New method for constructing extracellular matrix bioscaffold derived from human cirrhotic liver tissues

Qiang XIONG; Yuhua DENG; Zhenzhen ZHANG; Mingman ZHANG; Yingcun LI; Quan KANG; Xiaoke DAI

doi:10.16016/j.1000-5404.201903204

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New method for constructing extracellular matrix bioscaffold derived from human cirrhotic liver tissues

Basic Medicine | 更新时间：2026-01-09

- New method for constructing extracellular matrix bioscaffold derived from human cirrhotic liver tissues
- Journal of Third Military Medical University Vol. 41, Issue 18, Pages: 1758-1762(2019)
- 作者机构：
  
  400014 重庆，重庆医科大学附属儿童医院肝胆外科，儿童发育疾病研究教育部重点实验，儿童发育重大疾病国家国际科技合作基地
- 作者简介：
- 基金信息：
  
  the National Natural Science Foundation for Young Scholars of China(81501736);the Medical Project funded jointly by Chongqing Science and Technology Commission and Chongqing Health Commission(2018ZDXM041);the Fourth Batch of Young and Middle-aged Senior Backup Talents of Chongqing Health Commission(2018年)
- DOI：10.16016/j.1000-5404.201903204
  CLC： R318.08;R322.47;R329-33
- Received：24 March 2019，
  
  Revised：2019-06-02，
  
  Published：30 September 2019
- 稿件说明：
移动端阅览
Qiang XIONG, Yuhua DENG, Zhenzhen ZHANG, et al. New method for constructing extracellular matrix bioscaffold derived from human cirrhotic liver tissues[J]. Journal of Third Military Medical University, 2019, 41(18): 1758-1762.
DOI：

Qiang XIONG, Yuhua DENG, Zhenzhen ZHANG, et al. New method for constructing extracellular matrix bioscaffold derived from human cirrhotic liver tissues[J]. Journal of Third Military Medical University, 2019, 41(18): 1758-1762. DOI： 10.16016/j.1000-5404.201903204.

摘要

目的

通过理化方法制备人源性肝硬化组织细胞外基质(extracelluar matrix，ECM)生物支架，并利用肝硬化组织ECM对人原代肝细胞(primary human hepatocytes，PHH)进行立体培养，建立以肝硬化组织ECM为基础的细胞立体培养平台。

方法

收集肝移植患者移除的硬化肝脏，肝素生理盐水灌洗去除血细胞，将肝脏切割成125 mm

组织块。依次将组织块置于超纯水、去污剂中反复振荡，破坏细胞膜、DNA及免疫源性物质，制备出肝硬化组织ECM。采用悬滴法将PHH植入ECM进行细胞培养，连续培养9 d后，组织学检测PHH植入生物支架情况。qPCR比较离体培养及平

面培养模型下肝细胞的特异性基因表达差异。

结果

去除细胞的肝硬化组织呈无色透明，HE染色及扫描电镜未发现残留细胞核，残余DNA鉴定显示DNA残留约为10 ng/mg，提示该法成功去除肝硬化组织原有细胞及基因组DNA。将人PHH接种入人源性肝硬化组织ECM，连续培养9 d后，HE染色发现PHH能够植入人源性肝硬化组织ECM中并生长良好，立体培养条件下，PHH特异性基因表达水平明显高于传统平面培养。

结论

本研究利用人源性肝硬化组织成功制备无细胞ECM生物支架，并证实其在细胞立体培养的可行性。

Abstract

Objective

To develop a novel decellularized extracellular matrix (ECM) bioscaffold derived from human cirrhotic liver tissues and construct a 3-dimensional cell culture system using this bioscaffold material.

Methods

Clinical samples of cirrhotic liver tissues were collected from patients undergoing liver transplantation for hepatic cirrhosis. The cirrhotic liver tissues were cleansed

cut into small tissue blocks (5 mm×5 mm×5 mm)

and agitated in deionized water and detergents to obtain decellularized cirrhotic liver scaffold. Human primary human hepatocytes (PHHs) were seeded into the prepared ECM scaffold and cultured continuously for 9 d. Histological studies of the seeded cells were performed to assess the cell engraftment

and qPCR was used to detect the expression of the liver-specific genes in the cultured cells.

Results

The cirrhotic liver tissue became transparent after decellularization

and HE staining and scanning electron microscopy showed no residual nuclei in the bioscaffold. Residual DNA identification showed that the scaffold material had a residue DNA level less than 10 ng/mg of the tissue. After 9 d of continuous culture in the ECM scaffold

the PHHs showed good engraftment as shown by HE staining; the results of qPCR showed that the cells cultured in this 3D culture system expressed higher levels of liver-specific genes as compared with the cells in conventional 2D culture.

Conclusion

We successfully construct decellularized ECM bioscaffold derived from human cirrhotic liver tissues and confirmed the feasibility of 3D cell culture using this new bioscaffold material.

关键词

Keywords

references

MOON H, CHO K, SHIN S, et al . High risk of hepato-cellular carcinoma development in fibrotic liver: role of the hippo-YAP/TAZ signaling pathway[J ] . Int J Mol Sci, 2019, 20(3): E581. DOI: 10.3390/ijms20030581.

ELPEK G Ö. Cellular and molecular mechanisms in the pathogenesis of liver fibrosis: An update[J]. World J Gastroenterol, 2014, 20(23): 7260-7276. DOI: 10.3748/wjg.v20.i23.7260.

JUNG Y K, YIM H J. Reversal of liver cirrhosis: current evidence and expectations[J]. Korean J Int Med, 2017, 32(2): 213-228. DOI: 10.3904/kjim.2016.268.

CRAPO P M, GILBERT T W, BADYLAK S F. An overview of tissue and whole organdecellularization processes[J]. Biomaterials, 2011, 32(12): 3233-3243. DOI:10.1016/j.biomaterials.2011.01.057.

MCCLELLAND R, WAUTHIER E, URONIS J, et al . Gradients in the liver's extracellular matrix chemistry from periportal to pericentral zones: influence on human hepatic progenitors[J ] . Tissue Eng Part A, 2008, 14(1): 59-70. DOI: 10.1089/ten.a.2007.0058.

SUZUKI A, IWAMA A, MIYASHITA H, et al . Role for growth factors and extracellular matrix in controlling differentiation of prospectively isolated hepatic stem cells[J ] . Development, 2003, 130(11): 2513-2524. DOI: 10.1242/dev.00459.

MAZZA G, AL-AKKAD W, TELESE A, et al . Rapid production of human liver scaffolds for functional tissue engineering by high shear stress oscillation-decellularization[J ] . Sci Rep, 2017, 7(1): 5534. DOI: 10.1038/s41598-017-05134-1.

SCHUPPAN D, RUEHL M, SOMASUNDARAM R, et al . Matrix as a modulator of hepatic fibrogenesis[J ] . Semin Liver Dis, 2001, 21(3): 351-372. DOI: 10.1055/s-2001-17556.

ENGLER A J, SEN S, SWEENEY H L, et al . Matrix elasticity directs stem cell lineage specification[J ] . Cell, 2006, 126(4): 677-689. DOI: 10.1016/j.cell.2006.06.044.

WELLS R G. The role of matrix stiffness in regulating cell behavior[J]. Hepatology, 2008, 47(4): 1394-1400. DOI:10.1002/hep.22193.

CHEN Y X, WONG P P, SJEKLOCHA L, et al . Mature hepatocytes exhibit unexpected plasticity by direct dedifferentiation into liver progenitor cells in culture[J ] . Hepatology, 2012, 55(2): 563-574. DOI: 10.1002/hep.24712.

BAPTISTA P M, SIDDIQUI M M, LOZIER G, et al . The use of whole organ decellularization for the generation of a vascularized liver organoid[J ] . Hepatology, 2011, 53(2): 604-617. DOI: 10.1002/hep.24067.

MAZZA G, ROMBOUTS K, RENNIE HALL A, et al . Decellularized human liver as a natural 3D-scaffold for liver bioengineering and transplantati on[J ] . Sci Rep, 2015, 5: 13079. DOI: 10.1038/srep13079.

MATTEI G, MAGLIARO C, PIRONE A, et al .Decellularized human liver is too heterogeneous for designing a generic extracellular matrix mimic hepatic scaffold[J ] . Artif Organs, 2017, 41(12): E347-E355. DOI: 10.1111/aor.12925.

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