A Standardized Collagen-Based Scaffold Improves Human Hepatocyte Shipment and Allows Metabolic Studies over 10 Days

Ruoß, Marc; Häussling, Victor; Schügner, Frank; damink, L.H.H. Olde; Lee, Serene M. L.; Ge, Liming; Ehnert, Sabrina; Nüssler, Andreas K.

doi:10.3390/bioengineering5040086

Cited by 30 publications

(35 citation statements)

References 70 publications

(106 reference statements)

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“…Next, we measured the CYP450 enzyme activity after incubation with epigenetic modifiers. Using LC /MS technology the metabolization of specific drugs was carried out [17]. The results of the activity measurements (Figure 6) showed only a modest increase of CYP3A4, CYP2D6 and CYP2C9 enzyme activities in HepG2 cells stimulated with 5-AZA and Vitamin C. Additional supplementation of insulin and hydrocortisone to the medium resulted in a further modest increase.…”

Section: Resultsmentioning

confidence: 99%

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Epigenetic Modifications of the Liver Tumor Cell Line HepG2 Increase Their Drug Metabolic Capacity

Ruoß

Damm

Vosough

et al. 2019

IJMS

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Although human liver tumor cells have reduced metabolic functions as compared to primary human hepatocytes (PHH) they are widely used for pre-screening tests of drug metabolism and toxicity. The aim of the present study was to modify liver cancer cell lines in order to improve their drug-metabolizing activities towards PHH. It is well-known that epigenetics is strongly modified in tumor cells and that epigenetic regulators influence the expression and function of Cytochrome P450 (CYP) enzymes through altering crucial transcription factors responsible for drug-metabolizing enzymes. Therefore, we screened the epigenetic status of four different liver cancer cell lines (Huh7, HLE, HepG2 and AKN-1) which were reported to have metabolizing drug activities. Our results showed that HepG2 cells demonstrated the highest similarity compared to PHH. Thus, we modified the epigenetic status of HepG2 cells towards ‘normal’ liver cells by 5-Azacytidine (5-AZA) and Vitamin C exposure. Then, mRNA expression of Epithelial-mesenchymal transition (EMT) marker SNAIL and CYP enzymes were measured by PCR and determinate specific drug metabolites, associated with CYP enzymes by LC/MS. Our results demonstrated an epigenetic shift in HepG2 cells towards PHH after exposure to 5-AZA and Vitamin C which resulted in a higher expression and activity of specific drug metabolizing CYP enzymes. Finally, we observed that 5-AZA and Vitamin C led to an increased expression of Hepatocyte nuclear factor 4α (HNF4α) and E-Cadherin and a significant down regulation of Snail1 (SNAIL), the key transcriptional repressor of E-Cadherin. Our study shows, that certain phase I genes and their enzyme activities are increased by epigenetic modification in HepG2 cells with a concomitant reduction of EMT marker gene SNAIL. The enhancing of liver specific functions in hepatoma cells using epigenetic modifiers opens new opportunities for the usage of cell lines as a potential liver in vitro model for drug testing and development.

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Section: Resultsmentioning

confidence: 99%

“…After the described incubation times, the supernatants were removed and frozen at −80 °C until measurement. The enzymatic activity was measured by the company Pharmacelsus, Saarbrücken, Germany using a LC/MS based methodology [17,43].…”

Section: Methodsmentioning

confidence: 99%

Epigenetic Modifications of the Liver Tumor Cell Line HepG2 Increase Their Drug Metabolic Capacity

Ruoß

Damm

Vosough

et al. 2019

IJMS

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“…By slightly modifying this formula the swelling ration can be determined. As described before, these formulas were used to characterize the scaffolds [36]. Using an analytical balance [g] the dry and wet (scaffolds immerse in water for 1 h) weight of the scaffolds was measured.…”

Section: Methodsmentioning

confidence: 99%

Impact of Four Protein Additives in Cryogels on Osteogenic Differentiation of Adipose-Derived Mesenchymal Stem Cells

et al. 2019

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Human adipose-derived mesenchymal stem/stromal cells (Ad-MSCs) have great potential for bone tissue engineering. Cryogels, mimicking the three-dimensional structure of spongy bone, represent ideal carriers for these cells. We developed poly(2-hydroxyethyl methacrylate) cryogels, containing hydroxyapatite to mimic inorganic bone matrix. Cryogels were additionally supplemented with different types of proteins, namely collagen (Coll), platelet-rich plasma (PRP), immune cells-conditioned medium (CM), and RGD peptides (RGD). The different protein components did not affect scaffolds’ porosity or water-uptake capacity, but altered pore size and stiffness. Stiffness was highest in scaffolds with PRP (82.3 kPa), followed by Coll (55.3 kPa), CM (45.6 kPa), and RGD (32.8 kPa). Scaffolds with PRP, CM, and Coll had the largest pore diameters (~60 µm). Ad-MSCs were osteogenically differentiated on these scaffolds for 14 days. Cell attachment and survival rates were comparable for all four scaffolds. Runx2 and osteocalcin levels only increased in Ad-MSCs on Coll, PRP and CM cryogels. Osterix levels increased slightly in Ad-MSCs differentiated on Coll and PRP cryogels. With differentiation alkaline phosphatase activity decreased under all four conditions. In summary, besides Coll cryogel our PRP cryogel constitutes as an especially suitable carrier for bone tissue engineering. This is of special interest, as this scaffold can be generated with patients’ PRP.

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“…While hydrogels such as the collagen sandwich or Matrigel consist of almost exclusive ECM proteins, they are often integrated directly into the scaffold matrix in scaffold-based 3D cultivation systems, or the scaffold is subsequently coated with them. In addition to the liver-specific ECM proteins, fibronectin, collagen type I and gelatin are often used [27,28]. Since gelatin is a byproduct of collagen hydrolysis, it contains the same RGD peptides.…”

Section: Physical and Biochemical Properties Of Scaffolds Used In LIVmentioning

confidence: 99%

Tissue Engineering in Liver Regenerative Medicine: Insights into Novel Translational Technologies

Heydari

Najimi

Mirzaei

et al. 2020

Cells

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Organ and tissue shortage are known as a crucially important public health problem as unfortunately a small percentage of patients receive transplants. In the context of emerging regenerative medicine, researchers are trying to regenerate and replace different organs and tissues such as the liver, heart, skin, and kidney. Liver tissue engineering (TE) enables us to reproduce and restore liver functions, fully or partially, which could be used in the treatment of acute or chronic liver disorders and/or generate an appropriate functional organ which can be transplanted or employed as an extracorporeal device. In this regard, a variety of techniques (e.g., fabrication technologies, cell-based technologies, microfluidic systems and, extracorporeal liver devices) could be applied in tissue engineering in liver regenerative medicine. Common TE techniques are based on allocating stem cell-derived hepatocyte-like cells or primary hepatocytes within a three-dimensional structure which leads to the improvement of their survival rate and functional phenotype. Taken together, new findings indicated that developing liver tissue engineering-based techniques could pave the way for better treatment of liver-related disorders. Herein, we summarized novel technologies used in liver regenerative medicine and their future applications in clinical settings.

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A Standardized Collagen-Based Scaffold Improves Human Hepatocyte Shipment and Allows Metabolic Studies over 10 Days

Cited by 30 publications

References 70 publications

Epigenetic Modifications of the Liver Tumor Cell Line HepG2 Increase Their Drug Metabolic Capacity

Epigenetic Modifications of the Liver Tumor Cell Line HepG2 Increase Their Drug Metabolic Capacity

Impact of Four Protein Additives in Cryogels on Osteogenic Differentiation of Adipose-Derived Mesenchymal Stem Cells

Tissue Engineering in Liver Regenerative Medicine: Insights into Novel Translational Technologies

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