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The clay nano-particles gel, a novel differentiation matrix for human liver cells

The clay nano-particles gel, a novel differentiation matrix for human liver cells
The clay nano-particles gel, a novel differentiation matrix for human liver cells
Background:
Growing liver cells in 3D cultures potentially mimics the natural liver cell-cell interactions which favour long term cell viability, functionality and genetic stability. Clay nanoparticle hydrogels offer excellent biocompatibility and permeability for water-soluble metabolites, with the potential for self-organization in situ to allow delivery of growth factors. We have investigated the clay hydrogel as a novel culture matrix for culture of hepatocytes and hepatic stem/progenitor cells (HPCs).
Methods:
Liver specimens were obtained from patients undergoing liver surgery. Hepatocytes were extracted by a two-step collagenase protocol and EpCAM+ HPCs were isolated from the non-parenchymal cell population using magnetic beads cell sorting. Cells were encapsulated in clay (Laponite) nanoparticles hydrogel in Williams’ E conditioned medium. HPCs medium was supplemented with EGF, HGH and IL6 (10ng/ml each). Characterisation of HPCs was performed by immunofluorescence and analysis of mRNA expression by rtPCR. The differentiation of HPCs in clay hydrogel was compared with cells cultured in HydroMatrix peptide hydrogel, or on collagen over 5 days. Cell culture supernatant were tested for albumin, urea and LDH concentrations and MTT assays was performed.
Results:
Primary hepatocytes cultured in a clay hydrogel demonstrated low levels of LDH and survived up to 15 days. In contrast, hepatocytes cultured on collagen released high levels of LDH in the early days of culture and their survival was about 31.5% by day 3. Interestingly, hepatocytes grown in clay gels demonstrated a marked increase in albumin secretion with time (mean 100.2 ± 44.12 ng ml-1 at day 15), but cells cultured on collagen rapidly lost their ability to produce albumin (mean 26.44 ± 21.17 ng ml-1 at day 15). No marked difference was observed in urea production between both cultures.
EpCAM+HPCs cultured in clay hydrogel (without differentiation factors) showed high albumin levels, with a mean of 44.4 ±10.82 ng ml-1, at day 5 compared to 27.79 ± 16.61 ng ml-1 at day 3. Lower levels of albumin were released from cells grown on collagen or in HydroMatrix (mean 8.23 ± 0.73 ng ml-1, P = 0.282 and 14.37 ± 0.12 ng ml-1, P = 0.0003 respectively). Addition of 1% DMSO induced a minor increase in albumin levels (a mean of 54.18 ± SEM 14 ng ml-1) in the clay gel as did the wnt-3a (mean 51.05 ± SEM 11.55 ng ml-1).
Conclusion:
We show that laponite clay gels provide support for culture of primary human hepatocytes, with improved hepatocyte survival and synthesis properties.
Khedr, M.E.M.S.
37c876ff-7226-4d51-bd20-dfe0bbca3d73
Messaoudi, Walid, Mohamed
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Jonnalagadda, Umesh
b190719a-6c0c-4ba7-88c5-ccb18b709189
Abdelmotelb, Ahmed
47ec298b-4ec1-48e4-8a19-39f2235de4a5
Glynne-Jones, Peter
6ca3fcbc-14db-4af9-83e2-cf7c8b91ef0d
Hill, Martyn
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Khakoo, Salim
6c16d2f5-ae80-4d9b-9100-6bfb34ad0273
Abu-Hilal, Mohammed
2b7464e5-6a59-4bd8-bfe4-27a1918a5c5a
Khedr, M.E.M.S.
37c876ff-7226-4d51-bd20-dfe0bbca3d73
Messaoudi, Walid, Mohamed
d66ce719-d255-4197-b11c-39886853669a
Jonnalagadda, Umesh
b190719a-6c0c-4ba7-88c5-ccb18b709189
Abdelmotelb, Ahmed
47ec298b-4ec1-48e4-8a19-39f2235de4a5
Glynne-Jones, Peter
6ca3fcbc-14db-4af9-83e2-cf7c8b91ef0d
Hill, Martyn
0cda65c8-a70f-476f-b126-d2c4460a253e
Khakoo, Salim
6c16d2f5-ae80-4d9b-9100-6bfb34ad0273
Abu-Hilal, Mohammed
2b7464e5-6a59-4bd8-bfe4-27a1918a5c5a

Khedr, M.E.M.S., Messaoudi, Walid, Mohamed, Jonnalagadda, Umesh, Abdelmotelb, Ahmed, Glynne-Jones, Peter, Hill, Martyn, Khakoo, Salim and Abu-Hilal, Mohammed (2018) The clay nano-particles gel, a novel differentiation matrix for human liver cells. The British Association for the Study of the Liver: Annual Meeting, University of York, United Kingdom. 18 - 21 Sep 2018.

Record type: Conference or Workshop Item (Poster)

Abstract

Background:
Growing liver cells in 3D cultures potentially mimics the natural liver cell-cell interactions which favour long term cell viability, functionality and genetic stability. Clay nanoparticle hydrogels offer excellent biocompatibility and permeability for water-soluble metabolites, with the potential for self-organization in situ to allow delivery of growth factors. We have investigated the clay hydrogel as a novel culture matrix for culture of hepatocytes and hepatic stem/progenitor cells (HPCs).
Methods:
Liver specimens were obtained from patients undergoing liver surgery. Hepatocytes were extracted by a two-step collagenase protocol and EpCAM+ HPCs were isolated from the non-parenchymal cell population using magnetic beads cell sorting. Cells were encapsulated in clay (Laponite) nanoparticles hydrogel in Williams’ E conditioned medium. HPCs medium was supplemented with EGF, HGH and IL6 (10ng/ml each). Characterisation of HPCs was performed by immunofluorescence and analysis of mRNA expression by rtPCR. The differentiation of HPCs in clay hydrogel was compared with cells cultured in HydroMatrix peptide hydrogel, or on collagen over 5 days. Cell culture supernatant were tested for albumin, urea and LDH concentrations and MTT assays was performed.
Results:
Primary hepatocytes cultured in a clay hydrogel demonstrated low levels of LDH and survived up to 15 days. In contrast, hepatocytes cultured on collagen released high levels of LDH in the early days of culture and their survival was about 31.5% by day 3. Interestingly, hepatocytes grown in clay gels demonstrated a marked increase in albumin secretion with time (mean 100.2 ± 44.12 ng ml-1 at day 15), but cells cultured on collagen rapidly lost their ability to produce albumin (mean 26.44 ± 21.17 ng ml-1 at day 15). No marked difference was observed in urea production between both cultures.
EpCAM+HPCs cultured in clay hydrogel (without differentiation factors) showed high albumin levels, with a mean of 44.4 ±10.82 ng ml-1, at day 5 compared to 27.79 ± 16.61 ng ml-1 at day 3. Lower levels of albumin were released from cells grown on collagen or in HydroMatrix (mean 8.23 ± 0.73 ng ml-1, P = 0.282 and 14.37 ± 0.12 ng ml-1, P = 0.0003 respectively). Addition of 1% DMSO induced a minor increase in albumin levels (a mean of 54.18 ± SEM 14 ng ml-1) in the clay gel as did the wnt-3a (mean 51.05 ± SEM 11.55 ng ml-1).
Conclusion:
We show that laponite clay gels provide support for culture of primary human hepatocytes, with improved hepatocyte survival and synthesis properties.

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More information

Published date: 19 September 2018
Venue - Dates: The British Association for the Study of the Liver: Annual Meeting, University of York, United Kingdom, 2018-09-18 - 2018-09-21

Identifiers

Local EPrints ID: 432934
URI: http://eprints.soton.ac.uk/id/eprint/432934
PURE UUID: 01dac334-ba48-4541-9da4-3cb3471c25f8
ORCID for M.E.M.S. Khedr: ORCID iD orcid.org/0000-0001-9942-4409
ORCID for Peter Glynne-Jones: ORCID iD orcid.org/0000-0001-5684-3953
ORCID for Martyn Hill: ORCID iD orcid.org/0000-0001-6448-9448

Catalogue record

Date deposited: 01 Aug 2019 16:30
Last modified: 18 Feb 2021 17:20

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Contributors

Author: M.E.M.S. Khedr ORCID iD
Author: Walid, Mohamed Messaoudi
Author: Umesh Jonnalagadda
Author: Ahmed Abdelmotelb
Author: Martyn Hill ORCID iD
Author: Salim Khakoo
Author: Mohammed Abu-Hilal

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