• Journal of Animal Science and Technology
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• v.50 no.5
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• pp.649-656
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• 2008

The current study was undertaken to investigate the mechanism of action of insulin-like growth factors (IGFs) on proliferation and differentiation of pig preadipocytes. The preadipocytes were isolated from the backfat of new-born female pigs and cultured in serum-deprived medium in the presence and absence of recombinant native IGFs or recombinant mutant IGFs that have reduced affinity for binding to both type-1 IGF receptors and insulin receptors. Fifty ng/ml of either IGF-I, [Leu60]IGF-I, IGF-Ⅱ or [Leu27]IGF-Ⅱ were included in the media in which preadipocytes were cultured for 4 days. IGF-I, [Leu60]IGF-I, IGF-Ⅱ and [Leu27]IGF-Ⅱ stimulated proliferation of pig preadipocytes by 39%, 8%, 25% and 2% respectively, as measured by increased numbers of cells. This indicates that both IGF-I and -II promote replication of pig preadipocytes by actions mediated either by type-1 IGF receptor or insulin receptor. IGF-I, [Leu60]IGF-I, IGF-Ⅱ and [Leu27]IGF-Ⅱ stimulated differentiation of pig preadipocytes by 50%, 17%, 37% and 30%, respectively, measured as glycerolphosphate dehydrogenase activity. Reducing the affinity of IGF-I for type-1 IGF receptors or insulin receptors significantly reduced the differentiation response. However, the differentiation response to [Leu27]IGF-II was not significantly different from the response to IGF-II. This shows that IGF-I and IGF-Ⅱ promote cell differentiation by different receptor-mediated mechanisms. IGF-II promotes differentiation of pig preadipocytes by actions that do not involve either type-1 IGF receptors or insulin receptors. These actions therefore appear to be mediated by binding of IGF-II to type-2 IGF receptors(also known as cation-independendent mannose-6-phosphate receptor[CIM6P/IGF2 receptor]). This is the first study to find evidence that IGF-II promotes differentiation of preadipocytes from any animal species by actions mediated by CIM6P/IGF2 receptors. In summary, this study shows that IGF-I and IGF-Ⅱ promote differentiation of pig preadipocytes by mechanisms that involve different cellular receptors.

• Development and Reproduction
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• v.3 no.1
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• pp.69-74
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• 1999

Insulin-like growth factors (IGF-1 and IGF-2) play an important regulatory role in premplantation embryonic development. To study the role of IGF-1 during premplantation embryonic development in mouse, the presence of mRNA transcripts for IGF-1 and IGF-lR in the oocytes and preimplantation embryos was examined. In this study, the transcripts of IGF-1 was detected in oocytes using primers for IGF-1. The PCR products were identified by Msp I restriction enzyme digest. We revealed that the transcripts of IGF-1 and IGF-1R were presented in the oocytes and preimplantation embryos. The highest mRNA levels in GV stage oocytes were decreased at 4- or 8-cell stage and then reincreased upto blastocyst. The presence of IGF-1 and IGF-lR in GV-oocytes suggests that the transcripts in the early stage embryos were derived from maternal genome. Additionally, the presence of IGF-1 and IGF-lR in the oocytes and preimplantation embryos suggests that IGF-1 plays an autocrine role during preimplantation embryonic development through IGF-lR as a signalling pathway.

• Journal of Animal Science and Technology
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• v.51 no.1
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• pp.9-14
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• 2009

Insulin-like growth factor 2 (IGF2) is the first identified imprinted gene, which is paternally expressed in multiple mammalian species. A paternally expressed QTL for muscle growth and backfat thickness (BFT) has previously been identified near the IGF2 locus on the distal tip of pig chromosome 2 (SSC2p). Therefore the IGF2 gene is considered an economically important candidate gene for pig industry. Herein, this study explored genetic variation of IGF2 for in3-G3072A, in7-G162C and a new SNP in intron7 (C1589T) in Korean native pig (KNP) and commercial pig breeds, and detected their linkage disequilibrium within these breeds. Furthermore we investigated the effect of in3-G3072A on IGF2 gene expression in post-natal muscle and backfat tissues. The real-time quantitative PCR results showed that animals inherited allele G from a KNP sire had significant higher IGF2 gene expression in backfat tissue than those inherited allele A from a Yorkshire sire, however opposite situation in muscle. These results demonstrated the allele 3072G is associated with a higher IGF2 gene expression in fat tissues, but low gene expression in muscle tissues when compared with the 3072A allele. These results suggest that KNP with lower muscle mass and higher fat deposition might be associated with a higher frequency of the 3072G allele, and selecting KNP based on IGF2 genotypes could result in an economic benefit to KNP producers.

• Korean Journal of Animal Reproduction
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• v.22 no.1
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• pp.89-94
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• 1998

A study was made to investigate the effects of concerning factors with IGF-I recovery on the final IGF-I concentration in the effluent and to establish recovery conditions of IGF-I using liquid emulsion membranes(LEM). D2EHPA was best carrier among Amberlite LA2, Aliquit 336 and D2EHPA for recovery rate of IGF-I. Recovery rate of IGF-I by D2EHPA volume in the oil phase was increased as increasing D2EHPA volume, and optimal volume of D2EHPA was 5% in this experiment. The recovery rate of IGF-I by D2EHPA was increased by the decreasing from pH 7 to pH 4 of external phase. Therefore, optimal pH value was 4.0. Optimal concentrations of sulfuric acid in internal phase, paraffin oil in oil phase and Span 80 for recovery rate of IGF-I were 0.1M, 2.0% and 5%, respectively, and optimal W/O rate was 2. These results suggested that optimal conditions for recovery of IGF-I were D2EHPA(5%) as carrier, pH 4.0, 0.1M sulfuric acid, 2% paraffin oil, 2.0 W/O rate and 5.0% Span 80.

• Journal of Genetic Medicine
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• v.2 no.1
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• pp.41-47
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• 1998

Human uniparental gestations such as gynogenetic ovarian teratomas provide a model to evaluate the integrity of parent-specific gene expression - i.e. imprinting - in the absence of a complementary parental genetic contribution. The few imprinted genes characterized so far include the insulin-like growth factor-2 gene (IGF2) coding for a fetal growth factor and H19 gene whose normal function is unknown but it is likely to act as an mRNA. IGF2 is expressed by the paternal allele and H19 by the maternal allele. This reciprocal expression is quite interesting because both H19 and IGF2 genes are located close to each other on chromosome 11p15.5. In situ RNA hybridization analysis has shown variable expression of the H19 and IGF2 alleles according to the tissue origin in 11 teratomas. Especially, Skin, derivative of ectoderm, is expressed conspicuously. We examined imprinting of H19 and IGF2 in teratomas using PCR and RT-PCR of exonic polymorphism. H19 and IGF2 transcript could be expressed either biallelically or monoallelically in the teratomas. Biallelic expression (i.e., loss of imprinting) of IGF2 occurred in 5 out of 6 mature teratomas and 1 out of 1 immature teratoma. Biallelic expression of H19 occurred in 4 out of 10 mature teratomas and 1 out of 1 immature teratoma. Expression levels of H19 and IGF2 transcript using the semi-quantitative RT-PCR had no relation between monoallelic and biallelic expression. Moreover, IGF2 biallelic expression did not affect allele-specificity or levels of H19 expression. These results demonstrate that both genes, H19 and IGF2, can be imprinted, expressed and regulated independently and individually of each other in ovarian teratoma.

• Journal of Life Science
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• v.21 no.10
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• pp.1385-1392
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• 2011

It is well known that both insulin-like growth factor-I and suppressor of cytokine signaling-3 (SOCS-3) are known to modulate various aspects of physiology in skeletal muscle cells. Furthermore, although SOCS-3 expression is related to insulin resistance in non-skeletal muscle cells and is known to interact with insulin-like growth factor-I receptor, the effect of IGF-I on SOCS-3 gene expression in skeletal muscle cells is presently unknown. C2C12 myotubes were treated with different concentrations (0-200 ng/ml) of IGF-I or for various periods of time (3-72 hr). Immunofluorescent staining image revealed that IGF-I induced SOCS-3 protein expression in a dose-dependent manner. Western blot data also showed that SOCS-3 proteins were induced by IGF-I (200 ng/ml) in C2C12 myotubes in a time-dependent manner. The level of SOCS-3 mRNA was also significantly increased after 3hr of IGF-I (10-100 ng/ml) treatment. However, the levels of SOCS-3 mRNA were significantly decreased after 24 and 48 hr of IGF-I (10-100 ng/ml) treatment compared to the control. In conclusion, SOCS-3 protein is induced by IGF-I treatment in C2C12 skeletal muscle cells and this induction is regulated pretranslationally. The modulating effect of IGF-I on SOCS-3 expression may be an important regulator of gene expression in skeletal muscle cells.

• Food Science of Animal Resources
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• v.26 no.1
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• pp.113-120
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• 2006

Insulin-like growth factor-I(IGF-I) rich fraction, which was obtained molecules ranged between 30 and 1 kDa, was fractionated by ultrafiltration from bovine colostral whey. IGF-I rich fraction was confirmed by SDS-PACE and western blotting and then the quantity of IGF-I was measured by sandwich ELISA. ICF-I concentration in IGF-I rich fraction was 10 ng/mg proteins. IGF-I rich fraction, standard IGF-I and colostral whey weie treated to murine peritoneal macrophages. And then we experimented that effect of immune activity on macrophage and splenocyte. As a result, in group treated with IGF-I rich fraction $1{mu}g/mL$, production of interleukin-6 and nitric oxide were 9.85 ng and $17.17{\mu}M$ and production of phagocytosis, tumor necrosis factor-${\alpha}\;and\;H_{2}O_{2}$ were 78.3, 34.5 and 6% compared to the control group. In splenocyte immune response, B cell and T cell proliferation and NK cell activity were 103, 126 and 22.2% in group treated with IGF-I rich fraction $1{\mu}g/mL$ to compared to the control, respectively.

• Journal of Nutrition and Health
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• v.36 no.3
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• pp.270-279
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• 2003

Conjugated linoleic acid (CLA) is a group of positional and geometric isomers of linoleic acid (LA) and exhibits anticarcinogenic activity in a variety of animal models. We have previously observed that CLA inhibited the growth of Caco-2 cells, a human colon adenocarcinoma cell line. The present study was performed to determine whether the growth inhibitory effect of CLA is related to change in secretion of IGF- II and/or IGF-binding proteins (IGFBPs) that have been shown to regulate Caco-2 cell proliferation by an autocrine mechanism. Cells were incubated in serum-free medium with various concentrations of CLA or linoleic acid (LA). Immunoblot analysis of 24-hours, serum-free, conditioned medium using a monoclonal anti-IGF-IIantibody revealed that Caco-2 cells secreted both mature 6,500 Mr and higher Mr forms of pro IGF-II. The levels of pro IGF-II and mature IGF-IIwere decreased by 43 $\pm$ 2% and 53 $\pm$ 6%, respectively by treatment with 50 $\mu$ M CLA. LA slightly increased pro IGF- II levels. Results from Northern blot analysis showed that CLA decreased IGF-II mRNA levels at 50 $\mu$ M concentration suggesting that CLA regulation of IGF-II protein expression occurs partly at the transcriptional level. Ligand blot analysis of conditioned media using 1251-IGF-II revealed that CLA slightly decreased IGFBP-2 levels and increased IGFBP-4 levels. We confirmed our previous results that CLA inhibited cell growth in a dose-dependent manner but LA slightly increased cell growth. Exogenous IGF-II mitigated the growth inhibitory effect of CLA. These results indicate that the growth inhibitory effect of CLA may be at least in part mediated by decreasing IGF-II and IGFBP-2 secretion and increasing IGFBP-4 secretion in Caco-2 cells.

• Journal of the Korean Association of Oral and Maxillofacial Surgeons
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• v.31 no.5
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• pp.363-369
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• 2005

Purpose: To determine the role of Insulin-like Growth Factor-I (IGF-I) in the regulation of Vascular Endothelial Growth Factor (VEGF) expression in MG-63 cells and then to find the mechanism b which this regulation occurs. Materials and methods: MG-63 cells were grown to confluence in 60-mm dishes. To determine the effects of IGF-I on expression of VEGF mRNA according to time and concentration, the cells were treated with 10 nM IGF-I, following isolation of total RNA and Northern blot analysis after 1, 2, 4, 8, 12, 24 hours and after 2 hours of treatment with 0.5, 2, 10, 25, 50 nM IGF-I respectively, isolation of total RNA and Northern blot analysis were followed. To determine the mechanism of action of IGF-I, inhibitors such as hydroxyurea $(76.1\;{\mu}g/ml)$, actinomycin D $(2.5\;{\mu}g/ml)$, cycloheximide $(10\;{\mu}g/ml)$ were added 1 hour after treatment of 10 nM IGF-I. Results: 1. the expression of VEGF mRNA was increased with treatment of IGF-I. 2. The expression of VEGF mRNA was increased according to time-and concentration dependent manner of IGF-I. 3. The effect of IGF-I was decreased by hydroxyuera, actinomycin D, but not by cycloheximide. Conclusion: IGF-I regulate the expression of VEGF mRNA in the level of DNA synthesis and transcription. These results could suggest that IGF-I plays an important role in angiogenesis in the process of new bone formation and remodeling.

• Molecules and Cells
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• v.28 no.3
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• pp.189-194
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• 2009

The modulating effect of IGF-I on the regulation of AR gene expression and activation in skeletal muscle cells remains poorly understood. In this study, the effects of IGF-I treatment on AR induction and activation in the absence of AR ligands were examined. Differentiating C2C12 cells were treated with different concentrations (0-250 ng/ml) of IGF-I or for various periods of time (0-60 min) of 250 ng/ml IGF-I. Treatment of C2C12 cells with IGF-I resulted in a dose- and time-dependent increase in total AR and phosphorylated AR (Ser 213). IGF-I treatment also led to significantly increased AR mRNA expression when compared with the control. The levels of skeletal ${\alpha}-actin$ and myogenin mRNA, known target genes of AR, were also significantly upregulated after 5 or 10 min of treatment with IGF-I. Confocal images revealed that IGF-I stimulated nuclear localization of AR in the absence of ligands. In addition, an electrophoretic mobility shift assay indicated that IGF-I stimulated the AR DNA binding activity in a time-dependent manner. The present results suggest that IGF-I stimulates the expression and activation of AR by ligand-independent mechanism in differentiating C2C12 mouse skeletal muscle cells.