RESUMO
Aristolochic acid (AA)-IIIa is an AA analog present in Aristolochiaceae plants. To evaluate the chronic toxicity of AA-IIIa, mice were intragastrically administered with media control, 1â¯mg/kg AA-IIIa, and 10â¯mg/kg AA-IIIa, and designated as the control (CTL), AA-IIIa low dose (AA-IIIa-L), and AA-IIIa high dose (AA-IIIa-H) groups, respectively. AA-IIIa was administered three times a week, every other day, for 24 weeks (24-week time point). Thereafter, some mice were sacrificed immediately, while others were sacrificed 29 or 50 weeks after AA-IIIa withdrawal (53- or 74-week time point). Serum and organs were collected for biochemical and pathological analyses, respectively. Whole-genome sequencing was performed on the kidney, liver, and stomach tissues of AA-IIIa-treated mice for single-nucleotide polymorphism (SNP) detection. AA-IIIa-H mice died at 66 weeks, and the remaining mice showed moribund conditions at the 69 weeks. AA-IIIa induced minor kidney tubule injury, fibroblast hyperplasia, and forestomach carcinoma in mice. Bladder, intestine, liver, heart, spleen, lung, and testis tissues were not pathologically altered by AA-IIIa. In addition, AA-IIIa increased the C:G > A:T mutation in the kidney; however, no SNP mutation changes were observed in the liver and forestomach tissues of AA-IIIa-H mice at the 24-week time point compared with control mice. Therefore, we suspect that AA-IIIa is potentially mutagenic for mice after overdose and long-term administration. On the other hand, the forestomach is a unique organ in mice, but it does not exist in humans; thus, we hypothesize that the stomach toxicity induced by AA-IIIa is not a suitable reference for toxicological evaluation in humans. We recommend that Aristolochiaceae plants containing AA-IIIa should be properly supervised, and overdosing and long-term administration of drugs containing AA-IIIa should be avoided.
Assuntos
Ácidos Aristolóquicos , Animais , Ácidos Aristolóquicos/toxicidade , Camundongos , Masculino , Rim/efeitos dos fármacos , Rim/patologia , Polimorfismo de Nucleotídeo Único , Feminino , Fígado/efeitos dos fármacos , Fígado/patologia , Estômago/efeitos dos fármacos , Estômago/patologia , Testes de Toxicidade Crônica/métodos , Relação Dose-Resposta a DrogaRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Kelisha capsules (KLS) are often used to treat acute diarrhoea, bacillary dysentery, heat stroke, and other diseases. One of its components, Asarum, contains aristolochic acid I which is both nephrotoxic and carcinogenic. However, the aristolochic acid (AA) content in KLS and its toxicity remain unclear. AIM OF THE STUDY: The aims of this study were to quantitatively determine the contents of five aristolochic acid analogues (AAAs) in Asarum and KLS, and systematically evaluate the in vivo toxicity of KLS in rats. MATERIALS AND METHODS: Ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) was used to determine the content of the five AAAs in Asarum and KLS. Sprague-Dawley rats were administered KLS at 0, 0.75, 1.5, and 3.0 g/kg respectively, and then sacrificed after 4 weeks of administration or after an additional 2 weeks of recovery. The endpoints assessed included body weight measurements, serum biochemistry and haematology indices, and clinical and histopathological observations. RESULTS: The AAAs content in Asarum sieboldii Miq. (HB-ESBJ) were much lower than those of the other Asarums. The contents of AA I, AA IVa, and aristolactam I in KLS were in the ranges of 0.03-0.06 µg/g, 1.89-2.16 µg/g, and 0.55-1.60 µg/g, respectively, whereas AA II and AA IIIa were not detected. None of the rats showed symptoms of toxic reactions and KLS was well tolerated throughout the study. Compared to the control group, the activated partial thromboplastin time values of rats in the 1.5 and 3.0 g/kg groups significantly reduced after administration (P < 0.05). In addition, the serum triglycerides of male rats in the 0.75 and 1.5 g/kg groups after administration, and the 0.75, 1.5, 3.0 g/kg groups after recovery were significantly decreased (P < 0.01 or P < 0.001). No significant drug-related toxicological changes were observed in other serum biochemical indices, haematology, or histopathology. CONCLUSIONS: The AA I content in KLS met the limit requirements (<0.001%) of the Chinese Pharmacopoeia. Therefore, it is safe to use KLS in the short-term. However, for safety considerations, attention should be paid to the effects of long-term KLS administration on coagulation function and triglyceride metabolism.
Assuntos
Rim , Ratos Sprague-Dawley , Animais , Masculino , Administração Oral , Rim/efeitos dos fármacos , Rim/patologia , Ratos , Asarum/química , Fígado/efeitos dos fármacos , Fígado/patologia , Cápsulas , Ácidos Aristolóquicos/toxicidade , Ácidos Aristolóquicos/administração & dosagem , Medicamentos de Ervas Chinesas/toxicidade , Medicamentos de Ervas Chinesas/administração & dosagem , Feminino , Espectrometria de Massas em TandemRESUMO
BACKGROUND: Acute lung injury (ALI) often leads to serious respiratory diseases with high incidence rates and mortality. For centuries, Xiebai San (XBS) has been a classical traditional Chinese medicine (TCM) about respiratory illness such as pneumonia in children. However, the related mechanism of XBS against ALI remains indistinct. PURPOSE: To reveal specific targets of XBS in lipopolysaccharide (LPS)-induced ALI mice using integrated pharmacology. STUDY DESIGN: The integrated method was to expound mechanism and targets of XBS inhibited ALI. METHODS: The primary components in XBS were identified by ultra high performance liquid chromatography-quadrupole time of flight-mass spectrometry (UHPLC-QTOF-MS). The potential drug targets were established using network pharmacology. The anti-ALI effect of XBS was evaluated in mice. Additionally, therapeutic targets were screened by integrating metabolome and transcriptome and verified in lung tissue. RESULTS: In total, 163 chemical components were identified in XBS, and a network of "3 drugs-18 components-86 targets" for XBS against ALI was constructed. In ALI mice, XBS alleviated lung inflammation by decreasing permeation and expression of neutrophils, tumor necrosis factor α (TNF-α), interleukin-6 (IL-6), and interleukin-1ß (IL-1ß) in bronchoalveolar lavage fluid (BALF), serum, and lung tissue. Next, the transcriptome of lung tissue was analyzed and enriched, indicating the importance of mitogen-activated protein kinase (MAPK), Janus kinase-signal transducer and activator of transcription (JAK-STAT), and others, which was consistent with network pharmacology prediction. Also, western blotting and immunohistochemistry results showed that XBS was against ALI mainly by inhibiting extracellular signal regulated kinase (ERK) and signal transducer and activator of transcription 3 (Stat3) phosphorylation. In addition, the metabolome of lung tissue revealed that XBS mainly regulated pathways involved in arachidonic acid, glycerophospholipid, and tryptophan metabolisms. The expression levels of leukotriene, phosphatidylcholine, kynurenine, and others were also verified. CONCLUSION: XBS alleviated inflammation of ALI by inhibiting the phosphorylation of the ERK/Stat3 pathway and regulating arachidonic acid, glycerophospholipid, and tryptophan metabolisms. This study will guide clinical precision medicine and promote modernization of XBS.
Assuntos
Lesão Pulmonar Aguda , Medicamentos de Ervas Chinesas , Fator de Transcrição STAT3 , Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/metabolismo , Animais , Fator de Transcrição STAT3/metabolismo , Medicamentos de Ervas Chinesas/farmacologia , Camundongos , Masculino , Fosforilação/efeitos dos fármacos , Lipopolissacarídeos , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Pulmão/efeitos dos fármacos , Pulmão/metabolismo , Camundongos Endogâmicos C57BL , Modelos Animais de Doenças , Farmacologia em Rede , Transdução de Sinais/efeitos dos fármacosRESUMO
Due to soared obesity population worldwide, hepatosteatosis is becoming a major risk factor for hepatocellular carcinoma (HCC). Undertaken molecular events during the progression of steatosis to liver cancer are thus under intensive investigation. In this study, we demonstrated that high-fat diet potentiated mouse liver AKT2. Hepatic AKT2 hyperactivation through gain-of-function mutation of Akt2 (Akt2E17K) caused spontaneous hepatosteatosis, injury, inflammation, fibrosis, and eventually HCC in mice. AKT2 activation also exacerbated lipopolysaccharide and D-galactosamine hydrochloride-induced injury/inflammation and N-Nitrosodiethylamine (DEN)-induced HCC. A positive correlation between AKT2 activity and SCD1 expression was observed in human HCC samples. Activated AKT2 enhanced the production of monounsaturated fatty acid which was dependent on SREBP1 upregulation of SCD1. Blockage of active SREBP1 and ablation of SCD1 reduced steatosis, inflammation, and tumor burden in DEN-treated Akt2E17K mice. Therefore, AKT2 activation is crucial for the development of steatosis-associated HCC which can be treated with blockage of AKT2-SREBP1-SCD1 signaling cascade.
Assuntos
Metabolismo dos Lipídeos , Neoplasias Hepáticas , Proteínas Proto-Oncogênicas c-akt , Estearoil-CoA Dessaturase , Proteína de Ligação a Elemento Regulador de Esterol 1 , Animais , Humanos , Masculino , Camundongos , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Carcinoma Hepatocelular/genética , Dieta Hiperlipídica/efeitos adversos , Fígado Gorduroso/metabolismo , Fígado Gorduroso/patologia , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Camundongos Endogâmicos C57BL , Proteínas Proto-Oncogênicas c-akt/metabolismo , Estearoil-CoA Dessaturase/metabolismo , Estearoil-CoA Dessaturase/genética , Proteína de Ligação a Elemento Regulador de Esterol 1/metabolismo , Proteína de Ligação a Elemento Regulador de Esterol 1/genéticaRESUMO
NASH is a highly prevalent metabolic syndrome that has no specific approved agents up to now. BBBP, which mainly contains bile acids, possess various pharmacological properties and some bile acids are available for NASH treatment. Herein, the therapeutic effects and underlying mechanisms of BBBP against NASH were systemically evaluated. In this study, mice received an HFHS diet over a 20-week period to induce NASH with or without BBBP intervention were used to evaluate the effect and underlying mechanisms of BBBP against NASH. Our results demonstrated that BBBP attenuated hepatic steatosis, reduced body weight gain and lipid concentrations, and improved sensitivity to insulin and tolerance to glucose in mice fed an HFHS diet. Metabolomics and transcriptomic analysis revealed that BBBP suppressed the arginine biosynthesis by up-regulating NOS3 expression and the PI3K-Akt signaling pathway was also regulated by BBBP, as indicated by 55 DEGs. Bioinformatic analysis predicted the regulatory effect of the FXR/PXR-PI3K-AKT-NOS3 axis on arginine biosynthesis-related metabolites. These results were further confirmed by the significantly increased mRNA and protein levels of NOS3, PI3K (Pik3r2), and AKT1. And the increased levels of arginine biosynthesis related-metabolites, such as urea, aspartic acid, glutamic acid, citrulline, arginine, and ornithine, were confirmed accurately based on targeted metabolomics analysis. Together, our study uncoded the complicated mechanisms of anti-NASH activities of BBBP, and provided critical evidence inspiring the discovery of innovative therapies based on BBBP in the treatment of NASH.
Assuntos
Hepatopatia Gordurosa não Alcoólica , Ursidae , Animais , Camundongos , Bile/metabolismo , Ácidos e Sais Biliares/metabolismo , Dieta , Fígado , Camundongos Endogâmicos C57BL , Hepatopatia Gordurosa não Alcoólica/metabolismo , Fosfatidilinositol 3-Quinases/metabolismo , Pós/farmacologia , Pós/uso terapêutico , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
Cheqianzi Decoction (CQD) is a Traditional Chinese Medicine (TCM) formula comprising four herbs and is recorded in the Ancient Materia Medica "Shengji Zonglu". Individually, these four herbs have been shown to reduce uric acid (UA) levels, to treat hyperuricemia (HUA), and alleviate kidney damage. However, the therapeutic efficacy of the CQD and related mechanism are not yet clear. In this study, high performance liquid chromatography (HPLC) analysis confirmed that the contents of the chemical components of the four herbal medicines were in accordance with the provisions of the Chinese Pharmacopoeia. A total of 99 potential targets were identified in the network pharmacology analysis of CQD, indicating its involvement in the regulation of inflammatory and apoptotic signaling pathways, and potential value for treating HUA and alleviating kidney injury. In vivo pharmacodynamic studies showed that compared with the Model group, significantly decreased levels of serum uric acid (SUA), serum creatinine (SCr), blood urea nitrogen (BUN) (all P < 0.05), and inflammatory factors (P < 0.01) were detected in the CQD group. Quantitative real-time PCR and Western blot analyses showed that compared with the Model group, adenosine triphosphate (ATP)-binding cassette efflux transporter G2 (ABCG2) expression in the CQD group was significantly upregulated (P < 0.01) at both the mRNA and protein levels, while mRNA expression of Caspase3 and NOD-like receptor family member 3 (NLRP3) (P < 0.05) and protein expression of NLRP3 (P < 0.01) were significantly downregulated. In conclusion, CQD promotes UA excretion by activating ABCG2, and induces inflammasome NLRP3-mediated reduction in inflammatory and apoptotic factors to achieve renal protection. Thus, our findings indicate the therapeutic potential of CQD in HUA with kidney injury.
RESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: The nephrotoxicity and carcinogenicity induced by traditional Chinese medicines (TCMs) containing aristolochic acids (AAs) and related compound preparations have greatly limited their clinical application. While the toxicity of AA-I and AA-II is relatively clear, there are marked differences in the toxic effects of different types of aristolochic acid analogues (AAAs). Thus, the toxicity of TCMs containing AAAs cannot be evaluated based on the toxicity of a single compound. AIM OF THE STUDY: To systematically investigate the toxicity induced by Zhushalian (ZSL), Madouling (MDL) and Tianxianteng (TXT) as representative TCMs derived from Aristolochia. MATERIALS AND METHODS: AAA contents in ZSL, MDL and TXT were determined using HPLC. Subsequently, mice were treated for 2 weeks with high (H) and low (L) dosages of TCMs containing total AAA contents of 3 mg/kg and 1.5 mg/kg, respectively. Toxicity was evaluated using biochemical and pathological examination and was based on organ indices. Correlations between AAA contents and induced toxicity were analysed using multiple methods. RESULTS: Of the total AAA content, ZSL contained mainly AA-I and AA-II (>90%, of which AA-I accounted for 49.55%). AA-I accounted for 35.45% in MDL. TXT mainly contained AA-IVa (76.84%) and other AAAs accounted for <10%. Short-term toxicity tests indicated that ZSL and high-dose MDL induced obvious renal interstitial fibrosis and gastric injury, whereas TXT (high and low dosages) caused only slight toxicity. Correlation analysis suggested that AA-I might be the critical hazard factor for toxicity. CONCLUSIONS: The toxicity of TCMs containing AAAs cannot be generalised. The toxicity of TXT is relatively low compared with those of ZSL and MDL. The toxicity of Aristolochia depends mainly on the AA-I content; therefore, control of AA-I levels in TCMs and related compound preparations is required to reduce the risk of toxicity associated with the use of Aristolochia herbs in clinical settings.
Assuntos
Aristolochia , Ácidos Aristolóquicos , Medicamentos de Ervas Chinesas , Nefropatias , Animais , Camundongos , Aristolochia/química , Ácidos Aristolóquicos/toxicidade , Nefropatias/induzido quimicamente , Cromatografia Líquida de Alta Pressão/métodos , Medicamentos de Ervas Chinesas/químicaRESUMO
BACKGROUND: The safety of herbs containing aristolochic acids (AAs) has become a widespread concern. Previous reports indicate that AAs are highly nephrotoxic and carcinogenic, although there are more than 170 analogues of aristolochic acid. Not all AAs have the same degree of nephrotoxicity or carcinogenicity. Previous studies have found that aristolochic acid IVa (AA-IVa), the principal component of AAs within members of the Aristolochiaceae family, especially Asarum, a commonly used herb in China, has essentially no significant nephrotoxicity. However, several studies, including ours, have shown that aristolochic acid I (AA-I) is clearly nephrotoxic. PURPOSE: The focus of the study was to elucidate the molecular mechanism responsible for the difference in nephrotoxicity between the AA-I and AA-IVa. STUDY DESIGN/METHOD: Mice were administered with AA-I or AA-IVa for 22 weeks through the oral route, followed by a 50-week recovery time. The kidney tissues of mice were extracted at the end of 22 weeks. Pathological examination and proteomic detection (tandem mass tagging (TMT) and phosphorylated proteomics) were performed on the kidney tissue to investigate the key signaling pathways and targets of AAs-induced renal interstitial fibrosis (RIF). The key signaling pathways and targets were verified by Western blot (WB), siRNA transfection, and luciferase assays. RESULTS: AA-I caused severe nephrotoxicity, high mortality, and extensive RIF. However, the same AA-IVa dosage exhibited almost no nephrotoxicity and does not trigger RIF. The activation of the p38-STAT3-S100A11 signaling pathway and upregulated expression of α smooth muscle actin (α-SMA) and Bcl2-associated agonist of cell death (Bad) proteins could be the molecular mechanism underlying AA-I-induced nephrotoxicity. On the other hand, AA-IVa did not regulate the activation of the p38-STAT3-S100A11 signaling pathway and had relatively little effect on the expression of α-SMA and Bad. Consequently, the difference in the regulation of p38-STAT3-S100A11 pathway, α-SMA, and Bad proteins between AA-I and AA-IVa may be responsible for the divergence in their level of nephrotoxicity. CONCLUSION: This is the first study to reveal the molecular mechanism underlying the difference in nephrotoxicity between AA-I and AA-IVa. Whether STAT3 is activated or not may be the key factor leading to the difference in nephrotoxicity between AA-I and AA-IVa.
Assuntos
Ácidos Aristolóquicos , Nefropatias , Camundongos , Animais , Ácidos Aristolóquicos/metabolismo , Ácidos Aristolóquicos/farmacologia , Proteômica , Nefropatias/metabolismo , Transdução de Sinais , Fibrose , Rim , Proteínas S100/metabolismo , Proteínas S100/farmacologiaRESUMO
BACKGROUND: The expression of pyruvate kinase muscle 2 (PKM2) is augmented in macrophages of patients with atherosclerotic coronary artery disease. The role of PKM2 in atherosclerosis is to be determined. METHODS: Global and myeloid cell-specific PKM2 knock-in mice with ApoE-/- background (ApoE-/- , PKM2KI/KI and Lyz2-cre, ApoE-/- , and PKM2flox/flox ) were produced to evaluate the clinical significance of PKM2 in atherosclerosis development. Wild-type and PKM2 knock-in macrophages were isolated to assess the function of PKM2 in macrophage phagocytosis. Atherosclerotic mice were treated with PKM2 inhibitor shikonin (SKN) to evaluate the therapeutic potential of PKM2 suppression in atherosclerosis. RESULTS: Oxidized low-density lipoprotein (oxLDL) upregulated PKM2 in macrophages. PKM2 in return promoted the uptake of oxLDL by macrophages. Overexpressed PKM2 accelerated atherosclerosis in mice. SKN blocked the progress of mouse atherosclerosis. CONCLUSIONS: PKM2 accelerates macrophage phagocytosis and atherosclerosis. Targeting PKM2 is a potential therapy for atherosclerosis.
Assuntos
Aterosclerose , Piruvato Quinase , Animais , Camundongos , Apolipoproteínas E/metabolismo , Aterosclerose/genética , Aterosclerose/metabolismo , Macrófagos/metabolismo , Músculos/metabolismo , Fagocitose , Piruvato Quinase/genética , Piruvato Quinase/metabolismoRESUMO
BACKGROUND: Largely due to incidental detection, asymptomatic pancreatic cystic lesions (PCLs) have become prevalent in recent years. Among them, intraductal papillary mucinous neoplasm (IPMN) infrequently advances to pancreatic ductal adenocarcinoma (PDAC). Conservative surveillance versus surgical intervention is a difficult clinical decision for both caregivers and PCL patients. Because RNF43 loss-of-function mutations and KRAS gain-of-function mutations concur in a subset of IPMN and PDAC, their biological significance and therapeutic potential should be elucidated. METHODS: Pancreatic Rnf43 knockout and Kras activated mice (Rnf43-/-; KrasG12D) were generated to evaluate their clinical significance in pancreatic pre-neoplastic initiation and malignant transformation. RESULTS: Loss of Rnf43 potentiated the occurrence and severity of IPMN and PDAC in oncogenic Kras mice. The Wnt/ß-catenin signaling pathway was activated in pancreatic KrasG12D and Rnf43 knockout mice and the PORCN inhibitor LGK974 blocked pancreatic IPMN initiation and progression to PDAC accordingly. CONCLUSIONS: Rnf43 is a tumor suppressor in the prevention of pancreatic malignant transformation. This genetically reconstituted autochthonous pancreatic Rnf43-/-; KrasG12D preclinical cancer model recapitulates the pathological process from pancreatic cyst to cancer in humans and can be treated with inhibitors of Wnt/ß-catenin signaling. Since the presence of RNF43 and KRAS mutations in IPMNs predicts future development of advanced neoplasia from PCLs, patients with these genetic anomalies warrant surveillance, surgery, and/or targeted therapeutics such as Wnt/ß-catenin inhibitors.
Assuntos
Carcinoma Ductal Pancreático , Neoplasias Intraductais Pancreáticas , Neoplasias Pancreáticas , Aciltransferases/genética , Animais , Carcinoma Ductal Pancreático/genética , Genes ras , Humanos , Proteínas de Membrana/genética , Camundongos , Neoplasias Intraductais Pancreáticas/genética , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas p21(ras)/genética , Ubiquitina-Proteína Ligases/genéticaRESUMO
Previous reports demonstrated that aristolochic acids (AAs) exposure-induced nephrotoxicity, mutations, and tumorigenesis are mainly due to aristolochic acid I (AAI). Notably, the chemical structure of aristolochic acid IVa (AAIVa), which exists at higher levels in many Aristolochiaceae herbs, is extremely similar to AAI. In lack of toxicological data, it is unknown whether AAIVa exposure leads to aristolochic acid nephropathy (AAN), mutations, and tumorigenesis as of AAI. To answer these questions, mice were administered AAIVa by single or repeated long-term gavage, while AAI was used as a positive control. We found that single gavage of 40 mg/kg of AAIVa exhibited no obvious toxicity. Also, there were no tumors or death in mice administrated with 1 and 10 mg/kg of AAIVa for 6 months followed by a 12-month recovery time. There were no noteworthy alterations in gene mutation frequency in the kidney, liver, and stomach between the AAIVa and control mice. Fascinatingly, AA-associated mutational signatures, adenine-to-thymine (A>T) transversions, were absent in AAIVa-treated mice. Nonetheless, 10 mg/kg of AAIVa triggered lymphocytic infiltration and slight fibrous hyperplasia in the kidney at the 6th month; however, these were alleviated at the 12th and 18th months. On the contrary, AAI (positive control) caused severe diffuse fibrosis, tubular atrophy, necrosis, tumors in the forestomach and kidney, and death after the 6th month. It seems that long-term AAIVa exposure induced mild renal lesions could be due to the activation of the canonical or noncanonical transforming growth factor-ß (TGFß) pathway. Overall, these findings suggest that the mutagenicity and carcinogenic risk of AAIVa are very low.
Assuntos
Ácidos Aristolóquicos/toxicidade , Nefropatias/induzido quimicamente , Animais , Ácidos Aristolóquicos/administração & dosagem , Ácidos Aristolóquicos/química , Carcinógenos/administração & dosagem , Carcinógenos/química , Carcinógenos/toxicidade , Relação Dose-Resposta a Droga , Feminino , Nefropatias/fisiopatologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Mutagênicos/administração & dosagem , Mutagênicos/química , Mutagênicos/toxicidade , Fatores de TempoRESUMO
Even though aberrant mechanistic target of rapamycin (mTOR) signaling is known to cause cardiomyopathy, its underlying mechanism remains poorly understood. Because augmentation of αB-crystallin and hspB2 was presented in the cortical tubers and lymphangioleiomyomatosis of tuberous sclerosis complex patients, we deciphered the role of αB-crystallin and its adjacent duplicate gene, hspB2, in hyperactive mTOR-induced cardiomyopathy. Cardiac Tsc1 deletion (T1-hKO) caused mouse mTOR activation and cardiomyopathy. Overexpression of αB-crystallin and hspB2 was presented in the hearts of these mice. Knockout of αB-crystallin/hspB2 reversed deficient Tsc1-mediated fetal gene expression, mTOR activation, mitochondrial damage, cardiomyocyte vacuolar degeneration, cardiomyocyte size, and fibrosis of T1-hKO mice. These cardiac-Tsc1; αB-crystallin; hspB2 triple knockout (tKO) mice had improved cardiac function, smaller heart weight to body weight ratio, and reduced lethality compared with T1-hKO mice. Even though activated mTOR suppressed autophagy in T1-hKO mice, ablation of αB-crystallin and hspB2 failed to restore autophagy in tKO mice. mTOR inhibitors suppressed αB-crystallin expression in T1-hKO mice and rat cardiomyocyte line H9C2. Starvation of H9C2 cells activated autophagy and suppressed αB-crystallin expression. Since inhibition of autophagy restored αB-crystallin expression in starved H9C2 cells, autophagy is a negative regulator of αB-crystallin expression. mTOR thus stimulates αB-crystallin expression through suppression of autophagy. In conclusion, αB-crystallin and hspB2 play a pivotal role in Tsc1 knockout-related cardiomyopathy and are therapeutic targets of hyperactive mTOR-associated cardiomyopathy.
Assuntos
Cardiomiopatias/metabolismo , Cristalinas/metabolismo , Proteínas de Choque Térmico HSP27/metabolismo , Proteínas de Choque Térmico/metabolismo , Miócitos Cardíacos/metabolismo , Animais , Cardiomiopatias/tratamento farmacológico , Cardiomiopatias/genética , Proteínas de Choque Térmico HSP27/efeitos dos fármacos , Proteínas de Choque Térmico HSP27/genética , Proteínas de Choque Térmico/efeitos dos fármacos , Inibidores de MTOR/farmacologia , Camundongos Knockout , Miócitos Cardíacos/efeitos dos fármacos , Regiões Promotoras Genéticas/genética , Serina-Treonina Quinases TOR/efeitos dos fármacos , Serina-Treonina Quinases TOR/metabolismoRESUMO
Context: Aristolochia manshuriensis Kom (Aristolochiaceae) (AMK) is known for toxicity and mutagenicity.Objective: The tumorigenic role of AMK has yet to be understood.Materials and methods: AMK extracts were extracted from root crude drug. SD (Sprague Dawley) rats underwent gavage with AMK (0.92 g/kg) every other day for 10 (AMK-10) or 20 (AMK-20) weeks. Stomach samples were gathered for histopathological evaluation, microarray and mRNA analysis.Results: The gastric weight to body weight ratio (GW/BW) is 1.7 in the AMK-10 cohort, and 1.8 in AMK-20 cohort compared to control (CTL) cohort. Liver function was damaged in AMK-10 and AMK-20 rats compared to CTL rats. There were no significant changes of CRE (creatinine) in AMK-10 and AMK-20 rats. Histopathological analysis revealed that rats developed dysplasia in the forestomach in AMK-10 rats, and became gastric carcinoma in AMK-20 rats. Genes including Mapk13, Nme1, Gsta4, Gstm1, Jun, Mgst2, Ggt6, Gpx2, Gpx8, Calml3, Rasgrp2, Cd44, Gsr, Dgkb, Rras, and Amt were found to be critical in AMK-10 and AMK-20 rats. Pik3cb, Plcb3, Tp53, Hras, Myc, Src, Akt1, Gnai3, and Fgfr3 worked in AMK-10 rats, and PDE2a and PDE3a played a pivotal role in AMK-20 rats.Discussion and conclusions: AMK induced benign or malignant gastric tumours depends on the period of AMK administration. Genes including Mapk13, Nme1, Gsta4, Gstm1, Jun, Mgst2, Ggt6, Gpx2, Gpx8, Calml3, Rasgrp2, Cd44, Gsr, Dgkb, Rras, Amt, Pik3cb, Plcb3, Tp53, Hras, Myc, Src, Akt1, Gnai3, Fgfr3, PDE2a, and PDE3a were found to be critical in aristolochic acid-induced gastric tumour process.
Assuntos
Aristolochia/química , Extratos Vegetais/toxicidade , Neoplasias Gástricas/induzido quimicamente , Animais , Ácidos Aristolóquicos/isolamento & purificação , Ácidos Aristolóquicos/toxicidade , Análise em Microsséries , Extratos Vegetais/administração & dosagem , Ratos , Ratos Sprague-Dawley , Neoplasias Gástricas/genética , Neoplasias Gástricas/patologia , Fatores de TempoRESUMO
A high incidence of hypersensitivity reactions (HSRs) largely limits the use of paclitaxel injection. Currently, these reactions are considered to be mediated by histamine release and complement activation. However, the evidence is insufficient and the molecular mechanism involved in paclitaxel injection-induced HSRs is still incompletely understood. In this study, a mice model mimicking vascular hyperpermeability was applied. The vascular leakage induced merely by excipients (polyoxyl 35 castor oil) was equivalent to the reactions evoked by paclitaxel injection under the same conditions. Treatment with paclitaxel injection could cause rapid histamine release. The vascular exudation was dramatically inhibited by pretreatment with a histamine antagonist. No significant change in paclitaxel injection-induced HSRs was observed in complement-deficient and complement-depleted mice. The RhoA/ROCK signaling pathway was activated by paclitaxel injection. Moreover, the ROCK inhibitor showed a protective effect on vascular leakage in the ears and on inflammation in the lungs. In conclusion, this study provided a suitable mice model for investigating the HSRs characterized by vascular hyperpermeability and confirmed the main sensitization of excipients in paclitaxel injection. Histamine release and RhoA/ROCK pathway activation, rather than complement activation, played an important role in paclitaxel injection-induced HSRs. Furthermore, the ROCK inhibitor may provide a potential preventive approach for paclitaxel injection side effects.
Assuntos
Antineoplásicos Fitogênicos/efeitos adversos , Hipersensibilidade a Drogas/etiologia , Hipersensibilidade a Drogas/metabolismo , Paclitaxel/efeitos adversos , Transdução de Sinais , Quinases Associadas a rho/metabolismo , Proteína rhoA de Ligação ao GTP/metabolismo , Animais , Antineoplásicos Fitogênicos/administração & dosagem , Biópsia , Ativação do Complemento/imunologia , Proteínas do Sistema Complemento/imunologia , Proteínas do Sistema Complemento/metabolismo , Modelos Animais de Doenças , Hipersensibilidade a Drogas/patologia , Feminino , Liberação de Histamina , Masculino , Camundongos , Paclitaxel/administração & dosagemRESUMO
ETHNOPHARMACOLOGICAL RELEVANCE: Aristolochia manshuriensis Kom (AMK), belonging to the Aristolochia family, is traditionally used in China to remove heart fire, promote dieresis, restore menstruation, and enhance milk secretion. The active constitutes of AMK are aristolochic acids (AAs, I and II) that are reported to cause serious side effects including nephrotoxicity and carcinogenicity. AIM OF THE STUDY: The tumorigenic role of AMK is far to be understood. We analyzed the toxicity reactions after long-term exposure of AMK in rats. MATERIALS AND METHODS: Sprague-Dawley rats underwent gavage with AMK doses of 51â¯mg/kg (AMK-1), 253â¯mg/kg (AMK-2), 508â¯mg/kg (AMK-3), 1029â¯mg/kg (AMK-4) or AAs of 15â¯mg/kg (AAs), and then sacrificed at the 6th, 10th, 14th, 18th, 22th, 26th and 30th weeks. Endpoint measurements included clinical observations, body weights, blood biochemistry, haematology and histomorphological observations. RESULTS: Body weight decreased after AMK or AAs treatment in rats. AMK destroyed renal function, and induced anemia in rats. AMK caused kidney, stomach, bladder and subcutaneous tumors in rats. In addition, primary hepatic carcinoma was not observed in rats. CONCLUSIONS: AMK had significant toxic effects in rats with regard to decreased body weight, diminished renal function, increased anemia and tumor incidence. Kidney, stomach, bladder and subcutaneous tissue are carcinogenic target organs of AMK or AAs, however liver is no- carcinogenic target organ of AMK or AAs in rats. AMK is carcinogenic in rats, and not be safe for humans.
Assuntos
Aristolochia , Carcinógenos/toxicidade , Neoplasias/induzido quimicamente , Extratos Vegetais/toxicidade , Administração Oral , Anemia/induzido quimicamente , Animais , Peso Corporal/efeitos dos fármacos , Feminino , Rim/efeitos dos fármacos , Rim/patologia , Rim/fisiologia , Fígado/efeitos dos fármacos , Fígado/patologia , Masculino , Neoplasias/patologia , Tamanho do Órgão/efeitos dos fármacos , Ratos Sprague-Dawley , Estômago/efeitos dos fármacos , Estômago/patologia , Testes de Toxicidade Crônica , Bexiga Urinária/efeitos dos fármacos , Bexiga Urinária/patologiaRESUMO
BACKGROUND: Myocardial infarction (MI) is a major disease burden. Wild-type p53-induced phosphatase 1 (Wip1) has been studied extensively in the context of cancer and the regulation of different types of stem cells, but the role of Wip1 in cardiac adaptation to MI is unknown. We investigated the significance of Wip1 in a mouse model of MI. METHODS: The study began in June 2014 and was completed in July 2016. We compared Wip1-knockout (Wip1-KO) mice and wild-type (WT) mice to determine changes in cardiac function and survival in response to MI. The heart weight/body weight (HW/BW) ratio and cardiac function were measured before MI. Mouse MI was established by ligating the left anterior descending (LAD) coronary artery under 1.5% isoflurane anesthesia. After MI, survival of the mice was observed for 4 weeks. Cardiac function was examined by echocardiography. The HW/BW ratio was analyzed, and cardiac hypertrophy was measured by wheat germ agglutinin staining. Hematoxylin and eosin (H&E) staining was used to determine the infarct size. Gene expression of interleukin-6 (IL-6), tumor necrosis factor-α (TNF-α), and interleukin-1ß (IL-1ß) was assessed by quantitative real-time polymerase chain reaction (qPCR), and the levels of signal transducers and activators of transcription 3 (stat3) and phosphor-stat3 (p-stat3) were also analyzed by Western blotting. Kaplan-Meier survival analysis, log-rank test, unpaired t-test, and one-way analysis of variance (ANOVA) were used for statistical analyses. RESULTS: Wip1-KO mice had a marginally increased HW/BW ratio and slightly impaired cardiac function before LAD ligation. After MI, Wip1-deficient mice exhibited increased mortality (57.14% vs. 29.17%; n = 24 [WT], n = 35 [Wip1-KO], P< 0.05), increased cardiac hypertrophy (HW/BW ratio: 7 days: 7.25 ± 0.36 vs. 5.84 ± 0.18, n = 10, P< 0.01, and 4 weeks: 6.05 ± 0.17 vs. 5.87 ± 0.24, n = 10, P > 0.05; cross-sectional area: 7 days: 311.80 ± 8.29 vs. 268.90 ± 11.15, n = 6, P< 0.05, and 4 weeks: 308.80 ± 11.26 vs. 317.00 ± 13.55, n = 6, P > 0.05), and reduced cardiac function (ejection fraction: 7 days: 29.37 ± 1.38 vs. 34.72 ± 1.81, P< 0.05, and 4 weeks: 19.06 ± 2.07 vs. 26.37 ± 2.95, P< 0.05; fractional shortening: 7 days: 13.72 ± 0.71 vs. 16.50 ± 0.94, P< 0.05, and 4 weeks: 8.79 ± 1.00 vs. 12.48 ± 1.48, P< 0.05; n = 10 [WT], n = 15 [Wip1-KO]). H&E staining revealed a larger infarct size in Wip1-KO mice than in WT mice (34.79% ± 2.44% vs. 19.55% ± 1.48%, n = 6, P< 0.01). The expression of IL-6 and p-stat3 was downregulated in Wip1-KO mice (IL-6: 1.71 ± 0.27 vs. 4.46 ± 0.79, n = 6, P< 0.01; and p-stat3/stat3: 1.15 ± 0.15 vs. 1.97 ± 0.23, n = 6, P< 0.05). CONCLUSION: The results suggest that Wip1 could protect the heart from MI-induced ischemic injury.
Assuntos
Infarto do Miocárdio/metabolismo , Infarto do Miocárdio/patologia , Proteína Fosfatase 2C/metabolismo , Animais , Ecocardiografia , Interleucina-1beta/metabolismo , Interleucina-6/metabolismo , Masculino , Camundongos , Camundongos Knockout , Infarto do Miocárdio/genética , Miócitos Cardíacos/metabolismo , Proteína Fosfatase 2C/deficiência , Proteína Fosfatase 2C/genética , Reação em Cadeia da Polimerase em Tempo Real , Fator de Necrose Tumoral alfa/metabolismo , Remodelação VentricularRESUMO
BACKGROUND: Cinobufacin injection, also known as huachansu, is a preparation form of Cinobufacini made from Cinobufacin extract liquid. Despite that Cinobufacin injection is shown to shrink liver and gastric tumors, improving patient survival and life quality, the effective components in Cinobufacin remain elusive. In this study, we aim to screen antitumor components from Cinobufacin injection to elucidate the most effective antitumor components for treatment of liver and gastric cancers. MATERIALS AND METHODS: High performance liquid chromatography (HPLC) and LC-MS/MS analysis were used to separate and determine the components in Cinobufacin injection. Inhibition rates of various components in Cinobufacin injection on liver and gastric cancer cells were determined with MTT assay; Hepatocellular carcinoma and gastric cancer models were used to assess the antitumor effect of the compounds in vivo. RESULTS: The major constituents in Cinobufacin injection include peptides, nucleic acids, tryptamines and bufotalins. MTT assay revealed that bufadienolides had the best antitumor activity, with peptides being the second most effective components. Bufadienolides showed significant inhibition rates on gastric and hepatocellular tumour growth in vivo. CONCLUSION: Bufadienolides are the most effective components in Cinobufacini injection for the treatment of liver and gastric cancers. This discovery can greatly facilitate further research in improving the therapeutic effects of Cinobufacin injection, meanwhile reducing its adverse reaction.
Assuntos
Venenos de Anfíbios/farmacologia , Antineoplásicos/farmacologia , Bufanolídeos/farmacologia , Neoplasias Experimentais/tratamento farmacológico , Venenos de Anfíbios/química , Animais , Antineoplásicos/química , Bufanolídeos/química , Células Hep G2 , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos ICR , Camundongos Nus , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Aristolochia manshuriensis Kom (AMK) is a member of the Aristolochiaceae family and is a well-known cause of aristolochic acid (AA) nephropathy. In this study, we investigated the potential of omeprazole (OM) to alleviate AMK-induced nephrotoxicity. We found that OM reduced mouse mortality caused by AMK and attenuated AMK-induced acute nephrotoxicity in rats. OM enhanced hepatic Cyp 1a1/2 and renal Cyp 1a1 expression in rats, as well as CYP 1A1 expression in human renal tubular epithelial cells (HKCs). HKCs with ectopic CYP 1A1 expression were more tolerant to AA than the control cells. Therefore, OM may alleviate AMK-mediated acute nephrotoxicity through induction of CYP 1A1. We suggest that the coadministration of OM might be beneficial for reducing of AA-induced nephrotoxicity.
Assuntos
Doença Aguda/terapia , Aristolochia/efeitos adversos , Nefropatias/tratamento farmacológico , Omeprazol/farmacologia , Animais , Ácidos Aristolóquicos , Citocromo P-450 CYP1A1/metabolismo , Células Epiteliais/efeitos dos fármacos , Células Epiteliais/metabolismo , Feminino , Humanos , Rim/efeitos dos fármacos , Rim/metabolismo , Nefropatias/metabolismo , Fígado/efeitos dos fármacos , Fígado/metabolismo , Fígado/microbiologia , Masculino , Camundongos , Camundongos Endogâmicos ICR , Ratos , Ratos Sprague-DawleyRESUMO
The mechanism of penicillin immediate hypersensitivity reactions has not been completely elucidated. These reactions are generally considered to be mediated by IgE, but penicillin-specific IgE could not be detected in most cases. This study demonstrated that penicillin was able to cause vascular hyperpermeability in a mouse model mimicking clinical symptoms of penicillin immediate hypersensitivity reactions. The first exposure to penicillin also induced immediate edema and exudative reactions in ears and lungs of mice in a dose-dependent manner. Vasodilation was noted in microvessels in ears. These reactions were unlikely to be immune-mediated reactions, because no penicillin-specific IgE was produced. Furthermore, penicillin treatment directly elicited rapid histamine release. Penicillin also led to F-actin reorganization in human umbilical vein endothelial cells and increased the permeability of the endothelial monolayer. Activation of the RhoA/ROCK signaling pathway was observed in ears and lungs of mice and in endothelial cells after treatment with penicillin. Both an anti-histamine agent and a ROCK inhibitor attenuated penicillin immediate hypersensitivity reactions in mice. This study presents a novel mechanism of penicillin immediate hypersensitivity reactions and suggests a potential preventive approach against these reactions.