An in vitro hepatic zonation model with a continuous oxygen gradient in a microdevice

Asako Sato, Kanae Kadokura, Hideyuki Uchida, Kosuke Tsukada

Research output: Contribution to journalArticle

12 Citations (Scopus)

Abstract

In a hepatic lobule, different sets of metabolic enzymes are expressed in the periportal (PP) and pericentral (PC) regions, forming a functional zonation, and the oxygen gradient is considered a determinant of zone formation. It is desirable to reproduce lobular microenvironment in vitro, but incubation of primary hepatocytes in conventional culture dishes has been limited at fixed oxygen concentrations due to technical difficulties. We designed a cell culture microdevice with an oxygen gradient to reproduce the hepatic microenvironment in vitro. The oxygen gradient during cell culture was monitored using a laser-assisted phosphorescence quenching method, and the cellular oxygen consumption rate could be estimated from changes in the gradient. Culture medium was continuously exchanged through microchannels installed in the device to maintain the oxygen gradient for a long term without transient hyper-oxygenation. The oxygen consumption rates of hepatocytes at 70.0 mmHg and 31.4 mmHg of partial oxygen pressure, which correspond to PP and PC regions in the microdevice, were 3.67 × 10-10 and 3.15 × 10-10 mol/s/106 cells, respectively. Antimycin A changed the oxygen gradient profile, indicating that cellular respiration can be estimated during cell culture. RT-PCR analysis of hepatocytes cultured under the oxygen gradient showed that mRNA expression of PEPCK and GK significantly increased in culture areas corresponding to PP and PC regions, respectively. These results indicate that the developed microdevice can reproduce the hepatic lobular microenvironment. The oxygen gradient in the microdevice can be closely controlled by changing the sizes of gas channels and the ambient oxygen concentration around the device; therefore, it could be expected to mimic the oxygen gradient of various organs, and it may be applicable to other pathological models.

Original languageEnglish
Pages (from-to)767-771
Number of pages5
JournalBiochemical and Biophysical Research Communications
Volume453
Issue number4
DOIs
Publication statusPublished - 2014 Oct 31

Fingerprint

Oxygen
Liver
Hepatocytes
Cell culture
Cell Culture Techniques
Oxygen Consumption
In Vitro Techniques
Cell Respiration
Antimycin A
Equipment and Supplies
Partial Pressure
Phosphorescence
Oxygenation
Culture Media
Microchannels
Lasers
Gases
Quenching
Polymerase Chain Reaction
Messenger RNA

Keywords

  • Hypoxia
  • Liver
  • Metabolic zonation
  • Microdevice
  • Oxygen gradient

ASJC Scopus subject areas

  • Biochemistry
  • Biophysics
  • Cell Biology
  • Molecular Biology
  • Medicine(all)

Cite this

An in vitro hepatic zonation model with a continuous oxygen gradient in a microdevice. / Sato, Asako; Kadokura, Kanae; Uchida, Hideyuki; Tsukada, Kosuke.

In: Biochemical and Biophysical Research Communications, Vol. 453, No. 4, 31.10.2014, p. 767-771.

Research output: Contribution to journalArticle

@article{d7e595c1987048ca934ba718219f8807,
title = "An in vitro hepatic zonation model with a continuous oxygen gradient in a microdevice",
abstract = "In a hepatic lobule, different sets of metabolic enzymes are expressed in the periportal (PP) and pericentral (PC) regions, forming a functional zonation, and the oxygen gradient is considered a determinant of zone formation. It is desirable to reproduce lobular microenvironment in vitro, but incubation of primary hepatocytes in conventional culture dishes has been limited at fixed oxygen concentrations due to technical difficulties. We designed a cell culture microdevice with an oxygen gradient to reproduce the hepatic microenvironment in vitro. The oxygen gradient during cell culture was monitored using a laser-assisted phosphorescence quenching method, and the cellular oxygen consumption rate could be estimated from changes in the gradient. Culture medium was continuously exchanged through microchannels installed in the device to maintain the oxygen gradient for a long term without transient hyper-oxygenation. The oxygen consumption rates of hepatocytes at 70.0 mmHg and 31.4 mmHg of partial oxygen pressure, which correspond to PP and PC regions in the microdevice, were 3.67 × 10-10 and 3.15 × 10-10 mol/s/106 cells, respectively. Antimycin A changed the oxygen gradient profile, indicating that cellular respiration can be estimated during cell culture. RT-PCR analysis of hepatocytes cultured under the oxygen gradient showed that mRNA expression of PEPCK and GK significantly increased in culture areas corresponding to PP and PC regions, respectively. These results indicate that the developed microdevice can reproduce the hepatic lobular microenvironment. The oxygen gradient in the microdevice can be closely controlled by changing the sizes of gas channels and the ambient oxygen concentration around the device; therefore, it could be expected to mimic the oxygen gradient of various organs, and it may be applicable to other pathological models.",
keywords = "Hypoxia, Liver, Metabolic zonation, Microdevice, Oxygen gradient",
author = "Asako Sato and Kanae Kadokura and Hideyuki Uchida and Kosuke Tsukada",
year = "2014",
month = "10",
day = "31",
doi = "10.1016/j.bbrc.2014.10.017",
language = "English",
volume = "453",
pages = "767--771",
journal = "Biochemical and Biophysical Research Communications",
issn = "0006-291X",
publisher = "Academic Press Inc.",
number = "4",

}

TY - JOUR

T1 - An in vitro hepatic zonation model with a continuous oxygen gradient in a microdevice

AU - Sato, Asako

AU - Kadokura, Kanae

AU - Uchida, Hideyuki

AU - Tsukada, Kosuke

PY - 2014/10/31

Y1 - 2014/10/31

N2 - In a hepatic lobule, different sets of metabolic enzymes are expressed in the periportal (PP) and pericentral (PC) regions, forming a functional zonation, and the oxygen gradient is considered a determinant of zone formation. It is desirable to reproduce lobular microenvironment in vitro, but incubation of primary hepatocytes in conventional culture dishes has been limited at fixed oxygen concentrations due to technical difficulties. We designed a cell culture microdevice with an oxygen gradient to reproduce the hepatic microenvironment in vitro. The oxygen gradient during cell culture was monitored using a laser-assisted phosphorescence quenching method, and the cellular oxygen consumption rate could be estimated from changes in the gradient. Culture medium was continuously exchanged through microchannels installed in the device to maintain the oxygen gradient for a long term without transient hyper-oxygenation. The oxygen consumption rates of hepatocytes at 70.0 mmHg and 31.4 mmHg of partial oxygen pressure, which correspond to PP and PC regions in the microdevice, were 3.67 × 10-10 and 3.15 × 10-10 mol/s/106 cells, respectively. Antimycin A changed the oxygen gradient profile, indicating that cellular respiration can be estimated during cell culture. RT-PCR analysis of hepatocytes cultured under the oxygen gradient showed that mRNA expression of PEPCK and GK significantly increased in culture areas corresponding to PP and PC regions, respectively. These results indicate that the developed microdevice can reproduce the hepatic lobular microenvironment. The oxygen gradient in the microdevice can be closely controlled by changing the sizes of gas channels and the ambient oxygen concentration around the device; therefore, it could be expected to mimic the oxygen gradient of various organs, and it may be applicable to other pathological models.

AB - In a hepatic lobule, different sets of metabolic enzymes are expressed in the periportal (PP) and pericentral (PC) regions, forming a functional zonation, and the oxygen gradient is considered a determinant of zone formation. It is desirable to reproduce lobular microenvironment in vitro, but incubation of primary hepatocytes in conventional culture dishes has been limited at fixed oxygen concentrations due to technical difficulties. We designed a cell culture microdevice with an oxygen gradient to reproduce the hepatic microenvironment in vitro. The oxygen gradient during cell culture was monitored using a laser-assisted phosphorescence quenching method, and the cellular oxygen consumption rate could be estimated from changes in the gradient. Culture medium was continuously exchanged through microchannels installed in the device to maintain the oxygen gradient for a long term without transient hyper-oxygenation. The oxygen consumption rates of hepatocytes at 70.0 mmHg and 31.4 mmHg of partial oxygen pressure, which correspond to PP and PC regions in the microdevice, were 3.67 × 10-10 and 3.15 × 10-10 mol/s/106 cells, respectively. Antimycin A changed the oxygen gradient profile, indicating that cellular respiration can be estimated during cell culture. RT-PCR analysis of hepatocytes cultured under the oxygen gradient showed that mRNA expression of PEPCK and GK significantly increased in culture areas corresponding to PP and PC regions, respectively. These results indicate that the developed microdevice can reproduce the hepatic lobular microenvironment. The oxygen gradient in the microdevice can be closely controlled by changing the sizes of gas channels and the ambient oxygen concentration around the device; therefore, it could be expected to mimic the oxygen gradient of various organs, and it may be applicable to other pathological models.

KW - Hypoxia

KW - Liver

KW - Metabolic zonation

KW - Microdevice

KW - Oxygen gradient

UR - http://www.scopus.com/inward/record.url?scp=84910011720&partnerID=8YFLogxK

UR - http://www.scopus.com/inward/citedby.url?scp=84910011720&partnerID=8YFLogxK

U2 - 10.1016/j.bbrc.2014.10.017

DO - 10.1016/j.bbrc.2014.10.017

M3 - Article

C2 - 25445587

AN - SCOPUS:84910011720

VL - 453

SP - 767

EP - 771

JO - Biochemical and Biophysical Research Communications

JF - Biochemical and Biophysical Research Communications

SN - 0006-291X

IS - 4

ER -