Insulin-like Growth Factor (IGF) I Down-Regulates Type 1 IGF Receptor (IGF 1R) and Reduces the IGF I Response in A549 Non-Small-Cell Lung Cancer and Saos-2/B-10 Osteoblastic Osteosarcoma Cells

The insulin-like growth factor type 1 receptor (IGF 1R) mediates the acute metabolic effects of IGF I as well as IGF I-stimulated cell proliferation and protection from apoptosis. IGF binding proteins (IGFBPs) can modulate these responses. We, therefore, investigated whether intrinsic IGFBPs interfere with IGF I-induced regulation of IGF 1R expression and with the biological response to IGF I in two human tumor cell lines, the non-small-cell lung cancer cell line A549 and the osteoblastic osteosarcoma cell line Saos-2/B-10. We compared the growth rates, IGFBP production, IGF I binding characteristics, IGF 1R protein and mRNA levels, and the acute IGF I response (stimulation of glycogen synthesis) after pretreatment of the cells in serum-free medium with or without added IGF I or medium supplemented with 5% fetal calf serum (FCS). In contrast to A549 cells, which produce IGF I and significant amounts of IGFBPs, survival and proliferation of Saos-2/B-10 cells, which do not produce IGF I or significant amounts of IGFBPs, depended on the addition of exogenous IGF I. IGF I increased the concentration of IGFBP-2 and -3 and decreased the concentration of IGFBP-4 in the medium of A549 cells. As compared to FCS, IGF I pretreatment in both cell lines decreased the number of specific IGF I binding sites, down-regulated total and membrane IGF 1R protein, and largely reduced or abolished the acute IGF I response without affecting IGF 1R mRNA levels. The data suggest that the IGF 1R protein of the two cell lines is translationally and/or posttranslationally down-regulated by its ligand in the presence and in the absence of locally produced IGFBPs and that the cell lines have retained this negative feedback to counteract IGF I stimulation.     

β2-肾上腺素受体在新生大鼠心肌成纤维细胞中的分布及其促增殖作用

为了明确β-肾上腺素受体(AR)亚型在新生大鼠心肌成纤维细胞中的分布及其在成纤维细胞增殖反应中的作用,采用放射配体结合实验和[3H]-thymidine掺人法检测了新生大鼠心肌成纤维细胞的β-AR密度和DNA合成速率。结果显示,在培养心肌细胞和心肌成纤维细胞中β-AR密度(Bmtax)和解离常数(Kp)无显著性差异;竞争抑制曲线分析结果提示,心肌成纤维细胞对CGP 20712A和ICI ll8551单位点拟合均显著优于两位点拟合(P＜0．01),表现为对选择性β1-AR拮抗剂CGP 20712A的低亲和性(IC50值：10．1μmol/L)和对选择性β2-AR拮抗剂ICI 118551的高亲和性(IC50值：0.147μmol/L)。异丙肾上腺素(ISO)促心肌成纤维细胞增殖作用可被ICI 118551和心得安(非选择性β-AR拮抗剂)完全抑制,而CGP20812A则无此作用。上述结果提示,在培养心肌成纤维细胞中β-AR亚型占绝对优势,并且ISO引起的心肌成纤维细胞增殖反应是由β2-AR介导的。     

VEGFR-2-mediated increased proliferation and survival in response to oxygen and glucose deprivation in PlGF knockout astrocytes

In hypoxic/ischemic conditions, astrocytes are involved in neuroprotection and angiogenesis. Vascular endothelial growth factor (VEGF) induces angiogenesis and exhibits neuroprotective and neurotrophic properties. However, the role of placental growth factor (PlGF), a VEGF homolog, in these processes is unclear. Therefore, proliferation and survival studies were performed on PlGF knockout (PlGF-/-) and wild-type (PlGF+/+) mouse astrocytes. A significant increase in cell proliferation and survival to oxygen and glucose deprivation (OGD) was observed in PlGF-/- compared to PlGF+/+ astrocytes. Interestingly, no PlGF protein expression was detected in PlGF+/+ astrocytes and no changes in VEGF protein levels were observed between the two genotypes. Real-time PCR and immunocytochemistry showed over-expression of VEGF receptor-2 (VEGFR-2) in PlGF-/- compared with PlGF+/+ astrocytes. Confocal microscopy revealed nuclear, membrane, and cytoplasmic localization of VEGFR-2. In vivo over-expression of VEGFR-2 mRNA was also detected in PlGF-/- compared with PlGF+/+ astrocytes. Stimulation with VEGF165 resulted in increased proliferation in PlGF-/- compared with PlGF+/+ astrocytes. This effect was blocked by the VEGFR-2 antagonist, VEGF165b. The enhanced proliferation of PlGF-/- astrocytes correlated with increased phospho-extracellular-signal-regulated kinase-1/2 levels, while the resistance to OGD was independent of the phosphatidylinositol 3'-kinase/Akt pathway. These results suggest that VEGFR-2 mediates the enhanced proliferative/OGD resistant phenotype observed in PlGF-/- astrocytes.     

Hypertrophy of cultured adult rat ventricular cardiomyocytes induced by antibodies against the insulin-like growth factor (IGF)-I or the IGF-I receptor is IGF-II-dependent

Antibodies against the insulin-like growth factor-I (IGF-I) or the IGF-I receptor (IGF-IR) directly initiate a rapid (within 6 h) hypertrophy of isolated adult rat ventricular cardiomyocytes cultured in the absence of serum. Further, cardiomyocytes treated with either of these agonistic antibodies upregulate the expression of their genes for insulin-like growth factor-II (IGF-II) and the IGF-II receptor (IGF-IIR). Genistein, an inhibitor of the tyrosine kinase IGF-IR, also induces the cardiomyocytes to hypertrophy. Anti-IGF-II antibody inhibits the cardiomyocyte hypertrophy induced by anti-IGF-I and anti-IGF-IR antibodies or by genistein. Results are consistent with a model in which local production of IGF-II is upregulated when the IGF-IR signaling pathway is blocked and in which an IGF-II-mediated pathway, likely involving the IGF-IIR, then stimulates hypertrophy of the cardiomyocytes.     

Analysis of polymorphisms in the insulin-like growth factor 1 receptor (IGF1R) gene from Japanese quail selected for body weight

Insulin-like growth factor 1 receptor ( IGF1R ) is essential for the signalling of growth. In this study, we performed single nucleotide polymorphism (SNP) detection in the Japanese quail IGF1R coding region and an association study between SNPs and body weight in two lines (SS and LL) selected for large and small body weight. Of 21 SNPs obtained, a SNP at position AB292766:c.2293G>A led to the replacement of a valine with an isoleucine (V765I). The two lines were fixed for alternate alleles, with allele encoding valine fixed in the LL line. A significant effect of the SNP genotype was found on 10-week body weight ( P  < 0.01) and on 4- to 10-week and 6- to 10-week average daily gain ( P  < 0.05) in the F₂ family obtained from lines LL and SS. In six populations maintained in Japan or France, the frequency of allele encoding valine was higher than the allele encoding isoleucine.     

Properties of secretin receptor internalization differ from those of the beta(2)-adrenergic receptor.

The endocytic pathway of the secretin receptor, a class II GPCR, is unknown. Some class I G protein-coupled receptors (GPCRs), such as the beta(2)-adrenergic receptor (beta(2)-AR), internalize in clathrin-coated vesicles and this process is mediated by G protein-coupled receptor kinases (GRKs), beta-arrestin, and dynamin. However, other class I GPCRs, for example, the angiotensin II type 1A receptor (AT(1A)R), exhibit different internalization properties than the beta(2)-AR. The secretin receptor, a class II GPCR, is a GRK substrate, suggesting that like the beta(2)-AR, it may internalize via a beta-arrestin and dynamin directed process. In this paper we characterize the internalization of a wild-type and carboxyl-terminal (COOH-terminal) truncated secretin receptor using flow cytometry and fluorescence imaging, and compare the properties of secretin receptor internalization to that of the beta(2)-AR. In HEK 293 cells, sequestration of both the wild-type and COOH-terminal truncated secretin receptors was unaffected by GRK phosphorylation, whereas inhibition of cAMP-dependent protein kinase mediated phosphorylation markedly decreased sequestration. Addition of secretin to cells resulted in a rapid translocation of beta-arrestin to plasma membrane localized receptors; however, secretin receptor internalization was not reduced by expression of dominant negative beta-arrestin. Thus, like the AT(1A)R, secretin receptor internalization is not inhibited by reagents that interfere with clathrin-coated vesicle-mediated internalization and in accordance with these results, we show that secretin and AT(1A) receptors colocalize in endocytic vesicles. This study demonstrates that the ability of secretin receptor to undergo GRK phosphorylation and beta-arrestin binding is not sufficient to facilitate or mediate its internalization. These results suggest that other receptors may undergo endocytosis by mechanisms used by the secretin and AT(1A) receptors and that kinases other than GRKs may play a greater role in GPCR endocytosis than previously appreciated.     

Genetic inactivation of the adenosine A(2A) receptor exacerbates brain damage in mice with experimental autoimmune encephalomyelitis

Studies with multiple sclerosis patients and animal models of experimental autoimmune encephalomyelitis (EAE) implicate adenosine and adenosine receptors in modulation of neuroinflammation and brain injury. Although the involvement of the A(1) receptor has been recently demonstrated, the role of the adenosine A(2A) receptor (A(2A) R) in development of EAE pathology is largely unknown. Using mice with genetic inactivation of the A(2A) receptor, we provide direct evidence that loss of the A(2A) R exacerbates EAE pathology in mice. Compared with wild-type mice, A(2A) R knockout mice injected with myelin oligodendroglia glycoprotein peptide had a higher incidence of EAE and exhibited higher neurological deficit scores and greater decrease in body weight. A(2A) R knockout mice displayed increased inflammatory cell infiltration and enhanced microglial cell activation in cortex, brainstem, and spinal cord. In addition, demyelination and axonal damage in brainstem were exacerbated, levels of Th1 cytokines increased, and Th2 cytokines decreased. Collectively, these findings suggest that extracellular adenosine acting at A(2A) Rs triggers an important neuroprotective mechanism. Thus, the A(2A) receptor is a potential target for therapeutic approaches to multiple sclerosis.     

Signaling by fibroblast growth factors (FGF) and fibroblast growth factor receptor 2 (FGFR2)-activating mutations blocks mineralization and induces apoptosis in osteoblasts

Fibroblast growth factors (FGF) play a critical role in bone growth and development affecting both chondrogenesis and osteogenesis. During the process of intramembranous ossification, which leads to the formation of the flat bones of the skull, unregulated FGF signaling can produce premature suture closure or craniosynostosis and other craniofacial deformities. Indeed, many human craniosynostosis disorders have been linked to activating mutations in FGF receptors (FGFR) 1 and 2, but the precise effects of FGF on the proliferation, maturation and differentiation of the target osteoblastic cells are still unclear. In this report, we studied the effects of FGF treatment on primary murine calvarial osteoblast, and on OB1, a newly established osteoblastic cell line. We show that FGF signaling has a dual effect on osteoblast proliferation and differentiation. FGFs activate the endogenous FGFRs leading to the formation of a Grb2/FRS2/Shp2 complex and activation of MAP kinase. However, immature osteoblasts respond to FGF treatment with increased proliferation, whereas in differentiating cells FGF does not induce DNA synthesis but causes apoptosis. When either primary or OB1 osteoblasts are induced to differentiate, FGF signaling inhibits expression of alkaline phosphatase, and blocks mineralization. To study the effect of craniosynostosis-linked mutations in osteoblasts, we introduced FGFR2 carrying either the C342Y (Crouzon syndrome) or the S252W (Apert syndrome) mutation in OB1 cells. Both mutations inhibited differentiation, while dramatically inducing apoptosis. Furthermore, we could also show that overexpression of FGF2 in transgenic mice leads to increased apoptosis in their calvaria. These data provide the first biochemical analysis of FGF signaling in osteoblasts, and show that FGF can act as a cell death inducer with distinct effects in proliferating and differentiating osteoblasts.     

Solution structure of intracellular signal-transducing peptide derived from human beta(2)-adrenergic receptor

The structure of intracellular third loop peptide (betaIII-2: RRSSKFCLKEKKALK) was studied by CD and NMR spectroscopy. According to the CD study, this peptide forms a helix in the TFE solution. The three-dimensional molecular structure in TFE was determined by the 2D (1)H NMR spectroscopy. The structure consists of a positive charge cluster on the C-terminal side of the peptide chain. This part will be an active site of the peptide interacting with the G-protein.     

c-myb stimulates cell growth by regulation of insulin-like growth factor (IGF) and IGF-binding protein-3 in K562 leukemia cells

c-myb plays an important role in the regulation of cell growth and differentiation, and is highly expressed in immature hematopoietic cells. The human chronic myelogenous leukemia cell K562, highly expresses IGF-I, IGF-II, IGF-IR, and IGF-induced cellular proliferation is mediated by IGF-IR. To characterize the impact of c-myb on the IGF-IGFBP-3 axis in leukemia cells, we overexpressed c-myb using an adenovirus gene transfer system in K562 cells. The overexpression of c-myb induced cell proliferation, compared to control, and c-myb induced cell growth was inhibited by anti-IGF-IR antibodies. c-myb overexpression resulted in a significant increase in the expression of IGF-I, IGF-II, and IGF-IR, and a decrease in IGFBP-3 expression. By contrast, disruption of c-myb function by DN-myb overexpression resulted in significant reduction of IGF-I, IGF-II, IGF-IR, and elevation of IGFBP-3 expression. In addition, exogenous IGFBP-3 inhibited the proliferation of K562 cells, and c-myb induced cell growth was blocked by IGFBP-3 overexpression in a dose-dependent manner. The growth-promoting effects of c-myb were mediated through two major intracellular signaling pathways, Akt and Erk. Activation of Akt and Erk by c-myb was completely blocked by IGF-IR and IGFBP-3 antibodies. These findings suggest that c-myb stimulates cell growth, in part, by regulating expression of the components of IGF-IGFBP axis in K562 cells. In addition, disruption of c-myb function by DN-myb may provide a useful strategy for treatment of leukemia.     

Signalling through the type 1 insulin-like growth factor receptor (IGF1R) interacts with canonical Wnt signalling to promote neural proliferation in developing brain

Signalling through the IGF1R [type 1 IGF (insulin-like growth factor) receptor] and canonical Wnt signalling are two signalling pathways that play critical roles in regulating neural cell generation and growth. To determine whether the signalling through the IGF1R can interact with the canonical Wnt signalling pathway in neural cells in vivo, we studied mutant mice with altered IGF signalling. We found that in mice with blunted IGF1R expression specifically in nestin-expressing neural cells (IGF1R^Nestin−KO mice) the abundance of neural β-catenin was significantly reduced. Blunting IGF1R expression also markedly decreased: (i) the activity of a LacZ (β-galactosidase) reporter transgene that responds to Wnt nuclear signalling (LacZ^TCF reporter transgene) and (ii) the number of proliferating neural precursors. In contrast, overexpressing IGF-I (insulin-like growth factor I) in brain markedly increased the activity of the LacZ^TCF reporter transgene. Consistently, IGF-I treatment also markedly increased the activity of the LacZ^TCF reporter transgene in embryonic neuron cultures that are derived from LacZ^TCF Tg (transgenic) mice. Importantly, increasing the abundance of β-catenin in IGF1R^Nestin−KO embryonic brains by suppressing the activity of GSK3β (glycogen synthase kinase-3β) significantly alleviated the phenotypic changes induced by IGF1R deficiency. These phenotypic changes includes: (i) retarded brain growth, (ii) reduced precursor proliferation and (iii) decreased neuronal number. Our current data, consistent with our previous study of cultured oligodendrocytes, strongly support the concept that IGF signalling interacts with canonical Wnt signalling in the developing brain to promote neural proliferation. The interaction of IGF and canonical Wnt signalling plays an important role in normal brain development by promoting neural precursor proliferation.     

Regulation of Ligand and Shear Stress-induced Insulin-like Growth Factor 1 (IGF1) Signaling by the Integrin Pathway

Mechanical loading of the skeleton, as achieved during daily movement and exercise, preserves bone mass and stimulates bone formation, whereas skeletal unloading from prolonged immobilization leads to bone loss. A functional interplay between the insulin-like growth factor 1 receptor (IGF1R), a major player in skeletal development, and integrins, mechanosensors, is thought to regulate the anabolic response of osteogenic cells to mechanical load. The mechanistic basis for this cross-talk is unclear. Here we report that integrin signaling regulates activation of IGF1R and downstream targets in response to both IGF1 and a mechanical stimulus. In addition, integrins potentiate responsiveness of IGF1R to IGF1 and mechanical forces. We demonstrate that integrin-associated kinases, Rous sarcoma oncogene (SRC) and focal adhesion kinase (FAK), display distinct actions on IGF1 signaling; FAK regulates IGF1R activation and its downstream effectors, AKT and ERK, whereas SRC controls signaling downstream of IGF1R. These findings linked to our observation that IGF1 assembles the formation of a heterocomplex between IGF1R and integrin β3 subunit indicate that the regulation of IGF1 signaling by integrins proceeds by direct receptor-receptor interaction as a possible means to translate biomechanical forces into osteoanabolic signals.     

IGF2 derived from SH-SY5Y neuroblastoma cells induces the osteoclastogenesis of human monocytic precursors

The insulin-like growth factors 2 (IGF2) is a peptide hormone that binds to the insulin-like growth factor 1 receptor (IGF1R) and is abundantly stored in bone. IGF1R is deeply involved in the pathogenesis of many cancers that growth within bone and is also involved in osteoclast biology. Among different cell lines representative of osteolytic tumors, we found a very high expression of IGF2 in SH-SY5Y cells derived from neuroblastoma (NB). We previously showed that NB cells induce an osteolytic process through the Osteoprotegerin/RANKL/RANK and the canonical Wnt pathway system. Here, we hypothesized that NB promotes osteoclastogenesis also via IGF2. First, we demonstrated the presence of IGF1R on the osteoclast basolateral membrane, and we observed a cyclic IGF1R activation along with the differentiation process, also when induced by SH-SY5Y. Moreover, we found that IGF2 mRNA expression in SH-SY5Y cells was further increased when co-cultured with mesenchymal stromal cells, suggesting that IGF2 is important for NB interaction with the bone microenvironment. Finally, the treatment of SH-SY5Y cells with an anti-IGF2 siRNA or the addition of anti-IGF1R molecules impaired NB-induced osteoclastogenesis, even though the chemoattraction of monocytes by NB cells was unaffected. Our findings suggest that in IGF2-producing osteolytic tumors IGF1R is a good candidate for targeted therapies in combination with conventional drugs.     

Key regulators of mitochondrial biogenesis are increased in kidneys of growth hormone receptor knockout (GHRKO) mice

The growth hormone receptor knockout (GHRKO) mice are remarkably long-lived and highly insulin sensitive. Alterations in mitochondrial biogenesis are associated with aging and various metabolic derangements. We have previously demonstrated increased gene expression of key regulators of mitochondriogenesis in kidneys, hearts and skeletal muscles of GHRKO mice. The aim of the present study was to quantify the protein levels of the following regulators of mitochondriogenesis: peroxisome proliferator-activated receptor γ co-activator 1α (PGC-1α), AMP-activated protein kinase α (AMPKα), phospho-AMPKα (p-AMPKα), sirtuin-3 (SIRT-3), endothelial nitric oxide synthase (eNOS), phospho-eNOS (p-eNOS), nuclear respiratory factor-1 (NRF-1) and mitofusin-2 (MFN-2) in skeletal muscles and kidneys of GHRKOs in comparison to normal mice. We also were interested in the effects of calorie restriction (CR) and visceral fat removal (VFR) on these parameters. Both CR and VFR improve insulin sensitivity and can extend life span. Results: The renal levels of PGC-1α, AMPKα, p-AMPKα, SIRT-3, eNOS, p-eNOS and MFN-2 were increased in GHRKOs. In the GHRKO skeletal muscles, only MFN-2 was increased. Levels of the examined proteins were not affected by CR (except for PGC-1α and p-eNOS in skeletal muscles) or VFR. Conclusion: GHRKO mice have increased renal protein levels of key regulators of mitochondriogenesis, and this may contribute to increased longevity of these knockouts.     

Retracted: Downregulation of long noncoding RNA H19 rescues hippocampal neurons from apoptosis and oxidative stress by inhibiting IGF2 methylation in mice with streptozotocin-induced diabetes mellitus

The diabetes mellitus (DM)-induced reduction of neurogenesis in the hippocampus is consequently accompanied by cognitive decline. The present study set out to define the critical role played by long noncoding RNA H19 (lncRNA H19) in the apoptosis of hippocampal neurons, as well as oxidative stress (OS) in streptozotocin (STZ)-induced DM mice through regulation of insulin-like growth factor 2 (IGF2) methylation. The expression of lncRNA H19 in the hippocampal neurons and surviving neurons were detected. Hippocampal neurons were cultured and transfected with oe-H19, sh-H19, oe-IGF2, or sh-IGF2, followed by detection of the expressions of IGF2 and apoptosis-related genes. Determination of the lipid peroxide and glutathione levels was conducted, while antioxidant enzyme activity was identified. The IGF2 methylation, the binding of lncRNA H19 to DNA methyltransferase, and the binding of lncRNA H19 to IGF2 promoter region were detected. DM mice exhibited high expressions of H19, as well as a decreased hippocampal neurons survival rate. Higher lncRNA H19 expression was found in DM. Upregulated lncRNA H19 significantly increased the expression of Bax and caspase-3 but decreased that of Bcl-2, thus promoting the apoptosis of hippocampal neuron. Besides, upregulation of lncRNA H19 induced OS. LncRNA H19 was observed to bind specifically to the IGF2 gene promoter region and promote IGF2 methylation by enriching DNA methyltransferase, thereby silencing IGF2 expression. Taken together, downregulated lncRNA H19 reduces IGF2 methylation and enhances its expression, thereby suppressing hippocampal neuron apoptosis and OS in STZ-induced (DM) mice.     

ErbB2 is necessary for induction of carcinoma cell invasion by ErbB family receptor tyrosine kinases

The epidermal growth factor (EGF) family of tyrosine kinase receptors (ErbB1, -2, -3, and -4) and their ligands are involved in cell differentiation, proliferation, migration, and carcinogenesis. However, it has proven difficult to link a given ErbB receptor to a specific biological process since most cells express multiple ErbB members that heterodimerize, leading to receptor cross-activation. In this study, we utilize carcinoma cells depleted of ErbB2, but not other ErbB receptor members, to specifically examine the role of ErbB2 in carcinoma cell migration and invasion. Cells stimulated with EGF-related peptides show increased invasion of the extracellular matrix, whereas cells devoid of functional ErbB2 receptors do not. ErbB2 facilitates cell invasion through extracellular regulated kinase (ERK) activation and coupling of the adaptor proteins, p130CAS and c-CrkII, which regulate the actin-myosin cytoskeleton of migratory cells. Overexpression of ErbB2 in cells devoid of other ErbB receptor members is sufficient to promote ERK activation and CAS/Crk coupling, leading to cell migration. Thus, ErbB2 serves as a critical component that couples ErbB receptor tyrosine kinases to the migration/invasion machinery of carcinoma cells.     

Modulation of the neuronal binding of the beta subunit of nerve growth factor (NGF) by the alpha-NGF subunit

The effect of the alpha subunit of the 7S-NGF on the binding of beta-NGF to its two classes of sites on target cells has been studied. The presence of microM concentrations of alpha-NGF causes the displacement of 125I-beta-NGF from one class of sites on dissociated dorsal root ganglia neurons from stage E9 chicken embryos. At 0.1 nM 125I-beta-NGF, increasing alpha-NGF concentrations produce a monotonic displacement curve with half-maximal displacement occurring at 10 microM alpha-NGF. The affinity and number of sites of the 125I-beta-NGF displaced by alpha-NGF are similar to those of beta-NGF that binds to the higher affinity (site I) receptors. The binding to the lower affinity class of sites (site II) is not affected by concentrations of alpha-NGF up to 30 microM. This modulation of 125I-beta-NGF binding does not occur with equivalent concentrations of serum albumin. No detectable neuronal binding of 125I-alpha-NGF was found, suggesting that the mechanism does not involve direct competition for receptor sites. The dissociation constant for the alpha-beta complex is in the microM range, and formation of this complex in solution can thus compete with the process of 125I-beta-NGF binding to neurons. A model accounting for these observations includes binding of the alpha-beta complex to the lower affinity but not to the higher affinity sites. We conclude that there are differences in the specificity of the two classes of receptors.     

The RYR1 g.1843C>T mutation is associated with the effect of the IGF2 intron3-g.3072G>A mutation on muscle hypertrophy

Muscle growth is a complex phenomenon regulated by many factors, whereby net growth results from the combined action of synthesis and turnover. In pigs, two quantitative trait nucleotides (QTN) are known to have an important influence on muscle growth and fat deposition: one QTN is located in the ryanodine receptor 1 (RYR1) gene (RYR1 g.1843C>T) and the other, a paternally expressed QTN, is in the insulin-like growth factor 2 (IGF2) gene (IGF2 intron3-g.3072G>A). The mutation in IGF2 abrogates in vitro interaction with a repressor, which leads to a threefold increase of IGF2 expression in post-natal muscle. The family of the calpains, a family of Ca(2+)-sensitive muscle endopeptidases, and their specific inhibitor calpastatin play an important role in post-natal protein degradation, also influencing muscle and carcass traits. This study investigated the possible interactions between the genotypes of the RYR1 and IGF2 QTN on IGF2 expression. Samples were taken from several muscles and from pigs at several ages, and messenger RNA expression levels were measured using a real-time quantification assay. IGF2 expression in m. longissimus dorsi of animals with mutations in both IGF2 and RYR1 was significantly lower than in animals that inherited the IGF2 mutation but were homozygous wildtype for RYR1.     

Multiplication stimulating activity (MSA) from the BRL 3A rat liver cell line: Relation to human somatomedins and insulin

The properties of multiplication stimulating activity (MSA), an insulin-like growth factor (somatomedin) purified from culture medium conditioned by the BRL 3A rat liver cell line are summarized. The relationship of MSA to somatomedins purified from human and rat plasma are considered. MSA appears to be the predominant somatomedin in fetal rat serum, but a minor component ot adult rat somatomedin. In vitro biological effects of MSA and insulin in adipocytes, fibroblasts and chondrocytes are examined to determine whether they are mediated by insulin receptors or insulin-like growth factor receptors. The possible relationship of a primary defect of insulin receptors observed in fibroblasts from a patient with the rare genetic disorder, leprechaunism, to intrauterine growth retardation is discussed.