Lactiplantibacillus plantarum ATCC8014 Alleviates Postmenopausal Hypercholesterolemia in Mice by Remodeling Intestinal Microbiota to Increase Secondary Bile Acid Excretion.

Hypercholesterolemia poses a significant cardiovascular risk, particularly in postmenopausal women. The anti-hypercholesterolemic properties of Lactiplantibacillus plantarum ATCC8014 (LP) are well recognized; however, its improving symptoms on postmenopausal hypercholesterolemia and the possible mechanisms have yet to be elucidated. Here, we utilized female ApoE-deficient (ApoE-/-) mice undergoing bilateral ovariectomy, fed a high-fat diet, and administered 109 colony-forming units (CFU) of LP for 13 consecutive weeks. LP intervention reduces total cholesterol (TC) and triglyceride (TG) accumulation in the serum and liver and accelerates their fecal excretion, which is mainly accomplished by increasing the excretion of fecal secondary bile acids (BAs), thereby facilitating cholesterol conversion. Correlation analysis revealed that lithocholic acid (LCA) is an important regulator of postmenopausal lipid abnormalities. LP can reduce LCA accumulation in the liver and serum while enhancing its fecal excretion, accomplished by elevating the relative abundances of Allobaculum and Olsenella in the ileum. Our findings demonstrate that postmenopausal lipid dysfunction is accompanied by abnormalities in BA metabolism and dysbiosis of the intestinal microbiota. LP holds therapeutic potential for postmenopausal hypercholesterolemia. Its effectiveness in ameliorating lipid dysregulation is primarily achieved through reshaping the diversity and abundance of the intestinal microbiota to correct BA abnormalities.

Application of CRISPR/Cas13a-based biosensors in serum marker detection.

The detection of serum markers is important for the early diagnosis and monitoring of diseases, but conventional detection methods have the problem of low specificity or sensitivity. CRISPR/Cas13a-based biosensors have the characteristics of simple detection methods and high sensitivity, which have a certain potential to solve the problems of conventional detection. This paper focuses on the research progress of CRISPR/Cas13a-based biosensors in serum marker detection, introduces the principles and applications of fluorescence, electrochemistry, colorimetric, and other biosensors based on CRISPR/Cas13a in the detection of serum markers, compares and analyzes the differences between the above CRISPR/Cas13a-based biosensors, and looks forward to the future development direction of CRISPR/Cas13a-based biosensors.

Attenuation of estrogen and its receptors in the post-menopausal stage exacerbates dyslipidemia and leads to cognitive impairment.

Cognitive dysfunction increases as menopause progresses. We previously found that estrogen receptors (ERs) contribute to dyslipidemia, but the specific relationship between ERs, dyslipidemia and cognitive dysfunction remains poorly understood. In the present study, we analyzed sequencing data from female hippocampus and normal breast aspirate samples from normal and Alzheimer's disease (AD) women, and the results suggest that abnormal ERs signaling is associated with dyslipidemia and cognitive dysfunction. We replicated a mouse model of dyslipidemia and postmenopausal status in LDLR-/- mice and treated them with ß-estradiol or simvastatin, and found that ovariectomy in LDLR-/- mice led to an exacerbation of dyslipidemia and increased hippocampal apoptosis and cognitive impairment, which were associated with reduced estradiol levels and ERα, ERß and GPER expression. In vitro, a lipid overload model of SH-SY-5Y cells was established and treated with inhibitors of ERs. ß-estradiol or simvastatin effectively attenuated dyslipidemia-induced neuronal apoptosis via upregulation of ERs, whereas ERα, ERß and GPER inhibitors together abolished the protective effect of simvastatin on lipid-induced neuronal apoptosis. We conclude that decreased estrogen and its receptor function in the postmenopausal stage promote neuronal damage and cognitive impairment by exacerbating dyslipidemia, and that estrogen supplementation or lipid lowering is an effective way to ameliorate hippocampal damage and cognitive dysfunction via upregulation of ERs.

Salvianolic acid B promotes angiogenesis and inhibits cardiomyocyte apoptosis by regulating autophagy in myocardial ischemia.

BACKGROUND: Myocardial ischemia (MI) can cause angina, myocardial infarction, and even death. Angiogenesis is beneficial for ensuring oxygen and blood supply to ischemic tissue, promoting tissue repair, and reducing cell damage. In this study, we evaluated the effects of Salvianolic acid B (Sal B) against myocardial ischemia and explored its underlying mechanism on autophagy. METHODS: The anti-apoptosis effect of Sal B was conducted by staining Annexin V-FITC/PI and Hoechst as well as evaluating apoptosis bio-markers at protein level in H9c2 cells at glucose deprivation condition. HUVECs were co-cultured with H9c2, and the tube formation assay was used to monitor Sal B's impact on angiogenesis. The MI model of mice was induced by intraperitoneal injection of isoproterenol (ISO). The effect of Sal B on MI mice was evaluated by HE, Masson, immunohistochemistry, WB and kits. In addition, Atg5 siRNA was applied to verify whether the protective effect of Sal B was regulated to autophagy. RESULTS: In H9c2, Sal B reduced the levels of lactate dehydrogenase (LDH), malondialdehyde (MDA) and reactive oxygen species (ROS), improved the levels of superoxide dismutase (SOD) and mitochondrial membrane potential, downregulated the expressions of Bax and cleaved-Caspase3, upregulated the expression of Bcl-2. Therefore, Sal B could significantly inhibit the damage of H9c2 caused by glucose deprivation. In the co-culture system of H9c2 and HUVECs, vascular endothelial growth factor (VEGF) level in the supernatant was dramatically raised by Sal B. Sal B upregulated the expressions of VEGF, platelet derived growth factor (PDGF) and endothelial marker CD31. It implied that Sal B exerted a significant pro-angiogenic effect. Moreover, Sal B increased the expression of LC3, Atg5, and Beclin1, while reducing the level of P62. When the expression of Atg5 was inhibited, the protective effects of Sal B on apoptosis and angiogenesis was reversed. CONCLUSIONS: Sal B inhibited cardiomyocyte apoptosis and promoted angiogenesis by regulating autophagy, thereby improving MI.

Excessive exogenous cholesterol activating intestinal LXRα-ABCA1/G5/G8 signaling pathway can not reverse atherosclerosis in ApoE-/- mice.

BACKGROUND: The long-term excessive intake of exogenous cholesterol can lead to abnormally elevated blood lipid levels and induce cardiovascular and cerebrovascular diseases. However, the influence and relevance of exogenous cholesterol on plasma cholesterol components were still unclear, and the influence on intestinal lipid metabolism targets needs to be further explored. METHODS: In vivo, the C57BL/6 + NF group and ApoE-/- + NF group mice were fed a normal specific pathogen-free (SPF) diet; the ApoE-/- + HF group mice were fed a high-cholesterol SPF diet. The plasma and jejunum tissue homogenate were obtained for non-targeted lipid metabolomics. The lipid droplets in tissues were observed by transmission electron microscope and oil red O staining. Jejunum tissue morphology was observed by HE staining. The kits were used to detect lipid content in plasma, tissues, intestinal contents, and cells. Western blot, RT-PCR, immunohistochemistry (IHC), and immunofluorescence (IF) were used to observe the key target of lipid metabolism. In vitro, the final concentration of cholesterol was 100 µmol/L in Caco-cells. Oil red O staining, western blot, RT-PCR and immunofluorescence (IF) were used to observe the changes of lipid metabolism. Finally, the influence of liver X receptor alpha (LXRα) on intestinal cholesterol metabolism was clarified by applying the LXRα inhibitor GSK2033 and siRNA targeting LXRα. RESULTS: The aortic arch and intestinal villi of the two groups of ApoE-/- mice showed apparent lesions and lipid accumulation, and there were significant changes in a variety of lipids in the plasma and jejunum. Additionally, jejunum LXRα was markedly activated. High cholesterol can significantly activate LXRα in Caco-2 cells. After LXRα was inhibited, the protein level of ATP-binding cassette transporter A1/G5/G8 (ABCA1/G5/G8) decreased, and the quantity and volume of intracellular lipids soared. CONCLUSION: In a high-cholesterol environment, the intestine promotes the excretion of cholesterol from the cell through the LXRα-ABCA1/G5/G8 pathway, reduces the intestinal intake of a variety of exogenous cholesterol, and reduces the risk of AS.

Activation of estrogen receptor α inhibits TLR4 signaling in macrophages and alleviates the instability of atherosclerotic plaques in the postmenopausal stage.

Acute cardiovascular events increase significantly in postmenopausal women. The relationship between estrogen receptor (ER) and plaque stability in the postmenopausal stage remains to be elucidated. We aimed to explore whether ERα activation improves plaque instability in the postmenopausal stage. Here, we report that postmenopausal women showed increased macrophage activation and plaque instability with increased MCP-1, MMP9, TLR4, MYD88 and NF-κB p65 and decreased ERα and TIMP1 expression in the vascular endothelium. Moreover, ovariectomy in LDLR-/- mice resulted in a significant increase in plaque area and necrotic core area, as well as a significant decrease in collagen content and an increase in macrophage accumulation in the artery. Ovariectomy also reduced serum estrogen levels and ERα expression and upregulated TLR4 and MMP9 expression in arteries in LDLR-/- mice. Estrogen or phytoestrogen therapy upregulated the expression level of ERα in ovariectomized mice and increased plaque stability by inhibiting macrophage accumulation and TLR4 signaling. In vitro, LPS incubation of RAW264.7 cells resulted in a significant decrease in ERα and TIMP1 expression and an increase in TLR4 activation, and estrogen or phytoestrogen treatment increased ERα and TIMP1 expression and inhibited TLR4 activation and MMP9 expression in LPS-treated RAW264.7 cells. Compared to control siRNA transfected RAW264.7 cells, TLR4 siRNA promoted TIMP1 expression in RAW264.7 cells with LPS incubation, but did not affect ERα expression in RAW264.7 cells with or without LPS treatment. The ERα inhibitor MPP abolished the regulatory effect of estrogen or phytoestrogen on LPS-induced RAW264.7 cells. In conclusion, the present study demonstrates that decreased ERα expression promotes macrophage infiltration and plaque instability in the postmenopausal stage, and activation of ERα in the postmenopausal stage alleviates atherosclerotic plaque instability by inhibiting TLR4 signaling and macrophage-related inflammation.

Pinellia ternata lectin induces inflammation through TLR4 receptor and mediates PI3K/Akt/mTOR axis to regulate NF-κB signaling pathway.

Pinellia ternata, a widely used traditional Chinese medicine, contains a strong mucosal irritant that is connected with Pinellia ternata lectin (PTL) in its tubers. The purpose of this study was to explore the mechanisms by which PTL induces inflammation. We found that in RAW264.7 cells, PTL activated the PI3K/Akt/mTOR and NF-κB pathways, which resulted in the release of proinflammatory cytokines. Flow cytometry and laser confocal microscopy analysis showed that FITC-labeled PTL bound to the macrophages' surface. Based on kinetic analyses and protein-protein docking simulations, PTL was shown to bind toll-like receptor 4 (TLR4).it was demonstrated that PTL binds highly to Toll-like receptor 4 (TLR4). TLR4 knock-down or knockout resulted in a decrease in both cytokine release and PI3K/Akt/mTOR and NF-κB pathway activation in PTL-stimulated macrophages or mice. RNA-seq analysis showed that genes involved in the PI3K/Akt/mTOR signaling pathway were strongly upregulated in response to PTL stimulation, confirming that the PI3K/Akt/mTOR pathway is linked to the inflammatory effect of PTL in RAW264.7 cells. These findings reveal that PTL can mediate inflammation through TLR4 and activating the PI3K/Akt/mTOR to regulate NF-κB signaling pathways.

Continuous treatment with abemaciclib leads to sustained and efficient inhibition of breast cancer cell proliferation.

Abemaciclib is an oral, selective cyclin-dependent kinase 4 & 6 inhibitor (CDK4 & 6i), approved for hormone receptor-positive (HR+), human epidermal growth factor receptor 2-negative (HER2-) advanced breast cancer (ABC) as monotherapy for endocrine refractory disease, and with endocrine therapy (ET) for initial treatment and after progression on ET. Abemaciclib has also shown clinical activity in combination with ET in patients with high risk early BC (EBC). Here, we examined the preclinical attributes of abemaciclib and other CDK4 & 6i using biochemical and cell-based assays. In vitro, abemaciclib preferentially inhibited CDK4 kinase activity versus CDK6, resulting in inhibition of cell proliferation in a panel of BC cell lines with higher average potency than palbociclib or ribociclib. Abemaciclib showed activity regardless of HER2 amplification and phosphatidylinositol 3-kinase (PI3KCA) gene mutation status. In human bone marrow progenitor cells, abemaciclib showed lower impact on myeloid maturation than other CDK4 & 6i when tested at unbound concentrations similar to those observed in clinical trials. Continuous abemaciclib treatment provided profound inhibition of cell proliferation, and triggered senescence and apoptosis. These preclinical results support the unique efficacy and safety profile of abemaciclib observed in clinical trials.

Alisol B 23-acetate adjusts bile acid metabolisim via hepatic FXR-BSEP signaling activation to alleviate atherosclerosis.

BACKGROUND: Postmenopausal women have a high incidence of atherosclerosis. Phytosterols have been shown to have cholesterol-lowering properties. Alisa B 23-acetate (AB23A) is a biologically active plant sterol isolated from Chinese herbal medicine Alisma. However, the atherosclerosis effect of AB23A after menopause and its possible mechanism have not been reported yet. PURPOSE: To explore whether AB23A can prevent atherosclerosis by regulating farnesoid X receptor and subsequently increasing fecal bile acid and cholesterol excretion to reduce plasma cholesterol levels. METHODS: Aortic samples from premenopausal and postmenopausal women with ascending aortic arteriosclerosis were analyzed, and bilateral ovariectomized (OVX) female LDLR-/- mice and free fatty acid (FFA)-treated L02 cells were used to analyze the effect of AB23A supplementation therapy. RESULTS: AB23A increased fecal cholesterol and bile acids (BAs) excretion dependent on activation of hepatic farnesoid X receptor (FXR) in ovariectomized mice. AB23A inhibited hepatic cholesterol 7α-hydroxylase (CYP7A1) and sterol 12α-hydroxylase (CYP8B1) via inducing small heterodimer partner (SHP) expression. On the other hand, AB23A increased the level of hepatic chenodeoxycholic acid (CDCA), and activated the hepatic BSEP signaling. The activation of hepatic FXR-BSEP signaling by AB23A in ovariectomized mice was accompanied by the reduction of liver cholesterol, hepatic lipolysis, and bile acids efflux, and reduced the damage of atherosclerosis. In vitro, AB23A fixed abnormal lipid metabolism in L02 cells and increased the expression of FXR, BSEP and SHP. Moreover, the inhibition and silencing of FXR canceled the regulation of BSEP by AB23A in L02 cells. CONCLUSION: Our results shed light into the mechanisms behind the cholesterol-lowering of AB23A, and increasing FXR-BSEP signaling by AB23A may be a potential postmenopausal atherosclerosis therapy.

Catalpol Inhibits Macrophage Polarization and Prevents Postmenopausal Atherosclerosis Through Regulating Estrogen Receptor Alpha.

Lacking estrogen increases the risk of atherosclerosis (AS) in postmenopausal women. Inflammation plays a vital role in the pathological process of AS, and macrophages are closely related to inflammation. Catalpol is an iridoid glucoside extracted from the fresh roots of the traditional Chinese herb Rehmanniae radix preparata. In this study, we aimed to evaluate the effects of catalpol on macrophage polarization and postmenopausal AS. In addition, we investigated whether the mechanism of catalpol was dependent on regulating the expression of estrogen receptors (ERs). In vitro, lipopolysaccharides (LPS) and interferon-γ (IFN-γ) were applied to induce M1 macrophage polarization. In vivo, the ApoE-/- mice were fed with a high-fat diet to induce AS, and ovariectomy was operated to mimic the estrogen cessation. We demonstrated catalpol inhibited M1 macrophage polarization induced by LPS and INF-γ, and eliminated lipid accumulation in postmenopausal AS mice. Catalpol not only suppressed the inflammatory response but also reduced the level of oxidative stress. Then, ERs (ERα and ERß) inhibitors and ERα siRNA were also applied in confirming that the protective effect of catalpol was mediated by ERα, rather than ERß. In conclusion, catalpol significantly inhibited macrophage polarization and prevented postmenopausal AS by increasing ERα expression.

Liuwei Dihuang formula protect endothelial cells from apoptosis by up-regulating expression of estrogen receptor-α.

OBJECTIVE: To evaluate the efficacy of Liuwei Dihuang formula ( LWDHF) on endothelial cells, and to study the mechanism behind the action of modulating expression of estrogen receptors. METHODS: Hydrogen peroxide (H2O2) was applied to induce the apoptosis of human umbilical vein endothelial cells (HUVECs). The concentration of nitric oxide (NO), endothelial nitric oxide synthase (eNOS) and inducible NOS (iNOS) were measured by assay kits. Western blot and real-time polymerase chain reaction (RT-PCR) were used to detect the expression of iNOS, eNOS, b-cell lymphoma-2 (Bcl-2), Bcl-2-associated X protein (Bax), estrogen receptor (ER) α and ERß. Also, small interfering RNA (siRNA) was involved to confirm whether the protective effects of LWDHF was medicated by ERs. In vivo, the female rats were ovariectomized to establish postmenopausal vascular injury model. Then the model rats were divided into three groups and treated with saline, estradiol and LWDHF respectively. The concentration of NO and NOS in serum were measured by assay kits, and the expression of Bax, Bcl-2, ERα and ERß were detected by western blot and immunohistochemistry. RESULTS: In vitro study, LWDHF significantly protected HUVECs from H2O2-induced apoptosis, with the increase of Bcl-2 and the decrease of Bax. The treatment with LWDHF inhibited concentration of NO and iNOS, and upregulated the expression of eNOS, ERα and ERß. In addition, ERα siRNA could block the protective effects of LWDHF, while ERß siRNA showed little influence. In vivo, the treatment with LWDHF suppressed the vascular injury and reduced the level of NO and NOS. LWDHF increased the expression of Bcl-2, ERα and ERß, as well as inhibiting the Bax expression. CONCLUSION: LWDHF could improve endothelial function and protect HUVECs from apoptosis via increasing the expression of ERα.

[Analysis of the Efficacy of Irinotecan in the Second-line Treatment of Refractory and Relapsed Small Cell Lung Cancer].

BACKGROUND: Among malignant tumors, lung cancer has the highest mortality rate. Small cell lung cancer (SCLC) is a kind of malignant lung cancer. Its doubling time is very fast. Patients are prone to drug resistance during treatment, and their condition often deteriorates rapidly after recurrence. Except for topotecan, there is a lack of effective second-line single-agent chemotherapy. This study aims to analysis the efficacy and safety of irinotecan (CPT-11) in the second-line treatment of refractory and relapsed SCLC. METHODS: A total of 107 SCLC patients were collected from the Department of Oncology, Jilin Guowen Hospital, who were diagnosed from April 2012 to March 2020, relapsed within 6 months after first-line treatment, and received second-line chemotherapy with single-agent CPT-11. Follow-up until November 2020, calculate the patient's progression free survival (PFS) and overall survival (OS), and summarize the effects and adverse reactions of CPT-11 chemotherapy. RESULTS: The patient's median PFS was 3.8 (3.4-4.4) months, median OS was 8.1 (6.5-10.9) months, objective response rate (ORR) was 16.82% (18/107), and DCR was 55.14% (59/107). The incidence of grade 3-4 adverse reactions in patients was relatively low. Among them, neutropenia was 13.08%, delayed diarrhea was 7.48%, nausea and vomiting was 17.76%, and liver function impairment was 6.54%. The influencing factors of PFS in single-agent CPT-11 second-line chemotherapy were gender (P=0.001), NSE (P=0.029), and effusion (P=0.040). While the influencing factors of OS were NSE level only (P=0.033). CONCLUSIONS: For patients with refractory relapsed SCLC, CPT-11 single-agent second-line chemotherapy has a certain effect, is well tolerated, and is worthy of promotion.
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The gut microbiota during the progression of atherosclerosis in the perimenopausal period shows specific compositional changes and significant correlations with circulating lipid metabolites.

Atherosclerosis (AS) is exacerbated in the perimenopausal period, which significantly increases the incidence rate of cardiovascular disease. The disruption of the gut microbiota has been associated with AS or menopause, but the specific changes of AS-associated gut microbiota in the perimenopausal period remain largely unknown. As lipid abnormalities are mainly responsible for AS, the relationship between lipid metabolism abnormalities and gut microbiota disruptions during menopause is rarely reported hitherto. In the present study, ApoE-/- mice fed with a high-fat diet (HFD) were subjected to ovariectomy and supplemented with estrogen. The ovariectomized HFD-fed ApoE-/- mice underwent significant AS damage, hepatic lipid damage, hyperlipidemia, and changes of lipid metabolism- and transport-related enzymes. There was significantly higher abundance of some lipid metabolites in the plasma of ovariectomized HFD-fed ApoE-/- mice than in non-ovariectomized ones, including cholesterol esters, triglycerides, phospholipids, and other types of lipids (free fatty acids, acylcarnitine, sphingomyelins, and ceramides). The administration of estrogen significantly reduced the contents of most lipid metabolites. The diversity and composition of gut microbiota evidently changed in ovariectomized HFD-fed ApoE-/- mice, compared to HFD-fed ApoE-/- mice without ovariectomy. In contrast, with estrogen supplementation, the diversity and composition of gut microbiota were restored to approach that of non-ovariectomized HFD-fed ApoE-/- mice, and the relative abundances of some bacteria were even like those of C57BL/6 mice fed with a normal diet. On the other hand, the transplantation of feces from C57BL/6 mice fed with normal diet to ovariectomized HFD-fed ApoE-/- mice was sufficient to correct the hyperlipidemia and AS damage, and to reverse the characteristics changing of lipid metabolomics in ovariectomized HFD-fed ApoE-/- mice. These phenomena were also been observed after transplantation of feces from estrogen-treated ovariectomized HFD-fed ApoE-/- mice to ovariectomized HFD-fed ApoE-/- mice. Moreover, the gut microbiota and lipid metabolites were significantly correlated, demonstrating that the changes of serum lipids may be associated with the gut microbiota disruptions in the perimenopausal period. In conclusion, the gut microbiota during the progression of AS in the perimenopausal period showed specific compositional changes and significant correlations with circulating lipid metabolites. Estrogen supplementation may exert beneficial effects on gut bacteria and lipid metabolism.

Novel insights: Dynamic foam cells derived from the macrophage in atherosclerosis.

Atherosclerosis can be regarded as a chronic disease derived from the interaction between disordered lipoproteins and an unsuitable immune response. The evolution of foam cells is not only a significant pathological change in the early stage of atherosclerosis but also a key stage in the occurrence and development of atherosclerosis. The formation of foam cells is mainly caused by the imbalance among lipids uptake, lipids treatment, and reverse cholesterol transport. Although a large number of studies have summarized the source of foam cells and the mechanism of foam cells formation, we propose a new idea about foam cells in atherosclerosis. Rather than an isolated microenvironment, the macrophage multiple lipid uptake pathways, lipid internalization, lysosome, mitochondria, endoplasmic reticulum, neutral cholesterol ester hydrolase (NCEH), acyl-coenzyme A-cholesterol acyltransferase (ACAT), and reverse cholesterol transport are mutually influential, and form a dynamic process under multi-factor regulation. The macrophage takes on different uptake lipid statuses depending on multiple uptake pathways and intracellular lipids, lipid metabolites versus pro-inflammatory factors. Except for NCEH and ACAT, the lipid internalization of macrophages also depends on multicellular organelles including the lysosome, mitochondria, and endoplasmic reticulum, which are associated with each other. A dynamic balance between esterification and hydrolysis of cholesterol for macrophages is essential for physiology and pathology. Therefore, we propose that the foam cell in the process of atherosclerosis may be dynamic under multi-factor regulation, and collate this study to provide a holistic and dynamic idea of the foam cell.

Estrogen prevent atherosclerosis by attenuating endothelial cell pyroptosis via activation of estrogen receptor α-mediated autophagy.

Excessive inflammation and the pyroptosis of vascular endothelial cells caused by estrogen deficiency is one cause of atherosclerosis in post-menopausal women. Because autophagy is highly regulated by estrogen, we hypothesized that estrogen can reduce vascular endothelial cell pyroptosis through estrogen receptor alpha (ERα)-mediated activation of autophagy to improve atherosclerosis in post-menopausal stage. Aortic samples from pro-menopausal and post-menopausal women with ascending aortic arteriosclerosis were analyzed, and bilateral ovariectomized (OVX) female ApoE-/- mice and homocysteine (Hcy)-treated HUVECs were used to analyze the effect of estrogen supplementation therapy. The aortic endothelium showed a decrease in ERα expression and autophagy, but presented an increase in inflammation and pyroptosis in female post-menopausal patients. Estrogen treatment accelerated autophagy and ameliorated cell pyroptosis in the cardiac aortas of OVX ApoE-/- mice and Hcy-treated HUVECs. Estrogen had therapeutic effect on atherosclerosis and improved the symptoms associated with lipid metabolism disorders in OVX ApoE-/- mice. Inhibition and silencing of ERα led to a reduction in the autophagy promoting ability of estrogen and aggravated pyroptosis. Moreover, the inhibition of autophagy promoted pyroptosis and abolished the protective effect of estrogen, but had no influence on ERα expression. Thus, the results of the present study demonstrated that post-menopausal women present decreased autophagy and ERα expression and excessive damage to the ascending aorta. In addition, in vitro and in vivo assay results demonstrated that estrogen prevents atherosclerosis by upregulating ERα expression and subsequently induces autophagy to reduce inflammation and pyroptosis.

Alisol B 23-acetate activates ABCG5/G8 in the jejunum via the LXRα/ACAT2 pathway to relieve atherosclerosis in ovariectomized ApoE-/- mice.

Phytosterols have been shown to improve blood lipid levels and treat atherosclerosis. This research investigated the effects of phytosterol Alisol B 23-acetate (AB23A) on jejunum lipid metabolism and atherosclerosis. The results show that intragastric administration of AB23A can significantly reduce atherosclerotic plaque area and lipid accumulation in the jejunum of ovariectomized ApoE-/- mice fed a high-fat diet and can also improve the lipid mass spectra of the plasma and jejunum. In vitro studies have shown that AB23A can increase cholesterol outflow in Caco-2 cells exposed to high fat concentrations and increase the expression of ATP-binding cassette transfer proteins G5/G8 (ABCG5/G8), the liver X receptor α (LXRα). Furthermore, inhibition of LXRα can significantly eliminate the active effect of AB23A on decreasing intracellular lipid accumulation. We also confirmed that AB23A has a negative effect on Acyl-CoA cholesterol acyltransferase 2 (ACAT2) in Caco-2 cells cultured in the high concentrations of fat, and we found that AB23A further reduces ACAT2 expression in cells treated with the ACAT2 inhibitor pyripyropene or transfected with ACAT2 siRNA. In conclusion, we confirmed that AB23A can reduce the absorption of dietary lipids in the jejunum by affecting the LXRα-ACAT2-ABCG5/G8 pathway and ultimately exert an anti-atherosclerotic effect.

Activation of estrogen receptor α (ERα) is required for Alisol B23-acetate to prevent post-menopausal atherosclerosis and reduced lipid accumulation.

The risk of atherosclerosis (AS) ascends among post-menopausal women, while current hormone replacement therapy exerts several adverse effects. Alisol B 23-acetate (AB23A), a tetracyclic triterpenoid isolated from the rhizome of Alisma orientale, was reported to show multiple physiological activities, including regulating lipid metabolism. According to molecular docking analysis, it was predicted to bind with estrogen receptor α (ERα). In this study, we aimed to observe the effect of AB23A on preventing post-menopausal AS and explore whether the mechanism was mediated by ERα. In vitro, free fatty acid (FFA) was applied to induce the abnormal lipid metabolism of L02 cells. In vivo, the ApoE-/- mice were ovariectomized to mimic the cessation of estrogen. The high-fat diet was also given to induce post-menopausal AS. We demonstrated AB23A attenuated the accumulation of total cholesterol and triglyceride induced by free fatty acids in hepatocytes. In high-fat diet-ovariectomy-treated ApoE-/- mice, AB23A eliminated lipids in blood and liver. AB23A not only reduced the synthesis of proprotein convertase subtilisin/kexin type 9 (PCSK9) through sterol-regulatory element binding proteins (SREBPs) but also suppressed the secretion of PCSK9 through silent information regulator 1 (SIRT1). Notably, AB23A promoted the expression of ERα in vivo and in vitro. The both ERα inhibitor and ERα siRNA were also applied in confirming whether the hepatic protective effect of AB23A was mediated by ERα. We found that AB23A significantly promoted the expression of ERα. AB23A could inhibit the synthesis and secretion of PCSK9 through ERα, lower the accumulation of triglyceride and cholesterol, and prevent post-menopausal AS.

ß-estradiol adjusts intestinal function via ERß and GPR30 mediated PI3K/AKT signaling activation to alleviate postmenopausal dyslipidemia.

Decreases in estrogen secretion and estrogen receptor function lead to an increase in the incidence of dyslipidemia and cardiovascular disease (CVD) in postmenopausal women. We previously reported that ß-estradiol has a significant regulatory effect on lipids in ApoE-/- mice with bilateral ovariectomy. In the present study, we investigated how ß-estradiol regulates intestinal function via estrogen receptors to alleviate postmenopausal dyslipidemia. Ovariectomized ApoE-/- mice were treated with ß-estradiol for 90 days, and we found that ß-estradiol reduced TC, TG, LDL-c, IL-1ß and IL-18 levels in serum and decreased lipid accumulation in the liver. ß-estradiol reduced injury and inflammation in the jejunum in ovariectomized mice, and promoted the expression of tight junction-related proteins. Moreover, ß-estradiol increased ERα, ERß, GPR30 and ABCG5 protein expression, and decreased the levels of NPC1L1 and SR-B1 in the jejunum of ovariectomized mice. In Caco-2 cells incubated with cholesterol, ß-estradiol up-regulated PI3K/AKT signaling, reduced cholesterol accumulation, suppressed inflammatory signaling, and increased the expression of tight junction-related proteins. ERß or GPR30 inhibition decreased the protective effect of ß-estradiol on cholesterol accumulation, tight junctions, and inflammation in cholesterol incubated Caco-2 cells, while silencing both ERß and GPR30 completely eliminated the protective effect of ß-estradiol. PI3K/AKT inhibition abolished the protective effect of ß-estradiol on cholesterol accumulation, tight junction-related protein expression, and inflammation, but had no influence on ERα, ERß or GPR30 expression in cholesterol incubated Caco-2 cells. Our results provide evidence that ß-estradiol regulates intestinal function via ERß and GPR30 mediated PI3K/AKT signaling activation to alleviate postmenopausal dyslipidemia.

The protective effects of liguzinediol on congestive heart failure induced by myocardial infarction and its relative mechanism.

BACKGROUND: Heart failure (HF) is one of the most common causes of cardiovascular diseases in the world. Currently, the drugs used to treat HF in the clinic may cause serious side effects. Liguzinediol, 2, 5-dimethyl-3, 6-dimethyl-pyrazine, is a compound synthesized after the structural modification of ligustrazine (one active ingredient of Szechwan Lovage Rhizome). We aimed to observe the effects of liguzinediol on preventing HF and explore the related mechanisms. METHODS: The ligation of left anterior descending coronary artery was operated to established the myocardial infarction (MI) model in Sprague-Dawley rats. Cardiac functions were recorded by echocardiography and hemodynamics. The changes in the Renin-Angiotensin-Aldosterone System (RAAS), inflammation, and oxidative stress were detected by radioimmunoassay and Elisa kits. Western blot and real-time PCR were applied to determine the expressions of the TGF-ß1/Smads pathway. RESULTS: Firstly, liguzinediol enhanced the systolic and diastolic functions of the heart in MI rats. Liguzinediol improved ventricular remodeling by reducing myocardial cell necrosis, as well as reducing collagen deposition and myocardial fibrosis. Then, liguzinediol suppressed the activation of RAAS, inhibited the synthesis of pro-inflammation factors, and reduced oxidative stress. In the end, liguzinediol also down-regulated the expressions of the TGF-ß1/Smads pathway. CONCLUSIONS: Liguzinediol could alleviate HF caused by MI in rats, and the protective effect was associated with the regulation of the TGF-ß1/Smads pathway.

Liuwei Dihuang prevents postmenopausal atherosclerosis and endothelial cell apoptosis via inhibiting DNMT1-medicated ERα methylation.

ETHNOPHARMACOLOGICAL RELEVANCE: The classical and traditional Chinese medicine prescription, Liuwei Dihuang (LWDH), has been commonly used to treat the menopausal syndrome. It has been reported that LWDH could improve estrogen receptor α (ERα) expression to prevent atherosclerosis (AS), while the mechanism of LWDH on regulating ERα expression was still unknown. AIM OF THE STUDY: To reveal the mechanism of LWDH on regulating the ERα expression. MATERIALS AND METHODS: The protective effect of LWDH on Hcy-induced apoptosis of human umbilical vein endothelial cells (HUVECs) was examined. The expression of ERα and DNA methyltransferases 1 (DNMT1) were detected by Western blot and real-time polymerase chain reaction (RT-PCR). The methylation rate of the ERα gene was assayed by the bisulfite sequencing PCR (BSP). High-performance liquid chromatography-tandem mass spectrometry (HPLC-MS) was applied to determine the level of S-Adenosyl methionine (SAM) and S-Adenosyl homocysteine (SAH). In vivo, the ApoE-/- mice were ovariectomized to establish postmenopausal atherosclerosis (AS) model. RESULTS: In vitro study showed that LWDH protects HUVECs from Hcy-induced apoptosis. Treatment with LWDH significantly increased the ERα expression and reduced the methylation rate of the ERα gene by inhibiting the DNMT1 expression. The level of main methyl donor SAM and the ration of SAM/SAH were reduced by LWDH. In vivo, LWDH prevented the formation of plaque and reduced the concentration of Hcy. In addition, LWDH upregulated the ERα expression, as well as inhibiting the expression of DNMT1 in atherosclerotic mice. CONCLUSIONS: LWDH exerted protective effects on postmenopausal AS mice, and HUVECs treated with Hcy. LWDH increased of ERα expression via inhibiting DNMT1-dependent ERα methylation.