Transforming growth factor β1 stimulation improves activation of mouse primary hepatic stellate cells obtained using the improved culture method

doi:10.3969/j.issn.2095-4344.2015.51.007

Chinese Journal of Tissue Engineering Research ›› 2015, Vol. 19 ›› Issue (51): 8234-8240.doi: 10.3969/j.issn.2095-4344.2015.51.007

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Transforming growth factor β1 stimulation improves activation of mouse primary hepatic stellate cells obtained using the improved culture method

Bi Xiao-juan^{1, 2}, Zhang Chuan-shan¹, Li Liang¹, Lv Guo-dong¹, Lin Ren-yong^{1, 2}

¹Clinical Medicine Research Institute, State Key Laboratory Incubation Base of Xinjiang Major Diseases Research, Xinjiang Key Laboratory of Echinococcosis, the First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, Xinjiang Uygur Autonomous Region, China; ²Department of Biochemistry and Molecular Biology, Preclinical Medicine College, Xinjiang Medical University, Urumqi 830011, Xinjiang Uygur Autonomous Region, China

Received:2015-11-02 Online:2015-12-10 Published:2015-12-10
Contact: Lin Ren-yong, M.D., Researcher, Clinical Medicine Research Institute, State Key Laboratory Incubation Base of Xinjiang Major Diseases Research, Xinjiang Key Laboratory of Echinococcosis, the First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, Xinjiang Uygur Autonomous Region, China; Department of Biochemistry and Molecular Biology, Preclinical Medicine College, Xinjiang Medical University, Urumqi 830011, Xinjiang Uygur Autonomous Region, China
About author:Bi Xiao-juan, Assistant researcher, Clinical Medicine Research Institute, State Key Laboratory Incubation Base of Xinjiang Major Diseases Research, Xinjiang Key Laboratory of Echinococcosis, the First Affiliated Hospital of Xinjiang Medical University, Urumqi 830054, Xinjiang Uygur Autonomous Region, China; Department of Biochemistry and Molecular Biology, Preclinical Medicine College, Xinjiang Medical University, Urumqi 830011, Xinjiang Uygur Autonomous Region, China
Supported by:
the National Natural Science Foundation of China, No. 81371838; the Natural Science Foundation of Xinjiang Uygur Autonomous Region, No. 2015211C084

Abstract

Abstract:

BACKGROUND: Hepatic stellate cells are the key cell type in the liver to secrete the extracellular matrix; therefore, it is important in the study of liver fibrosis.
OBJECTIVE: To improve the primary culture method of hepatic stellate cells so as to obtain an adequate amount of cells that are subjected to transforming growth factor β1 stimulation to study the expression of fibrogenic factors.
METHODS: Mice were anesthetized using ketamine to isolate the liver using in situ liver perfusion through puncture via the portal vein. Then, hepatic stellate cells were isolated using the density gradient centrifugation method. Primary hepatic stellate cells were identified by morphological observation and immunofluorescence staining. At 24 hours after culture, hepatic stellate cells were processed with 10 μg/L transforming growth factor
β1 (experimental group) and PBS (control group) for 48 hours, respectively.
RESULTS AND CONCLUSION: Mouse primary hepatic stellate cells were successfully harvested by using in situ liver perfusion and enzymatic digestion methods. The positive rate of cells for trypan blue staining was (97.2±0.8)%, and the purity of cells for immunofluorescence staining was (90.4±1.2)%. The total cell count was about 2.5×106 per mouse. Real-time quantitative PCR assay showed that compared with the control group, the mRNA expressions of smooth muscle actin α, type I collagen, transforming growth factor β receptor 1 and transforming growth factor β receptor 2 were significantly increased in the experimental group (P < 0.05). These findings indicate that the use of improved culture method has more efficient access to high-quality primary hepatic stellate cells, and transforming growth factor β1 stimulation can lead to the activation of hepatic stellate cells to secrete fibrogenic factors.

中国组织工程研究杂志出版内容重点：组织构建；骨细胞；软骨细胞；细胞培养；成纤维细胞；血管内皮细胞；骨质疏松；组织工程

CLC Number:

R318

Bi Xiao-juan, Zhang Chuan-shan, Li Liang, Lv Guo-dong, Lin Ren-yong . Transforming growth factor β1 stimulation improves activation of mouse primary hepatic stellate cells obtained using the improved culture method[J]. Chinese Journal of Tissue Engineering Research, 2015, 19(51): 8234-8240.

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Transforming growth factor β1 stimulation improves activation of mouse primary hepatic stellate cells obtained using the improved culture method

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