Primary cultured hepatocytes of the bony fish, Oreochromis mossambicus, the tilapia: a valid tool for physiological studies on IGF-I expression in liver

Abstract

In spite of the importance of IGF-I for growth and development, knowledge about regulation of its production in submammalian species is rather limited. In order to create a tool for investigation of direct regulatory effects on the expression of IGF-I in bony fish liver, a primary cell culture of hepatocytes from Oreochromis mossambicus, the tilapia, was established. The cells were viable for up to 3 days and IGF-I mRNA synthesis was detected by northern blot and semiquantitative reverse transcriptase (RT)-PCR. Northern blot analysis of the primary cultured hepatocytes revealed four different IGF-I transcripts, 0·5, 1·9, 3·9 and 6·0 kb in size, which were identical to those in liver tissue. However, the expression rate was weaker than that in liver. The direct effects of recombinant tilapia (rt) growth hormone (GH) and salmon (s) IGF-I on the expression of IGF-I in primary cultured hepatocytes were investigated in time-course and doseresponse experiments. In untreated cultures, IGF-I mRNA decreased with time. Hepatocytes treated with 100 nM rtGH resulted in a pronounced stimulation of IGF-I mRNA expression throughout the experiment. Treatment with rtGH in concentrations ranging from 0·1 nM to 1 µM caused a clear dose-dependent increase in the amount of IGF-I mRNA. Significant stimulation was obtained even with 0·1 nM, reaching a plateau with 10 nM. Neither significant inhibitory nor stimulatory effects were detected by adding sIGF-I from 0·1 nM to 1 µM to the hepataocytes. Our results indicate that the established primary cell culture of tilapia hepatocytes is a useful system in which to study direct effects of potential regulators of bony fish liver cell function.

Differential regulation of Igf1 and Igf2 mRNA levels in tilapia hepatocytes: effects of insulin and cortisol on GH sensitivity

“…Both igf1 and igf2 mRNA levels were stimulated by GH in tilapia primary hepatocytes, as has been found in previous in vitro studies in teleost fishes and elasmobranches (Shamblott et al 1995, Schmid et al 2000, Pierce et al 2004, Leung et al 2008, Moriyama et al 2008a. Recent in vivo studies have shown that GH treatment increases liver igf2 mRNA levels in a wide variety of fish species (see Introduction for citations), which is consistent with our results.…”

“…5!10 K9 M) to the medium increased mRNA levels of both igfs versus time-matched controls. Basal and GH-stimulated igf1 decreased with time in previous studies with primary tilapia and salmon hepatocytes (Schmid et al 2000, Pierce et al 2004. Based on the results for these and other transcripts, an incubation time of 6 h was selected, giving a total culture age of 10 h.…”

Metabolic hormones modulate the effect of growth hormone (GH) on insulin-like growth factor-I (IGF-I) mRNA level in primary culture of salmon hepatocytes

“…GH consistently stimulated IGF-I mRNA expression in salmon hepatocytes, as in previous studies using salmonid hepatocyte culture , Shamblott et al 1995, Pierce et al 2004a) and tilapia hepatocyte culture (Schmid et al 2000). Consistent stimulation of IGF-I mRNA expression by GH in fish hepatocyte culture confirms that this functional relationship in the GH/IGF axis is conserved in fishes (Nicoll et al 1999, PerezSanchez & Le Bail 1999, Mommsen 2001).…”

<i>In vitro</i> Developed Spontaneously Contracting Cardiomyocytes from Rainbow Trout as a Model System for Human Heart Research

“…Model systems, devoid of the existing endogenous influences in the organism, are suitable tools for studying physiological mechanisms in vitro for the first testing period [1]. They are therefore useful in vitro systems to examine adaptive mechanisms of cardiomyocytes to varied environmental influences.…”