Bie Jia Jian pill enhances the amelioration of bone mesenchymal stem cells on hepatocellular carcinoma progression.

BACKGROUND: The therapeutic efficiency of Traditional Chinese Medicine (TCM) in suppressing the recurrence and metastasis of hepatocellular carcinoma (HCC) has been well proved. OBJECTIVE: The aim of this study is to investigate the role of Bie Jia Jian pill (BJJP) combined with bone mesenchymal stem cells (BMSCs) in HCC progression. METHODS: Flow cytometry was used to identify BMSCs isolated from BALB/c mice. The expressions of biomarkers and apoptosis rate of cancer stem cells (CSCs) enriched from Huh7 cells were also measured. The osteogenic differentiation and adipogenic differentiation ability of isolated BMSCs was determined by oil red O staining and Alizarin Red Staining. CSCs were used to establish the orthotopic HCC model. Histological changes in the liver tissues were examined by hematoxylin-eosin (H&E) staining and Van Gieson (VG) staining. The cell apoptotic rate in the cancer tissues was detected by TUNEL staining. The cell proliferation antigen Ki67 in the cancer tissues were detected by immunofluorescence assay and PCR, respectively. The levels of CSCs cellular surface markers (CD24, CD133 and EpCAM) and Wnt/ß-catenin signal pathway related proteins were detected by PCR and western blot. RESULTS: Treatment of BJJP or BMSCs both improved the morphology induced by HCC and suppressed the differentiation ability of CSCs, as evidenced by down-regulated expressions of CD24, CD133, EpCAM and Ki67. The protective effect of BJJP or BMSCs in cancer tissues can be enhanced by the combination of BJJP and BMSCs. In addition to that, BJJP or BMSCs alone was found to increase the expression of miR-140 and promote cell apoptosis in CSCs, while down-regulation of miR-140 partially reversed the protective effect of BMSCs or BJJP + BMSCs on cancer tissues. BJJP + BMSCs treatment together also can down-regulate the expressions of Wnt3a and ß-catenin. CONCLUSIONS: These results proved the inhibitory role of BJJP + BMSCs in HCC development through regulating miR-140 and Wnt/ß-catenin signal pathway.

Bie Jia Jian Pill Combined with Bone Mesenchymal Stem Cells Regulates microRNA-140 to Suppress Hepatocellular Carcinoma Stem Cells.

BACKGROUND AND OBJECTIVES: Cancer stem cells (CSCs) with tumorigenic potential are reported as the crucial factors of hepatocellular carcinoma (HCC) recurrence and therapy resistance. Bone mesenchymal stem cells (BMSCs) are documented to play an important role in the protection of hepatocytes. Bie Jia Jian pill (BJJP), a Traditional Chinese Medicine, has been used to treat liver fibrosis and liver cancer. This study aimed to explore the potential role of combined use of BJJP with BMSCs in HCC cell lines. METHODS AND RESULTS: Flow cytometry was used to identify BMSCs isolated from BALB/c mice and CSCs enriched from Huh7 cells by measuring CD24, CD133, CD44, CD73, CD105, CD166, CD29, CD14 and CD34. Differentiation potential of BMSCs was also determined. Cell viability and proliferation ability of CSCs were determined by CCK-8 assay and clone formation assay. The expressions of CSCs biomarkers and Wnt/ß-catenin signal pathway related proteins were determined by PCR and western blot. TOP-Flash/FOP-Flash luciferase assay was applied to measure the activity of ß-catenin/TCF. Compared with untreated CSCs, BJJP or BMSCs treatment alone on CSCs lead to increased miR-140 expression and cell apoptosis, as well as decreased expressions of CD24, CD133, EpCAM and cell viability. Downregualted expressions of Wnt/ß-catenin signal pathway related proteins, Wnt3a and ß-catenin were found in response to BJJP or BMSCs treatment alone. The combination of BJJP＋BMSCs treatment on CSCs could further enhance the suppressive effect on CSCs. Down-regulation of miR-140 in CSCs partially blocked the effects of BMSCs or BMSCs＋BJJP on the expressions of Wnt3a and ß-catenin as well as the cell viability and apoptosis of CSCs. Reversed expression pattern was found in CSCs transfected with miR-140 overexpression. CONCLUSIONS: Taken together, we demonstrate that BJJP＋BMSCs together could further enhance the suppressive effect on CSCs through regulating miR-140 and suppressing Wnt/ß-catenin signal pathway. This study demonstrated the potential of BJJP＋BMSCs in therapeutic treatment of HCC.

Function of JNK Signaling Pathway in Induction of Brain Ischemic Tolerance and Regulation of Shu-Xue Tong-Mai Capsule Pretreatment / 世界科学技术-中医药现代化

This study was aimed to explore the function of c-Jun N-terminal kinase (JNK) signaling pathway in the induction of brain ischemic tolerance, and observe the function of Shu-Xue Tong-Mai (SXTM) capsule pretreatment. Ischemic preconditioning was performed for 3 min on rats to induce cerebral ischemic tolerance. Rat model of cere-bral ischemia reperfusion (the ischemia pretreatment group, I/R group) was established 24 h later. Western blot was used to detect the protein expression of JNK and phosphorylation of c-Jun N-terminal kinase (P-JNK), comparing to the expression with the sham operation group, I/R group and SXTM capsule group. Tunel method was applied to de-tect the apoptosis of neurons. Relationship between expression of JNK, P-JNK and apoptosis of neurons was also studied. The results showed that compared with the model group, expressions of P-JNK in ischemia preconditioning group and SXTM group were declined significantly (P < 0.05); and the apoptosis of neurons quantity was also de-clined (P< 0.05). It was concluded that ischemia preconditioning can decrease the apoptosis of neurons in cerebral ischemia reperfusion, and improve neurologic function. Its mechanism related to the inhibition of JNK signaling path-way. SXTM capsule pretreatment can protect the cerebral by inhibiting the JNK signaling pathway.

Changes of protein kinase-like endoplasmic reficulum kinase and glucose-regulated protein 78 expression in rats after focal ischemic preconditioning / 中华神经科杂志

ObjectiveTo investigate the effect of focal ischemic preconditioning (IPC) on the expression of protein kinase-like endoplasmic reticulum kinase ( PERK ) and glucose-regulated protein 78 (GRP78) mRNA and protein after focal cerebral ischemia/reperfusion (I/R) in rats.MethodsAll 120 male SD rats were randomly divided into three groups: sham-operation group,middle cerebral artery occlusion (MCAO) group and brain ischemia preconditioning (BIP) group.Each group was further divided into 4 subgroups according to 12 h,1,2 and 3 d after I/R.The IPC models were made in order to measure the expression of PERK,GRP78 mRNA and protein by in situ hybridization and Western blot,and the apoptosis rate of neuron by flow cytometry. Results ①The expression of PERK mRNA increased and reached the peak at 12 h,then decreased continuously after 1 d.BIP could decrease its expression.The expression of PERK protein increased at 12 h and reached the peak at 24 h,then decreased continuously after 2 d.BIP could decrease its expression.②The expression both of GRP78 mRNA and its protein all increased and reached the peak at 12 h,then decreased continuously.BIP could increase their expression (mRNA:12 h: 136.70±9.53,F=32.265; 24 h:147.54 ±9.97,F=54.920; 2 d:158.16 ±9.44,F=45.374; 3d: 165.85±10.26,F=16.493,P＜0.05; protein:12 h: 1.319±0.116,F=5.619,P＜0.05; 24 h: 1.226±0.108,F=33.742,P＜0.01; 2 d:1.183 ±0.112,F =46.556,P ＜0.01; 3 d:1.115± 0.098,F =11.730,P＜0.05).③The rate of apoptosis neuron of rats in MCAO increased markedly at 12 h after reperfusion,and reached the peak at 1 d,then decreased continuously.BIP could decrease the rate of apoptosis neuron. Conclusion BIP can protect neurons through inhibiting the expression of PERK and inducing the expression of GRP78 after endoplasmic reticulum stress in rats.