PhD Scientific Days 2025

Pharmaceutical Sciences and Health Technologies I.

Functional Maintenance of Phase I Enzyme and Hepatobiliary Transporter Expression in Long-term Human Hepatocyte Culture

Előadó neve

Ms. Pacsuta Johanna, MSc

Neptun code

Q2OPF7

Előadó munkahelye

Semmelweis University/ Charles River Laboratories Hungary

Előadó telefonszáma

+36202183241

Előadó e-mail címe

pacsuta.johanna@phd.semmelweis.hu

Az előadás címe

Functional Maintenance of Phase I Enzyme and Hepatobiliary Transporter Expression in Long-term Human Hepatocyte Culture

Szerző(k) neve és munkahelye

Pacsuta Johanna^1,2,3, Mártonné-Tóth Beáta², Bárány Nándor², Gáborik Zsuzsanna², Jemnitz Katalin^2,3

1: Semmelweis University
2: Charles River Laboratories Hungary
3: HUN-REN TTK

Bemutatás módja

Szóbeli

Szekció

Pharmaceutical Sciences and Health Technologies I.

Language of the presentation

English

Preferred session

Pharmaceutical Sciences and Health Technologies

Összefoglaló szövege

The high number of drug withdrawals because of hepatotoxic effects had resulted in a strong need for better in vitro DILI (drug-induced liver injury) predictions, for which PHH (primary human hepatocyte) model has been recognized as the gold standard. But the rapid dedifferentiation and loss of functions of these cells in culture limits their predictivity, so recently great effort is focusing on the functional maintenance of PHH in vitro. In this regard, using a small molecule cocktail, namely 5C, has been described to inhibit dedifferentiation processes in PHH.
Our goal was to combine the advantages of 5C-treatment with culturing hepatocytes in sandwich configuration (SPHH) that would result in a more stable in vitro hepatic model suitable for long-term toxicology studies and high-throughput screening of drug candidates. In addition, we aimed to provide direct functional measurements, next to the gene expression profiling.
SPHH, SPHH-5C and PHH-5C were compared based on the gene expression of relevant Phase I CYP enzymes and hepatic transporters, measured by qPCR, while the metabolic activity and the uptake clearance of the cells were assessed by LC-MS/MS. The formation of biliary pockets was visualized using fluorescent substrates of biliary transporters and measured by High Content Imaging.
SPHH-5C cells showed stable mRNA expression of almost all CYP enzymes examined, while SPHH cells rapidly lost their function. Similarly, the hepatic transporter mRNA levels were best maintained in 5C-treated cultures. Also, CYP activity and hepatic clearance was the best preserved in SPHH-5C until 21 days. Furthermore, the bile canaliculi in SPHH-5C cultures had better efflux capacity than that in SPHH cultures.
The comprehensive findings from our research confirm that the optimized culturing conditions exerts a great influence on the stability of gene expression and function of vital CYP enzymes and hepatic transporters, making this model applicable for future developments. Herein, we provided an easy-to-culture, highly reliable, high-throughput compatible method for long-term maintenance of hepatic functions.
This research was supported by the 2024-2.1.2-EKÖP-KDP new National Excellence Program of the Ministry for Culture and Innovation from the source of the National Research, Development an Innovation Fund and was executed at Charles River Laboratories Hungary.

University

Semmelweis University

Supervisor

Jemnitz Katalin

Publication of my abstract

I do not give consent to the publication of my abstract on the website of the congress.

phd.section.field

in doctoral studies before complex exam (PhD)

Kind

Szabad

Status

elfogadva

Accepted presentation method

szóbeli

Előadás fájl jóváhagyás

nem hagyta jóvá

Előadó

9009

Start

15:00

End

15:15