The protective effect of the earthworm active ingredients on hepatocellular injury induced by endoplasmic reticulum stress.

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.

[Apoptotic mechanism of gecko crude peptides on human liver carcinoma cell line HepG2].

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.

[Inhibitory effects of Gecko alcohol extract on human esophageal squamous carcinoma cell line EC-109 proliferation and associated mechanism].

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.

Antitumor effect and mechanism of Gecko on human esophageal carcinoma cell lines in vitro and xenografted sarcoma 180 in Kunming mice.

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.