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1.
J Cell Biochem ; 120(3): 4255-4267, 2019 03.
Article in English | MEDLINE | ID: mdl-30269356

ABSTRACT

Mitochondrial dysfunction plays a central role in hepatic ischemia-reperfusion injury (IRI). The significance of mitophagy in hepatic IRI remains poorly understood. The mechanisms that cause IRI are complex, and many factors are involved in the injury formation process. The miR-330-3p mediates cell proliferation, cell death, and metabolism in various organisms. In this study, the levels of miR-330-3p were significantly downregulated in hepatic IRI, and the number of autophagosomes was increased in response to IRI as obtained under both in vivo and in vitro conditions. These results demonstrate that a reduction in miR-330-3p expression represents an important factor involved with promoting hepatic IRI. Moreover, we found that miR-330-3p interacted with phosphoglycerate mutase family member 5 (PGAM5) to regulate mitophagy. In specific, an overexpression of miR-330-3p diminished PGAM5 levels, which promoted mitophagy in response to IRI. In contrast, a downregulation of miR-330-3p was associated with increased PGAM5 levels leading to increased mitophagy. In conclusion, miR-330-3p suppresses PGAM5-induced mitophagy to alleviate hepatic IRI. Such findings not only reveal some of the mechanistic basis for this microRNA in liver injury, but also provide a foundation for new therapeutic approaches in the treatment of this condition.


Subject(s)
Liver/injuries , MicroRNAs/metabolism , Mitophagy , Phosphoprotein Phosphatases/metabolism , Reperfusion Injury/metabolism , Animals , Cell Line , Disease Models, Animal , Hepatocytes/metabolism , Male , Mice , Mice, Inbred C57BL , MicroRNAs/genetics , Transfection
2.
Biomed Pharmacother ; 82: 304-11, 2016 Aug.
Article in English | MEDLINE | ID: mdl-27470367

ABSTRACT

The earthworm is a widely used Chinese herbal medicine. There are more than 40 prescriptions including earthworms in the "Compendium of Materia Medica". TCM theory holds that earthworms exert antispasmodic and antipyretic effects through the liver meridian to calm the liver. However, the clinical effect of earthworms on liver injury has not been clearly demonstrated. We have previously established a method to extract the active ingredients from earthworms (hereinafter referred to as EWAs) [1]. In the present study, we observed protective effect of the EWAs on tunicamycin-induced ERS (endoplasmic reticulum stress) model in human hepatic L02 cells. The results showed that the EWAs promote proliferation and reduced apoptosis of ERS model in L02 cells (P<0.01). The up-regulation of ERS-related proteins, including PERK (protein kinase RNA-like endoplasmic reticulum kinase), eIF2a (eukaryotic translation initiation factor 2a), ATF4 (activating transcription factor 4) and CHOP (CCAAT/enhancer binding protein homologous protein), in L02 cell under ERS was inhibited by treatment of the EWAs (P<0.01). In summary, our data suggest the EWAs can significant attenuate ERS-induced hepatocyte injury via PERK-eIF2a-ATF4 pathway.


Subject(s)
Endoplasmic Reticulum Stress/drug effects , Liver/injuries , Liver/pathology , Oligochaeta/chemistry , Protective Agents/pharmacology , Activating Transcription Factor 4/genetics , Activating Transcription Factor 4/metabolism , Animals , Apoptosis/drug effects , Cell Line , Cell Proliferation/drug effects , Down-Regulation/drug effects , Eukaryotic Initiation Factor-2/genetics , Eukaryotic Initiation Factor-2/metabolism , Humans , Liver/drug effects , Models, Biological , RNA, Messenger/genetics , RNA, Messenger/metabolism , Transcription Factor CHOP/genetics , Transcription Factor CHOP/metabolism , Tunicamycin/pharmacology , Up-Regulation/drug effects , eIF-2 Kinase/metabolism
3.
Zhong Yao Cai ; 35(6): 863-6, 2012 Jun.
Article in Chinese | MEDLINE | ID: mdl-23236815

ABSTRACT

OBJECTIVE: To investigate the effect of Gecko crude peptides (GCPS) on human liver carcinoma HepG2 cells and its mechanism. METHODS: MTT assay was used to analyze the effect of the GCPS on the proliferation of HepG2 Cell; Nucleus change of HepG2 treated with GCP was observed by Hoechst33258 fluorescence staining, and BAX and BCL-2 were detected with western-blot assay. RESULTS: GCPS could inhibit the proliferation of HepG2 Cell in a time and dosage dependent way, and its half-maximal inhibitory concentration (IC50) was 1.2 mg/mL; HepG2 pretreated with GCPS showed apoptotic morphological changes. GCPS (1.6 mg/mL, 0.8 mg/mL) could decrease the expression of BCL-2 protein, and increase the expression of BAX protein. CONCLUSION: GCPS can inhibit the proliferation of HepG2 cell. The mechanism may be related to the induction apoptosis of HepG2.


Subject(s)
Antineoplastic Agents/pharmacology , Apoptosis/drug effects , Cell Proliferation/drug effects , Lizards , Materia Medica/pharmacology , Peptides/pharmacology , Animals , Blotting, Western , Dose-Response Relationship, Drug , Hep G2 Cells , Humans , Proto-Oncogene Proteins c-bcl-2/metabolism , Staining and Labeling/methods , Time Factors , bcl-2-Associated X Protein/metabolism
4.
J Physiol Sci ; 62(1): 29-43, 2012 Jan.
Article in English | MEDLINE | ID: mdl-22125186

ABSTRACT

Heat stress will stimulate cells of living organisms to generate heat shock proteins (Hsps). In the mouse liver, impacts of heat stress on hepatocyte proliferation, apoptosis and metabolism have not been studied systematically at different temperatures. In this research, the test mice were heated to 40, 42, 44 and 46°C, respectively, for 20 min and recovered at room temperature for 8 h in normal feeding conditions; the control animals were kept at room temperature without heat stress. The expression levels of Hsp70, Pcna, Bax, Bcl2, cytochrome P450 1A2 (CYP1A2), CYP2E1 and analog of CYP3A4 (not reported in mouse before), the parameters reflecting stress strength, cell proliferation, apoptosis and metabolism, were detected by western blotting, immunohistochemistry and semi-quantitative RT-PCR in test and control mice. Haematoxylin-eosin (H&E) staining and TUNEL analysis were further used to study the impacts of heat stress at different temperatures on hepatocellular necrosis and apoptosis. Serum AST and ALT levels, the markers of liver injury, were measured after heat stress at different temperatures. The data show that Hsp70 expression was significantly increased when temperature increased (P < 0.05). At lower temperatures (40 or 42°C), expression of Pcna, CYP1A2 and analog of CYP3A4 were considerably increased (P < 0.05) while hepatocyte necrosis and apoptosis were not induced (P > 0.05). At higher temperatures (44 or 46°C), expression of Pcna was decreased while hepatocyte necrosis and apoptosis were induced (P < 0.05). Expressions of CYP1A2 and analog of CYP3A4 were decreased especially at 46°C (P < 0.05). Expression of CYP2E1 could not be detected to increase at 40°C but was at high levels at 42, 44 and 46°C (P < 0.05). Expressions of AST and ALT were not different between the test mice and control mice at 40°C while they were significantly higher in the test mice than those in the control mice at 42 (P < 0.05), 44 and 46°C (P < 0.01). In conclusion, heat stress at lower temperatures promotes hepatocyte proliferation and improves the metabolic efficiency in mouse liver while heat stress at higher temperatures inhibits hepatocyte proliferation, promotes hepatocyte apoptosis and induces hepatocyte necrosis. This may give a hint to understanding human liver injury in high temperatures. Moreover, it is the first time that the analog of CYP3A4 was detected in mouse hepatocellular cytoplasm. It is worthwhile to dissect its function in future work.


Subject(s)
Apoptosis , Cell Proliferation , Heat Stress Disorders/physiopathology , Hepatocytes/metabolism , Hepatocytes/pathology , Alanine Transaminase/blood , Animals , Aspartate Aminotransferases/blood , Cytochrome P-450 CYP1A2/biosynthesis , Cytochrome P-450 CYP2E1/biosynthesis , Cytochrome P-450 CYP3A/biosynthesis , Female , HSP70 Heat-Shock Proteins/biosynthesis , Hot Temperature , In Situ Nick-End Labeling , Liver , Male , Mice , Mice, Inbred BALB C , Necrosis , Proliferating Cell Nuclear Antigen/biosynthesis , Proto-Oncogene Proteins c-bcl-2/biosynthesis , RNA, Messenger/metabolism , bcl-2-Associated X Protein/biosynthesis
5.
Zhong Yao Cai ; 34(7): 1020-3, 2011 Jul.
Article in Chinese | MEDLINE | ID: mdl-22066391

ABSTRACT

OBJECTIVE: To explore the proliferation inhibition effects of Gecko alcohol extract (GAE) on human esophageal squamous carcinoma cell line EC-109 and its mechanism. METHODS: The inhibitory effects of GAE on proliferation of EC-109 cells were measured by MTT. Nucleolus change of apoptotic cells was observed by Hoechest33342 fluorescence staining. Apoptosis rate of EC-109 cells was detected by flow cytometry. The expressions of apoptosis protein Caspase-3 and FAS in EC-109 cells were investigated by immunohistochemistry. RESULTS: GAE had the inhibition effects on the proliferation of esophageal carcinoma cell EC-109. The apoptosis rate of EC-109 cell treated with GAE(3.0 mg/mL, 4.0 mg/mL) for 48h was 20.63% and 39.73%, respectively. Compared with control group,the expression of Fas and Caspase-3 was significantly up-regulated in GAE treated group. CONCLUSION: GAE can inhibit the proliferation of esophageal carcinoma EC-109 cells and induce them apoptosis which may be correlated with increasing expression of protein Fas and Caspase-3.


Subject(s)
Antineoplastic Agents/pharmacology , Cell Proliferation/drug effects , Lizards , Materia Medica/pharmacology , Animals , Antineoplastic Agents/administration & dosage , Antineoplastic Agents/chemistry , Apoptosis/drug effects , Carcinoma, Squamous Cell/pathology , Caspase 3/metabolism , Cell Line, Tumor , Cell Survival/drug effects , Esophageal Neoplasms/pathology , Ethanol/chemistry , Flow Cytometry , Humans , Immunohistochemistry , Materia Medica/administration & dosage , Materia Medica/chemistry , Up-Regulation , fas Receptor/metabolism
6.
Parasitol Res ; 104(2): 429-35, 2009 Jan.
Article in English | MEDLINE | ID: mdl-18923843

ABSTRACT

Actin gene of Trypanosoma evansi (STIB 806) was cloned and expressed in Escherichia coli. The predicted amino acid sequence of T. evansi actin shows 100%, 98.7%, and 93.1%, homology with Trypanosoma equiperdum, Trypanosoma brucei brucei, and Trypanosoma cruzi. Recombinant actin was expressed as inclusion bodies in E. coli. It was purified and renatured for immunological studies. Mice immunized with the renatured recombinant actin were protected from lethal challenge with T. evansi STIB 806, T. equiperdum STIB 818, and T. b. brucei STIB 940, showing 63.3%, 56.7%, and 53.3% protection, respectively. Serum collected from the rabbit immunized with recombinant actin inhibited the growth of T. evansi, T. equiperdum, and T. b. brucei in vitro cultivation. Serum from mice and rabbits immunized with recombinant actin only recognized T. evansi actin but not mouse actin. The results of this study suggest that the recombinant T. evansi actin induces protective immunity against T. evansi, T. equiperdum, and T. b. brucei infection and may be useful in the development of a vaccine with other cytoskeletal proteins to prevent animal trypanosomiasis caused by these three trypanosome species.


Subject(s)
Actins/immunology , Protozoan Proteins/immunology , Protozoan Vaccines/immunology , Trypanosomiasis/prevention & control , Actins/genetics , Animals , Antibodies, Protozoan/immunology , Cloning, Molecular , Escherichia coli/genetics , Gene Expression , Mice , Mice, Inbred BALB C , Molecular Sequence Data , Protozoan Proteins/genetics , Rabbits , Survival Analysis , Vaccines, Subunit/immunology , Vaccines, Synthetic/immunology
7.
World J Gastroenterol ; 14(25): 3990-6, 2008 Jul 07.
Article in English | MEDLINE | ID: mdl-18609682

ABSTRACT

AIM: To investigate the anti-tumor effect of Chinese medicine Gecko on human esophageal carcinoma cell lines and xenografted sarcoma 180 in Kunming mice and its mechanism. METHODS: The serum pharmacological method was used in vitro. The growth rates of the human esophageal carcinoma cells (EC9706 or EC1) were measured by a modified 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The transplanted tumor model of the mouse S180 sarcoma was established. Fifty mice were randomly divided into five groups (n=10). Three Gecko groups were treated respectively with oral administration of Gecko powder at a daily dose of 13.5 g/kg, 9 g/kg, and 4.5 g/kg. The negative group (NS group) was treated with oral administration of an equal volume of saline and the positive group (CTX group) was treated with 100 mg/kg Cytoxan by intraperitoneal injection at the first day. After 2 wk of treatment, the anti-tumor activity was evaluated by tumor tissue weighing. The impact on immune organ was detected based on the thymus index, spleen index, phagocytic rate and phagocytic index. The protein expression of vascular endothelial growth factor (VEGF) and basic fibroblast growth factor (bFGF) were detected by immunohistochemistry. The cell apoptotic rate was detected by terminal deoxynucleotidyl transferase (TdT)-mediated dUTP nick end labeling (TUNEL) assay. RESULTS: The OD value in each group treated with Gecko after 72 h was reduced significantly in EC9706 and in EC1. The tumor weight in each group of Gecko was decreased significantly (1.087+/-0.249 vs 2.167+/-0.592; 1.021+/-0.288 vs 2.167+/-0.592; 1.234+/-0.331 vs 2.167+/-0.592; P<0.01, respectively). However, the thymus index and Spleen index of mice in Gecko groups had no significant difference compared with the NS group. The immunoreactive score of VEGF and bFGF protein expression of each Gecko group by immunohistochemical staining were lowered significantly. The apoptosis index (AI) of each group was increased progressively with increase of dose of Gecko by TUNEL. CONCLUSION: Gecko has anti-tumor effects in vitro and in vivo; induction of tumor cell apoptosis and the down-regulation of protein expression of VEGF and bFGF may be contributed to anti-tumor effects of Gecko.


Subject(s)
Antineoplastic Agents/pharmacology , Cell Proliferation/drug effects , Esophageal Neoplasms/pathology , Lizards , Medicine, Chinese Traditional , Sarcoma 180/drug therapy , Administration, Oral , Animals , Antineoplastic Agents/administration & dosage , Apoptosis/drug effects , Cell Line, Tumor , Cell Survival/drug effects , Dose-Response Relationship, Drug , Down-Regulation , Female , Fibroblast Growth Factor 2/metabolism , Humans , Mice , Rats , Rats, Sprague-Dawley , Sarcoma 180/metabolism , Sarcoma 180/pathology , Spleen/drug effects , Thymus Gland/drug effects , Time Factors , Vascular Endothelial Growth Factor A/metabolism , Xenograft Model Antitumor Assays
8.
Article in Chinese | MEDLINE | ID: mdl-21162200

ABSTRACT

AIM: To elucidate the location and effects of transcription factor-nuclear factor-kappaB (NF-kappaB) in lung tissues of rats with chronic obstructive pulmonary disease (COPD). METHODS: Fourteen male Wistar rats were randomly divided into COPD model and control groups equally. The COPD model was established by intratracheal instillation of lipopolysaccharide twice and exposure to cigarette smoke daily. We detected the NF-kappaB p65 protein in lung by immunohistochemical method, and the expression of NF-kappaB p65 mRNA in lung by in situ hybridization. RESULTS: Immunohistochemistry, the expression of NF-kappaB p65 protein in alveolar, bronchiolar epithelium and arteriolar endothelium was significantly higher in the COPD group (0.426 +/- 0.007, 0.434 +/- 0.012 and 0.313 +/- 0.007, respectively) than those of the control group (0.115 +/- 0.006, 0.116 +/- 0.005 and 0.095 +/- 0.007, respectively, all P < 0.01). In situ hybridization showed that the expressions of NF-kappaB p65 mRNA in alveolar epithelium (0.203 +/- 0.008), bronchiolar and arteriolar smooth muscle cell (0.208 + 0. 010 and 0.206 + 0.007) of rats in the COPD group were stronger than those in the control group (0.100 +/- 0.006, 0.102 +/- 0.002 and 0.103 +/- 0.003 respectively) by semiquantitative analysis (all P < 0.01). CONCLUSION: The expression and nuclear translocation of NF-kappaB may be the basis event of gene expression of many cytokines and inflammatory mediators, which may positively regulate gene expression of many cytokines and inflammatory mediators in various cell lines.


Subject(s)
Lung/metabolism , Pulmonary Disease, Chronic Obstructive/metabolism , Transcription Factor RelA/metabolism , Animals , Lung/pathology , Male , Pulmonary Disease, Chronic Obstructive/pathology , Rats , Rats, Wistar
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