Coagulant activity of recombinant human factor VII produced by lentiviral human F7 gene transfer in immortalized hepatocyte-like cell line

Abstract

Human mesenchymal stem cells (hMSCs) have the potential to differentiate into hepatocyte-like cells, indicating that these cells may be the new target cell of interest to produce biopharmaceuticals. Our group recently established a hMSC-derived immortalized hepatocyte-like cell line (imHC) that demonstrates several liver-specific phenotypes. However, the ability of imHC to produce coagulation factors has not been characterized. Here, we examined the potential for imHC as a source of coagulation protein production by investigating the ability of imHC to produce human factor VII (FVII) using a lentiviral transduction system. Our results showed that imHC secreted a low amount of FVII (~22 ng/mL) into culture supernatant. Moreover, FVII from the transduced imHC (0.11 ± 0.005 IU/mL) demonstrated a similar coagulant activity compared with FVII from transduced HEK293T cells (0.12 ± 0.004 IU/mL) as determined by chromogenic assay. We demonstrate for the first time, to the best of our knowledge, that imHC produced FVII, albeit at a low level, indicating the unique characteristic of hepatocytes. Our study suggests the possibility of using imHC for the production of coagulation proteins.

Site-directed mutagenesis of tissue factor pathway inhibitor–binding exosite D60A on factor VII results in a new factor VII variant with lower coagulant activity

“…FVII wild type (WT) was constructed as described previously [ 48 ]. Human FVII WT complementary DNA (cDNA) was ligated into the pTG19-T polymerase chain reaction cloning vector (Vivantis Technologies), which was used as a template for mutant FVII D60A cloning.…”

Role of Nrf2 signaling in development of hepatocyte-like cells

“…Fortunately, unlike most other organs, the introduction of exogenous hepatocytes into the liver parenchyma is a relatively simple undertaking, suggesting that the liver is highly amenable to tissue therapy using induced pluripotent stem cell-derived hepatocytes (8,9). Several studies showed that human mesenchymal stem cells (MSCs) derived from adipose tissue, bone marrow, or umbilical cord blood are capable of differentiating into hepatocyte-like cells (HLCs) in specialized in vitro culture conditions (10)(11)(12)(13)(14). Recent studies also demonstrated that hepatocytes derived from human adipose stem cells (hADSCs) are a potentially scalable and applicable alternative to human hepatocytes (13,(15)(16)(17)(18).…”

Role of Nrf2 Signaling in Development of Hepatocyte-Like Cells

“…Fortunately, unlike most other organs, the introduction of exogenous hepatocytes into the liver parenchyma is a relatively simple undertaking, suggesting that the liver is highly amenable to tissue therapy using induced pluripotent stem cell-derived hepatocytes 8,9 . Several studies showed that human mesenchymal stem cells (MSCs) derived from adipose tissue, bone marrow, or umbilical cord blood are capable of differentiating into hepatocyte-like cells (HLCs) in specialized in vitro culture conditions [10][11][12][13][14] . Recent studies also demonstrated that hepatocytes derived from human adipose stem cells (hADSCs) are a potentially scalable and applicable alternative to human hepatocytes 13,15−18 .…”