Insulin-like Growth factor-I Peptide Research Articles

Insulin-like growth factor I (IGF-I) in Chinese alligator, Alligator sinensis: Molecular characterization, tissue distribution and mRNA expression changes during the active and hibernating periods

The Chinese alligator Alligator sinensis is an endangered species endemic to China, up to date, little is known about the regulation of its growth and development. Insulin-like growth factor I (IGF-I) plays a vital role in regulating vertebrate growth and development. In this study, the full-length cDNA of IGF-I in Chinese alligator (caIGF-I) was obtained for the first time, it contains 890-bp nucleotides encoding a 153-amino acid precursor, the mature caIGF-I consists of 70 amino acids by cleaving the signal peptide and C-terminal extension (E domain). The caIGF-I contains all the features of IGF-I peptide with B, C, A, and D domains and the six conservative cysteine residues involved in the stable tertiary structure. Multiple alignment analysis showed that the amino acid sequence of caIGF-I shares high identity with American alligator Alligator mississippiensis (100%) and birds (95–97%). Phylogenetic tree analysis of the IGF-I amino acid sequences indicated that alligators cluster into the bird branch. Real-time quantitative PCR technique showed that caIGF-I is widely expressed in all the examined tissues with the highest expression level in liver, higher in pancreas and oviduct while lower in heart, spleen, lung, kidney, stomach, intestines, ovary and muscles. During hibernation, the caIGF-I expression level decreased significantly in liver, pancreas, oviduct and kidney, while did not significantly change in heart, spleen, lung, stomach, small intestine, ovary and muscles. The mRNA expression changes during the two periods implicate that caIGF-I might play an important role in the regulation of feeding and growth in the Chinese alligator.

Potassium diformate influences gene expression of GH/IGF-I axis and glucose homeostasis in weaning piglets

The objective of this study was to determine the effects of potassium diformate (KDF) on growth performance of weaning piglets and genes expression related with growth axis and hepatic gluconeogenesis. A total of 180 piglets weaned at 28d of age were allocated randomly into two groups, with 6 pens in each group and 15 piglets in each pen. Piglets in the control group were fed basal diet, whereas the KDF-treated group was fed basal diet supplemented with 10g/kg KDF. After 5 weeks feeding, KDF-treated piglets showed higher average daily feed intake (ADFI) and average daily bodyweight gain (ADG) than those of the control (P<0.05), whereas the feed conversion ratio (FCR) was lower than that of the control (P=0.02). The apparent digestibility of dry matter and crude protein of the KDF-treated group were higher than those of the control group (P<0.01). KDF treatment did not change the blood indices level of insulin-like growth factor-I (IGF-I), high density lipoprotein (HDL), low density lipoprotein (LDL), total cholesterol (TC), triglyceride (TG) and blood glucose (BG) in plasma. KDF treatment did not influence the mRNA expression of growth hormone releasing hormone (GHRH) and somatostatin (SS) in hypothalamus, and the levels of growth hormone receptor (GHR) and IGF-I receptor (IGF-IR) mRNA in pituitary. Whereas, it increased the abundance of GH mRNA in pituitary, and the mRNA of GHR, IGF-I, IGF-IR, glucose-6-phosphatase (G6PC), fructose-1,6-bisphosphatase (FBP) and phosphoenolpyruvate carboxykinase (PCK) in liver (P<0.05). Furthermore, KDF treatment increased the concentration of IGF-I peptide in liver (P=0.000). These results suggest that, the beneficial effect of KDF on the growth performance may relate to the regulation of the GH axis and glucose homeostasis in piglets.

The Complexity of the IGF1 Gene Splicing, Posttranslational Modification and Bioactivity

The insulinlike growth factor-I (IGF-I) is an important factor which regulates a variety of cellular responses in multiple biological systems. The IGF1 gene comprises a highly conserved sequence and contains six exons, which give rise to heterogeneous mRNA transcripts by a combination of multiple transcription initiation sites and alternative splicing. These multiple transcripts code for different precursor IGF-I polypeptides, namely the IGF-IEa, IGF-IEb and IGF-IEc isoforms in humans, which also undergo posttranslational modifications, such as proteolytic processing and glycosylation. IGF-I actions are mediated through its binding to several cell-membrane receptors and the IGF-I domain responsible for the receptor binding is the bioactive mature IGF-I peptide, which is derived after the posttranslational cleavage of the pro-IGF-I isoforms and the removal of their carboxy-terminal E-peptides (that is, the Ea, Eb and Ec). Interestingly, differential biological activities have been reported for the different IGF-I isoforms, or for their E-peptides, implying that IGF-I peptides other than the IGF-I ligand also possess bioactivity and, thus, both common and unique or complementary pathways exist for the IGF-I isoforms to promote biological effects. The multiple peptides derived from IGF-I and the differential expression of its various transcripts in different conditions and pathologies appear to be compatible with the distinct cellular responses observed to the different IGF-I peptides and with the concept of a complex and possibly isoform-specific IGF-I bioactivity. This concept is discussed in the present review, in the context of the broad range of modifications that this growth factor undergoes which might regulate its mechanism(s) of action.

Cloning, recombinant expression and purification ofturbot (&lt;I&gt;Scophthalmus maximus&lt;/I&gt;) mature IGF-I

Insulin-like growth factor-I (IGF-I) is a conserved peptide expressed ubiquitously,which shows highly homologous diverse effects on development,growth,and metabolism. With RT-PCR,the fragment encoding the turbot (Scophthalmus maximus) mature IGF-I peptide was amplified. It was predicted that the mature peptide was composed of 70 amino acids including 6 cysteines which may form 3 disulfide bonds. The target fragment was then successfully subcloned into the express vector pGEX-4T-1 and was highly expressed in E.coli BL21(DE3)plysS. The result of SDS-PAGE showed that the fusion protein expressed in the form of inclusion bodies with molecular weight of 34 ku and maximally amounted to 59 % of the whole protein in the E.coli cell 4 hours after being induced with IPTG. The western blotting indicated that recombinant protein had the antigenicity to anti-GST antibody. The inclusion bodies were dissolved in 6 mol/Lguanidine chloride followed by pulse renaturation in refolding buffer containing 0.5 mol/L L-Arginine,1.0 mol/L GSH and 0.2 mol/L GSSG. Then the renatured recombinant protein was purified by GSTrap FF affinity chromatography. The effect of purified GST-IGF-I on turbot kidney cells was analysed,and it indicated that the recombinant GST-IGF-I can stimulate the proliferation of the cells.

IGF-I in epithelial ovarian cancer and its role in disease progression

Insulin-like growth factor-I (IGF-I) is known to be involved in the development and progression of several types of solid tumors including ovarian cancer. IGF-I levels in local tissue is subject to both endocrine and paracrine/autocrine regulation. To investigate which regulation is more importantly involved in IGF-I action in ovarian cancer regarding tumor progression, we analyzed IGF-I mRNA expression (assuming only from paracrine/autocrine regulation) and peptide concentration (subject to both endocrine and paracrine/autocrine regulation) as well as a genetic polymorphism (CA dinucleotide repeats) in 215 epithelial ovarian cancer patients. Genomic DNA, total RNA and cytosol proteins were extracted from fresh tumor samples. Two alternatively spliced IGF-I transcripts (IGF-IA and IGF-IB) were analyzed using real-time PCR. Cytosol levels of free and total IGF-I were measured with enzyme-linked immunosorbent assay. DNA sizing analysis was performed to determine the CA polymorphism. The study showed that the CA polymorphism had a weak influence on IGF-I expression, but no effect on tumor progression. High levels of free, not total, IGF-I peptide were associated with elevated risk of disease progression (HR = 2.06; 95%CI: 1.22–3.50), and the association was independent of clinicopathologic features of the disease. One of the IGF-I transcripts (IGF-IA) had a similar but less significant impact on disease progression. Women with high IGF-I mRNA and peptide were at greater risk for disease progression compared to those with low in both (HR = 2.13; 95%CI: 1.13–3.95). These findings support the notion that IGF-I is involved in ovarian cancer progression and free IGF-I plays a more important role in the disease. The study also suggests that both endocrine and paracrine/autocrine are involved in the regulation of IGF-I activity in ovarian cancer.

Expression and localization of insulin-like growth factor-I in four parts of the red deer antler

The expression and localization of insulin-like growth factor-I (IGF-I) in the four parts (tip, upper, mid and base) of the red deer antler has been extensively investigated. We used reverse transcriptase polymerase chain reaction (RT-PCR) and real-time reverse transcriptase polymerase chain reaction (real time RT-PCR), in situ hybridization, immunohistochemistry and Western blot techniques to localize IGF-I messenger ribonucleic acid (mRNA) and IGF-I peptide in the four parts of the antler. The specific sequence encoding IGF-I was detected by RT-PCR in all of the four specimens, and the 395 bp IGF-I sequence from the red deer antler was shown to have very high homology with human, goat and mouse IGF-I. In situ hybridization and immunohistochemistry results demonstrated that the expression of IGF-I occurred in chondrocytes and osteoblasts in the tip and upper parts of the antler. However, IGF-I was only detectable in osteoblasts around the bone in the mid and base parts. There were significant differences in the intensity of the signal obtained with the IGF-I probe in the tip, upper, mid and base tissues. The Western blot analysis also provided evidence that IGF-I expression was localized differentially in the four parts of the deer antler. This study indicates that antler tissue is an essential part of the IGF system, which is involved in the regulation of the growth of red deer antlers. The specific expression of IGF-I in the four parts of the deer antler suggests that the IGF-I molecule is present at significantly different levels throughout the deer antler development and regeneration processes. Localization of IGF-I in chondrocytes and osteoblasts suggests that IGF-I may play an important role in cartilage and bone formation. In addition, it may have a variety of biophysical effects that influence the rapid growth of deer antlers.

Organ-specific expression of IGF-I during early development of bony fish as revealed in the tilapia, Oreochromis niloticus, by in situ hybridization and immunohistochemistry: indication for the particular importance of local IGF-I

The cellular sites of insulin-like growth factor I (IGF-I) synthesis in the early developing tilapia (0-140 days post fertilization, DPF) were investigated. IGF-I mRNA and peptide appeared in liver as early as 4 DPF and in gastro-intestinal epithelial cells between 5-9 DPF. In exocrine pancreas, the expression of IGF-I started at 4 DPF and continued until 90 DPF. IGF-I production was detected in islets at 6 DPF in non-insulin cells and occurred throughout life. In renal tubules and ducts, IGF-I production started at 8 DPF. IGF-I production in chondrocytes had its onset at 4 DPF, was more pronounced in growing regions and was also found in adults. IGF-I mRNA and peptide appeared in the cytoplasm of skeletal muscle cells at 4 DPF. In gill chloride cells, IGF-I production started at 6 DPF. At 13 DPF, IGF-I was detected in cardiac myocytes. IGF-I-producing epidermal cells appeared at 5 DPF. In brain and ganglia, IGF-I was expressed in virtually all neurones from 6 to 29 DPF, their number decreasing with age. Neurosecretory IGF-I-immunoreactive axons were first seen in the neurohypophysis around 17 DPF. Endocrine cells of the adenohypophysis exhibited IGF-I mRNA at 28 DPF and IGF-I immunoreactivity at 40 DPF. Thus, IGF-I appeared early (4-5 DPF), first in liver, the main source of endocrine IGF-I, and then in organs involved in growth or metabolism. The expression of IGF-I was more pronounced during development than in juvenile and adult life. Local IGF-I therefore seems to have a high functional impact in early growth, metabolism and organogenesis.

Intranasal administration of IGF-I improves behavior and Purkinje cell pathology in SCA1 mice

Spinocerebellar ataxia type 1 (SCA1) is a neurodegenerative disease caused by the expansion of polyglutamine repeat within ataxin-1 protein. Cerebellar Purkinje cells are the primary targets of SCA1 pathology. These cells synthesize insulin-like growth factor-I (IGF-I) and express its receptors during their entire life. The aim of present study was to determine if intranasally administered IGF-I to SCA1 transgenic mice suppresses toxic effects of ataxin-1. Two-week old SCA1 heterozygous animals were randomly divided into two treatment groups of IGF-I (30 and 60 μg IGF-I/animal) and a vehicle-treated control group. The wildtype animals served as normal controls. IGF-I or vehicle was administered at 48 h intervals for the total of 10 doses. Animals were then subjected to rotarod test, sacrificed, brains removed and processed for immunohistochemical and Western blot analysis. Radiolabeled IGF-I and bioactive TAT peptide accumulated in the brains of SCA1 mice following intranasal administration validating the use of intranasal route. SCA1 mice showed SCA1 pathology with impaired motor function and downregulation of calcium binding proteins as compared to wildtype mice. However, 30 and 60 μg IGF-I-treated animals showed improved performance on the rotarod as compared to vehicle-treated SCA1 mice with significant improvement ( p < 0.05) on day 3 in 60 μg IGF-I group. The immunohistochemical data further showed partial recovery in the expression of calbindin D28k and protein kinase C-γ in Purkinje cells in IGF-I-treated SCA1 animals. Our results indicate that suppression of ataxin-1-mediated adverse effects by intranasal IGF-I treatment may be of a therapeutic value to treat SCA1.

Differential expression of IGF-I mRNA and peptide in the male and female gonad during early development of a bony fish, the tilapia Oreochromis niloticus

Insulin-like growth factor I (IGF-I) plays a key role in the complex system that regulates bony fish growth, differentiation, and reproduction. The major source of circulating IGF-I is liver, but IGF-I-producing cells also occur in other organs, including the gonads. Because no data are available on the potential production sites of IGF-I in gonad development, developmental stages of monosex breedings of male and female tilapia from 0 day postfertilization (DPF) to 90 DPF were investigated for the production sites of IGF-I at the peptide (immunohistochemistry) and mRNA (in situ hybridization) level. IGF-I mRNA first appeared in somatic cells of the male and female gonad anlage at 7 DPF followed by IGF-I peptide around 9–10 DPF. Gonad anlagen were detected from 7 DPF. Starting at 7 DPF, IGF-I peptide but no IGF-I mRNA was observed in male and female primordial germ cells (PGCs) provided that IGF-I mRNA was not under the detection level, this observation may suggest that IGF-I originates from the somatic cells and is transferred to the PGCs or is of maternal origin. While in female germ cells IGF-I mRNA and peptide appeared at 29 DPF, in male germ cells both were detected as late as at 51–53 DPF. It is assumed that the production of IGF-I in the germ cells is linked to the onset of meiosis that in tilapia ovary starts at around 28 DPF and in testes at around 52–53 DPF. In adult testis, IGF-I mRNA and peptide occurred in the majority of spermatogonia and spermatocytes as well as in Leydig cells, the latter indicating a role of IGF-I in the synthesis of male sex steroids. In adult ovary, IGF-I mRNA and IGF-I peptide were always present in small and previtellogenic oocytes but only IGF-I peptide infrequently occurred in oocytes at the later stages. IGF-I expression appeared in numerous granulosa and some theca cells of follicles at the lipid stage and persisted in follicles with mature oocytes. The results suggest a crucial role of local IGF-I in the formation, differentiation and function of tilapia gonads.

IGF-I and vasoactive intestinal peptide (VIP) regulate cAMP-response element-binding protein (CREB)-dependent transcription via the mitogen-activated protein kinase (MAPK) pathway in pituitary cells: requirement of Rap1

In previous studies we demonstrated that vasoactive intestinal peptide (VIP) mediation, and interactions between mitogen-activated protein kinase (MAPK) and cAMP/protein kinase A (PKA) signaling pathways are implicated in insulin-like growth factor I (IGF-I)- and VIP-induced lactotroph proliferation. These facts led us to investigate the intracellular mechanisms involved in IGF-I- and VIP-induced lactotroph proliferation. Exposure of cultured male rat pituitary cells to IGF-I (10(-7) M) or VIP (10(-7) M) stimulated the MAPK cascade. Studies in GH4C1 cells, with an expression vector for Rap1 GTPase-activating protein (Rap1 GAP1), demonstrated reduced VIP-induced MAPK activation, indicating that VIP-dependent activation of the extracellular signal-regulated kinase (ERK) pathway requires PKA-Rap1 signaling. IGF-I induced cAMP-response element (CRE)-binding protein (CREB) phosphorylation through the Ras-MAPK pathway, whereas VIP phosphorylated CREB directly via PKA. The mechanisms that regulate IGF-I-and VIP-CREB-dependent gene transcription were examined using GH4C1 cells transiently transfected with a CRE reporter gene. IGF-I and VIP stimulation of CRE-mediated transcription required activation of both Ras-MAPK and cAMP/PKA signaling. This activation was blocked in the presence of Rap1 GAP1. In summary, we showed that IGF-I and VIP stimulated MAPK activity and the phosphorylation of CREB in pituitary cells. Furthermore, VIP-dependent activation of PKA-Rap1-ERK pathways mediated VIP and IGF-I effects on CREB-dependent transcription in GH4C1 cells. Thus, it is possible that VIP- and IGF-I-induced lactotroph proliferation may involve Rap1.

Influence of varying degrees of malnutrition on IGF-I expression in the rat diaphragm.

This study evaluated the impact of varying degrees of prolonged malnutrition on the local insulin-like growth factor-I (IGF-I) system in the costal diaphragm muscle. Adult rats were provided with either 60 or 40% of usual food intake over 3 wk. Nutritionally deprived (ND) animals (i.e., ND60 and ND40) were compared with control (Ctl) rats fed ad libitum. Costal diaphragm fiber types and cross-sectional areas were determined histochemically. Costal diaphragm muscle IGF-I mRNA levels were determined by RT-PCR. Serum and muscle IGF-I peptide levels were determined by using a rat-specific radioimmunoassay. The body weights of Ctl rats increased by 5%, whereas those of ND60 and ND40 animals decreased by 16 and 26%, respectively. Diaphragm weights were reduced by 17 and 27% in ND60 and ND40 animals, respectively, compared with Ctl. Diaphragm fiber proportions were unaffected by either ND regimen. Significant atrophy of both type IIa and IIx fibers was noted in the ND60 group, whereas atrophy of all three fiber types was observed in the diaphragm of ND40 rats. Serum IGF-I levels were reduced by 62 and 79% in ND60 and ND40 rats, respectively, compared with Ctl. Diaphragm muscle IGF-I mRNA levels in both ND groups were similar to those noted in Ctl. In contrast, IGF-I concentrations were reduced by 36 and 42% in the diaphragm muscle of ND60 and ND40 groups, respectively, compared with Ctl. We conclude that the local (autocrine/paracrine) muscle IGF-I system is affected in our models of prolonged ND. We propose that this contributes to disordered muscle protein turnover and muscle cachexia with atrophy of muscle fibers. This is particularly so in view of recent data demonstrating the importance of the autocrine/paracrine system in muscle growth and maintenance of fiber size.

Growth hormone (GH) hypersecretion and GH receptor resistance in streptozotocin diabetic mice in response to a GH secretagogue.

The growth hormone (GH) and insulin-like growth factor I (IGF-I) axis were studied in streptozotocin (STZ) diabetic and nondiabetic female mice following intravenous (IV) injection of the GH secretagogue (GHS) ipamorelin or saline. On day 14, blood samples were obtained before and 10 minutes after the injection. Livers were removed and frozen for determination of the mRNA expressions of the GH receptor, GH-binding protein, and IGF-I, and hepatic IGF-I peptide. Serum samples were analyzed for GH and IGF-I. Following ipamorelin injection, the GH levels were found to be 150 ± 35 μg/L and 62 ± 11 μg/L in the diabetic compared to the nondiabetic mice (P < .05). Serum IGF-I levels were lower in diabetic than in nondiabetic animals, and rose after stimulation only in the nondiabetic animals. Furthermore, hepatic GH resistance and IGF-I mRNA levels and IGF-I peptide were increased in nondiabetic animals in response to GH stimulation, whereas the low levels per se of all these parameters in diabetic mice were unaffected. The study shows that STZ diabetic mice demonstrate a substantial part of the clinical features of type 1 diabetes in humans, including GH hypersecretion and GH resistance. Accordingly, it is proposed that STZ diabetic mice may be a better model of the perturbations of the GH/IGF-I axis in diabetes than STZ diabetic rats.

Strategic advantages of insulin-like growth factor-I expression for cardioprotection.

Insulin-like growth factor-I (IGF-I) peptide has beneficial effects on cardiomyocyte function and survival, many of which are mediated through the serine-threonine kinase, Akt. However, concerns about systemic effects of IGF-I peptide limit its clinical application. The present study tested whether local IGF-I expression could mediate cardioprotection without elevating serum [IGF-I]. The ability of a recombinant adenovirus encoding IGF-IB (Ad.IGF-I) to activate Akt and protect cardiomyocytes from hypoxia-induced apoptosis in vitro was compared with the effects of IGF-I peptide or expression of constitutively active Akt (myr-Akt). In vivo, cardiac IGF-I gene transfer was performed prior to ischemia-reperfusion injury (IRI). Effects on the ischemic and infarcted areas were assessed while serum [IGF-I] was measured by radioimmunoassay. Compared with IGF-I peptide, Ad.IGF-I achieved more sustained activation of Akt and reduced hypoxia-induced apoptosis at lower media IGF-I concentrations. In a co-culture system, Ad.IGF-I protected both infected and uninfected cells from hypoxic injury, while myr-Akt protected only infected cells. In vivo cardiac injection of Ad.IGF-I mediated significant local IGF-I expression, without affecting serum [IGF-I] levels. After IRI, Ad.IGF-I did not affect the ischemic area but reduced infarct size approximately 50% (32 +/- 13 vs. 64 +/- 14% AAR in Ad.GFP rats, p < 0.003), although the transgene was expressed in only approximately 15% of the ischemic region, consistent with possible paracrine benefit. Somatic gene transfer of IGF-I may offer strategic advantages over both systemic delivery of IGF-I peptide and expression of cell autonomous cardioprotective transgenes such as Akt by mediating autocrine and paracrine cardiomyocyte protection without elevating serum [IGF-I] levels.

Local Expression of Insulin-Like Growth Factor-I Affects Angiogenesis in Colorectal Cancer

Insulin-like growth factor-I (IGF-I) has potent mitogenic and anti-apoptotic effects that give it a critical role in the regulation of rapidly renewing epithelial cell populations such as those found in the colon. Recent evidence has implicated circulating IGF-I levels as an important determinant of colorectal cancer risk, but the role, if any, of its autocrine/paracrine expression remains unexplored. Therefore, we investigated the local expression of IGF-I and IGF-I type I receptor (IGF-IR) in 50 paired normal colon and carcinoma samples. IGF-IR mRNA was present in all samples, whereas IGF-I mRNA was detected in only 30 normal (60%) and 27 tumour (54%) biopsies. Samples that did not express IGF-I mRNA had no IGF-I peptide detectable by immunocytochemistry. The absence of local IGF-1 expression was associated with significantly reduced mRNA levels specifying the proliferating cell nuclear antigen and c-myc, as well as Cox-2, and vascular endothelial growth factor – gene products that regulate angiogenesis. The biological relevance of this finding is suggested by the significant association between local IGF-I mRNA levels and microvessel density in the colorectal cancers.

Effects of short- and long-term rat hind limb immobilization on spinal cord insulin-like growth factor-I and its receptor

In this study we investigated changes in the spinal cord insulin-like growth factor-I peptide (IGF-I) and its receptors (IGF-IR) after hind limb immobilization for 5 days, 2, 4, and 8 weeks. Moreover, effects on IGF-I and nicotinic cholinergic receptors (nAChRs) in two types of skeletal muscle were also investigated. IGF-I levels were measured by radioimmunoassay (RIA) whereas IGF-IR and nAChRs were measured by quantitative receptor autoradiography. Spinal cord IGF-I levels decreased significantly after 5 days, 2 and 4 weeks of immobilization, whereas IGF-IR increased significantly after 4 and 8 weeks compared to controls. In skeletal muscles, nAChRs increased significantly after 5 days and 2 weeks in the soleus (SOL) and tibialis anterior (TIB) muscles, respectively, and continued up to 8 weeks in both muscles. IGF-I concentration decrease significantly after 4 and 8 weeks in the SOL and TIB muscles, respectively. Despite the normal levels of IGF-I in both muscles at the early time points (5 days and 2 weeks), low levels of IGF-I were observed concurrently in the spinal cord ipsilateral to the immobilized limb. Our findings suggest that the early decrease in the IGF-I level and the late uppregulation in the IGF-IR in the spinal cord might represent a nervous system response to disuse.

Cyclic Adenosine 3',5'-Monophosphate Inhibits Insulin-Like Growth Factor I Gene Expression in Rat Glioma Cell Lines: Evidence for Regulation of Transcription and Messenger Ribonucleic Acid Stability

cAMP inhibits growth and stimulates differentiation in glioma cells. We examined the effect of cAMP on insulin-like growth factor I (IGF-I) gene expression in the C6 cell line, a rat glioma cell line previously reported to grow in response to autocrine IGF-I. cAMP potently inhibited IGF-I messenger RNA (mRNA) and peptide secretion in C6 cells, associated with an attenuation of DNA synthesis. Exogenous IGF-I peptide at least partially prevented the inhibition of DNA synthesis, suggesting that the reduction in IGF-I biosynthesis may contribute to the inhibitory effect of cAMP on C6 cell growth. cAMP also inhibited IGF-I mRNA in rat RG2 glioma cells, but not in three other nonglioma tumor cell lines. The nuclear IGF-I pre-mRNA level and the half-life of mature IGF-I mRNA were both reduced by cAMP in C6 cells, suggesting effects on gene transcription and mRNA stability. However, cAMP had no effect on the activities of IGF-I exon 1 promoter-luciferase constructs. Protein synthesis inhibition partially reduced the inhibition of IGF-I mRNA by cAMP. Inhibition of cAMP-activated protein kinase A activity by H89 did not alter the inhibition of IGF-I gene expression in response to cAMP, suggesting that protein kinase A does not mediate the cAMP inhibitory effect on IGF-I gene expression.

Cyclic adenosine 3',5'-monophosphate inhibits insulin-like growth factor I gene expression in rat glioma cell lines: evidence for regulation of transcription and messenger ribonucleic acid stability.

cAMP inhibits growth and stimulates differentiation in glioma cells. We examined the effect of cAMP on insulin-like growth factor I (IGF-I) gene expression in the C6 cell line, a rat glioma cell line previously reported to grow in response to autocrine IGF-I. cAMP potently inhibited IGF-I messenger RNA (mRNA) and peptide secretion in C6 cells, associated with an attenuation of DNA synthesis. Exogenous IGF-I peptide at least partially prevented the inhibition of DNA synthesis, suggesting that the reduction in IGF-I biosynthesis may contribute to the inhibitory effect of cAMP on C6 cell growth. cAMP also inhibited IGF-I mRNA in rat RG2 glioma cells, but not in three other nonglioma tumor cell lines. The nuclear IGF-I pre-mRNA level and the half-life of mature IGF-I mRNA were both reduced by cAMP in C6 cells, suggesting effects on gene transcription and mRNA stability. However, cAMP had no effect on the activities of IGF-I exon 1 promoter-luciferase constructs. Protein synthesis inhibition partially reduced the inhibition of IGF-I mRNA by cAMP. Inhibition of cAMP-activated protein kinase A activity by H89 did not alter the inhibition of IGF-I gene expression in response to cAMP, suggesting that protein kinase A does not mediate the cAMP inhibitory effect on IGF-I gene expression.

Changes in tissue levels of growth hormone, insulin-like growth factor-I, and somatostatin in the femurs of hind-limb immobilized rats.

Immobilization of an extremity causes skeletal muscle atrophy and a dramatic increase in bone resorption. Growth hormone (GH) is known to play an important role in bone remodeling mediated in part by local insulin-like growth factor-I (IGF-I). In this study, we investigated changes in the levels of GH and IGF-I peptide in bone extracts from the femur after hind-limb immobilization for 5 days, 2, 4, and 8 weeks. The levels of somatostatin, which interacts with GH, were also measured in the bone extracts. GH levels increased after 8 weeks of hind-limb immobilization whereas the IGF-I concentrations increased after 2 weeks, but returned to control levels at 4 weeks, and decreased after 8 weeks of immobilization. The somatostatin levels in the bone extracts increased only after 8 weeks of hind-limb immobilization. Our findings suggest that, after hind-limb immobilization, changes in the concentrations of GH, IGF-I, and somatostatin in bone may mediate bone resorption either directly or through interaction with other factors.

Effect of Growth Hormone and Insulin-Like Growth Factor I (IGF-I) on the Expression of IGF-I Messenger Ribonucleic Acid and Peptide in Rat Tibial Growth Plate and Articular Chondrocytes in Vivo

Effect of Growth Hormone and Insulin-Like Growth Factor I (IGF-I) on the Expression of IGF-I Messenger Ribonucleic Acid and Peptide in Rat Tibial Growth Plate and Articular Chondrocytes in Vivo

Effect of growth hormone and insulin-like growth factor I (IGF-I) on the expression of IGF-I messenger ribonucleic acid and peptide in rat tibial growth plate and articular chondrocytes in vivo.

Conflicting data exist as to whether insulin-like growth factor I (IGF-I) messenger RNA (mRNA) and peptide are expressed within chondrocytes. This question is pertinent to the mode of GH action on longitudinal bone growth. We have, therefore, investigated this issue in normal rats and in hypophysectomized rats treated for 24 h with GH or IGF-I using in situ hybridization and immunohistochemistry. Serum IGF-I, body weight, and tibial growth plate, but not articular cartilage, height increased with both treatments. Both IGF-I mRNA and IGF-I immunoreactivity occurred in all chondrocyte layers of growth plate and articular cartilage. The percentage of cells with IGF-I mRNA correlated well with IGF-I immunoreactivity under all experimental conditions. In normal rats, IGF-I expression was highest in the upper hypertrophic zone in growth plate (68-71%) and articular cartilage (32-34%). Hypophysectomy, GH, or IGF-I did not significantly affect this percentage. In the stem cell and proliferative and lower hypertrophic zones of growth plate, hypophysectomy dramatically reduced the percentage of labeled chondrocytes, and GH restored it. IGF-I increased IGF-I mRNA and immunoreactivity only in the proliferative zone. In articular cartilage, both remained unchanged under all experimental conditions. Together with our previous finding that GH infusion of hypophysectomized rats enhances chondrocyte maturation at all differentiation stages, the present results are compatible with the idea that IGF-I produced by all chondrocyte layers under the influence of GH mediates chondrocyte maturation and thus longitudinal bone growth in an autocrine/paracrine manner.