OBJECTIVE: To observe the effects of electroacupuncture (EA) at "Shenting" (GV24) and "Baihui" (GV20) on mitochondrial autophagy in hippocampal neurons and silent information regulator sirtuin 1 (Sirt1)/forkhead box O3 (FOXO3)/PTEN-inducible kinase 1 (PINK1)/Parkin pathway in rats with learning-memory impairment after cerebral ischemia reperfusion injury. METHODS: A total of 35 male SD rats were randomly divided into a sham operation group (9 rats) and a modeling group (26 rats). In the modeling group, middle cerebral artery occlusion method was used to establish the middle cerebral artery ischemia-reperfusion (MCAO/R) model, and 18 rats of successful modeling were randomly divided into a model group and an EA group, 9 rats in each one. EA was applied at "Shenting" (GV24) and "Baihui" (GV20) in the EA group, 30 min a time, once a day for 14 days. After modeling and on 7th and 14th days of intervention, neurologic deficit score was observed; the learning-memory ability was detected by Morris water maze test; the morphology of neurons in CA1 area of hippocampus was detected by Nissl staining; the mitochondrial morphology was observed by transmission electron microscopy; the protein expression of Beclin-1, microtubule-associated protein 1 light chain 3B (LC3B), P62, Sitrt1, FOXO3, PINK1 and Parkin was detected by Western blot. RESULTS: After modeling, the neurologic deficit scores in the model group and the EA group were higher than that in the sham operation group (P<0.001); on 7th and 14th days of intervention, the neurologic deficit scores in the model group were higher than those in the sham operation group (P<0.001), the neurologic deficit scores in the EA group were lower than those in the model group (P<0.05, P<0.01). After modeling, the escape latency in the model group and the EA group was prolonged compared with that in the sham operation group (P<0.001); on 9th-13th days of intervention, the escape latency in the model group was prolonged compared with that in the sham operation group (P<0.001), the escape latency in the EA group was shortened compared with that in the model group (P<0.05, P<0.01, P<0.001). The number of crossing plateau in the model group was less than that in the sham operation group (P<0.001); the number of crossing plateau in the EA group was more than that in the model group (P<0.05). In the model group, in CA1 area of hippocampus, the number of neurons was less, with sparse arrangement, nuclear fixation, deep cytoplasmic staining, and reduction of Nissl substance; the morphology of mitochondrion was swollen, membrane structure was fragmented, and autophagic lysosomes were formed. Compared with the model group, in the EA group, in CA1 area of hippocampus, the number of neurons was increased, the number of cells of abnormal morphology was decreased, and the number of Nissl substance was increased; the morphology of mitochondrion was more intact and the number of autophagic lysosomes was increased. Compared with the sham operation group, in the model group, the protein expression of Beclin-1, FOXO3, PINK1, Parkin and the LC3BⅡ/Ⅰ ratio in hippocampus were increased (P<0.01, P<0.001), while the protein expression of P62 was decreased (P<0.05). Compared with the model group, in the EA group, the protein expression of Beclin-1, Sirt1, FOXO3, PINK1, Parkin and the LC3BⅡ/Ⅰratio in hippocampus were increased (P<0.001, P<0.01), while the protein expression of P62 was decreased (P<0.001). CONCLUSION EA at "Shenting" (GV24) and "Baihui" (GV20) can relieve the symptoms of neurological deficits and improve the learning-memory ability in MCAO/R rats, its mechanism may relate to the modulation of Sirt1/FOXO3/PINK1/Parkin pathway and the enhancement of mitochondrial autophagy.
Animals ; Electroacupuncture ; Male ; Rats, Sprague-Dawley ; Rats ; Forkhead Box Protein O3/genetics* ; Reperfusion Injury/metabolism* ; Ubiquitin-Protein Ligases/genetics* ; Brain Ischemia/complications* ; Mitochondria/genetics* ; Autophagy ; Protein Kinases/genetics* ; Sirtuin 1/genetics* ; Humans ; Memory Disorders/psychology* ; Signal Transduction

Heart failure (HF) is a common end-stage clinical manifestation of cardiovascular diseases, imposing substantial health-related burdens worldwide. With its high mortality rates and poor long-term prognosis, there is a pressing need for novel therapies. SIRT1, a nicotinamide adenine dinucleotide (NAD⁺)-dependent deacetylase, has anti-cardiovascular aging properties and other cardioprotective effects, attracting much research attention in recent years. In addition, SIRT1 plays an important role in HF pathophysiology. This review summarized the roles of SIRT1 and its activators in HF, the changes of SIRT1 gene expression in cardiac tissues from animal models and HF patients, and the current status of clinical trials investigating SIRT1 activators as potential therapies for HF. This will provide new ideas for further exploration of pathological mechanisms and the development of clinical prevention strategies for HF.
Heart Failure/metabolism* ; Sirtuin 1/genetics* ; Humans ; Animals

This study aims to explore the mechanism of Huanglian Jiedu Decoction(HJD) in treating acute liver injury(ALI) in the mouse model of sepsis induced by lipopolysaccharide(LPS). Fifty-four male C57BL/6 mice were randomized into six groups: blank group, model group, low-, medium-, and high-dose group HJD, and dexamethasone group. The mouse model of sepsis was established by intraperitoneal injection of LPS after 7 days of gavage with HJD, and dexamethasone(0.2 mL) was injected intraperitoneally 1.5 h after modeling. The murine sepsis score(MSS) was recorded 12 h after modeling. The levels of alanine aminotransferase(ALT) and aspartate aminotransferase(AST) in the liver tissue and tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6) in the serum were measured by ELISA. Hematoxylin-eosin(HE) staining was used to observe the pathological changes of the mouse liver. The content of light chain 3 of microtubule-associated protein 1(LC3) was detected by immunofluorescence, and that of sirtuin 1(SIRT1) was detected by immunohistochemistry. The mRNA levels of adenosine 5'-monophosphate-activated protein kinase(AMPK), LC3, and P62 were detected by RT-PCR. Western blot was employed to determine the protein levels of AMPK, p-AMPK, and SIRT1 in the liver tissue. The results showed that compared with model group, drug interventions decreased the MSS and liver injury indicators, lowered the levels of inflammatory cytokines, improved the liver tissue structure, upregulated the protein levels of of p-AMPK/AMPK and SIRT1 and the mRNA levels of AMPK and LC3, and downregulated the mRNA level of P62. To sum up, HJD can regulate the autophagy level and reduce inflammation to ameliorate acute liver injury in septic mice by activating the AMPK/SIRT1 autophagy pathway.
Animals ; Drugs, Chinese Herbal/administration & dosage* ; Sirtuin 1/genetics* ; Male ; Mice ; Sepsis/metabolism* ; Mice, Inbred C57BL ; Autophagy/drug effects* ; AMP-Activated Protein Kinases/genetics* ; Liver/metabolism* ; Humans ; Signal Transduction/drug effects* ; Disease Models, Animal ; Tumor Necrosis Factor-alpha/genetics*

This article investigated the effect and mechanism of salidroside(SAL) on the expression of adhesion molecules in oxidized low-density lipoprotein(oxLDL)-induced mouse aortic endothelial cell(MAEC). The oxLDL-induced endothelial cell injury model was constructed, and the safe concentration and action time of SAL were screened. The cells were divided into control group, oxLDL group, low and high concentration groups of SAL, and ferrostatin-1(Fer-1) group. The cell viability was detected by CCK-8 assay; lactate dehydrogenase(LDH) leakage was measured by colorimetry; the expression of intercellular adhesion molecule 1(ICAM-1) and recombinant vascular cell adhesion molecule 1(VCAM-1) were detected by immunofluorescence; Fe~(2+),glutathione(GSH),malondialdehyde(MDA),and 4-hydroxynonenal(4-HNE) levels were detected by kit method; reactive oxygen species(ROS) was detected by DCFH-DA probe; the levels of glutathione peroxidase 4(GPX4),silent mating type information regulation 2 homolog 1(SIRT1), and nuclear factor erythroid 2-related factor 2(Nrf2) were determined by using Western blot. The inhibitors of Nrf2 and SIRT1 were used, and endothelial cell were divided into control group, oxLDL group, SAL group, ML385 group(Nrf2 inhibitor), and EX527 group(SIRT1 inhibitor). The ultrastructure of mitochondria was observed by electron microscope; mitochondrial membrane potential(MMP) was detected by flowcytometry; the expressions of SIRT1,Nrf2,solute carrier family 7 member 11(SLC7A11),GPX4,ferroportin 1(FPN1),ferritin heavy chain 1(FTH1),ICAM-1, and VCAM-1 were detected by Western blot. The results showed that similar to Fer-1,low and high concentrations of SAL could improve cell viability, inhibit LDH release and the expression of ICAM-1 and VCAM-1 in oxLDL-induced endothelial cells(P<0.05 or P<0.01). It was related to increase in GSH level, decrease in Fe~(2+),ROS,MDA, and 4-HNE level, and up-regulation of SIRT1,Nrf2, and GPX4 expression to inhibit ferroptosis(P<0.05 or P<0.01). The intervention effect of high concentration SAL was the most significant. ML385 and EX527 could partially offset the protection of SAL on mitochondrial structure and MMP and reverse the ability of SAL to up-regulate the expression of SIRT1,Nrf2,SLC7A11,GPX4,FPN1, and FTH1 and down-regulate the expression of ICAM-1 and VCAM-1(P<0.05 or P<0.01).To sum up, SAL could reduce the expression of ICAM-1 and VCAM-1 in oxLDL-induced endothelial cell, which may relate to activation of SLC7A11/GPX4 antioxidant signaling pathway mediated by SITR1/Nrf2, up-regulation of FPN1 and FTH1 expression, and inhibition of ferroptosis.
Sirtuin 1/genetics* ; Animals ; Ferroptosis/drug effects* ; Lipoproteins, LDL/metabolism* ; NF-E2-Related Factor 2/genetics* ; Mice ; Endothelial Cells/cytology* ; Glucosides/pharmacology* ; Phenols/pharmacology* ; Cell Adhesion Molecules/genetics* ; Reactive Oxygen Species/metabolism* ; Intercellular Adhesion Molecule-1/genetics* ; Vascular Cell Adhesion Molecule-1/genetics* ; Cell Survival/drug effects*

Objective To investigate the effects of ROCK-siRNA transfection on endothelial cell senescence and endothelial microparticles (EMPs) induced by angiotensin II (Ang II). Methods Human umbilical vein endothelial cells (HUVECs) were treated with Ang II (1.0 μmo/L) to induce cellular senescence models, followed by transfection with ROCK-siRNA. The cells were divided into four groups: control group, model group, negative transfection control group (Ang II combined with NC-siRNA), and ROCK-siRNA transfection group (Ang II combined with ROCK-siRNA). Cellular senescence was assessed by SA-β-Gal staining. EMP levels in cell supernatants and intracellular reactive oxygen species (ROS) levels were assessed using flow cytometry. The expression levels of silenced information regulator 1(SIRT1) and p53 protein in each group were analyzed by Western blotting. Results Following ROCK-siRNA transfection, the number of senescent cells induced by Ang II was significantly reduced, accompanied by decreased CD31⁺ EMP levels and suppressed intracellular ROS levels. Meanwhile, the expression levels of SIRT1 were up-regulated, while the expression levels of p53 were down-regulated. Conclusion Silencing ROCK expression suppresses EMP release, reduces ROS generation, regulates the expression of SIRT1 and p53, and ultimately attenuates Ang II-induced endothelial cell senescence.
Humans ; Angiotensin II/pharmacology* ; Cellular Senescence/genetics* ; Human Umbilical Vein Endothelial Cells/cytology* ; RNA, Small Interfering/metabolism* ; Reactive Oxygen Species/metabolism* ; Sirtuin 1/genetics* ; Transfection ; Tumor Suppressor Protein p53/genetics* ; Cell-Derived Microparticles/drug effects* ; rho-Associated Kinases/metabolism* ; Endothelial Cells/metabolism* ; Cells, Cultured

The activation of the sirtuin1 (SIRT1)/nuclear factor erythroid 2-related factor 2 (Nrf2)/heme oxygenase 1 (HO-1) pathway has been shown to mitigate oxidative stress-induced apoptosis and mitochondrial damage by reducing reactive oxygen species (ROS) levels. Clinical trials have demonstrated that Zhongfeng Xingnao Liquid (ZFXN) ameliorates post-stroke cognitive impairment (PSCI). However, the underlying mechanism, particularly whether it involves protecting mitochondria and inhibiting apoptosis through the SIRT1/Nrf2/HO-1 pathway, remains unclear. This study employed an oxygen-glucose deprivation (OGD) cell model using SH-SY5Y cells and induced PSCI in rats through modified bilateral carotid artery ligation (2VO). The effects of ZFXN on learning and memory, neuroprotective activity, mitochondrial function, oxidative stress, and the SIRT1/Nrf2/HO-1 pathway were evaluated both in vivo and in vitro. Results indicated that ZFXN significantly increased the B-cell lymphoma 2 (Bcl2)/Bcl2-associated X (Bax) ratio, reduced terminal deoxynucleotidyl transferase-mediated dUTP nick-end-labeling (TUNEL)⁺ cells, and markedly improved cognition, synaptic plasticity, and neuronal function in the hippocampus and cortex. Furthermore, ZFXN exhibited potent antioxidant activity, evidenced by decreased ROS and malondialdehyde (MDA) content and increased superoxide dismutase (SOD), catalase (CAT), and glutathione (GSH) levels. ZFXN also demonstrated considerable enhancement of mitochondrial membrane potential (MMP), Tom20 fluorescence intensity, adenosine triphosphate (ATP) and energy charge (EC) levels, and mitochondrial complex I and III activity, thereby inhibiting mitochondrial damage. Additionally, ZFXN significantly increased SIRT1 activity and elevated SIRT1, nuclear Nrf2, and HO-1 levels. Notably, these effects were substantially counteracted when SIRT1 was suppressed by the inhibitor EX-527 in vitro. In conclusion, ZFXN alleviates PSCI by activating the SIRT1/Nrf2/HO-1 pathway and preventing mitochondrial damage.
Sirtuin 1/genetics* ; Animals ; NF-E2-Related Factor 2/genetics* ; Cognitive Dysfunction/genetics* ; Male ; Rats, Sprague-Dawley ; Rats ; Humans ; Signal Transduction/drug effects* ; Drugs, Chinese Herbal/administration & dosage* ; Heme Oxygenase-1/genetics* ; Stroke/complications* ; Oxidative Stress/drug effects* ; Apoptosis/drug effects* ; Mitochondria/metabolism* ; Reactive Oxygen Species/metabolism* ; Neuroprotective Agents

Objective To investigate the role of proanthocyanidins (PC) in lipopolysaccharide (LPS)-induced inflammatory response and its possible mechanism in RAW264.7 macrophages. Methods RAW264.7 macrophages were cultured and treated with PBS and different concentrations of PC for 24 hours, followed by 1 μg/mL LPS for 6 hours. Real-time PCR was used to detect the mRNA expression of interleukin1β (IL-1β), IL-6, monocyte chemoattractant protein 1 (MCP-1), tumor necrotic factor α (TNF-α), IL-4 and arginase 1 (Arg1) in RAW264.7 macrophages. Flow cytometry was used to detect the effects of PBS group, LPS group and PC combined with LPS group on M1 and M2 polarization of macrophages. The protein expressions of silenced information regulator 1 (SIRT1), nuclear factor kappa B p65(NF-κB p65) and acetylated NF-κB p65 (Ace-p65) were detected by Western blot analysis after different concentrations of PC treatment. Co-immunoprecipitation assay was used to detect the binding effect of SIRT1 to NF-κB p65 in macrophages treated with PC. Results Compared with PBS group, the mRNA expression of macrophage pro-inflammatory cytokines IL-1β, IL-6, MCP-1 and TNF-α decreased and the mRNA expression of anti-inflammatory factors IL-4 and Arg1 increased in PC group. Compared with LPS group, PC combined with LPS group could significantly inhibit M1 polarization and promote M2 polarization of macrophages. With the increase of PC concentration, the expression of SIRT1 was up-regulated, and NF-κB p65 protein did not change significantly. The expression of Ace-p65 protein decreased significantly when treated with high concentration of PC. Conclusion PC can significantly alleviate the LPS-induced inflammatory response by up-regulating the expression of SIRT1 and inhibiting NF-κB pathway in RAW264.7 macrophages.
Animals ; Mice ; Interleukin-4 ; Interleukin-6 ; Lipopolysaccharides ; Macrophages ; NF-kappa B ; Proanthocyanidins ; RNA, Messenger ; Sirtuin 1/genetics* ; Tumor Necrosis Factor-alpha ; RAW 264.7 Cells

OBJECTIVE: To explore the possible effects and mechanism of Zhizhu Decoction (ZZD) on the pathophysiology of slow transit constipation (STC). METHODS: A total of 54 C57BL/6 mice was randomly divided into the following 6 groups by a random number table, including control, STC model (model), positive control, and low-, medium- and high-doses ZZD treatment groups (5, 10, 20 g/kg, namely L, M-, and H-ZZD, respectively), 9 mice in each group. Following 2-week treatment, intestinal transport rate (ITR) and fecal water content were determined, and blood and colon tissue samples were collected. Hematoxylin-eosin and periodic acid-Schiff staining were performed to evaluate the morphology of colon tissues and calculate the number of goblet cells. To determine intestinal permeability, serum levels of lipopolysaccharide (LPS), low-density lipoprotein (LDL) and mannose were measured using enzyme-linked immunosorbent assay (ELISA). Western blot analysis was carried out to detect the expression levels of intestinal tight junction proteins zona-occludens-1 (ZO-1), claudin-1, occludin and recombinant mucin 2 (MUC2). The mRNA expression levels of inflammatory cytokines including tumor necrosis factor (TNF)-α, interleukin (IL)-1β, IL-6, IL-4, IL-10 and IL-22 were determined using reverse transcription-quantitative reverse transcription reaction. Colon indexes of oxidative stress were measured by ELISA, and protein expression levels of colon silent information regulator 1/forkhead box O transcription factor 1 (SIRT1/FoxO1) antioxidant signaling pathway were detected by Western blot. RESULTS: Compared with the model group, ITR and fecal moisture were significantly enhanced in STC mice in the M-ZZD and H-ZZD groups (P<0.01). Additionally, ZZD treatment notably increased the thickness of mucosal and muscular tissue, elevated the number of goblet cells in the colon of STC mice, reduced the secretion levels of LPS, LDL and mannose, and upregulated ZO-1, claudin-1, occludin and MUC2 expressions in the colon in a dose-dependent manner, compared with the model group (P<0.05 or P<0.01). In addition, ZZD significantly attenuated intestinal inflammation and oxidative stress and activated the SIRT1/FoxO1 signaling pathway (P<0.05 or P<0.01). CONCLUSION ZZD exhibited beneficial effects on the intestinal system of STC mice and alleviated intestinal inflammation and oxidative stress via activating SIRT1/FoxO1 antioxidant signaling pathway in the colon.
Mice ; Animals ; Sirtuin 1/genetics* ; Antioxidants ; Occludin ; Lipopolysaccharides ; Claudin-1 ; Mannose ; Mice, Inbred C57BL ; Constipation/drug therapy* ; Inflammation ; Signal Transduction

This study aimed to explore the possible effect of Xixin Decoction(XXD) on the learning and memory ability of Alzheimer's disease(AD) model senescence-accelerated mouse-prone 8(SAMP8) and the related mechanism in enhancing neuroprotective effect and reducing neuroinflammation. Forty SAMP8 were randomly divided into a model group(10 mL·kg~(-1)·d~(-1)), a probiotics group(0.39 g·kg~(-1)·d~(-1)), a high-dose group of XXD granules(H-XXD, 5.07 g·kg~(-1)·d~(-1)), a medium-dose group of XXD granules(M-XXD, 2.535 g·kg~(-1)·d~(-1)), and a low-dose group of XXD granules(L-XXD, 1.267 5 g·kg~(-1)·d~(-1)). Eight senescence-accelerated mouse-resistant 1(SAMR1) of the same age and strain were assigned to the control group(10 mL·kg~(-1)·d~(-1)). After ten weeks of intragastric administration, the Morris water maze was used to test the changes in spatial learning and memory ability of mice after treatment. Meanwhile, immunofluorescence staining was used to detect the positive expression of receptor for advanced glycation end products(AGER), Toll-like receptor 1(TLR1), and Toll-like receptor 2(TLR2) in the hippocampal CA1 region of mice. Western blot was employed to test the protein expression levels of silencing information regulator 2 related enzyme 1(SIRT1), AGER, TLR1, and TLR2 in the hippocampus of mice. Enzyme linked immunosorbent assay(ELISA) was applied to assess the levels of Aβ_(1-42) in the hippocampus of mice and the levels of nuclear factor κB p65(NF-κB p65), NOD-like receptor protein 3(NLRP3), tumor necrosis factor-α(TNF-α), and interleukin-1β(IL-1β) in the serum and hippocampus of mice. Compared with the model group, XXD significantly improved the spatial learning and memory ability of SAMP8, increased the expression of neuroprotective factors in the hippocampus, decreased the levels of neuroinflammatory factors, and inhibited the expression of Aβ_(1-42). In particular, H-XXD significantly increased the expression of SIRT1 in the hippocampus of mice, reduced the expression levels of NF-κB p65, NLRP3, TNF-α, and IL-1β in the serum and hippocampus of mice, and decreased the expression of AGER, TLR1, and TLR2 in the hippocampus of mice(P<0.05 or P<0.01). XXD may improve the spatial learning and memory ability of AD model SAMP8 by enhancing the neuroprotective effect and inhibiting neuroinflammation.
Humans ; Neuroprotective Agents/therapeutic use* ; Sirtuin 1/metabolism* ; Toll-Like Receptor 2/metabolism* ; NF-kappa B/metabolism* ; Tumor Necrosis Factor-alpha/metabolism* ; Neuroinflammatory Diseases ; NLR Family, Pyrin Domain-Containing 3 Protein/metabolism* ; Toll-Like Receptor 1/metabolism* ; Alzheimer Disease/genetics* ; Hippocampus

This study aims to investigate the role of slient mating-type information regulation 2 homolog 1(SIRT1)/tuberous sclerosis complex 2(TSC2)/mammalian target of rapamycin(mTOR) signaling pathways in the Periplaneta americana extract CⅡ-3-induced senescence of human leukemia K562 cells. K562 cells were cultured in vitro and treated with 0(control), 5, 10, 20, 40, 80, and 160 μg·mL~(-1) of P. americana extract CⅡ-3. Cell counting kit-8(CCK-8) and flow cytometry were employed to examine the proliferation and cell cycle of the K562 cells. Senescence-associated β-galactosidase stain kit(SA-β-gal) was used to detect the positive rate of senescent cells. Mitochondrial membrane potential was detected by flow cytometry. The relative mRNA level of telomerase reverse transcriptase(TERT) was determined by fluorescence quantitative PCR. The mRNA and protein levels of SIRT1, TSC2, and mTOR were determined by fluorescence quantitative PCR and Western blot, respectively. The results showed that CⅡ-3 significantly inhibited the proliferation of K562 cells and the treatment with 80 μg·mL~(-1) CⅡ-3 for 72 h had the highest inhibition rate. Therefore, 80 μg·mL~(-1) CⅡ-3 treatment for 72 h was selected as the standard for subsequent experiments. Compared with the control group, CⅡ-3 increased the proportion of cells arrested in G_0/G_1 phase, decreased the proportion of cells in S phase, increased the positive rate of SA-β-Gal staining, elevated the mitochondrial membrane potential and down-regulated the mRNA expression of TERT. Furthermore, the mRNA expression of SIRT1 and TSC2 was down-regulated, while the mRNA expression of mTOR was up-regulated. The protein expression of SIRT1 and p-TSC2 was down-regulated, while the protein expression of p-mTOR was up-regulated. The results indicated that P. americana extract CⅡ-3 induced the senescence of K562 cells via the SIRT1/mTOR signaling pathway.
Humans ; Animals ; Periplaneta ; Sirtuin 1/genetics* ; K562 Cells ; Signal Transduction ; TOR Serine-Threonine Kinases/genetics* ; RNA, Messenger ; Mammals