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1.
Artigo em Chinês | WPRIM | ID: wpr-1030935

RESUMO

ObjectiveTo observe the effect of Scutellariae Radix-Coptidis Rhizoma on plaque stability in atherosclerotic (AS) mice and to explore its possible mechanism of action based on the Toll-like receptor 4 (TLR4)/myeloid differentiation factor 88 (MyD88)/nuclear factor kappa B (NF-κB) signaling pathway. MethodTen normal C57BL/6J mice were used as the normal group, and the same strain of ApoE knockout (ApoE-/-) mice were fed with a high-fat diet for 12 weeks to construct an atherosclerosis model. Mice were randomly divided into five groups, namely the model group, the atorvastatin group, and the Scutellariae Radix-Coptidis Rhizoma low-, medium-, and high-dose groups, with ten mice in each group. Then normal and model groups were given equal volume of saline gavage, and the low-, medium-, high-dose Scutellariae Radix-Coptidis Rhizoma groups were given 1.95, 3.9, 7.8 g·kg-1 of the drug by gavage for 8 weeks, respectively. The general state of mice was observed. Hematoxylin-eosin staining was utilized to observe the pathology of aortic root plaques and calculate the percentage of plaque area. Masson staining and oil red O staining combined with immunohistochemistry of F4/80 and α-SMA were used to detect the plaque components of aortic root plaques and calculate the plaque vulnerability index. Enzyme-linked immunosorbent assay was adopted to detect the expression levels of serum tumor necrosis factor-α (TNF-α) and interleukin-6 (IL-6). Western blot was applied to detect the protein expression levels of matrix metalloproteinase-2 (MMP-2), matrix metalloproteinase-9 (MMP-9), TLR4, MyD88, NF-κB p65, and phosphorylation (p) -NF-κB p65 in the aortic tissues of mice in each group. Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR) assay was employed to detect the expression levels of MMP-2, MMP-9, TLR4, and MyD88, NF-κB p65 mRNA. ResultCompared with the model group, the general state of the mice in each medication group was improved, and no obvious side effects were observed. Compared with the model group, the percentage of plaque area in the aortic root of AS mice was significantly reduced in the medium- and high-dose Scutellariae Radix-Coptidis Rhizoma groups (P<0.05). The content of collagen fibers and smooth muscle cells in the plaques of the high-dose Scutellariae Radix-Coptidis Rhizoma group was significantly increased (P<0.01), and the content of lipids and macrophages was significantly reduced (P<0.05), the plaque vulnerability index of each dose group of Scutellariae Radix-Coptidis Rhizoma was significantly reduced, with significant reduction of the medium- and high-dose groups (P<0.01). MMP-2 and MMP-9 protein and mRNA expression levels in aortic tissues were significantly reduced in medium- and high-dose Scutellariae Radix-Coptidis Rhizoma groups (P<0.05). The serum levels of TNF-α and IL-6 were significantly reduced in AS mice in medium- and high-dose Scutellariae Radix-Coptidis Rhizoma groups (P<0.05). In the medium- and high-dose Scutellariae Radix-Coptidis Rhizoma groups, the levels of TLR4, MyD88 protein, and mRNA expression in aortic tissues were significantly reduced (P<0.05), and the level of NF-κB p65 phosphorylation in aortic tissues was significantly reduced (P<0.05). ConclusionScutellariae Radix-Coptidis Rhizoma may play an anti-inflammatory and stabilizing role by inhibiting the TLR4/MyD88/NF-κB signaling pathway.

2.
Artigo em Chinês | WPRIM | ID: wpr-1011450

RESUMO

ObjectiveTo investigate the therapeutic effect of Scutellariae Radix-Coptidis Rhizoma (SRCR) on atherosclerosis (AS) in mice and the effect of SRCR on macrophage pyroptosis in plaques via NOD-like receptor thermal protein domain-associated protein 3 (NLRP3) inflammasomes. MethodApoE-/- mice were fed with a high-fat diet for the modeling of AS and randomized into model, atorvastatin (5 mg·kg-1), and low-, medium-, and high-dose (1.95, 3.9, 7.8 g·kg-1, respectively) SRCR groups. Normal C57BL/6J mice were selected as the control group. After 8 weeks of administration, hematoxylin-eosin staining was used to observe the pathological status of the aortic plaque. The lipid accumulation in aortic plaque was observed by oil red O staining. The serum levels of total cholesterol (TC), triglyceride (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C) in mice were measured. Immunofluorescence double staining was employed to detect the co-localized expression of EGF-like module-containing mucin-like hormone receptor-like 1 (EMR1)/NLRP3 and EMR1/gasdermin D (GSDMD). The serum levels of interleukin-1β (IL-1β) and interleukin-18 (IL-18) were determined by enzyme-linked immunosorbent assay (ELISA). The protein levels of NLRP3, apoptosis-associated speck-like protein (ASC), Caspase-1, cleaved Caspase-1, GSDMD, N-terminus of GSDMD (GSDMD-NT), pro-IL-1β, IL-1β, and IL-18 were determined by Western blot, and the mRNA levels of NLRP3, ASC, Caspase-1, GSDMD, IL-1β, and IL-18 were determined by Real-time fluorescence quantitative polymerase chain reaction (Real-time PCR). ResultCompared with the control group, the model group showed obvious plaques, elevated serum levels of TG, TC, LDL-C, IL-1β, and IL-18 (P<0.01), lowered serum level of HDL-C (P<0.01), and up-regulated expression of NLRP3 inflammasomes and molecules related to pyroptosis in the aortic plaques (P<0.01). Compared with the model group, SRCR, especially at the medium and high doses, alleviated the plaque pathology, reduced the lipid content in plaques (P<0.05, P<0.01), recovered the serum lipid levels (P<0.05), reduced the macrophage recruitment (P<0.01), activation of NLRP3 inflammasomes, and pyroptosis in aortic root plaques (P<0.05), lowered the serum IL-1β and IL-18 levels (P<0.01), and down-regulated the protein levels of NLRP3, ASC, Caspase-1, cleaved Caspase-1, GSDMD, GSDMD-NT, pro-IL-1β, IL-1β, and IL-18 (P<0.05) and the mRNA levels of NLRP3, ASC, Caspase-1, GSDMD, IL-1β, and IL-18 in the aortic tissue (P<0.05). ConclusionSRCR exerts a therapeutic effect on high-fat diet-induced AS in mice by inhibiting the activation NLRP3 inflammasomes and reducing the pyroptosis of macrophages in plaques.

3.
Chinese Journal of Biotechnology ; (12): 702-710, 2015.
Artigo em Chinês | WPRIM | ID: wpr-240607

RESUMO

To explore the anti-tumor proliferation activity of human interleukin-29 (hIL-29) variant and based on bioinformatics analyzed data of hIL-29, a mutant gene hIL-29(mut33,35) was amplified by site-directed mutagenesis and megaprimer PCR. The hIL-29(mut33,35) was inserted into an eukaryotic expression plasmid pPIC9K and successfully expressed in Pichia pastoris GS115. A recombinant variant protein (rhIL-29(mut33,35)) was purified from the ferment supernatant of the engineering GS115. To observe the antineoplastic activity of the variant rhIL-29(mut33,35), a CCK-8 reagent was used to detect the anti-proliferation effect. Results show that it has strong anti-proliferation effect when acted on liver cancer cell BEL7402, colon cancer cell HCT8 and gastric cancer cell SGC7901. The inhibition ratios of the three tumor cells were (30.99 ± 1.58)%, (22.47 ± 1.37)% and (32.05 ± 2.02)%, respectively. In high dose group, the anti-proliferation effect of the rhIL-29(mut33,35) was stronger than that of wild type rhIL-29 (P < 0.01). This indicates the variant rhIL-29(mut33,35) has potential development value for medicine.


Assuntos
Humanos , Antineoplásicos , Farmacologia , Carcinoma Hepatocelular , Patologia , Linhagem Celular Tumoral , Interleucinas , Farmacologia , Neoplasias Hepáticas , Patologia , Mutagênese Sítio-Dirigida , Pichia , Plasmídeos , Reação em Cadeia da Polimerase , Proteínas Recombinantes , Farmacologia
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