 (TLR4)/nuclear factor kappa B (NFκB)-induced neuroinflammation and up-regulated the expression of Sirt 1 to protect hippocampal neurons. At the same time, the probiotic combination regulated the intestinal microbiota, reduced the relative abundance of Alistipes and Prevotella in SAMP8 mice, inhibited TLR4/NFκB-induced intestinal inflammation, and increased the expression of intestinal permeability related proteins zonula occludens-1 (ZO-1) and Occuldin. The anti-aging effects of the probiotic combination may be through the regulating intestinal microbiota and inhibiting TLR4/NFκB-induced inflammation. This research provides the basis and technical support for the future production and application of the probiotic combination.

Palmitoylethanolamide Counteracts Enteric Inflammation and Bowel Motor Dysfunctions in a Mouse Model of Alzheimer's Disease.

Palmitoylethanolamide (PEA), an endogenous lipid mediator, is emerging as a promising pharmacological agent in multiple neurodegenerative disorders for its anti-inflammatory and neuroprotective properties. However, its effects on enteric inflammation and colonic dysmotility associated with Alzheimer's disease (AD) are lacking. This study was designed to investigate the beneficial effect of PEA administration in counteracting the enteric inflammation and relieving the bowel motor dysfunctions in an AD mouse model, SAMP8 mice. In addition, the ability of PEA in modulating the activation of enteric glial cells (EGCs), pivotally involved in the pathophysiology of bowel dysfunctions associated with inflammatory conditions, has also been examined. SAMP8 mice at 4 months of age were treated orally with PEA (5 mg/kg/day) for 2 months. SAMR1 animals were employed as controls. At the end of treatment, parameters dealing with colonic motility, inflammation, barrier integrity and AD protein accumulation were evaluated. The effect of PEA on EGCs was tested in cultured cells treated with lipopolysaccharide (LPS) plus ß-amyloid 1-42 (Aß). SAMP8 treated with PEA displayed: 1) an improvement of in vitro colonic motor activity, citrate synthase activity and intestinal epithelial barrier integrity and 2) a decrease in colonic Aß and α-synuclein (α-syn) accumulation, S100-ß expression as well as enteric IL-1ß and circulating LPS levels, as compared with untreated SAMP8 mice. In EGCs, treatment with PEA counteracted the increment of S100-ß, TLR-4, NF-κB p65 and IL-1ß release induced by LPS and Aß. These results suggest that PEA, under a condition of cognitive decline, prevents the enteric glial hyperactivation, reduces AD protein accumulation and counteracts the onset and progression of colonic inflammatory condition, as well as relieves intestinal motor dysfunctions and improves the intestinal epithelial barrier integrity. Therefore, PEA represents a viable approach for the management of the enteric inflammation and motor contractile abnormalities associated with AD.

The effect and pharmacology of Yizhi Sheng Hui decoction on Alzheimer's disease.

BACKGROUND: Alzheimer's disease (AD) has gradually increased as society has aged and is now a serious social problem. In the clinical treatment of AD, patients show improvement in cognitive function after treatment with the traditional Chinese medicine compound, Yizhi Sheng Hui (YZSH) decoction. This study systematically investigates the effect and pharmacology of the YZSH decoction on AD. METHODS: In this study, 24 SAMP8 AD model mice were randomly divided into three groups: an untreated control group, a group treated with the YZSH decoction, and a positive control group treated with donepezil hydrochloride. Eight SAMR1 mice were placed in the normal control group. A Morris water maze test and a step-down test were conducted at 8 and 13 weeks after continuous intragastric administration of the two drugs. After 13 weeks of administration, the hippocampal expression of Aß1-42 and tau protein were measured. RESULTS: There was no change in the latent period duration and the number of platform crossings in each group after 8 weeks of administration, but after 13 weeks of administration, the latent period of the treatment group and the positive control group were significantly shorter than the untreated control group. Initially, the SAMP8 mice showed a lower spatial exploration ability than the SAMR1 mice. However, after 13 weeks of administration, the treatment group and the control group exhibited a better exploration ability. Compared with the SAMR1 mice, the on-stage evasion time and step-down errors significantly increased in the untreated group. Compared with the untreated group, mice in the treatment group and the positive control group showed a shorter latent period after 8 weeks of administration, and the on-stage evasion time for both groups was significantly reduced after 13 weeks of administration. The treatment group showed fewer instances of electric shock. Hippocampal expression of Aß1-42 was high in the untreated group, was much lower in the positive control group, and no Aß1-42 expression was observed in the treatment group. CONCLUSIONS: The YZSH decoction improved the learning and memory of mice with AD, related to the inhibition of Aß1-42 expression.