Hydroxysafflor yellow A inhibits endothelial cell ferroptosis in diabetic atherosclerosis mice by regulating miR-429/SLC7A11.

CONTEXT: Ferroptosis may play an essential role in lipid peroxidation and endothelial dysfunction of aortic endothelial cells (ECs) in type 2 diabetes mellitus (T2DM) with atherosclerosis (AS). Hydroxysafflor yellow A (HSYA) has shown substantial antioxidant stress and anti-ferroptosis. OBJECTIVE: This study confirms whether HSYA improves symptoms in a mouse model of T2DM/AS and elucidates the underlying mechanisms. MATERIALS AND METHODS: ApoE-/- mice were fed with high fat combined with 30 mg/kg streptozotocin to establish a T2DM/AS model. Then mice were treated with intraperitoneal injections of 2.25 mg/kg HSYA for 12 weeks. Human Umbilical Vein Endothelial cells (HUVEC) induced by 33.3 mM d-glucose +100 µg/mL ox-LDL were used to construct a high lipid and high glucose cell model treated with 25 µM HSYA. The changes in oxidative stress- and ferroptosis-related markers were detected, and the regulatory effect of HSYA on the miR-429/SLC7A11 was also verified. Normal ApoE-/- mice or HUVEC cells were used as the control group. RESULTS: HSYA effectively reduced atherosclerotic plaque formation in the T2DM/AS mouse model and inhibited HUVEC ferroptosis, such as upregulating GSH-Px, SLC7A11 and GPX4, but inhibited ACSL4. Furthermore, HSYA also downregulated the expression of miR-429, which further regulated SLC7A11 expression. After miR-429 mimic or SLC7A11 siRNA transfection in the HUVEC, the antioxidative stress and anti-ferroptosis effects of HSYA were significantly abolished. CONCLUSIONS: HSYA is expected to become an important health drug to prevent the occurrence and development of T2DM/AS.

[Effects of Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination on inflammatory responses in atherosclerotic mice].

The study aims to observe the effects and explore the mechanisms of Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination in the treatment of the inflammatory response of mice with atherosclerosis(AS) via the Toll-like receptor 4(TLR4)/myeloid differentiation primary response protein 88(MyD88)/nuclear factor-κB(NF-κB) signaling pathway. Male ApoE~(-/-) mice were randomly assigned into a model group, a Buyang Huanwu Decoction group, an Astragali Radix-Angelicae Sinensis Radix combination group, and an atorvastatin group, and male C57BL/6J mice of the same weeks old were used as the control group. Other groups except the control group were given high-fat diets for 12 weeks to establish the AS model, and drugs were administrated by gavage. Aortic intimal hyperplasia thickness, blood lipid level, plasma inflammatory cytokine levels, M1/M2 macrophage markers, and expression levels of proteins in TLR4/MyD88/NF-κB pathway in the vessel wall were measured to evaluate the effects of drugs on AS lesions and inflammatory responses. The results showed that the AS model was successfully established with the ApoE~(-/-) mice fed with high-fat diets. Compared with the control group, the model group showed elevated plasma total cholesterol(TC), triglyceride(TG), and low-density lipoprotein cholesterol(LDL-c) levels(P<0.05), thickened intima(P<0.01), and increased plasma tumor necrosis factor-α(TNF-α) and interleukin-6(IL-6) levels(P<0.01). Moreover, the model group showed increased expression of vascular cell adhesion molecule-1(VCAM-1) and inducible nitric oxide synthase(iNOS)(P<0.01), inhibited expression of endothelial nitric oxide synthase(eNOS) and cluster of differentiation 206(CD206)(P<0.01), and up-regulated mRNA and protein levels of TLR4, MyD88, NF-κB inhibitor alpha(IκBα), and NF-κB in the vessel wall(P<0.05). Compared with the model group, Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination lowered the plasma TC and LDL-c levels(P<0.01), alleviated the intimal hyperplasia(P<0.01), and reduced the plasma TNF-α and IL-6 levels(P<0.05). Moreover, the two interventions promoted the expression of eNOS and CD206(P<0.05), inhibited the expression of VCAM-1 and iNOS(P<0.01), and down-regulated the mRNA and protein levels of TLR4, MyD88, IκBα, and NF-κB(P<0.05) in the vessel wall. This study indicated that Buyang Huanwu Decoction and Astragali Radix-Angelicae Sinensis Radix combination could delay the progression of AS, inhibit the polarization of vascular wall macrophages toward M1 type, and attenuate vascular inflammatory response by inhibiting the activation of TLR4/MyD88/NF-κB signaling pathway in the vascular wall. Astragali Radix and Angelicae Sinensis Radix were the main pharmacological substances in Buyang Huanwu Decoction for alleviating the AS vascular inflammatory response.

[Study on effect of "Trichosanthis Fructus-Allii Macrostemonis Bulbus" on atherosclerosis in ApoE~(-/-) mice based on liver metabonomics].

In this study, ultra-performance liquid chromatography coupled with quadrupole time-of-flight tandem mass spectrometry(UPLC-Q-TOF-MS)-based liver metabolomics approach was used to explore the mechanism of "Trichosanthis Fructus-Allii Macrostemonis Bulbus" in improving atherosclerosis(AS) of mice with apolipoprotein E gene knockout(ApoE~(-/-)). AS mouse model was induced by high-fat diet. The pathological and biochemical indexes such as the histopathological changes, body weight, liver weight, blood lipid level and inflammatory factors in the liver of mice were determined. The metabolic profiling of mice liver samples was performed with UPLC-Q-TOF-MS. Multiple statistical analysis methods including partial least squares discriminant analysis(PLS-DA) and orthogonal partial least squares discriminant analysis(OPLS-DA) were employed to screen and identify biomarkers. The levels of related enzymes including LCAT, sPLA2, EPT1 and ACER1 were detected. The results showed that "Trichosanthis Fructus-Allii Macrostemonis Bulbus" significantly reduced the areas of aortic plaque and fat vacuoles of liver in AS mice and decreased the accumulation of lipid droplets and liver coefficient. "Trichosanthis Fructus-Allii Macrostemonis Bulbus" also regulated the levels of blood lipid and inflammatory injury in the liver. The metabolites of the control group, the model group and the "Trichosanthis Fructus-Allii Macrostemonis Bulbus" group could be distinguished significantly. Fifteen potential biomarkers related to AS were discovered and preliminarily identified, seven of which could be regulated by "Trichosanthis Fructus-Allii Macrostemonis Bulbus" in a trend of returning to normal. Metabolic pathway analysis screened out two major metabolic pathways. "Trichosanthis Fructus-Allii Macrostemonis Bulbus" obviously regulated the levels of LCAT, sPLA2, EPT1 and ACER1. It was inferred that "Trichosanthis Fructus-Allii Macrostemonis Bulbus" could play a major role in AS treatment by regulating glycerophospholipid and sphingolipid metabolism disorders in the liver, with the mechanism probably relating to the intervention of the expression of LCAT, sPLA2, EPT1 and ACER1.

Immunohistochemical biomarkers and distribution of telocytes in ApoE-/- mice.

Telocytes had been identified as a peculiar stromal cell type implicated in tissue homeostasis and the development and pathophysiology of diseases. Telocyte existed in most organs and tissues in humans and animals. However, few studies have examined telocytes in ApoE gene deficient mice. In our studies, we verified the existence, the morphology and immunohistochemical characteristics of telocytes in critical organs of the ApoE-/- mice. Male adult ApoE-/- mice were selected as an experimental model. Immunohistochemical bio-markers, such as CD34, CD117, CD28, Vimentin and PDGFR-α were utilized to determine the distribution and morphology of telocytes in the heart, liver and kidney. Telocyte expressed positively for CD34 and CD117, and partial telocyte and telopode expressed positively for PDGFR-α in heart and liver, but negatively in kidney. Double immunofluorescence assays for CD28/Vimentin, CD34/CD117 and CD34/PDGFR-α were used to demonstrate the biochemistry speciality of telocytes, respectively. The evidence of telocytes in the ApoE-/- mice is the first step of our sturdy, which aims to demonstrate changes in telocytes in atherosclerosis in this animal model.

How radiation influences atherosclerotic plaque development: a biophysical approach in ApoE⁻/⁻ mice.

Atherosclerosis is the development of lipid-laden plaques in arteries and is nowadays considered as an inflammatory disease. It has been shown that high doses of ionizing radiation, as used in radiotherapy, can increase the risk of development or progression of atherosclerosis. To elucidate the effects of radiation on atherosclerosis, we propose a mathematical model to describe radiation-promoted plaque development. This model distinguishes itself from other models by combining plaque initiation and plaque growth, and by incorporating information from biological experiments. It is based on two consecutive processes: a probabilistic dose-dependent plaque initiation process, followed by deterministic plaque growth. As a proof of principle, experimental plaque size data from carotid arteries from irradiated ApoE[Formula: see text] mice was used to illustrate how this model can provide insight into the underlying biological processes. This analysis supports the promoting role for radiation in plaque initiation, but the model can easily be extended to include dose-related effects on plaque growth if available experimental data would point in that direction. Moreover, the model could assist in designing future biological experiments on this research topic. Additional biological data such as plaque size data from chronically-irradiated mice or experimental data sets with a larger variety in biological parameters can help to further unravel the influence of radiation on plaque development. To the authors' knowledge, this is the first biophysical model that combines probabilistic and mechanistic modeling which uses experimental data to investigate the influence of radiation on plaque development.

The Effects of Aerobic Exercise on Plasma Adiponectin Level and Adiponectin-related Protein Expression in Myocardial Tissue of ApoE(-/-) Mice.

Numerous reports have confirmed the effect of ApoE knockout in the induction of cardiovascular diseases and the protective effect of adiponectin against the progression of cardiovascular diseases. The aim of this study was to reveal the roles of adiponectin signaling in the progression of cardiovascular diseases induced by ApoE knockout and to analyze the healthy effects of aerobic exercise on ApoE knockout mice (ApoE(-/-) mice) through observing the changes of adiponectin signaling caused by ApoE knockout and aerobic exercise. A twelve-week aerobic exercise program was carried out on the male ApoE(-/-) mice and the C57BL / 6J mice (C57 mice) of the same strain. Results show that the body weights, blood lipid level, plasma adiponectin level and adiponectin-related proteins in myocardial tissue were all significantly changed by ApoE knockout. A twelve-week aerobic exercise program exerted only minimal effects on the body weights, blood lipid levels, and plasma adiponectin levels of ApoE(-/-) mice, but increased the expressions of four adiponectin-related proteins, AdipoR1, PPARα, AMPK and P-AMPK, in the myocardial tissue of the ApoE(-/-) mice. In summary, adiponectin signaling may play an import role in the progression of cardiovascular diseases induced by ApoE knockout, and the beneficial health effects of aerobic exercise on ApoE(-/-) mice may be mainly from the increased adiponectin-related protein expression in myocardial tissue. Key pointsA twelve-week aerobic exercise program exerted only limited effects on the body weights and the plasma adiponectin levels of both the normal mice and the ApoE(-/-) mice but did effectively regulate the blood lipid levels of the normal mice (but not the ApoE(-/-) mice).After 12 weeks of aerobic exercise, expression of the adiponectin-related proteins in the myocardial tissue of the ApoE(-/-) and normal mice was increased, but the increased amplitudes of these proteins in the ApoE(-/-) mice were much larger in the ApoE(-/-) mice than in the normal mice.Aerobic exercise might not alter the plasma adiponectin levels and blood lipid levels of ApoE(-/-) mice, but improve myocardial energy metabolism and relieve cardiovascular disease symptoms by increasing adiponectin-related protein expression in myocardial tissue.

A combined transcriptomics and proteomics approach to reveal the mechanism of AEE relieving hyperlipidemia in ApoE-/- mice.

Hyperlipidemia caused by abnormal lipid metabolism has reached epidemic proportions. This phenomenon is also common in companion animals. Previous studies showed that AEE significantly improves abnormal blood lipids in hyperlipidemia rats and mice, but its mechanism is still not clear enough. In this study, the mechanism and potential key pathways of AEE on improving hyperlipidemia in mice were investigated through the transcriptome and proteome study of ApoE-/- mice liver and the verification study on high-fat HepG2 cells. The results showed that AEE significantly decreased the serum TC and LDL-C levels of hyperlipidemia ApoE-/- mice, and significantly increased the enzyme activity of CYP7A1. After AEE intervention, the results of mice liver transcriptome and proteome showed that differential genes and proteins were enriched in lipid metabolism-related pathways. The results of RT-qPCR showed that AEE significantly regulated the expression of genes related to lipid metabolism in mice liver tissue. AEE significantly upregulated the protein expression of CYP7A1 in hyperlipidemia ApoE-/- mice liver tissue. The results in vitro showed that AEE significantly decreased the levels of TC and TG, and improved lipid deposition in high-fat HepG2 cells. AEE significantly increased the expression of CYP7A1 protein in high-fat HepG2 cells. AEE regulates the expression of genes related to lipid metabolism in high-fat HepG2 cells, mainly by FXR-SHP-CYP7A1 and FGF19-TFEB-CYP7A1 pathways. To sum up, AEE can significantly improve the hyperlipidemia status of ApoE-/- mice and the lipid deposition of high-fat HepG2 cells, and its main pathway is probably the bile acid metabolism-related pathway centered on CYP7A1.

DHA dietary intervention caused different hippocampal lipid and protein profile in ApoE-/- and C57BL/6J mice.

BACKGROUND: Changes in protein and lipid levels may occur in the Alzheimer's disease brain, and DHA can have beneficial effects on it. To investigate the impact of DHA dietary intervention on brain protein and lipid profile in ApoE-/- mice and C57 mice. METHOD: Three-month-old ApoE-/- mice and C57 mice were randomly divided into two groups respectively, and fed with control diet and DHA-fortified diet for five months. Cortical TC, HDL-C and LDL-C levels and cholesterol metabolism-related protein expression were measured by ELISA or immunohistochemistry methods. Hippocampus were collected for proteomic and lipidomics analysis by LC-MS/MS and differential proteins and lipid metabolites were screened and further analyzed by GO functional annotation and KEGG pathway enrichment analysis. RESULTS: DHA intervention decreased cortical TC level in both C57 and ApoE-/- mice (P < 0.05), but caused different change of cortical HDL-C, LDL-C level and LDL-C/HDL-C ratio in C57 and ApoE-/- mice (P < 0.05). Discrepant cortical and hippocampal LDLR, ABCG1, Lox1 and SORT1 protein expression was found between C57 and ApoE-/- mice (P < 0.05), and DHA treatment caused different changes of these proteins in C57 and ApoE-/- mice (P < 0.05). Differential hippocampal proteins and lipids profile were found in C57 and ApoE-/- mice before and after DHA treatment, which were mainly involved in vesicular transport and phospholipid metabolic pathways. CONCLUSION: ApoE genetic defect caused abnormal cholesterol metabolism, and affected protein and lipid profile, as well as discrepant response of hippocampal protein and lipids profile in the brain of mice given DHA fortified diet intervention.

Role of Apolipoprotein E in the Hippocampus and Its Impact following Ionizing Radiation Exposure.

Apolipoprotein E (ApoE) is a lipid carrier in both the peripheral and the central nervous systems (CNSs). Lipid-loaded ApoE lipoprotein particles bind to several cell surface receptors to support membrane homeostasis and brain injury repair. In the brain, ApoE is produced predominantly by astrocytes, but it is also abundantly expressed in most neurons of the CNS. In this study, we addressed the role of ApoE in the hippocampus in mice, focusing on its role in response to radiation injury. To this aim, 8-week-old, wild-type, and ApoE-deficient (ApoE-/-) female mice were acutely whole-body irradiated with 3 Gy of X-rays (0.89 Gy/min), then sacrificed 150 days post-irradiation. In addition, age-matching ApoE-/- females were chronically whole-body irradiated (20 mGy/d, cumulative dose of 3 Gy) for 150 days at the low dose-rate facility at the Institute of Environmental Sciences (IES), Rokkasho, Japan. To seek for ApoE-dependent modification during lineage progression from neural stem cells to neurons, we have evaluated the cellular composition of the dentate gyrus in unexposed and irradiated mice using stage-specific markers of adult neurogenesis. Our findings indicate that ApoE genetic inactivation markedly perturbs adult hippocampal neurogenesis in unexposed and irradiated mice. The effect of ApoE inactivation on the expression of a panel of miRNAs with an established role in hippocampal neurogenesis, as well as its transcriptional consequences in their target genes regulating neurogenic program, have also been analyzed. Our data show that the absence of ApoE-/- also influences synaptic functionality and integration by interfering with the regulation of mir-34a, mir-29b, and mir-128b, leading to the downregulation of synaptic markers PSD95 and synaptophysin mRNA. Finally, compared to acute irradiation, chronic exposure of ApoE null mice yields fewer consequences except for the increased microglia-mediated neuroinflammation. Exploring the function of ApoE in the hippocampus could have implications for developing therapeutic approaches to alleviate radiation-induced brain injury.

The Potential Association of TFR1/SLC11A2/GPX4 with Ferroptosis in Mediating Lipid Metabolism Disorders in Atherosclerosis.

PURPOSE: Atherosclerosis is the most common and significant form of arterial disease, characterized primarily by lipid accumulation and inflammatory cell infiltration as its main pathological basis. This study aims to investigate the molecular mechanisms and associated pathways by which iron accumulation may be involved in lipid metabolism abnormalities in atherosclerotic mice. METHODS: Relying on ApoE-/- mouse body position observation, blood biochemical analysis, oxidative stress test and aortic tissue sectioning techniques, the effects of ferroptosis on lipid metabolism in atherosclerotic mice were analyzed. Use RT-PCR analysis and transcriptomics tests to understand the specific molecular mechanism. RESULTS: Our analysis reveals a correlation between Ferroptosis and elevated levels of TC, TG, ALT, AST, IL-1ß, and TNF-α in the blood of atherosclerotic model mice. At the same time, it exacerbates the pathological changes of mouse aorta tissue. Our results suggest a potential link between ferroptosis and the dysregulation of TFR1/SLC11A2/GPX4 expression, along with the presence of oxidative stress, in the progression of AS. Transcriptomics results indicate that ferroptosis- mediated deterioration of atherosclerosis in ApoE-/- mice is potentially associated with cell phagocytosis, apoptosis involving TNF-α, and the expression of atherosclerotic and other process-related genes. CONCLUSION: Ferroptosis exacerbated the lipid metabolism disorder in atherosclerotic mice. The core mechanism of its effect is that ferroptosis activates the TFR1/SLC11A2/GPX4 signaling pathway, which leads to the up-regulation of oxidative stress in ApoE-/- mice, and ultimately aggravates the abnormal lipid metabolism in ApoE-/- mice.

The Chinese medicine Xin-tong-tai granule protects atherosclerosis by regulating oxidative stress through NOX/ROS/NF-κB signal pathway.

BACKGROUND: Xin-tong-tai Granule (XTTG), a traditional Chinese medicine, has been used to treat atherosclerosis (AS), but its mechanism is poorly understood. Intriguingly, oxidative stress has been recognized as vital factors in the treatment of atherosclerosis. PURPOSE: This study aims to explore the potential mechanism of XTTG for treating AS. METHODS: An in-vivo model of AS was established by feeding ApoE-/- mice with a high-fat diet (HFD), and the Human Aortic Vascular Smooth Muscle Cells (HAVSMCs) were induced by oxidized low-density lipoprotein (ox-LDL) in vitro. After treatment, the blood lipid levels and pathological aortic changes of each group were observed, and the cell proliferation and lipid droplet aggregation in each group were evaluated. The oxidative stress indicators such as malondialdehyde (MDA) and superoxide dismutase (SOD) levels and related NOX/ROS/NF-κB signaling pathway indicators were observed. RESULTS: XTTG improved blood lipid levels and pathological aortic changes of ApoE-/- mice with HFD feeding, inhibited HAVSMCs proliferation and lipid droplet aggregation induced by ox-LDL, reduced MDA content, increased SOD content, inhibited NOX4 and p22phox protein expression, downregulated ROS content, inhibited IKK-α, IKK-ß, NF-κB protein and mRNA expression and the phosphorylation of NF-κB. CONCLUSION: XTTG can inhibit NOX/ROS/NF-κB signaling pathway, reduce damages caused by oxidative stress, and exert anti-AS effects.

Untargeted lipidomics and metagenomics reveal the mechanism of aspirin eugenol ester relieving hyperlipidemia in ApoE-/- mice.

Hyperlipidemia is induced by abnormal lipid metabolism, which can cause the occurrence of cardiovascular diseases and lead to grievous injury to health. Studies showed that AEE had a significant therapeutic effect on hyperlipidemia and is likely to be associated with the up-regulation of cholesterol 7-alpha hydroxylase (CYP7A1), the key enzyme for cholesterol conversion to bile acids, but no research confirmed whether the effect of AEE on hyperlipidemia was related to the gut microbiota and liver lipids. At the same time, more and more studies have shown that gut microbiota and lipids are closely related to hyperlipidemia. Hence, in this study, we investigated the effects of AEE on liver lipids through LC-MS-based untargeted lipidomics and the effects of AEE on gut microbiota based on cecal contents metagenomics by Illumina sequencing in HFD-induced hyperlipidemia ApoE-/- mice at the overall level. The results of lipidomics showed that AEE relieved hyperlipidemia by decreasing the concentration of 10 PEs and 12 SMs in the liver and regulating the pathways of glycerophospholipid metabolic pathway, sphingolipid signaling pathway, and NF-kB signaling pathway. The results of metagenomics concluded that AEE treatment changed the composition of gut microbiota and regulated the functions of lipid transport and metabolism, as well as the metabolism of bile acids and secondary bile acids. The results of the joint analysis between lipidomics and metagenomics showed that the abundance of Verrucomicrobia, Verrucomicrobiales, Candidatus_Gastranaerophilales, and Candidatus_Melainabacteria was significantly positively correlated with the concentration of SM (d18:1/18:0) and PE (16:0/18:1) in the process of AEE alleviating hyperlipidemia in mice. In conclusion, these results suggested that the effect of AEE on hyperlipidemia was closely related to the gut microbiota by the change of bile acids and liver lipids.

Unraveling the molecular mechanisms of hyperlipidemia using integrated lncRNA and mRNA microarray data.

Long non-coding RNAs (lncRNAs) have key roles in various diseases; however, their functions in hyperlipidemia (HLP) have remained elusive. In the present study, microarray technology was utilized to analyze the differential expression of lncRNAs and mRNAs in liver tissues of apolipoprotein E-/- mice as a model of HLP compared with control mice. A total of 104 and 96 differentially expressed lncRNAs and mRNAs, respectively, were identified. Differentially expressed genes were significantly enriched in biological processes such as nitric oxide biosynthesis, innate immune response and inflammatory response. Finally, two pairs of target genes and 38 transcription factors with regulatory functions in HLP were predicted based on the lncRNA and mRNA co-expression network. The lncRNA expression profile was significantly altered in liver tissues of the mouse model of HLP and may provide novel targets for research into treatments.

Various bioactive peptides in collagen hydrolysate from salmo salar skin and the combined inhibitory effects on atherosclerosis in vitro and in vivo.

Atherosclerosis (AS) is the underlying condition in most cardiovascular diseases, which is blood vessel inflammation participated by many factors. Collagen hydrolysate from salmo salar skin (SCH) obtained in this study showed strong anti-inflammatory activity, protection of endothelial cell injury, antioxidant activity, and anti-platelet aggregation activity in vitro, exhibiting a great potential of attenuating AS. In this study, multifunctional peptides FAGPPGGDGQPGAK and IAGPAGPRGPSGPA, which mainly showed strong anti-inflammatory activity, were identified from SCH after separation of ultrafiltration and column chromatography. Moreover, SCH (contained anti-platelet peptides and anti-inflammatory peptides) was observed to inhibit arterial intima thickening and plaques formation in apolipoprotein E-deficient (ApoE-/-) mice fed with high-fat diets without side effects, exhibiting a comparable effect with aspirin. SCH showed combined effect on regulating serum biomarkers of inflammation (IL-6 and TNF-α), endothelial injury (MCP-1), platelet activation (TXB2 and PF4) and oxidative stress (MDA and CAT). This research suggested SCH as a potential dietary supplement for the primary prevention of AS.

Varying Dietary Component Ratios and Lingonberry Supplementation May Affect the Hippocampal Structure of ApoE-/- Mice.

OBJECTIVE: This study aimed to investigate and compare the morphological and biochemical characteristics of the hippocampus and the spatial memory of young adult ApoE-/- mice on a standard chow diet, a low-fat diet (LFD), a high-fat diet (HFD), and an HFD supplemented with lingonberries. METHODS: Eight-week-old ApoE-/- males were divided into five groups fed standard chow (Control), an LFD (LF), an HFD (HF), and an HFD supplemented with whole lingonberries (HF+WhLB) or the insoluble fraction of lingonberries (HF+InsLB) for 8 weeks. The hippocampal cellular structure was evaluated using light microscopy and immunohistochemistry; biochemical analysis and T-maze test were also performed. Structural synaptic plasticity was assessed using electron microscopy. RESULTS: ApoE-/- mice fed an LFD expressed a reduction in the number of intact CA1 pyramidal neurons compared with HF+InsLB animals and the 1.6-3.8-fold higher density of hyperchromic (damaged) hippocampal neurons relative to other groups. The LF group had also morphological and biochemical indications of astrogliosis. Meanwhile, both LFD- and HFD-fed mice demonstrated moderate microglial activation and a decline in synaptic density. The consumption of lingonberry supplements significantly reduced the microglia cell area, elevated the total number of synapses and multiple synapses, and increased postsynaptic density length in the hippocampus of ApoE-/- mice, as compared to an LFD and an HFD without lingonberries. CONCLUSION: Our results suggest that, in contrast to the inclusion of fats in a diet, increased starch amount (an LFD) and reduction of dietary fiber (an LFD/HFD) might be unfavorable for the hippocampal structure of young adult (16-week-old) male ApoE-/- mice. Lingonberries and their insoluble fraction seem to provide a neuroprotective effect on altered synaptic plasticity in ApoE-/- animals. Observed morphological changes in the hippocampus did not result in notable spatial memory decline.

Combination of folic acid with nifedipine is completely effective in attenuating aortic aneurysm formation as a novel oral medication.

Aortic aneurysms are prevalent and severe vascular diseases with high mortality from unpredicted ruptures, while the only treatment option is surgical correction of large aneurysms with considerable risk. We have shown that folic acid (FA) is highly effective in alleviating development of aneurysms although not sufficient to completely attenuate aneurysm formation. Here, we examined therapeutic effects on aneurysms of combining FA with Nifedipine as novel and potentially more effective oral medication. Oral administration with FA (15 mg/kg/day) significantly reduced incidence of AAA from 85.71% to 18.75% in Ang II-infused apolipoprotein E (apoE) null mice, while combination of FA with Nifedipine (1.5, 5.0 or 20 mg/kg/day) substantially and completely further reduced incidence of AAA to 12.5%, 11.76% and 0.00% respectively in a dose-dependent manner. The combinatory therapy substantially and completely further alleviated enlargement of abdominal aortas defined by ultrasound, vascular remodeling characterized by elastin degradation and adventitial hypertrophy, as well as aortic superoxide production and eNOS uncoupling activity also in a dose-dependent manner, with combination of FA with 20 mg/kg/day Nifedipine attenuating all of these features by 100% to control levels. Aortic NO and H4B bioavailabilities were also dose-dependently further improved by combining FA with Nifedipine. These data establish entirely innovative and robust therapeutic regime of FA combined with Nifedipine for the treatment of aortic aneurysms. The comminatory therapy can serve as a first-in-class and most effective oral medication for aortic aneurysms, which can be rapidly translated into clinical practice to revolutionize management of the devastating vascular diseases of aortic aneurysms known as silent killers.

Berberine attenuates atherosclerotic lesions and hepatic steatosis in ApoE-/- mice by down-regulating PCSK9 via ERK1/2 pathway.

BACKGROUND: It has been demonstrated that berberine (BBR), a kind of alkaloid derived from Chinese herbal medicine, has multiple pharmacological effects on human's diseases including anti-atherosclerosis action. However, although the previous studies showed that the beneficial impact of BBR on atherosclerosis might be associated with proprotein convertase subtilisin/kexin type 9 (PCSK9), the exact underlying mechanism are not fully determined. The present study aimed to investigate potential mechanisms of anti-atherosclerosis by BBR using ApoE-/- mice. METHODS: The eight-week mice were divided into five groups: group 1 (wild type C57BL/6J mice with normal diet), group 2 (ApoE-/- mice with normal diet), group 3 [ApoE-/- mice with high-fat diet (HFD)], group 4 (ApoE-/- mice with HFD, and treatment with low dose BBR of 50 mg/kg/d), and group 5 (ApoE-/- mice with HFD, and treatment with high dose BBR of 100 mg/kg/d). After a 16-week treatment, the blood sample, aorta and liver were collected for lipid analysis, hematoxylin-eosin (HE) or oil red O staining, and Western blotting respectively. Besides, HepG2 Cells were cultured and treated with different concentrations of BBR (0, 5, 25 and 50 µg/mL) for 24 hours. Subsequently, cells were collected for real-time PCR or western blotting assays. Finally, the expression levels of PCSK9, LDL receptor (LDLR), ATP-binding cassette transporter A1 (ABCA1), ATP-binding cassette transporter G1 (ABCG1), and scavenger receptor class B type I (SR-BI) were examined. RESULTS: Fifty mg/kg/d and 100 mg/kg/d of BBR decreased total cholesterol (TC), triglyceride (TG), low-density lipoprotein (LDL) cholesterol (LDL-C), and increased high-density lipoprotein cholesterol (HDL-C) level. Moreover, BBR reduced aorta atherosclerotic plaque, and ameliorated lipid deposition in ApoE-/- mice fed with HFD. Finally, in vitro study showed that BBR promoted intracellular cholesterol efflux, up-regulated LDLR and down-regulated PCSK9 expression via the ERK1/2 pathway in cultured HepG2 cells. CONCLUSIONS: Data indicated that BBR significantly attenuated lipid disorder, reduced aortic plaque formation, and alleviated hepatic lipid accumulation in ApoE-/- mice fed with HFD, which was associated with down-regulation of PCSK9 through ERK1/2 pathway.

Pyk2/MCU Pathway as a New Target for Reversing Atherosclerosis.

Objective: Multiple mechanisms including vascular endothelial cell damage have a critical role in the formation and development of atherosclerosis (AS), but the specific molecular mechanisms are not exactly clarified. This study aims to determine the possible roles of proline-rich tyrosine kinase 2 (Pyk2)/mitochondrial calcium uniporter (MCU) pathway in AS mouse model and H2O2-induced endothelial cell damage model and explore its possible mechanisms. Approach and Results: The AS mouse model was established using apolipoprotein E-knockout (ApoE-/-) mice that were fed with a high-fat diet. It was very interesting to find that Pyk2/MCU expression was significantly increased in the artery wall of atherosclerotic mice and human umbilical vein endothelial cells (HUVECs) attacked by hydrogen peroxide (H2O2). In addition, down-regulation of Pyk2 by short hairpin RNA (shRNA) protected HUVECs from H2O2 insult. Furthermore, treatment with rosuvastatin on AS mouse model and H2O2-induced HUVEC injury model showed a protective effect against AS by inhibiting the Pyk2/MCU pathway, which maintained calcium balance, prevented the mitochondrial damage and reactive oxygen species production, and eventually inhibited cell apoptosis. Conclusion: Our results provide important insight into the initiation of the Pyk2/MCU pathway involved in AS-related endothelial cell damage, which may be a new promising target for atherosclerosis intervention.

Exercise reduces hyperlipidemia-induced kidney damage in apolipoprotein E-deficient mice.

Hyperlipidemia is a risk factor of kidney damage that can lead to chronic kidney disease. Studies have shown that exercise reduces kidney damage; however, the specific mechanisms underlying the protective effects of exercise remain unclear. For 12 weeks, 8-week-old male apolipoprotein E-deficient (ApoE-/-) mice were randomly divided into four treatment groups (n=7/group) as follows: Mice fed a normal diet (ND group); mice fed a ND and exercised (ND + E group); mice fed a high-fat diet (HD group); and mice fed a HD and exercised (HD + E group). Exercise training consisted of swimming for 40 min, 5 days/week. Metabolic parameters, such as low-density lipoprotein-cholesterol, total cholesterol and creatinine levels were higher in the ApoE-/- HD mice compared with those in the ApoE-/- HD + E mice. Serum levels of glutathione peroxidase and superoxide dismutase were significantly decreased in the HD group compared with those in the HD + E group. Significant pathological changes were observed in the HD + E group compared with in the HD group. Immunohistochemistry and immunoblotting revealed increased levels of oxidative stress (nuclear factor erythroid-2-related factor 2) and fibrosis (Smad3 and TGF-ß) markers in the ApoE-/- HD group; however, the expression levels of these markers were significantly decreased in the ApoE-/- HD + E group. Furthermore, NF-κB expression in the HD + E group was significantly lower compared with that in the HD group. These results suggested that exercise may exert protective effects against kidney damage caused by hyperlipidemia.

Telocytes in the atherosclerotic carotid artery: Immunofluorescence and TEM evidence.

Telocytes, which possess distinct body shapes and long telopodes, are allocated in the vascular wall. As a fundamental cell type, telocytes construct a three-dimensional network to form a support structure for the artery. This study aims to characterize the morphology and ultrastructure of telocytes in atherosclerotic arteries. ApoE gene-deficient mice were selected as the atherosclerosis animal model and fed a high-fat diet for at least 12 weeks, and immunofluorescence assays and transmission electron microscopy techniques were used to observe changes in telocytes in atherosclerotic arteries. By immunofluorescence staining, CD34, CD117 and PDGFR-α were positive compared with negative CD28/vimentin in telocytes in the atherosclerotic carotid artery, and they were distributed in the tunica intima and tunica adventitia. Under transmission electron microscopy, the bodies of telocytes became larger, while telopodes became shorter compared with their normal condition, and a mass of lipidosomes was present during the progression of atherosclerosis. These results demonstrate that immunofluorescence with TEM is the critical method for identifying TCs and that steatosis of TCs is a reason for atherosclerotic artery dysfunction.