[Effect of Erchen Decoction on liver mitochondrial function by inhibiting mTORC1/SREBP1/CAV1 pathway in mice with high-fat diet].

This study aims to investigate the effect of Erchen Decoction(ECD) on liver mitochondrial function in mice with a high-fat diet and its possible mechanism. A total of sixty C57BL/6J mice were randomly divided into a normal group, high-fat group, ECD group, mTORC1 activator(MHY) group, ECD+MHY group, and polyene phosphatidyl choline(PPC) group, with 10 rats in each group. The normal group was given a normal diet, and the other groups were fed a high-fat diet for 20 weeks. At the 17th week, the ECD group and ECD+MHY group were given ECD(8.7 g·kg~(-1)) daily, and the PPC group was given PPC(0.18 g·kg~(-1)) daily, while the remaining groups were given normal saline(0.01 mL·g~(-1)) daily for four weeks. In the 19th week, the MHY group and ECD+MHY group were injected intraperitoneally with MHY(5 mg·kg~(-1)) every other day for two weeks. During the experiment, the general conditions of the mice were observed. The contents of triglyceride(TG) and total cholesterol(TC) in serum were measured. Morphological changes in liver tissue were examined through HE and oil red O staining. The content of adenosine triphosphate(ATP) was determined using chemiluminescence, and mitochondrial membrane potential was assessed using a fluorescence probe(JC-1). Western blot was performed to detect the expression of rapamycin target protein complex 1(mTOR1), ribosomal protein S6 kinase B1(S6K), sterol regulatory element binding protein 1(SREBP1), and caveolin 1(CAV1). RESULTS:: revealed that compared with the normal group, the mice in the high-fat group exhibited significant increases in body weight and abdominal circumference(P<0.01). Additionally, there were significant increases in TG and TC levels(P<0.01). HE and oil red O staining showed that the boundaries of hepatic lobules were unclear; hepatocytes were enlarged, round, and irregularly arranged, with obvious lipid droplet deposition and inflammatory cell infiltration. The liver ATP content and mitochondrial membrane potential decreased significantly(P<0.01). The expression of p-mTOR, p-S6K, and n-SREBP1 increased significantly(P<0.01), while the expression of CAV1 decreased significantly(P<0.01). Compared with the high-fat group, the body weight and TG content of mice in the ECD group and PPC group decreased significantly(P<0.05). Improvements were observed in hepatocyte morphology, lipid deposition, and inflammatory cell infiltration. Furthermore, there were significant increases in ATP content and mitochondrial membrane potential(P<0.05 or P<0.01). The expression of p-mTOR, p-S6K, and n-SREBP1 decreased significantly in the ECD group(P<0.01), while CAV1 expression increased significantly(P<0.01). However, the indices mentioned above did not show improvement in the MHY group. When the ECD+MHY group was compared with the MHY group, there were significant reductions in body weight and TG contents(P<0.05). The morphological changes of hepatocytes, lipid deposition, and inflammatory cell infiltration were recovered. Moreover, there were significant increases in liver ATP content and mitochondrial membrane potential(P<0.05 or P<0.05). The expression of p-mTOR, p-S6K, and n-SREBP1 decreased significantly(P<0.01), while CAV1 expression increased significantly(P<0.01). In conclusion, ECD can improve mitochondrial function by regulating the mTORC1/SREBP1/CAV1 pathway. This mechanism may be involved in the resolution of phlegm syndrome and the regulation of lipid metabolism.

Erchen decoction alleviates the progression of NAFLD by inhibiting lipid accumulation and iron overload through Caveolin-1 signaling.

ETHNOPHARMACOLOGICAL RELEVANCE: A combination of 6 different Chinese herbs known as Erchen decoction (ECD) has been traditionally used to treat digestive tract diseases and found to have a protective effect against nonalcoholic fatty liver disease (NAFLD). Despite its efficacy in treating NAFLD, the precise molecular mechanism by which Erchen Decoction regulated iron ion metabolism to prevent disease progression remained poorly understood. AIM OF STUDY: Our study attempted to confirm the specific mechanism of ECD in reducing lipid and iron in NAFLD from the perspective of regulating the expression of Caveolin-1 (Cav-1). STUDY DESIGN: In our study, the protective effect of ECD was investigated in Palmitic Acid + Oleic Acid-induced hepatocyte NAFLD model and high-fat diet-induced mice NAFLD model. To investigate the impact of Erchen Decoction (ECD) on lipid metabolism and iron metabolism via mediating Cav-1 in vitro, Cav-1 knockdown cell lines were established using lentivirus-mediated transfection techniques. MATERIALS AND METHODS: We constructed NAFLD model by feeding with high-fat diet for 12 weeks in vivo and Palmitic Acid + Oleic Acid treatment for 24 h in vitro. The regulation of Lipid and iron metabolism results by ECD were detected by serological diagnosis, immunofluorescent and immunohistochemical staining, and western blotting. The binding ability of 6 small molecules of ECD to Cav-1 was analyzed by molecular docking. RESULTS: We demonstrated that ECD alleviated the progression of NAFLD by inhibiting lipid accumulation, nitrogen oxygen stress, and iron accumulation in vivo and in vitro experiments. Furthermore, ECD inhibited lipid and iron accumulation in liver by up-regulating the expression of Cav-1, which indicated that Cav-1 was an important target for ECD to exert its curative effect. CONCLUSIONS: In summary, our study demonstrated that ECD alleviated the accumulation of lipid and iron in NAFLD through promoting the expression of Cav-1, and ECD might serve as a novel Cav-1 agonist to treat NAFLD.

YuPingFeng (YPF) upregulates caveolin-1 (CAV1) to alleviate pulmonary fibrosis through the TGF-ß1/Smad2 pathway.

ETHNOPHARMACOLOGICAL RELEVANCE: Traditional Chinese medicine (TCM) is considered a valuable asset in China's medical tradition. YPF is a classic prescription that has been derived from the "Jiu Yuan Fang" formula and consists of three herbs: Huangqi (Astragalus membranaceus Bunge), Baizhu (Atractylodes rubra Dekker), and Fangfeng (Saposhnikovia divaricata (Turcz.) Schischk). This prescription is widely acclaimed for its exceptional pharmacological properties, including potent antioxidant effects, hormone regulation, and immune modulation effects. AIM OF THE STUDY: Previous research provides evidence suggesting that YPF may have therapeutic effects on pulmonary fibrosis. Further exploration is essential to confirm its effectiveness and elucidate the fundamental processes. MATERIALS AND METHODS: First, the active components and target genes of YPF were extracted from the Traditional Chinese Medicine Systems Pharmacology (TCMSP) database. Next, the GSE53845 dataset, which contains information on pulmonary fibrosis, was downloaded from the GEO database. Network informatics methods was then be utilized to identify target genes associated with pulmonary fibrosis. A YPF-based network of protein-protein interactions was constructed to pinpoint possible target genes for pulmonary fibrosis treatment. Additionally, an in vitro model of pulmonary fibrosis induced by bleomycin (BLM) was established to further investigate and validate the possible mechanisms underlying the effectiveness of YPF. RESULTS: In this study, a total of 24 active ingredients of YPF, along with 178 target genes associated with the treatment, were identified. Additionally, 615 target genes related to pulmonary fibrosis were identified. Functional enrichment analysis revealed that 18 candidate genes (CGs) exhibited significant responses to tumor necrosis factor, NF-kB survival signaling, and positive regulation of apoptosis processes. Among these CGs, CAV1, VCAM1, and TP63 were identified as key target genes. Furthermore, cell experiments confirmed that the expression of CAV1 protein and RNA expression was increased in pulmonary fibrosis, but significantly decreased after treatment with YPF. Additionally, the expression of pSmad2, α-SMA, TGF-ß1, and TNF-α was also decreased following YPF treatment. CONCLUSIONS: Network pharmacology analysis revealed that YPF exhibits significant potential as a therapeutic intervention for pulmonary fibrosis by targeting various compounds and pathways. This study emphasizes that the efficacy of YPF in treating pulmonary fibrosis may be attributed to its ability to up-regulate CAV1 expression and inhibiting pulmonary fibrosis via modulation of the TGF-ß1/Smad2 signaling pathway. These findings underscore the promising role of YPF and its ability to potentially alleviate pulmonary fibrosis.

Chemical characterization and effects of volatile oil of Alpinia zerumbet on the quality of collagen deposition and caveolin-1 expression in a muscular fibrosis murine model / Caracterização química e efeitos do óleo volátil de Alpinia zerumbet na qualidade da deposição do colágeno e expressão da caveolina-1 em modelo murino de fibrose muscular

Abstract This study evaluated the effect of the volatile oil of Alpinia zerumbet (VOAz) on caveolin-1 gene expression and muscular fibrosis. The rats were immobilized to induce fibrosis of the gastrocnemius muscle, and they were treated with VOAz. Collagen quality was assessed by histology and the expression of the caveolin-1 (CAV-1) gene was evaluated using qPCR. Histomorphological analysis indicated a significant reduction in the perimeter, width, and intensity of collagen in the treated groups, thus showing that the oil was effective in regulating the quality of collagen at the three concentrations. The results of expression levels suggested a decrease in the lesioned group and in two treatment groups (0.0115 µg/g and 0.009 µg/g). However, with the lowest concentration (0.0065 µg/g), no significant difference was observed, with levels similar to those found in healthy tissue. Therefore, the results showed that VOAz has the potential to be a non-invasive and low-cost alternative to aid in the treatment of muscular fibrosis.

Resumo Este estudo avaliou o efeito do óleo volátil de Alpinia zerumbet (OVAz) na expressão do gene da caveolina-1 e na fibrose muscular. Os ratos foram imobilizados para induzir a fibrose do músculo gastrocnêmio, e foram tratados com OVAz. A qualidade do colágeno foi avaliada com histologia e à expressão do gene caveolina-1 (CAV-1) foi avaliada usando qPCR. A análise histomorfológica indicou uma redução significativa no perímetro, largura e intensidade do colágeno nos grupos tratados. Os resultados dos níveis de expressão sugeriram diminuição nos grupos de lesão e em dois grupos de tratamento (0,0115 µg/g e 0,009 µg/g). No entanto, com a menor concentração (0,0065 µg/g), não foi observada diferença significativa, apresentando níveis semelhantes aos encontrados em tecido saudável. O uso do OVAz foi eficaz para reverter as alterações do colágeno causadas pela fibrose, e sua menor concentração apresentou uma possível tendência de aumento na expressão do CAV-1. Portanto, os resultados mostraram que o OVAz tem potencial para ser uma alternativa não invasiva e de baixo custo para auxiliar no tratamento da fibrose muscular.

Caveolin-mediated cytosolic delivery of spike nanoparticle enhances antitumor immunity of neoantigen vaccine for hepatocellular carcinoma.

Rationale: Although neoantigen-based cancer vaccines have shown promise in various solid tumors, limited immune responses and clinical outcomes have been reported in patients with advanced disease. Cytosolic transport of neoantigen and adjuvant is required for the activation of intracellular Toll-like receptors (TLRs) and cross-presentation to prime neoantigen-specific CD8+T cells but remains a significant challenge. Methods: In this study, we aimed to develop a virus-like silicon vaccine (V-scVLPs) with a unique spike topological structure, capable of efficiently co-delivering a hepatocellular carcinoma (HCC)-specific neoantigen and a TLR9 agonist to dendritic cells (DCs) to induce a robust CD8+T cell response to prevent orthotopic tumor growth. We evaluated the antitumor efficacy of V-scVLPs by examining tumor growth and survival time in animal models, as well as analyzing tumor-infiltrating CD8+T cells and cytokine responses in the tumor microenvironment (TME). To evaluate the synergistic efficacy of V-scVLPs in combination with α-TIM-3 in HCC, we used an orthotopic HCC mouse model, a lung metastasis model, and a tumor rechallenge model after hepatectomy. Results: We found that V-scVLPs can efficiently co-deliver the hepatocellular carcinoma (HCC)-specific neoantigen and the TLR9 agonist to DCs via caveolin-mediated endocytosis. This advanced delivery strategy results in efficient lymph node draining of V-scVLPs to activate lymphoid DC maturation for promoting robust CD8+T cells and central memory T cells responses, which effectively prevents orthotopic HCC tumor growth. However, in the established orthotopic liver tumor models, the inhibitory receptor of TIM-3 was significantly upregulated in tumor-infiltrating CD8+T cells after immunization with V-scVLPs. Blocking the TIM-3 signaling further restored the antitumor activity of V-scVLPs-induced CD8+T cells, reduced the proportion of regulatory T cells, and increased the levels of cytokines to alter the tumor microenvironment to efficiently suppress established orthotopic HCC tumor growth, and inhibit lung metastasis as well as recurrence after hepatectomy. Conclusion: Overall, the developed novel spike nanoparticles with efficient neoantigen and adjuvant intracellular delivery capability holds great promise for future clinical translation to improve HCC immunotherapy.

Impact of NB-UVB phototherapy on Caveolin-1 expression in chronic plaque psoriasis.

BACKGROUND: Caveolin-1 (Cav-1) is a significant structural and regulatory constituent of cell membranes that has been implicated in cell kinetics and inflammation. OBJECTIVE: To assess Cav-1 expression in psoriasis before and after phototherapy. PATIENTS AND METHODS: Thirty psoriasis cases and 30 healthy controls were recruited. Cases were managed with narrow band-ultraviolet B (NB-UVB) phototherapy at frequency three times per week for 12 weeks. From every case, two biopsy specimens were gained from psoriatic lesions (pre and post phototherapy), in addition to one from apparently normal skin of psoriasis cases. Regarding the control group, one biopsy was taken from a matched site. All were studied for Cav-1 antibody immuno-expression. RESULTS: There was a significant decrease in Cav-1 expression in psoriatic lesions compared to both the apparently normal skin of psoriasis patients and standard control skin of healthy individuals. After NB-UVB phototherapy, significant upregulation of Cav-1 immunostaining score was observed in previously psoriatic skin when compared to that before treatment. In addition, there were significant negative correlations between Cav-1 immunostaining score and the clinical scores of psoriasis severity including; the erythema, scaling, and induration (ESI) score and the patient psoriasis area and severity index (PASI) score. CONCLUSION: Induction of Cav-1 expression may be a likely pathway for the effectiveness of NB-UVB in psoriasis. Cav-1 may be a useful marker for evaluation of psoriasis severity, disease progression, and therapeutic efficacy.

Biochemical and Biophysical Characterization of the Caveolin-2 Interaction with Membranes and Analysis of the Protein Structural Alteration by the Presence of Cholesterol.

Caveolin-2 is a protein suitable for the study of interactions of caveolins with other proteins and lipids present in caveolar lipid rafts. Caveolin-2 has a lower tendency to associate with high molecular weight oligomers than caveolin-1, facilitating the study of its structural modulation upon association with other proteins or lipids. In this paper, we have successfully expressed and purified recombinant human caveolin-2 using E. coli. The structural changes of caveolin-2 upon interaction with a lipid bilayer of liposomes were characterized using bioinformatic prediction models, circular dichroism, differential scanning calorimetry, and fluorescence techniques. Our data support that caveolin-2 binds and alters cholesterol-rich domains in the membranes through a CARC domain, a type of cholesterol-interacting domain in its sequence. The far UV-CD spectra support that the purified protein keeps its folding properties but undergoes a change in its secondary structure in the presence of lipids that correlates with the acquisition of a more stable conformation, as shown by differential scanning calorimetry experiments. Fluorescence experiments using egg yolk lecithin large unilamellar vesicles loaded with 1,6-diphenylhexatriene confirmed that caveolin-2 adsorbs to the membrane but only penetrates the core of the phospholipid bilayer if vesicles are supplemented with 30% of cholesterol. Our study sheds light on the caveolin-2 interaction with lipids. In addition, we propose that purified recombinant caveolin-2 can provide a new tool to study protein-lipid interactions within caveolae.

Tiao-bu-fei-shen formula promotes downregulation of the caveolin 1-p38 mapk signaling pathway in COPD - Associated tracheobronchomalacia cell model.

ETHNOPHARMACOLOGICAL RELEVANCE: The Tiao-bu-fei-shen (TBFS) formula, extensively used in Traditional Chinese Medicine (TCM), can enhance therapeutic efficacy and reduce the frequency of acute exacerbations of lung-kidney Qi deficiency in patients with chronic obstructive pulmonary disease (COPD). According to both TCM theory and long-term observation of practice, TBFS has become an effective treatment for COPD-associated tracheobronchomalacia (TBM). AIM OF THE STUDY: To investigate the mechanism of the TBFS formula in treating COPD-associated TBM based on caveolin 1-p38 MAPK signaling and apoptosis. MATERIALS AND METHODS: A rat COPD model was prepared by exposure to smoking combined with tracheal lipopolysaccharide injection. The trachea or bronchus chondrocytes from COPD rats were isolated, cultured, and treated with 10 ng/mL IL-1ß for 24 h to develop a model of COPD-associated TBM. Normal rats were administered TBFS to prepare drug-containing serum, and CCK8 assays were used to screen the optimal drug-containing serum concentration and SB203580 dose. TBFS drug-containing serum and SB203580 were processed separately for the control, model, drug-containing serum, blocker, and drug-containing serum combined with blocker groups. Flow cytometry and CCK8 assays were used to detect apoptosis and proliferative activity. Toluidine blue staining and immunohistochemistry were used to analyze the chondrocyte proteoglycan and type II collagen content. Western blotting was used to detect the expression of caveolin 1, p-p38 MAPK, TNF-α, IL-1ß, MMP-13, Bax, and Bcl-2 proteins. Quantitative PCR was used to detect the expression of caveolin 1, p38 MAPK, IL-1ß, MMP-13, Bax, Bcl-2, and miR-140-5p. RESULTS: The isolation and identification of bronchial chondrocytes from COPD rats revealed that 10 ng/mL IL-1ß can produce a stable COPD-associated TBM model. Screened via the CCK8 method, fourth-generation bronchial chondrocytes were determined as the optimal cells, and 5 µM SB203580 and 5% low-dose drug-containing serum were the optimal intervention doses. The experimental chondrocytes of each group were treated separately for 48 h. Toluidine blue staining and immunohistochemical analysis revealed that TBFS drug-containing serum, SB203580, and TBFS drug-containing serum combined with SB203580 can effectively increase the proteoglycan and type II collagen content after chondrocyte degradation. Flow cytometry of cells treated with SB203580 and TBFS drug-containing serum combined with SB203580 revealed significantly reduced cell apoptosis and enhanced cell proliferation activity. Western blot and qPCR analyses revealed that the TBFS drug-containing serum, SB203580, and TBFS drug-containing serum combined with SB203580 effectively inhibit the expression of caveolin 1, p-p38 MAPK, MMP-13, IL-1ß, TNF-α, and Bax proteins while promoting Bcl -2 protein expression. Treatment with TBFS drug-containing serum and SB203580 effectively inhibited the expression of MMP-13, p38 MAPK, caveolin 1, and Bax genes, and promoted the expression of Bcl-2 and miR-140-5p genes. CONCLUSIONS: A concentration of 10 ng/mL of IL-1ß can generate a stable COPD-associated TBM cell model. TBFS can improve the proteoglycan and type II collagen content, increase cell activity, and reduce the amount of chondrocyte apoptosis. The role of TBFS may be related to mechanisms of inhibiting the expression of the key signaling molecules caveolin 1 and p-p38 MAPK in the caveolin 1-p38 MAPK signaling pathway, thereby reducing the expression of the downstream effector products MMP-13, IL-1ß, and TNF-α, while inhibiting the expression of the apoptotic gene Bax and improving the expression of Bcl-2 and miR-140-5p genes.

Isoliquiritigenin alleviates liver fibrosis through caveolin-1-mediated hepatic stellate cells ferroptosis in zebrafish and mice.

BACKGROUND: Liver fibrosis is a major disease that threatens people's health around the world. However, there is a lack of effective treatment to completely reverse liver fibrosis. Liver transplantation is currently the only curative option for patients with advanced cirrhosis. Ferroptosis is a newly discovered type of cell death and plays an important role in the process of liver fibrosis, but the specific mechanism needs to be clarified. HYPOTHESIS/PURPOSE: To explore the regulatory mechanism of isoliquiritigenin (ISL) in the process of liver fibrosis and the relationship between Cav-1 and ferroptosis. METHODS: In this research, zebrafish, HSC-T6 cells, and mice were used as the research object. Different ROS probes to visually detect the content and distribution of ROS in live zebrafish and cells. Lentivirus and siRNA-mediated transfection techniques were used for the construction of Cav-1 overexpression and knockdown cell lines to verify the important role of Cav-1 in vitro. RESULTS: Generally, we first elucidated that ISL relieved liver fibrosis by inducing hepatic stellate cells (HSCs) ferroptosis through repressing GPX4 expression and increasing the expression of TFR and DMT1, thus producing a large number of ROS, we also found that Cav-1 exerted its anti-hepatic fibrosis effect by promoting HSCs ferroptosis. CONCLUSION: Our results have shown that Cav-1-mediated HSCs ferroptosis is necessary for ISL to play an anti-fibrotic effect in vitro and in vivo.

Chemical characterization and effects of volatile oil of Alpinia zerumbet on the quality of collagen deposition and caveolin-1 expression in a muscular fibrosis murine model.

This study evaluated the effect of the volatile oil of Alpinia zerumbet (VOAz) on caveolin-1 gene expression and muscular fibrosis. The rats were immobilized to induce fibrosis of the gastrocnemius muscle, and they were treated with VOAz. Collagen quality was assessed by histology and the expression of the caveolin-1 (CAV-1) gene was evaluated using qPCR. Histomorphological analysis indicated a significant reduction in the perimeter, width, and intensity of collagen in the treated groups, thus showing that the oil was effective in regulating the quality of collagen at the three concentrations. The results of expression levels suggested a decrease in the lesioned group and in two treatment groups (0.0115 µg/g and 0.009 µg/g). However, with the lowest concentration (0.0065 µg/g), no significant difference was observed, with levels similar to those found in healthy tissue. Therefore, the results showed that VOAz has the potential to be a non-invasive and low-cost alternative to aid in the treatment of muscular fibrosis.

Notoginsenoside R1 intervenes degradation and redistribution of tight junctions to ameliorate blood-brain barrier permeability by Caveolin-1/MMP2/9 pathway after acute ischemic stroke.

BACKGROUND: The leakage of blood-brain barrier (BBB) is main pathophysiological change in acute stage of ischemic stroke, which not only deteriorates neurological function, but also increases the risk of hemorrhagic transformation after thrombolysis. PURPOSE/STUDY DESIGN: This article investigates the efficacy of Notoginsenoside R1, an active ingredient of Panax notoginseng, on BBB permeability and explores related mechanisms after acute ischemic stroke. METHODS: In vivo, male Sprague-Dawley rats (260-280 g) were selected and randomly divided into 6 groups: sham group, model group, low, middle and high doses of Notoginsenoside R1 groups and positive drug Dl-3-n-Butylphthalide group. Except for sham group, rats were performed with permanent middle cerebral artery occlusion model in each group. Twelve hours later, rats were evaluated for Bederson neurological function, and BBB integrity by Evans blue leak imaging; Triphenyltetrazolium chloride staining was used to detect the volume of cerebral infarction. Frozen sections of rats' brain tissue were prepared for detection of MMPs activity in situ zymography. Peripheral tissue of cerebral infarction was collected and tested the expression of MMP2, 9 and tight junction proteins (zo1, claudin5, occludin) by western blot. In vitro, transwell endothelial barrier model was established by bEnd.3 cells. Oxygen glucose deprivation (OGD) was chosen to simulate the hypoxic environment. Suitable OGD stimulation time as well as Notoginsenoside R1 and Dl-3-n-Butylphthalide optimal dose concentrations were determined through transwell leakage and CCK8 assay. Furthermore, endothelial subcellular component proteins were extracted. The change of zo1, claudin5, occludin and caveolin1 was detected by western blot. RESULTS: Notoginsenoside R1 treatment significantly reduced BBB leakage and cerebral infarction volume, weakened neurological deficits in post-stroke rats. Moreover, it inhibited the activity of MMPs in infarcted cortex and striatum, down-regulated MMP2, 9 and up-regulated zo1 and claudin5 expressions in penumbra. In vitro, Notoginsenoside R1 treatment decreased OGD-induced endothelial barrier permeability, restored expressions of zo1, claudin5 on cellular membrane and cytoplasm, as well as mediated membrane redistribution of occludin and caveolin1 from actin cytoskeletal fraction. CONCLUSIONS: Notoginsenoside R1 treatment attenuates BBB permeability, cerebral infarction volume and neurological impairments in rats with acute cerebral ischemia. The mechanisms might be related to intervening degradation and redistribution of zo1, caludin5 and occludin by caveolin1/ MMP2/9 pathway. More effects and mechanisms of Notoginsenoside R1 on rehabilitation of stroke are worthy to be explored in the future.

Targeting the HIF-1α/Cav-1 Pathway with a Chicory Extract/Daidzein Combination Plays a Potential Role in Retarding Hepatocellular Carcinoma.

BACKGROUND: Hepatocellular carcinoma (HCC) is one of the most rapidly growing solid cancers, that is characterized by hypoxia. Hypoxia-inducible factor-1α (HIF-1α) is a transcription factor that regulates tumor proliferation and metastasis. It induces caveolin-1 (Cav-1) expression, a glycoprotein found on the membrane surface, then Cav-1 triggers angiogenesis and metastasis in HCC. OBJECTIVE: We hypothesize that targeting HIF-1α and consequently, Cav-1 using the antioxidant natural compound such as chicoric acid and a Cav-1 inhibitor daidzein (DAZ) could be a useful approach in the management of HCC. This study was conducted to investigate the possible therapeutic efficacy of standardized chicory leaf extract (SCLE) and DAZ via modulation of HIF-1α and Cav-1 in HCC rats. METHODS: Diethyl nitrosamine (DENA) was used for HCC induction. After the induction period, four groups (10 rats for each) were treated with SCLE, DAZ, a combination of both, as well as sorafenib, all compared to the non-treated control. We assessed hepatic HIF-1α protein expression, Cav-1 gene expression, serum level of AFP, hepatic tissue content of VEGF, MMP-9, oxidative stress markers MDA and SOD. RESULTS: DAZ, SCLE, and their combination, significantly down-regulated the expression of HIF-1α, Cav-1, and consequently dampened MMP-9, VEGF, hepatic content. It has been observed that the combination treatment showed a synergistic effect compared to either treatment alone. Importantly, the combination treatment exhibited a significantly more potent effect than sorafenib. CONCLUSION: This study showed the potential role of the HIF-1α/Cav-1 pathway in HCC progression, moreover, SCLE and DAZ showed a potent efficacy in retarding HCC via modulation of this pathway.

VEGF-R2/Caveolin-1 Pathway of Undifferentiated ARPE-19 Retina Cells: A Potential Target as Anti-VEGF-A Therapy in Wet AMD by Resvega, an Omega-3/Polyphenol Combination.

Age-related macular degeneration (AMD) is one of the main causes of deterioration in vision in adults aged 55 and older. In spite of therapies, the progression of the disease is often observed without reverse vision quality. In the present study, we explored whether, in undifferentiated ARPE-19 retinal cells, a disruption of the VEGF receptors (VEGF-R)/caveolin-1 (Cav-1)/protein kinases pathway could be a target for counteracting VEGF secretion. We highlight that Resvega®, a combination of omega-3 fatty acids with an antioxidant, resveratrol, inhibits VEGF-A secretion in vitro by disrupting the dissociation of the VEGF-R2/Cav-1 complex into rafts and subsequently preventing MAPK activation. Moreover, DNA ChIP analysis reveals that this combination prevents the interaction between AP-1 and vegf-a and vegf-r2 gene promoters. By these pathways, Resvega could present a potential interest as nutritional complementation against AMD.

A novel mitochondrial Kv1.3-caveolin axis controls cell survival and apoptosis.

The voltage-gated potassium channel Kv1.3 plays an apparent dual physiological role by participating in activation and proliferation of leukocytes as well as promoting apoptosis in several types of tumor cells. Therefore, Kv1.3 is considered a potential pharmacological target for immunodeficiency and cancer. Different cellular locations of Kv1.3, at the plasma membrane or the mitochondria, could be responsible for such duality. While plasma membrane Kv1.3 facilitates proliferation, the mitochondrial channel modulates apoptotic signaling. Several molecular determinants of Kv1.3 drive the channel to the cell surface, but no information is available about its mitochondrial targeting. Caveolins, which are able to modulate cell survival, participate in the plasma membrane targeting of Kv1.3. The channel, via a caveolin-binding domain (CDB), associates with caveolin 1 (Cav1), which localizes Kv1.3 to lipid raft membrane microdomains. The aim of our study was to understand the role of such interactions not only for channel targeting but also for cell survival in mammalian cells. By using a caveolin association-deficient channel (Kv1.3 CDBless), we demonstrate here that while the Kv1.3-Cav1 interaction is responsible for the channel localization in the plasma membrane, a lack of such interaction accumulates Kv1.3 in the mitochondria. Kv1.3 CDBless severely affects mitochondrial physiology and cell survival, indicating that a functional link of Kv1.3 with Cav1 within the mitochondria modulates the pro-apoptotic effects of the channel. Therefore, the balance exerted by these two complementary mechanisms fine-tune the physiological role of Kv1.3 during cell survival or apoptosis. Our data highlight an unexpected role for the mitochondrial caveolin-Kv1.3 axis during cell survival and apoptosis.

Oxymatrine improves blood-brain barrier integrity after cerebral ischemia-reperfusion injury by downregulating CAV1 and MMP9 expression.

BACKGROUND: Ischemic stroke (IS) is a major neurological condition associated with extremely high morbidity and mortality worldwide. Oxymatrine (OMT), a quinolizidine alkaloid extracted from the root of Sophora flavescens, has neuroprotective properties and protects against IS. However, whether its protective effect involves alterations in the integrity of the blood-brain barrier (BBB) is unknown. PURPOSE: Here, we used in vivo and in vitro models of IS to evaluate the protective effects of OMT and to establish whether its effects are mediated via the modulation of the BBB function. METHODS: We assessed the effects of OMT by using neurological function scores, triphenyltetrazolium chloride staining, Nissl staining, and terminal deoxynucleotidyl transferase dUTP nick end labeling. RESULTS: OMT significantly prevented cellular damage, improved neurological function, and reduced BBB permeability in a mouse model of cerebral ischemia-reperfusion. Additionally, OMT protected the function of the tight junctions of bEend.3 cells against the consequences of oxygen-glucose deprivation. Furthermore, intracranial lentivirus injection of short hairpin RNA targeting Cav1 decreased caveolin-1 expression and inhibited the neuroprotective effects of OMT. CONCLUSIONS: OMT attenuated ischemia-reperfusion injury-induced damage to the BBB, and this neuroprotective action was at least partially dependent on the expression levels of CAV1 and MMP9 proteins. Therefore, OMT may offer effective protection against BBB injury induced by ischemia-reperfusion episodes.

Icariin modulates eNOS activity via effect on post-translational protein-protein interactions to improve erectile function of spontaneously hypertensive rats.

BACKGROUND: Type 5 phosphodiesterase inhibitor (PDE5I) has become the first-line treatment for erectile dysfunction (ED). However, its effective rate for hypertension ED is only 60%-70%. How to improve the efficacy of ED treatment is the focus of current research. OBJECTIVE: To explore whether icariin can improve the erectile function of spontaneously hypertensive rats (SHR) by affecting post-translational protein-protein interactions to regulate endothelial nitric oxide synthetase (eNOS) activity. METHOD: Twelve-week-old healthy male SHR rats and Wistar-Kyoto rats (WKY) were randomly divided into four groups: SHR control group, SHR + icariin (10 mg/kg·d gavage) treatment group, WKY control group, and WKY + icariin (10 mg/kg·d gavage) treatment group (n = 5). After 4 weeks, the maximum penile intracavernous pressure/mean arterial pressure (ICPmax/MAP), the expression of heat-shock protein 90 (Hsp90), caveolin-1, calmodulin, p-eNOS, and eNOS in penile cavernous tissue and the content of nitric oxide (NO) and cGMP were measured. The interaction between eNOS and Hsp90, caveolin-1, and calmodulin were detected by immunoprecipitation. RESULT: The ICPmax/MAP in the SHR + icariin treatment group (0.08 ± 0.01, 0.23 ± 0.07, 0.40 ± 0.05) was significantly higher than the SHR group (0.03 ± 0.01, 0.13 ± 0.03, 0.21 ± 0.02) under 3V and 5V electrical stimulations (P < .05). Compared with the SHR group, the expression of HSP90, calmodulin, P-eNOS, eNOS, and P-eNOS/eNOS in the penile cavernous tissue of rats in the WKY group and the SHR + icariin treatment group were significantly increased (P < .05), and the expression of caveolin-1 was significantly decreased (P < .05). The NO content (2.16 ± 0.22 µmol/g) and cGMP concentration (3.69 ± 0.12 pmol/mg) in the SHR + icariin treatment group were significantly higher than those in the SHR group (1.01 ± 0.14 µmol/g, 2.31 ± 0.22 pmol/mg) (P < .05). Compared with the SHR group, the interaction between eNOS and HSP90 in the cavernosa of the rats in the SHR + icariin treatment group was significantly increased (P < .05), the interaction between eNOS and caveolin-1 was significantly decreased (P < .01), and the interaction between eNOS and calmodulin did not significantly change. DISCUSSION AND CONCLUSION: Up-regulating the expression of HSP90 and calmodulin and inhibiting caveolin-1 in SHR corpus cavernosum, promoting the interaction between eNOS and HSP90, inhibiting the interaction between eNOS and caveolin-1, increasing p-eNOS/eNOS, may be the mechanism of icariin that improves SHR erectile function.

Kirenol inhibits RANKL-induced osteoclastogenesis and prevents ovariectomized-induced osteoporosis via suppressing the Ca2+-NFATc1 and Cav-1 signaling pathways.

BACKGROUND: Osteoporosis is a threat to aged people who have excessive osteoclast activation and bone resorption, subsequently causing fracture and even disability. Inhibiting osteoclast differentiation and absorptive functions has become an efficient approach to treat osteoporosis, but osteoclast-targeting inhibitors available clinically remain rare. Kirenol (Kir), a bioactive diterpenoid derived from an antirheumatic Chinese herbal medicine Herba Siegesbeckiae, can treat collagen-induced arthritis in vivo and promote osteoblast differentiation in vitro, while the effects of Kir on osteoclasts are still unclear. PURPOSE: We explore the role of Kir on RANKL-induced osteoclastogenesis in vitro and bone loss in vivo. METHODS: The in vitro effects of Kir on osteoclast differentiation, bone resorption and the underlying mechanisms were evaluated with bone marrow-derived macrophages (BMMs). In vivo experiments were performed using an ovariectomy (OVX)-induced osteoporosis model. RESULTS: We found that Kir remarkably inhibited osteoclast generation and bone resorption in vitro. Mechanistically, Kir significantly inhibited F-actinring formation and repressed RANKL-induced NF-κB p65 activation and p-p38, p-ERK and c-Fos expression. Moreover, Kir inhibited both the expression and nuclear translocation of NFATc1. Ca2+ oscillation and caveolin-1 (Cav-1) were also reduced by Kir during osteoclastogenesis in vitro. Consistent with these findings, 2-10 mg/kg Kir attenuated OVX-induced osteoporosis in vivo as evidenced by decreased osteoclast numbers and downregulated Cav-1 and NFATc1 expression. CONCLUSIONS: Kir suppresses osteoclastogenesis and the Cav-1/NFATc1 signaling pathway both in vitro and in vivo and protects against OVX-induced osteoporosis. Our findings reveal Kir as a potential safe oral treatment for osteoporosis.

Caveolin-1 stabilizes ATP7A, a copper transporter for extracellular SOD, in vascular tissue to maintain endothelial function.

Caveolin-1 (Cav-1) is a scaffolding protein and a major component of caveolae/lipid rafts. Previous reports have shown that endothelial dysfunction in Cav-1-deficient (Cav-1-/-) mice is mediated by elevated oxidative stress through endothelial nitric oxide synthase (eNOS) uncoupling and increased NADPH oxidase. Oxidant stress is the net balance of oxidant generation and scavenging, and the role of Cav-1 as a regulator of antioxidant enzymes in vascular tissue is poorly understood. Extracellular SOD (SOD3) is a copper (Cu)-containing enzyme that is secreted from vascular smooth muscle cells/fibroblasts and subsequently binds to the endothelial cells surface, where it scavenges extracellular [Formula: see text] and preserves endothelial function. SOD3 activity is dependent on Cu, supplied by the Cu transporter ATP7A, but whether Cav-1 regulates the ATP7A-SOD3 axis and its role in oxidative stress-mediated vascular dysfunction has not been studied. Here we show that the activity of SOD3, but not SOD1, was significantly decreased in Cav-1-/- vessels, which was rescued by re-expression of Cav-1 or Cu supplementation. Loss of Cav-1 reduced ATP7A protein, but not mRNA, and this was mediated by ubiquitination of ATP7A and proteasomal degradation. ATP7A bound to Cav-1 and was colocalized with SOD3 in caveolae/lipid rafts or perinucleus in vascular tissues or cells. Impaired endothelium-dependent vasorelaxation in Cav-1-/- mice was rescued by gene transfer of SOD3 or by ATP7A-overexpressing transgenic mice. These data reveal an unexpected role of Cav-1 in stabilizing ATP7A protein expression by preventing its ubiquitination and proteasomal degradation, thereby increasing SOD3 activity, which in turn protects against vascular oxidative stress-mediated endothelial dysfunction.

Salidroside-Mediated Autophagic Targeting of Active Src and Caveolin-1 Suppresses Low-Density Lipoprotein Transcytosis across Endothelial Cells.

Subendothelial retention of apolipoprotein B100-containing lipoprotein, such as low-density lipoprotein (LDL), is the initial step of atherogenesis. Activation of autophagy exhibits beneficial effects for the treatment of atherosclerosis. In our previous study, we demonstrated that hyperglycemia suppressed autophagic degradation of caveolin-1, which in turn resulted in acceleration of caveolae-mediated LDL transcytosis across endothelial cells and lipid retention. Therefore, targeting the crossed pathway in autophagy activation and LDL transcytosis interruption may be a promising antiatherosclerotic strategy. In metabolic diseases, including atherosclerosis, salidroside, a phenylpropanoid glycoside compound (3,5-dimethoxyphenyl) methyl-ß-glucopyranoside), is the most important compound responsible for the therapeutic activities of Rhodiola. However, whether salidroside suppresses LDL transcytosis to alleviate atherosclerosis has not yet been elucidated. In the present study, we demonstrated that salidroside significantly decreased LDL transcytosis across endothelial cells. Salidroside-induced effects were dramatically blocked by AMPK (adenosine monophosphate-activated protein kinase) inhibitor (compound c, AMPKα siRNA) and by overexpression of exogenous tyrosine-phosphorylated caveolin-1 using transfected cells with phosphomimicking caveolin-1 on tyrosine 14 mutant plasmids (Y14D). Furthermore, we observed that salidroside promoted autophagosome formation via activating AMPK. Meanwhile, the interaction between caveolin-1 and LC3B-II, as well as the interaction between active Src (indicated by the phosphorylation of Src on tyrosine 416) and LC3B-II, was significantly increased, upon stimulation with salidroside. In addition, both bafilomycin A1 (a lysosome inhibitor) and an AMPK inhibitor (compound c) markedly prevented salidroside-induced autophagic degradation of p-Src and caveolin-1. Moreover, the phosphorylation of caveolin-1 on tyrosine 14 was disrupted due to the downregulation of p-Src and caveolin-1, thereby directly decreasing LDL transcytosis by attenuating the number of caveolae on the cell membrane and by preventing caveolae-mediated LDL endocytosis released from the cell membrane. In ApoE-/- mice, salidroside significantly delayed the formation of atherosclerotic lesions. Meanwhile, a significant increase in LC3B, accompanied by attenuated accumulation of the autophagy substrate SQSTM1, was observed in aortic endothelium of ApoE-/- mice. Taken together, our findings demonstrated that salidroside protected against atherosclerosis by inhibiting LDL transcytosis through enhancing the autophagic degradation of active Src and caveolin-1.

[Inhibitory effect of paeonol on aortic endothelial inflammation in atherosclerotic rats by up-regulation of caveolin-1 expression and suppression of NF-κB pathway].

To explore whether paeonol can play an anti-atherosclerotic role by regulating the expression of aortic caveolin-1 and affecting NF-κB pathway, so as to inhibit the inflammatory response of vascular endothelium in atherosclerotic rats. The atherosclerotic model of rats was induced by high-fat diet and vitamin D_2. The primary culture of vascular endothelial cells(VECs) was carried out by tissue block pre-digestion and adherent method. The injury model of VECs was induced by lipopolysaccharide(LPS), and filipin, a small concave protein inhibitor, was added for control. HE staining was used to observe pathological changes of aorta. TNF-α, IL-6 and VCAM-1 were detected by ELISA. Western blot assay was used to detect the protein expression levels of caveolin-1 and p65 in aorta and VECs. The results showed that as compared with model group, paeonol significantly reduced aortic plaque area and lesion degree in rats, decreased the level of serum TNF-α, IL-6 and VCAM-1 in the rats and enhanced the relative expression level of caveolin-1, decreased p65 expression conversely(P<0.05 or P<0.01). In vitro, as compared to model group, paeonol obviously improved cell morphology, decreased the secretion of TNF-α, IL-6 and VCAM-1 in VECs, increased caveolin-1 expression, and decreased p65 protein expression(P<0.05 or P<0.01). Furthermore, filipin could reverse the effect of paeonol on expression of inflammatory factors and proteins(P<0.05 or P<0.01). According to the results, it was found that paeonol could play the role of anti-atherosclerosis by up-regulating the expression of caveolin-1 and inhibiting the activation of NF-κB pathway to reduce vascular inflammation in atherosclerotic rats.