In vitro and in vivo bioactivity of recombinant canine hepatocyte growth factor

Abstract

Hepatocyte growth factor (HGF) is crucial for the development and regeneration of the liver and offers a possible new therapeutic strategy for the treatment of canine liver disease. In this study, the in vitro and in vivo bioactivity of recombinant canine HGF (rcHGF) produced with a baculoviral expression system in insect cells was measured.In vitro rcHGF induced mitogenesis, motogenesis, and phosphorylated the HGF receptor c-MET and its downstream mediators PKB and ERK1/2 in two canine epithelial cell lines. After a partial hepatectomy (phx) in dogs, rcHGF increased phosphorylation of c-MET, PKB and ERK1/2. A moderate increase was seen with the cell cycle protein PCNA in rcHGF treated livers, but no HGF-induced increase in liver weight was seen 7 days after phx. Furthermore, rcHGF treated livers showed lower levels of the key mediator of apoptosis, caspase-3, at 7 days after phx. It is concluded that rcHGF is a biologically active protein in vitro and in vivo and the baculoviral expression system supplies sufficient amounts of rcHGF for future clinical studies in dogs.

Hepatic progenitor cells in canine and feline medicine: potential for regenerative strategies

“…However, immunohistochemical stainings for PCNA or Ki67 in various canine liver diseases show prominent proliferation of hepatocytes after experimental PHx and mild acute hepatitis, with moderate proliferation in chronic hepatitis. Conversely, HPC response was pronounced in chronic hepatitis, moderate in mild acute hepatitis and non-existent after PHx (Figure 2) [6,30]. The response pattern of HPCs to various types of liver disease in the dog appears to be comparable to human pathology and rodent experimental findings, and recent studies suggest a similar comparison for feline HPC response [6,7,31], [Unpublished observations section: Valtolina et al].…”

“…After partial hepatectomy (PHx), liver regeneration occurs through hepatocyte proliferation (many PCNA positive hepatocytes indicated by arrows) but the HPC remains quiescent (few K7 positive cells). In chronic hepatitis the proliferative capacity of hepatocytes is exhausted indicated by a few Ki67 positive hepatocytes and a prominent ductular reaction (K7 positive, indicated by arrows) [6,30]. …”

Recombinant hepatocyte growth factor treatment in a canine model of congenital liver hypoplasia

“…Furthermore, the mitogenic activity of recombinant feline HGF is similar to recombinant human HGF and recombinant canine HGF in canine liver cells in vitro (supporting information, Fig. S1) (18).…”

“…For RNA isolation and real‐time quantitative polymerase chain reaction (QPCR) measurements, liver biopsies were fixed in RNAlater (Applied Biosystems, Nieuwerkerk a/d IJssel, the Netherlands) for 24 h and then stored at −70 °C until use. RNA isolation and QPCR were performed as described previously (18). Gene expressions were normalized with the average gene expressions of the endogenous references β‐2‐microglobulin ( B2M ), hypoxanthine guanine phosphoribosyl transferase‐1 ( HPRT1 ), ribosomal protein S5 ( RPS5 ) and ribosomal protein S19 ( RPS19 ) (24).…”

Inhibition of the ubiquitin ligase activity improves the production of biologically active fusion protein HSA-HGF in Chinese hamster ovary cells

“…1 In vivo, HGF is a pleiotropic factor that stimulates proliferation, survival, motility, scattering and differentiation during growth and development. [2][3][4] Knock out mice have demonstrated that HGF is necessary for normal embryogenesis. 5 In clinically treatment, HGF exhibits pleiotropic effects and contributes to mitogenic, anti-inflammatory, anti-apoptotic and anti-fibrogenetic activities.…”