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We elucidated the molecular mechanism of cancer-associated fibroblast (CAF)-associated gene insulin-like growth factor binding protein-2 (IGFBP2)-induced M2 macrophage polarization in the tumor microenvironment involved in glioma progression. The Cancer Genome Atlas (TCGA) and Chinese Glioma Genome Atlas (CGGA) provided bulk RNA-sequencing datasets, ESTIMATE scores for glioma stromal cells, and overall survival-clinicopathological correlation analyses. TIMER provided CAF abundance in the TCGA glioma-related dataset, differential gene analysis was performed for high- and low-CAF groups, and weighted gene coexpression network analysis identified CAF-related genes. Univariate and multifactorial cyclooxygenase (COX) regression analyses created the CAF risk models single sample gene set enrichment analysis, CIBERSORT, and GSE84465. Mice were implanted with gliomas, and Western blot and RT-quantitative PCR showed IGFBP2 in tumor tissues. Adeno-associated virus (AAV) decreased IGFBP2, flow cytometry measured M1 and M2 macrophage ratios, and immunohistochemistry detected markers. TCGA and CGGA transcriptome data showed malignant gliomas had higher stromal cell scores and worse prognoses. Low- and high-CAF TCGA gliomas were detected, and differential expression, WGCNA, and multifactorial COX identified 132 CAF-related genes and seven high-risk genes (CPQ, EFEMP2, IGFBP2, RAB42, TNFRSF12A, and VASN). Neither CAF risk score, grade, nor 1p/19q affected glioma prognosis. CAF only enriched EFEMP2 and IGFBP2. Gene Expression Profiling Interactive Analysis compared EFEMP2 and IGFBP2 expression in normal brain tissue and gliomas. Low-grade glioma and malignant glioblastoma highly expressed IGFBP2 and EFEMP2. GSEA raised IGFBP2. CIBERSORT linked M2 macrophage infiltration to TCGA glioma immune cell subpopulation IGFBP2 expression. IGFBP2 knockdown stopped mouse glioma and M2 macrophage polarization. CAF plays a procarcinogenic role in glioma, and the CAF-related gene IGFBP2 could promote glioma progression by inducing M2 macrophage polarization. NEW & NOTEWORTHY: The cancer-associated fibroblast (CAF)-related gene insulin-like growth factor binding protein-2 (IGFBP2) is highly expressed in gliomas and is associated with poor prognosis. CAF-related gene IGFBP2 promotes glioma progression by inducing polarization of M2 macrophages. This study provides a new basis for an in-depth investigation of the functional mechanisms of the glioma tumor microenvironment and the search for key genes involved in immune regulation in CAF. [ABSTRACT FROM AUTHOR]

Brown bears conserve muscle and bone mass during 6 mo of inactive hibernation. The molecular mechanisms underlying hibernation physiology may have translational relevance for human therapeutics. We hypothesize that protective mechanisms involve increased tissue availability of insulin-like growth factors (IGFs). In subadult Scandinavian brown bears, we observed that mean plasma IGF-1 and IGF-2 levels during hibernation were reduced to 36 ± 10% and 56 ± 15%, respectively, compared with the active state (n = 12). Western ligand blotting identified IGF-binding protein (IGFBP)-3 as the major IGFBP in the active state, whereas IGFBP-2 was codominant during hibernation. Acid labile subunit (ALS) levels in hibernation were reduced to 41±16% compared with the active state (n = 6). Analysis of available grizzly bear RNA sequencing data revealed unaltered liver mRNA IGF-1, IGFBP-2, and IGFBP-3 levels, whereas ALS levels were significantly reduced during hibernation (n = 6). Reduced ALS synthesis and circulating levels during hibernation should prompt a shift from ternary IGF/IGFBP/ALS to smaller binary IGF/IGFBP complexes, thereby increasing IGF tissue availability. Indeed, size-exclusion chromatography of bear plasma demonstrated a shift to lower molecular weight IGF-containing complexes in the hibernating versus the active state. Furthermore, we note that the major IGF-2 mRNA isoform expressed in livers in both Scandinavian brown bears and grizzly bears was an alternative splice variant in which Ser29 is replaced with a tetrapeptide possessing a positively charged Arg residue. Homology modeling of the bear IGF-2/IGFBP-2 complex showed the tetrapeptide in proximity to the heparin-binding domain involved in bone-specific targeting of this complex. In conclusion, this study provides data which suggest that increased IGF tissue availability combined with tissue-specific targeting contribute to tissue preservation in hibernating bears. [ABSTRACT FROM AUTHOR]

This investigation examined the influence of 12-week ballistic resistance training programs on the IGF-I system in circulation, interstitial fluid, and skeletal muscle, at rest and in response to acute exercise. Seventeen college-aged subjects (11 women/6 men; 21.7 ± 3.7 yr) completed an acute ballistic exercise bout before and after the training program. Blood samples were collected pre-, mid-, and postexercise and analyzed for serum total IGF-I, free IGF-I, and IGF binding proteins (IGFBPs) 1–4. Dialysate and interstitial free IGF-I were analyzed in vastus lateralis (VL) interstitial fluid collected pre- and postexercise via microdialysis. Pre- and postexercise VL muscle biopsies were analyzed for IGF-I protein expression, IGF-I receptor phosphorylation (p-IGF-IR), and AKT phosphorylation (p-AKT). Following training, basal serum IGF-I, free IGF-I, IGFBP-2, and IGFBP-3 decreased whereas IGFBP-1 and IGFBP-4 increased. Training reduced basal dialysate and interstitial free IGF-I but had no effect on basal skeletal muscle IGF-I, p-IGF-IR, or p-AKT. Acute exercise elicited transient changes in IGF-I system concentrations and downstream anabolic signaling both pre- and posttraining; training did not affect this acute exercise response. Posttraining, acute exercise-induced changes in dialysate/interstitial free IGF-I were strongly correlated with the changes in intramuscular IGF-I expression, p-IGF-IR, and p-AKT. The divergent influence of resistance training on circulating/interstitial and skeletal muscle IGF-I demonstrates the importance of concurrent, multiple biocompartment analysis when examining the IGF-I system. As training elicited muscle hypertrophy, these findings indicate that IGF-I’s anabolic effects on skeletal muscle are mediated by local, rather than systemic mechanisms. [ABSTRACT FROM AUTHOR]

Insulin and insulin-like growth factor-1 (IGF-1) are fetal hormones critical to establishing normal fetal growth. Experimentally elevated IGF-1 concentrations during late gestation increase fetal weight but lower fetal plasma insulin concentrations. We therefore hypothesized that infusion of an IGF-1 analog for 1 wk into late gestation fetal sheep would attenuate fetal glucose-stimulated insulin secretion (GSIS) and insulin secretion in islets isolated from these fetuses. Late gestation fetal sheep received infusions with IGF-1 LR3 (IGF-1, n = 8), an analog of IGF-1 with low affinity for the IGF binding proteins and high affinity for the IGF-1 receptor, or vehicle control (CON, n = 9). Fetal GSIS was measured with a hyperglycemic clamp (IGF-1, n = 8; CON, n = 7). Fetal islets were isolated, and insulin secretion was assayed in static incubations (IGF-1, n = 8; CON, n = 7). Plasma insulin and glucose concentrations in IGF-1 fetuses were lower compared with CON (P = 0.0135 and P = 0.0012, respectively). During the GSIS study, IGF-1 fetuses had lower insulin secretion compared with CON (P = 0.0453). In vitro, glucose-stimulated insulin secretion remained lower in islets isolated from IGF-1 fetuses (P = 0.0447). In summary, IGF-1 LR3 infusion for 1 wk into fetal sheep lowers insulin concentrations and reduces fetal GSIS. Impaired insulin secretion persists in isolated fetal islets indicating an intrinsic islet defect in insulin release when exposed to IGF-1 LR3 infusion for 1 wk. We speculate this alteration in the insulin/IGF-1 axis contributes to the long-term reduction in b-cell function in neonates born with elevated IGF-1 concentrations following pregnancies complicated by diabetes or other conditions associated with fetal overgrowth. [ABSTRACT FROM AUTHOR]

Currently, there is no effective treatment for placental dysfunction in utero. In a ligated mouse model of fetal growth restriction (FGR), nanoparticle-mediated human insulin-like 1 growth factor (hIGF1) gene delivery (NP-Plac1-hIGF1) increased hIGF1 expression and maintained fetal growth. However, whether it can restore fetal growth remains to be determined. Using the endothelial nitric oxide synthase knockout (eNOS-/-) mouse model, a genetic model of FGR, we found that despite inducing expression of hIGF1 in the placentas treated with NP-Plac1-hIGF1 (P = 0.0425), FGR did not resolve. This was associated with no change to the number of fetal capillaries in the placental labyrinth; an outcome which was increased with NP-Plac1-hIGF1 treatment in the ligated mouse model, despite increased expression of angiopoietin 1 (P = 0.05), and suggested IGF1 signaling in the placenta requires eNOS to modulate placenta angiogenesis. To further assess this hypothesis, BeWo choriocarcinoma cell line and human placental explant cultures were treated with NP-Plac1-hIGF1, oxidative stress was induced with hydrogen peroxide (H2O2), and NOS activity was inhibited using the inhibitor NG-monomethyl-L-arginine (L-NMMA). In both BeWo cells and explants, the protective effect of NP-Plac1-hIGF1 treatment against H2O2-induced cell death/lactate dehydrogenase release was prevented by eNOS inhibition (P = 0.003 and P < 0.0001, respectively). This was associated with an increase in mRNA expression of oxidative stress markers hypoxia inducing factor 1 α (HIF1 α; P < 0.0001) and ADAM10 (P = 0.0002) in the NP-Plac1-hIGF1 + H2O2 + L-NMMA-treated BeWo cells. These findings show for the first time the requirement of eNOS/NOS in IGF1 signaling in placenta cells that may have implications for placental angiogenesis and fetal growth. [ABSTRACT FROM AUTHOR]

Klotho interacts with various membrane proteins such as receptors for transforming growth factor-β (TGF-β) and insulin-like growth factor (IGF). Renal expression of klotho is diminished in polycystic kidney disease (PKD). In the present study, the effects of klotho supplementation on PKD were assessed. Recombinant human klotho protein (10 μg·kg-1·day-1) or a vehicle was administered daily by subcutaneous injection to 6-wk-old mice with PKD (DBA/2-pcy). Blood pressure was measured using tail-cuff methods. After 2 mo, mice were killed, and the kidneys were harvested for analysis. Exogenous klotho protein supplementation reduced kidney weight, cystic area, systolic blood pressure, renal angiotensin II levels, and 8-epi-PGF2α excretion (P < 0.05). Klotho protein supplementation enhanced glomerular filtration rate, renal expression of superoxide dismutase, and klotho itself (P < 0.05). Klotho supplementation attenuated renal expressions of TGF-α and collagen type I and diminished renal abundance of Twist, phosphorylated Akt, and mammalian target of rapamycin (P < 0.05). Pathological examination revealed that klotho decreased the fibrosis index and nuclear staining of Smad in PKD kidneys (P < 0.05). Our data indicate that klotho protein supplementation ameliorates the reninangiotensin system, reducing blood pressure in PKD mice. Furthermore, the present results implicate klotho supplementation in the suppression of Akt/mammalian target of rapamycin signaling, slowing cystic expansion. Finally, our findings suggest that klotho protein supplementation attenuated fibrosis at least partly by inhibiting epithelial mesenchymal transition in PKD. [ABSTRACT FROM AUTHOR]

Circulating myostatin-attenuating agents are being developed to treat muscle-wasting disease despite their potential to produce serious off-target effects, as myostatin/activin receptors are widely distributed among many nonmuscle tissues. Our studies suggest that the myokine not only inhibits striated muscle growth but also regulates pituitary development and growth hormone (GH) action in the liver. Using a novel myostatin-null label-retaining model (Jekyll mice), we determined that the heterogeneous pool of pituitary stem, transit-amplifying, and progenitor cells in Jekyll mice depletes more rapidly after birth than the pool in wild-type mice. This correlated with increased levels of GH, prolactin, and the cells that secrete these hormones, somatotropes and lactotropes, respectively, in Jekyll pituitaries. Recombinant myostatin also stimulated GH release and gene expression in pituitary cell cultures although inhibiting prolactin release. In primary hepatocytes, recombinant myostatin blocked GHstimulated expression of two key mediators of growth, insulin-like growth factor (IGF)1 and the acid labile subunit and increased expression of an inhibitor, IGF-binding protein-1. The significance of these findings was demonstrated by smaller muscle fiber size in a model lacking myostatin and liver IGF1 expression (LID-o-Mighty mice) compared with that in myostatin-null (Mighty) mice. These data together suggest that myostatin may regulate pituitary development and function and that its inhibitory actions in muscle may be partly mediated by attenuating GH action in the liver. They also suggest that circulating pharmacological inhibitors of myostatin could produce unintended consequences in these and possibly other tissues. [ABSTRACT FROM AUTHOR]

Enhancing placental insulin-like growth factor (IGF) availability appears to be an attractive strategy for improving outcomes in fetal growth restriction (FGR). Our approach was the novel use of [Leu27]IGF-II, a human IGF-II analog that binds the IGF-II clearance receptor IGF-IIR in fetal growth-restricted (FGR) mice. We hypothesized that the impact of [Leu27]IGF-II infusion in C57BL/6J (wild-type) and endothelial nitric oxide synthase knockout (eNOS-/-; FGR) mice would be to enhance fetal growth and investigated this from mid- to late gestation; 1 mgµkg-1µday-1 [Leu27]IGF-II was delivered via a subcutaneous min-iosmotic pump from E12.5 to E18.5. Fetal and placental weights recorded at E18.5 were used to generate frequency distribution curves; fetuses <5th centile were deemed growth restricted. Placentas were harvested for immunohistochemical analysis of the IGF system, and maternal serum was collected for measurement of exogenously administered IGF-II. In WT pregnancies, [Leu27]IGF-II treatment halved the number of FGR fetuses, reduced fetal(P = 0.028) and placental weight variations (P = 0.0032), and increased the numbers of pups close to the mean fetal weight (131 vs. 112 pups within 1 SD). Mixed-model analysis confirmed litter size to be negatively correlated with fetal and placental weight and showed that [Leu27] IGF-II preferentially improved fetal weight in the largest litters, as defined by number. Unidirectional 14CMeAIB transfer per gram placenta (System A amino acid transporter activity) was inversely correlated with fetal weight in [Leu27]IGF-II-treated WT animals (P < 0.01). In eNOS-/- mice, [Leu27]IGF-II reduced the number of FGR fetuses(1 vs. 5 in the untreated group). The observed reduction in FGR pup numbers in both C57 and eNOS-/- litters suggests the use of this analog as a means of standardizing and rescuing fetal growth, preferentially in the smallest offspring. [ABSTRACT FROM AUTHOR]

The igf1 gene is alternatively spliced as IGF-IEa and IGF-IEc variants in humans. In fibrostenotic Crohn's disease, the fibrogenic cytokine TGF-β1 induces IGF-IEa expression and IGF-I production in intestinal smooth muscle and results in muscle hyperplasia and collagen I production that contribute to stricture formation. Mechano-growth factor (MGF) derived from IGF-IEc induces skeletal and cardiac muscle hypertrophy following stress. We hypothesized that increased IGF-IEc expression and MGF production mediated smooth muscle hypertrophy also characteristic of fibrostenotic Crohn's disease. IGFIEc transcripts and MGF protein were increased in muscle cells isolated from fibrostenotic intestine under regulation by endogenous TGF-β1. Erk5 and MEF2C were phosphorylated in vivo in fibrostenotic muscle; both were phosphorylated and colocalized to nucleus in response to synthetic MGF in vitro. Smooth muscle-specific protein expression of γ-smooth muscle actin, γ-smooth muscle actin, and smoothelin was increased in affected intestine. Erk5 inhibition or MEF2C siRNA blocked smooth muscle-specific gene expression and hypertrophy induced by synthetic MGF. Conditioned media of cultured fibrostenotic muscle induced muscle hypertrophy that was inhibited by immunoneutralization of endogenous MGF or pro-IGFIEc. The results indicate that TGF-β1-dependent IGF-IEc expression and MGF production in patients with fibrostenotic Crohn's disease regulates smooth muscle cell hypertrophy a critical factor that contributes to intestinal stricture formation. [ABSTRACT FROM AUTHOR]

The somatotropic signaling pathway has been implicated in aging and longevity studies in mice and other species. The physiology and lifespans of a variety of mutant mice, both spontaneous and genetically engineered, have contributed to our current understanding of the role of growth hormone and insulin-like growth factor I on aging-related processes. Several other mice discovered to live longer than their wild-type control counterparts also exhibit differences in growth factor levels; however, the complex nature of the phenotypic changes in these animals may also impact lifespan. The somatotropic axis impacts several pathways that dictate insulin sensitivity, nutrient sensing, mitochondrial function, and stress resistance as well as others that are thought to be involved in lifespan regulation. [ABSTRACT FROM AUTHOR]

Glutamate acts as the universal agonist at ionotropic glutamate receptors in part because of its high degree of conformational flexibility. Other amino acids and small peptides, however, can activate N-methyl-D-aspartate (NMDA) receptors, albeit usually with lower affinity and efficacy. Here, we examined the action of glycine-prolineglutamate (GPE), a naturally occurring tripeptide formed in the brain following cleavage of IGF-I. GPE is thought to have biological activity in the brain, but its mechanism of action remains unclear. With its flanking glutamate and glycine residues, GPE could bind to either the agonist or coagonist sites on NMDA receptors, however, this has not been directly tested. Using whole cell patch-clamp recordings in combination with rapid solution exchange, we examined both steady-state currents induced by GPE as well as the effects of GPE on synaptically evoked currents. High concentrations of GPE evoked inward currents, which were blocked either by NMDA receptor competitive antagonists or the voltage-dependent channel blocker Mg2+. GPE also produced a slight attenuation in the NMDA- and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)- mediated excitatory postsynaptic currents without altering the pairedpulse ratio. Our results suggest that GPE can activate NMDA receptors but at concentrations well above the expected concentration of GPE in the brain. Therefore, it is unlikely that endogenous GPE interacts with glutamate receptors under normal conditions. [ABSTRACT FROM AUTHOR]

Accumulation of visceral fat, more so than subcutaneous fat, is strongly associated with severe metabolic complications. However, the factors regulating depot-specific adipogenesis are poorly understood. In this study, we show differential expression of pregnancy-associated plasma protein-A (PAPP-A), a secreted regulator of local insulin-like growth factor (IGF) action, in adipose tissue of mice. PAPP-A mRNA expression was fivefold higher in visceral (mesenteric) fat compared with subcutaneous (inguinal, subscapular), perirenal, and brown fat of mice. To investigate the possible role of depot-specific PAPP-A expression in fat accumulation, wild-type (WT) and PAPP-A knockout (KO) mice were fed a high-fat diet (HFD) for up to 20 wk. Adipocyte size increased in subcutaneous and perirenal depots similarly in WT and PAPP-A KO mice. However, fat cell size and in vivo lipid uptake were significantly reduced in mesenteric fat of PAPP-A KO compared with WT mice. After 20 wk on HFD, phosphorylation of AKT, a downstream signaling intermediate of IGF-I and insulin receptor activation, was significantly decreased by 50% in mesenteric compared with subcutaneous fat in WT mice, but was significantly increased threefold in mesenteric compared with subcutaneous fat in PAPP-A KO mice. This appeared to be because of enhanced insulinstimulated signaling in mesenteric fat of PAPP-A KO mice. These data establish fat depot-specific expression of PAPP-A and indicate preferential impact of PAPP-A deficiency on visceral fat in the mouse that is associated with enhanced insulin receptor signaling. Thus, PAPP-A may be a potential target for treatment and/or prevention strategies for visceral obesity and related morbidities. [ABSTRACT FROM AUTHOR]

To establish a causal role for locally produced IGF-I in the mechanical strain response in the bone, we have generated mice with conditional disruption of the insulin-like growth factor (IGF) I gene in type 1±2 collagen-expressing cells using the Cre-loxP approach. At 10 wk of age, loads adjusted to account for bone size difference were applied via four-point bending or axial loading (AL) in mice. Two wk of bending and AL produced significant increases in bone mineral density and bone size at the middiaphysis of wild-type (WT), but not knockout (KO), mice. In addition, AL produced an 8-25% increase in trabecular parameters (bone volume-tissue volume ratio, trabecular thickness, and trabecular bone mineral density) at the secondary spongiosa of WT, but not KO, mice. Histomorphometric analysis at the trabecular site revealed that AL increased osteoid width by 60% and decreased tartrate-resistance acidic phosphatase-labeled surface by 50% in the WT, but not KO, mice. Consistent with the in vivo data, blockade of IGF-I action with inhibitory IGF-binding protein (IGFBP4) in vitro completely abolished the fluid flow stress-induced MC3T3-E1 cell proliferation. One-way ANOVA revealed that expression levels of EFNB1, EFNB2, EFNA2, EphB2, and NR4a3 were different in the loaded bones of WT vs. KO mice and may, in part, be responsible for the increase in bone response to loading in the WT mice. In conclusion, IGF-I expressed in type 1 collagen-producing bone cells is critical for converting mechanical signal to anabolic signal in bone, and other growth factors cannot compensate for the loss of local IGF-I. [ABSTRACT FROM AUTHOR]

Somatolactin (SL), a member of the growth hormone/prolactin family, is a pituitary hormone unique to fish models. Although SL is known to have diverse functions in fish, the mechanisms regulating its secretion and synthesis have not been fully characterized. Using grass carp pituitary cells as a model, here we examined the role of insulin-like growth factor (IGF) in SL regulation at the pituitary level. As a first step, the antisera for the two SL isoforms expressed in the carp pituitary, SLa and SLβ, were produced, and their specificity was confirmed by antiserum preabsorption and immunohistochemical staining in the carp pituitary. Western blot using these antisera revealed that grass carp SLα and SLβ could be N-linked glycosylated and their basal secretion and cell content in carp pituitary cells could be elevated by IGF-I and -II treatment. These stimulatory effects occurred with parallel rises in SLα and SLβ mRNA levels, and these SL gene expression responses were not mimicked by insulin but blocked by IGF-I receptor inactivation. In carp pituitary cells, IGF-I and -II could induce rapid phosphorylation of IGF-I receptor, MEK1/2, ERK1/2, MKK3/6, and p38 MAPK; and SLα and SLβ secretion, protein production, and mRNA expression caused by IGF-I and -II stimulation were negated by inactivating MEK1/2 and p38 MAPK. Parallel inhibition of PI3K and Akt, however, were not effective in these regards. These results, taken together, provide evidence that IGF can upregulate SL secretion and synthesis at the pituitary level via stimulation of MAPK- but not PI3K/Akt-dependent pathways. [ABSTRACT FROM AUTHOR]

Eels are unique in that they maintain lower plasma SO42- concentration in SO42--rich (~30 mM) seawater (SW) than in SO42--poor (<0.3 mM) freshwater (FW), showing drastic changes in SO42- regulation between FW and SW. We previously showed that the expression of renal SO42- transporter genes, FW-specific Slc13a1 and SW-specific Slc26a6a, changes profoundly after transfer of FW eels to SW, which results in the decrease in plasma SO42- concentration after 3 days in SW. In this study, we attempted to identify the environmental factor(s) that trigger the switching of SO42- regulation using changes in plasma and urine SO42- concentrations and expression of the transporter genes as markers. Transfer of FW eels to 30 mM SO42- or transfer of SW eels to SO42--free SW did not change the SO42- regulation. Major divalent cations in SW, Mg2+ (50 mM) and Ca2+ (10 mM), were also ineffective, but 50 mM NaCl was effective for switching the SO42- regulation. Further analyses using choline-Cl and Na-gluconate showed that Cl- is a primary factor and Na2+ is permissive for the Cl- effect. Since plasma SO42- and Cl- concentrations were inversely correlated, we injected various solutions into the blood and found that Cl- alone triggered the switching from FW to SW-type regulation. Furthermore, the inhibitor of Na-Cl cotransporter (NCC) added to media significantly impaired the expression of SW-specific Slc26a6a in 150 mM NaCl. In summary, it appears that Cl- ions in SW are taken up into the circulation via the NCC together with Na2+, and the resultant increase in plasma Cl- concentration enhances SO42- excretion by the kidney through downregulation of absorptive Slc13a1 and upregulation of excretory Slc26a6a, resulting in low plasma SO42- concentration in SW. [ABSTRACT FROM AUTHOR]

This study tested the hypothesis that transdermal fluid (TDF) provides a more sensitive and accurate measure of exercise-induced increases in insulin-like growth factor-I (IGF-I) than serum, and that these increases are detectable proximal, but not distal, to the exercising muscle. A novel, noninvasive methodology was used to collect TDF, followed by sampling of total IGF-I (tIGF-I) and free IGF-I (fIGF-I) in TDF and serum following an acute bout of exercise. Experiment 1: eight men (23 ± 3 yrs, 79 ± 7 kg) underwent two conditions (resting and 60 min of cycling exercise at 60% V̇o2peak) in which serum and forearm TDF were collected for comparison. There were no significant changes in tIGF-I or fIGF-I in TDF obtained from the forearm or from serum following exercise (P > 0.05); however, the proportion of fIGF-I to tIGF-I in TDF was approximately fourfold greater than that of serum (P ≤ 0.05). These data suggest that changes in TDF IGF-I are not evident when TDF is sampled distal from the working tissue. To determine whether exercise-induced increases in local IGF-I could be detected when TDF was sampled directly over the active muscle group, we performed a second experiment. Experiment 2: fourteen subjects (22 ± 4 yr, 68 ± 11 kg) underwent an acute plyometric exercise condition consisting of 10 sets of 10 plyometric jumps with 2-min rest between sets. We observed a significant increase in TDF tIGF-I following exercise (P ≤ 0.05) but no change in serum tIGF-I (P > 0.05). Overall, these data suggest that TDF may provide a noninvasive means of monitoring acute exercise-induced changes in local IGF-I when sampled in proximity to exercising muscles. Moreover, our finding that the proportion of free to tIGF-I was greater in TDF than in serum suggests that changes in local IGF-I may be captured more readily using this system. [ABSTRACT FROM AUTHOR]

Arterial smooth muscle cells enter the cell cycle and proliferate in conditions of disease and injury, leading to adverse vessel remodeling. In the pulmonary vasculature, diverse stimuli cause proliferation of pulmonary artery smooth muscle cells (PASMCs), pulmonary artery remodeling, and the clinical condition of pulmonary hypertension associated with significant health consequences. PASMC proliferation requires extracellular Ca2+ influx that is intimately linked with intracellular Ca2+ homeostasis. Among the primary sources of Ca2+ influx in PASMCs is the low-voltage-activated family of T-type Ca2+ channels; however, up to now, mechanisms for the action of T-type channels in vascular smooth muscle cell proliferation have not been addressed. The Cav3.1 T-type Ca2+ channel mRNA is upregulated in cultured PASMCs stimulated to proliferate with insulin-like growth factor-I (IGF-I), and this upregulation depends on phosphatidylinositol 3-kinase/Akt signaling. Multiple stimuli that trigger an acute rise in intracellular Ca2+ in PASMCs, including IGF-I, also require the expression of Cav3.1 Ca2+ channels for their action. IGF-I also led to cell cycle initiation and proliferation of PASMCs, and, when expression of the Cav3.1 Ca2+ channel was knocked down by RNA interference, so were the expression and activation of cyclin D, which are necessary steps for cell cycle progression. These results confirm the importance of T-type Ca2+ channels in proper progression of the cell cycle in PASMCs stimulated to proliferate by IGF-I and suggest that Ca2+ entry through Cav3.1 T-type channels in particular interacts with Ca2+-dependent steps of the mitogenic signaling cascade as a central component of vascular remodeling in disease. [ABSTRACT FROM AUTHOR]

The podocyte secretory proteome may influence the phenotype of adjacent podocytes, endothelial cells, parietal epithelial cells, and tubular epithelial cells but has not been systematically characterized. We have initiated studies to characterize this proteome, with the goal of further understanding the podocyte cell biology. We cultured differentiated conditionally immortalized human podocytes and subjected the proteins in conditioned medium to mass spectrometry. At a false discovery rate of <3%, we identified 111 candidates from conditioned medium, including 44 proteins that have signal peptides or are described as secreted proteins in the UniProt database. As validation, we confirmed that one of these proteins, insulin-like growth factor-binding protein-related protein-I (IGFBP-rPI), was expressed in mRNA and protein of cultured podocytes. In addition, transforming growth factor-J31 stimulation increased IGFBP-rPI in conditioned medium. We analyzed IGFBP-rP1 glomerular expressio in a mouse model of human immunodeficiency virus-associated nephropathy. IGFBP-rP 1 was absent from podocytes of normal mice and was expressed in podocytes and pseudocrescents of transgenic mice, where it was coexpressed with desmin, a podocyte injury marker. We conclude that IGFBP-rP1 may be a product of injured podocytes. Further analysis of the podocyte secretory proteomc may identify biomarkers of podocyte injury. [ABSTRACT FROM AUTHOR]

Both mechanical and humoral triggers have been put forward to explain the hypertrophic response of the challenged cardiomyocyte. The aim of the present study was to investigate whether cyclic equibiaxial stretch is a direct stimulus for isolated adult rabbit cardiomyocytes to develop hypertrophy and to explore the potential involvement of the autocrine/ paracrine factors ANG II, transforming growth factor (TGF)-β1, and IGF-I in this process. Isolated cardiomyocytes were exposed to 10% cyclic equibiaxial stretch (1 Hz) for up to 48 h or treated with ANG 11(100 nM), TGF-β1 (5 ng/ml), IGF-I (100 ng/ml), ANG II type 1 (AT1) receptor blockers, or conditioned medium of stretched fibroblasts. Cyclic stretch significantly increased cell surface area (+3.1%), protein synthesis (+21%), and brain natriuretic peptide (BNP) mRNA expression (6-fold) in cardiomyocytes. TGF-β1 expression increased (+42%) transiently at 4 h, whereas cardiomyocyte IGF-I expression was not detectable under all experimental conditions. The AT1 receptor blockers candesartan and irbesartan (100 nM) did not prevent the stretch-induced hypertrophic response. Direct exposure to ANG II, TGF-β1, or IGF-I did not enhance cardiomyocyte BNP expression. In cardiac fibroblasts, stretch elicited a significant approximately twofold increase in TGF-β1 and IGF-I expression. Conditioned medium of stretched fibroblasts increased BNP expression in cardiomyocytes (∼2-fold, P = 0.07). This study clearly indicates that cyclic stretch is a strong, direct trigger to induce hypertrophy in fully differentiated rabbit cardiomyocytes. The present findings do not support the notion that stretch-mediated hypertrophy of adult rabbit cardiomyocytes involves autocrine/paracrine actions of ANG II, TGF-β1, or IGF-I. [ABSTRACT FROM AUTHOR]

This study compared alternating current and pulsed current electrical muscle stimulation (EMS) for torque output, skin temperature (Tsk), blood lactate and hormonal responses, and skeletal muscle damage markers. Twelve healthy men (23-48 yr) received alternating current EMS (2.5 kHz delivered at 75 Hz, 400 µs) for the knee extensors of one leg and pulsed current (75 Hz, 400 µs) for the other leg to induce 40 isometric contractions (on-off ratio 5-15 s) at the knee joint angle of 100° (0°: full extension). The use of the legs for each condition was counter-balanced among subjects, and the two EMS bouts were separated by 2 wk. The current amplitude was consistently increased to maximally tolerable level, and the torque and perceived intensity were recorded over 40 isometric contractions. Tsk of the stimulated and contralateral knee extensors were measured before, during, and for 30 min after EMS. Blood lactate, growth hormone, testosterone, insulin-like growth factor 1, testosterone, and cortisol were measured before, during, and for 45 min following EMS. Muscle damage markers included maximal voluntary isometric contraction torque, muscle soreness with a 100-mm visual analog scale, and plasma creatine kinase (CK) activity, which were measured before and 1, 24, 48, 72, and 96 h after EMS. No significant differences in the torque induced during stimulation (∼30% maximal voluntary isometric contraction) and perceived intensity were found, and changes in Tsk, blood lactate, and hormones were not significantly different between conditions. However, all of the measures showed significant (P < 0.05) changes from baseline values. Skeletal muscle damage was evidenced by prolonged strength loss, development of muscle soreness, and increases in plasma CK activity; however, the changes in the variables were not significantly different between conditions. It is concluded that acute effects of alternating and pulsed current EMS on the stimulated muscles are similar. [ABSTRACT FROM AUTHOR]

Insulin-like growth factor I (IGF-1) coordinates proliferation and differentiation in a wide variety of cell types. The igfl gene not only produces IGF-I, but also generates multiple carboxy-terminal extensions, the E-peptides, through alternative splicing leading to different isoforms. It is not known if the IGF-I isoforms share a common pathway for their actions, or if there are specific actions of each protein. Viral administration of murine IGF-IA, IGF-IB, and mature IGF, which lacked an E-peptide extension, was utilized to identify IGF-I isoform-specific responsive genes in muscles of young growing mice. Microarray analysis revealed responses that were driven by increased IGF-I regardless of the presence `of E-peptide, such as Bcl-XL. In contrast, distinct expression patterns were observed after viral delivery of IGF-IA or IGF-IB, which included matrix metalloproteinase 13 (MMPI3). Expression of Bcl-XL was prevented when viral administration of the IGF-I isoforms was performed into muscles of MKR mice, which lack functional IGF-I receptors on the muscle fibers. However, MMPI3 expression persisted under the same conditions after viral injection of IGF-IB. At 4 mo after viral delivery, expression of IGF-IA or IGF-IB promoted muscle hypertrophy, but viral delivery of mature IGF-1 failed to increase muscle mass. These studies provide evidence that local production of IGF-I requires the E-peptides to drive hypertrophy in growing muscle and that both common and unique pathways exist for the IGF-I isoforms to promote biological effects. [ABSTRACT FROM AUTHOR]

Progastrin and insulin-like growth factors (IGFs) stimulate hyperproliferation of intestinal epithelial cells (IECs) via endocrine/paracrine routes; hyperproliferation is a known risk factor for colon carcinogenesis. In the present study, inhibitory potency of curcumin in the presence or absence of progastrin andlor IGF-II was examined. Progastrin and JGF-II significantly increased proliferation of an immortalized IEC cell line, IEC-18, whereas curcumin decreased the proliferation in a dose-dependent manner. IGF-II was significantly more effective than progastrin in reversing antiproliferative effects of curcumin and reversed proapoptotic effects of curcumin by >80%; progastrin was relatively ineffective toward reversing proapoptotic effects of curcumin. IEC-18 clones were generated to overexpress either progastrin (IEC-PG) or hIGF-II (IEC-IGF). Proliferation of IEC-PG and IEC-IGF clones was increased, compared with that of control clones. Curcumin significantly reduced proliferation of IEC-PG, but not IEC-IGF, clones. Similarly, a human colon cancer cell line, Caco-2 (which expresses autocrine IGF-II), was relatively resistant to inhibitory effects of curcumin. However, Caco-2 cells treated with antiIGF-II-antibodies were rendered sensitive to inhibitory effects of curcumin. Significant differences in inhibitory potency of curcumin against PGvs. IGF-II-stimulated growth of IEC-18 cells were not reflected by differences in curcumin-mediated inhibition of activated (phosphorylated) ERKs/IKKα/β/p65NF-κB and c-Src in wild-type (wt)IEC-18 cells, in response to the two growth factors. Surprisingly, curcumin was almost ineffective in reducing IGF-II-stimulated activation of p38MAPK but significantly reduced progastrin-stimulated phosphorylation of p38. Treatment with a p38MAPK inhibitor resulted in loss of protective effects of IGF-II against inhibitory effects of curcumin. These novel findings suggest that growth factor profile of patients and tumors may dictate inhibitory potency of curcumin and that combination of curcumin + p38MAPK inhibitor may be required for reducing hyperproliferative or tumorigenic response of IECs to endocrine and autocrine IGFs. [ABSTRACT FROM AUTHOR]

Normal childhood growth is determined by ultradian and infradian variations in GH secretion, yet OH treatment of children with short stature is restricted to daily fixed doses. We have used OH-deficient dwarf rats to determine whether variable OH dose regimens promote growth more effectively than fixed doses. Animals were treated with saline or 4.2 mg of recombinant bovine GH given as 1) 700 μg/wk in 100 μg/day doses, 2) alternating weekly doses of 966 (138 μg/day) or 434 μg (62 μg/day), or 3) 700 μg/wk in randomized daily doses (5-250 μg/day). Body weight and length were measured weekly. Femur and tibia lengths and internal organ, fat pad, and muscle weights were recorded at the end of the study (6 wk); blood was collected for IGF axis measurements. GH promoted femur [F(3,60) 14.67, P < 0.05], tibia [F(3,60) = 14.90, P < 0.051, muscle [F(3,60) = 10.37, P < 0.051, and organ growth [liver: F(3,60) = 9.30, P < 0.05; kidney: F(3,60) = 2.82, P < 0.051 and an increase in serum IGF-I [F(3,60) = 9.18, P < 0.05] and IGFBP-3 [F(3,60) = 6.70, P < 0.051 levels. IGF-I levels correlated with final weight (r = 0.45, P < 0.05) and length (r = 0.284, P < 0.05) in the whole cohort, but within each group, growth parameters correlated with serum IGF-I only in animals treated with random GH doses. The variable regimens promoted femur length (P < 0.05) and muscle (P < 0.05) and kidney (P < 0.05) weight more effectively than treatment with the fixed regimen. This study demonstrates that aspects of growth are improved following introduction of infradian variation to GH treatment in a GH-deficient model. The data suggest that varying the pattern of OH doses administered to children may enhance growth performance without increasing the overall GH dose. [ABSTRACT FROM AUTHOR]

The effects of insulin-like growth factor-I (IGF-I), insulin, and leucine on protein turnover and pathways that regulate proteolytic gene expression and protein polyubiquitination were investigated in primary cultures of 4-day-old rainbow trout myocytes. Supplementing media with 100 nM IGF-I increased protein synthesis by 13% (P < 0.05) and decreased protein degradation by 14% (P < 0.05). Treatment with 1 μM insulin increased protein synthesis by 13% (P < 0.05) and decreased protein degradation by 17% (P < 0.05). Supplementing media containing 0.6 mM leucine with an additional 2.5 mM leucine did not increase protein synthesis rates but reduced rates of protein degradation by 8% (P < 0.05). IGF-I (1 nM-100 nM) and insulin (1 nM1 μM) independently reduced the abundance of ubiquitin ligase mRNA in a dose-dependent manner, with maximal reductions of ∼70% for muscle atrophy F-box (Fbx) 32, 40% for Fbx25, and 25% for muscle RING finger-1 (MuRF1, P < 0.05). IGF-I and insulin stimulated phosphorylation of FOXO1 and FOXO4 (P < 0.05), which was inhibited by the phosphatidylinositol 3-kinase (PI 3-kinase) inhibitor wortmannin, and decreased the abundance of polyubiquitinated proteins by 10-20% (P < 0.05). Supplementing media with leucine reduced Fbx32 expression by 25% (P < 0.05) but did not affect Fbx25 nor MuRF1 transcript abundance. Serum deprivation decreased rates of protein synthesis by 60% (P < 0.05), increased protein degradation by 40% (P < 0.05), and increased expression of all ubiquitin ligases. These data suggest that, similar to mammals, the inhibitory effects of IGF-I and insulin on proteolysis occur via PI 3-kinase/protein kinase B signaling and are partially responsible for the ability of these compounds to promote protein accretion. [ABSTRACT FROM AUTHOR]

This study was conducted to examine telomere biology in terms of improving insulin sensitivity in a type 2 diabetic animal model: Otsuka Long-Evans Tokushima fatty (OLETF) rats. To improve insulin sensitivity, pioglitazone (PG; 10 mg∙kg-1∙day-1) was administrated to OLETF rats from 20 to 40 wk of age, and the effects of treatment were compared with those in untreated OLETF or control Long-Evans Tokushima Otsuka fatty rats. At the end of the study, the homeostasis model assessment of insulin resistance significantly increased in OLETF rats but decreased in OLETF rats treated with PG. No shortening of telomere length was observed in the heart tissue of OLETF rats, whereas telomerase activity was decreased in OLETF heart tissue. The mRNA expression of both telomerase reverse transcriptase and telomere repeat binding factor 2 was downregulated in the hearts of OLETF rats. The protein expression of phospho-Akt, insulin-like growth factor, and endothelial nitric oxide synthase was reduced in OLETF rats. On the other hand, myocardial matrix metalloproteinase-9 expression was elevated in OLETF rats. The changes observed in OLETF rats were inhibited by PG treatment. However, protein and mRNA expression of Sirti, a lifespan modulator, were attenuated in OLETF rat hearts, although they were enhanced in OLETF rats with PG treatement. Myocardial fibrosis was less extensive and diastolic dysfunction more greatly ameliorated in PG-treated OLETF rats than in OLETF rats. These findings suggest that improving insulin sensitivity via the activation of peroxisom proliferator-activated receptor-y may exert regulatory effects on cardiac telomere biology and may have desirable morphological and functional effects on the diabetic heart. [ABSTRACT FROM AUTHOR]

The relationship between maternal plasma volume (PV) expansion and fetal growth is well established, but the underlying mechanisms remain unclear. Here, we examined the influence of maternal body weight and fetoplacental mass on gestational PV increment in the rat. Because IGF-l and IGF-II have growth-promoting and vasoactive properties, their relationship to PV expansion and fetoplacental growth was also studied. In normal rats, the gradual expansion of PV (+ 35% at day 22, i.e., term) was accompanied by a rise in circulating IGF-II (+45%) and a considerable drop in IGF-I (-73%). Increased maternal body weight induced by an obesogenic diet did not influence PV and circulating IGFs compared with rats on the standard diet. Combining the results from both diets, circulating IGF-II was the principal correlate of PV. A second experiment examined the effect of fetoplacental mass reduction by surgically removing half of the gestational sacs at day 16. This procedure reduced maternal PV and circulating IGF-II at term by 14% and 20%, respectively. We then investigated the effect of a constant infusion of IGF-II (1 mg·kg-1·day-1) from day 16, which raised circulating IGF-II by 38% and found increased PV (+ 19%) and a larger placental trophospongial area (+29%) at term. Our results indicate that the placenta, the primary source of IGF-II synthesis in pregnancy, drives PV expansion, and that IGF-II is among the regulatory factors of the gestational PV increment. Further studies should clarify whether IGF-II directly affects vascular function and/or indirectly promotes the secretion of placenta-derived vasoactive substances. [ABSTRACT FROM AUTHOR]

This study determined the effect of genetic variation, feed deprivation, and insulin-like growth factor-I (IGF-I) on weight loss, plasma IGF-I and growth hormone, and indexes of protein degradation in eight full-sibling families of rainbow trout. After 2 wk of feed deprivation, fish treated with IGF-I lost 16% less (P < 0.05) wet weight than untreated fish. Feed deprivation increased growth hormone (P < 0.05) and decreased IGF-I (P < 0.05), but hormone levels were not altered by IGF-I. Plasma 3-methylhistidine concentrations were not affected by IGF-I but were decreased after 2 wk (P < 0.05) and increased after 4 wk (P < 0.05) of feed deprivation. In white muscle, transcript abundance of genes in the ubiquitin-proteasome, lysosomal, and calpain- and caspase-dependent pathways were affected by feed deprivation (P < 0.05). IGF-I prevented the feed deprivation-induced upregulation of MAFbx (F-box) and cathepsin transcripts and reduced abundance of proteasomal mRNAs (P < 0.05), suggesting that reduction of protein degradation via these pathways may be partially responsible for the IGF-I-induced reduction of weight loss. Family variations in gene expression, IGF-I concentrations, and weight loss during fasting suggest genetic variation in the fasting response, with considerable impact on regulation of proteolytic pathways. These data indicate that nutrient availability, IGF-I, and genetic variation affect weight loss, in part through alterations of proteolytic pathways in rainbow trout, and that regulation of genes within these pathways is coordinated in a way that supports a similar physiological response. [ABSTRACT FROM AUTHOR]

This study assessed the potential therapeutic efficacy of adipose-derived stem cells (ASCs) on infarcted hearts. Myocardial infarction was induced in rat hearts by occlusion of the left anterior descending artery (LAD). One week after LAD occlusion, the rats were divided into three groups and subjected to transplantation of ASCs or transplantation of cell culture medium (CCM) or remained untreated. During a 1-mo recovery period, magnetic resonance imaging showed that the ASCtreated hearts had a significantly greater left ventricular (LV) ejection fraction and LV wall thickening than did the CCM-treated and untreated hearts. The capillary density in infarct border zone was significantly higher in the ASC-treated hearts than in the CCM-treated and untreated hearts. However, only 0.5% of the ASCs recovered from the ASC-treated hearts were stained positive for cardiac-specific fibril proteins. It was also found that ASCs under a normal culture condition secreted three cardiac protective growth factors: vascular endothelial growth factor, hepatocyte growth factor, and insulin-like growth factor-1. Results of this study suggest that ASCs were able to improve cardiac function of infarcted rat hearts. Paracrine effect may be the mechanism underlying the improved cardiac function and increased capillary density. [ABSTRACT FROM AUTHOR]

We examined postprandial changes in circulating growth hormone (GH), insulin, insulin-like growth factor (IGF)-I, and IGF-binding proteins (IGFBPs) in yearling coho salmon under different feeding regimes. Fish were initially fasted for 1 day, 1 wk, or 3 wk. Fasted fish were then fed, and blood was collected at 4-h intervals over 26 h. After the various periods of fasting, basal levels of insulin were relatively constant, whereas those of IGF-I, IGFBPs and GH changed in proportion to the duration of the fast. A single meal caused a rapid, large increase in the circulating insulin levels, but the degree of the increase was influenced by the fasting period. IGF-I showed a moderate increase 2 h after the meal but only in the regularly fed fish. Plasma levels of 41-kDa IGFBP were increased in all groups within 6 h after the single meal. The fasting period did not influence the response of 41-kDa IGFBP to the meal. IGFBP- 1 and GH decreased after the meal to the same extent among groups regardless of the fasting period. The present study shows that insulin and IGF-I respond differently to long (weeks)- and short (hours)-term nutritional changes in salmon; insulin maintains its basal level but changes acutely in response to food intake, whereas IGF-I adjusts its basal levels to the long-term nutritional status and is less responsive to acute nutritional input. IGFBPs maintain their sensitivity to food intake, even after prolonged fasting, suggesting their critical role in the nutritional regulation of salmon growth. [ABSTRACT FROM AUTHOR]

Protein tyrosine phosphatase-α (PTPα) is a widely expressed receptor-type phosphatase that functions in multiple signaling systems. The actions of PTPα can be regulated by its phosphorylation on serine and tyrosine residues, although little is known about the conditions that promote PTPα phosphorylation. In this study, we tested the ability of several extracellular factors to stimulate PTPα tyrosine phosphorylation. The growth factors IGF-I and acidic FGF induced the highest increase in PTPα phosphorylation at tyrosine 789, followed by PMA and lysophosphatidic acid, while EGF had little effect. Further investigation of IGF-I-induced PTPα tyrosine phosphorylation demonstrated that this occurs through a novel Src family kinase-independent mechanism that does not require focal adhesion kinase, phosphatidylinositol 3-kinase, or MEK. We also show that PTPα physically interacts with the IGF-I receptor. In contrast to IGF-I-induced PTPα phosphorylation, this association does not require IGF-I. The interaction of PTPα and the IGF-I receptor is independent of PTPα catalytic activity, and expression of exogenous PTPα does not promote IGF-T receptor tyrosine dephosphorylation, indicating that PTPα does not act as an IGF-I receptor phosphatase. However, PTPα mediates IGF-I signaling, because IGF- I-stimulated fibroblast migration was reduced by ∼50% in cells lacking PTPα or in cells with mutant PTPα lacking the tyrosine 789 phosphorylation site. Our results suggest that PTPα tyrosine phosphorylation can occur in response to diverse stimuli and can be mediated by various tyrosine kinases. In the case of IGF-I, we propose that IGF-I-induced tyrosine 789 phosphorylation of PTPα, possibly catalyzed by the PTPα-associated IGF-I receptor tyrosine kinase, is required for efficient cell migration in response to this growth factor. [ABSTRACT FROM AUTHOR]

Endogenous insulin-like growth factor-I (IGF-I) regulates intestinal smooth muscle growth by concomitantly stimulating proliferation and inhibiting apoptosis. IGF-I-stimulated growth is augmented by the αvβ3 integrin ligands vitro-nectin and fibronectin. IGF-I expression in smooth muscle is increased in both TNBS-induced colitis and Crohn's disease. We hypothesized that intestinal inflammation increased vitronectin and fibronectin expression by smooth muscle and, along with IGF-I upregulation, increased intestinal muscle growth. Intestinal smooth muscle cells were examined 7 days following the induction of TNBS-induced colitis. Although αvβ3 integrin expression was not altered by TNBS-induced colitis, vitronectin and fibronectin levels were increased by 80 ± 10% and 90 ± 15%, above control levels, respectively. Basal IGF-I receptor phosphorylation in inflamed muscle from TNBS-treated rats was increased by 86 ± 8% over vehicle-treated controls. Basal ERK1/2, p70S6 kinase, and GSK-3β phosphorylation in muscle cells of TNBS-treated rats were also increased by 140-180%. TNBS treatment increased basal muscle cell proliferation by 130 ± 15% and decreased apoptosis by 20 ± 2% compared with that in vehicle-treated controls. The changes in proliferation and apoptosis were reversed by an IGF-l receptor tyrosine kinase inhibitor or an αvβ3 integrin antagonist. The results suggest that smooth muscle hyperplasia in TNBS-induced colitis partly results from the upregulation of endogenous IGF-I and ligands of αvβ3 integrin that mediate increased smooth muscle cell proliferation and decreased apoptosis. This paper has identified one mechanism regulating smooth muscle hyperplasia, a feature of stricture formation that occurs in the chronically inflamed intestine of TNBS-induced colitis and potentially Crohn's disease. [ABSTRACT FROM AUTHOR]

The insulin-like growth factor (IGF) signaling pathway is a conserved pathway that regulates animal development, growth, metabolism, reproduction, and aging. The biological actions of IGFs are modulated by IGF-binding proteins (IGFBPs). Although the structure and function of fish IGFBP-1, -2, -3, and -5 have been elucidated, there is currently no report on the full-length structure of a fish IGFBP-4 nor its biological action. In this study, we cloned and characterized the IGFBP-4 gene from fugu. Sequence comparison, phylogenetic, and synteny analyses indicate that its chromosomal location, gene, and protein structure are similar to its mammalian orthologs. Fugu IGFBP-4 mRNA was easily detectable in all adult tissues examined with the exception of spleen. Older animals tended to have higher levels of IGFBP-4 mRNA in the muscle and eyes compared with younger animals. Starvation resulted in significant increases in IGFBP-4 mRNA abundance in the muscle, liver, gallbladder, and brain. Overexpression of fugu and human IGFBP-4 in zebrafish embryos caused a significant decrease in body size and somite number, suggesting that fugu IGFBP-4 inhibits growth and development, possibly by binding to IGFs and inhibiting their binding to the IGF receptors. These results provide new information about the structural and functional conservation, expression patterns, and physiological regulation of the IGFBP-4 gene in a teleost fish. [ABSTRACT FROM AUTHOR]

We recently demonstrated that interferon (IFN)-β has a more potent antitumor activity than IFN-α in BUN cells, a neuroendocrine tumor (NET) cell line. The present study showed the role of type I IFNs in the modulation of the insulin-like growth factor (IGF) system in NETs. BUN cells expressed IGF-I, IGF-II, IGF-I receptor, and insulin receptor mRNA. In addition, IGF-I and IGF-II stimulated the proliferation of BUN cells and induced an inhibition of DNA fragmentation (apoptosis). As evaluated by quantitative RT-PCR, treatment with IFN-α (100 lU/mi) or IFN-β (100 IU/ml) inhibited the expression of IGF-II mRNA (-42% and -65%, respectively, both P < 0.001), whereas IGF-I receptor mRNA was significantly upregulated by IFN-α (+28%, P < 0.00 1) and downregulated by IFN-β (-47%, P < 0.001). Immunoreactive IGF-II concentration decreased in the conditioned medium during IFN-α (-16%, P < 0.05) and IFN- β (-69%, P < 0.001) treatment. Additionally, IGF-I receptor bioactivity was reduced (-54%) after IFN-β treatment. Scatchard analysis of 125I- labeled IGF-I binding to cell membrane of BUN cells revealed a dramatic suppression of maximum binding capacity only in the presence of IFN-β. Finally, the proapoptotic activity of IFN-β was partially counteracted by the coadministration of IGF-I and IGF-II (both at 50 nM). In conclusion, these data demonstrate that the IGF system has an important role in autocrine/paracrine growth of BUN cells. The more potent antitumor activity of IFN-β compared with IFN-β could be explained by several effects on this system: 1) both IFNs inhibit the transcription of IGF-II, but the suppression is significantly higher after IFN-β than IFN-α and 2) only IFN-β inhibits the expression of IGF-I receptor. [ABSTRACT FROM AUTHOR]

Intrafetal insulin-like growth factor (IGF)-I promotes cardiac hypertrophy in the late-gestation fetal sheep; whether these effects are sustained is unknown. IGF-I was infused for 4 days at 80 μg/h from 121 to 125 days of gestation, and its effects at 128 days, 3 days after the infusion stopped, were determined by comparison with untreated fetal sheep. After IGF-I treatment, fetal weights were similar to those in control fetuses but kidney weights were bigger (P < 0.05), as were spleen weights of male fetuses (P < 0.05). Cardiac myocytes were larger in female than male fetal sheep (P < 0.001). IGF-I increased male (P < 0.001) but not female myocyte volumes. IGF-I did not alter the proportions of uni- or binucleated right or left ventricular myocytes. Female fetal sheep had a greater proportion of binucleated cardiac myocytes than males (P < 0.05). IGF-I-treated fetuses had a slightly greater proportion of right ventricular nuclei in cell cycle phase G2/M and a reduced proportion of G0/G1 phase nuclei (P < 0.1). Therefore, evidence for IGF-l-stimulated cardiac cell hyperplasia in fetal sheep in late gestation was limited. In conclusion, the greater sizes and larger proportion of binucleated cardiac myocytes in female fetal sheep suggest that myocyte maturation may occur earlier in females than in males. This may explain in part the male sex-specific responsiveness of cardiac hypertrophy to IGF-I in late gestation. If IGF-I-stimulated cardiomyocyte growth is accompanied by maturation of contractile function, IGF-I may be a potential therapeutic agent for maintaining cardiac output in preterm males. [ABSTRACT FROM AUTHOR]

Glucagon-like peptide-2 (GLP-2) is a nutrient-dependent, intestinotrophic hormone derived from posttranslational processing of proglucagon in the distal bowel. GLP-2 is thought to act through indirect mediators, such as IGF-I. We investigated whether intestinal expression of GLP-2 and IGF-I system components are increased with the mucosal growth induced by enteral nutrient (EN) and/or a low dose of GLP-2 in parenterally fed rats. Rats were randomized to four treatment groups using a 2X2 design and maintained with parenteral nutrition (PN) for 7 days: PN alone, EN, GLP-2, and EN+GLP-2; n = 7-9. The two main treatment effects are ±GLP-2 (100 µg·kg body wt-1·day-1) and ±EN (43% of energy needs, days 4-6). Combination treatment with EN+GLP-2 induced synergistic intestinal growth in ileum, resulting in greater mucosal cellularity, sucrase segmental activity, and gain of body weight (ENXGLP-2, P < 0.04). In addition, EN+GLP-2 induced a significant 28% increase in plasma concentration of bioactive GLP-2, a significant 102% increase in ileal proglucagon mRNA with no change in ileal dipeptidyl peptidase-IV (DPP-IV) specific activity, and significantly reduced plasma DPP-IV activity compared with GLP-2. This indicates that EN potentiates the intestinotrophic action of GLP-2. Proliferation of enterocytes due to GLP-2 infusion was associated with greater expression of ileal proglucagon, GLP-2 receptor, IGF-I, IGF binding protein-3 mRNAs, and greater IGF-I peptide concentration in ileum (P < 0.032). Ileal IGF-I mRNA was positively correlated with expression of proglucagon, GLP-2R, and IGFBP-5 mRNAs (R² = 0.43-0.56, P < 0.0001). Our findings support the hypothesis that IGF-I is one of the downstream mediators of GLP-2 action in a physiological model of intestinal growth. [ABSTRACT FROM AUTHOR]

It is generally well accepted that the pubertal surge in estrogen is responsible for the rapid bone accretion that occurs during puberty and that this effect is mediated by an estrogen-induced increase in growth hormone (GH)/insulin-like growth factor (IGF) action. To test the cause and effect relationship between estrogen and GH/IGF, we evaluated the consequence of ovariectomy (OVX) in prepubertal mice (C57BL/6J mice at 3 wk of age) on skeletal changes and the GH/IGF axis during puberty. Contrary to our expectations, OVX increased body weight (12-18%), bone mineral content (11%), bone length (4%), bone size (3%), and serum, liver, and bone IGF-I (30-50%) and decreased total body fat (18%) at 3 wk postsurgery. To determine whether estrogen is the key ovarian factor responsible for these changes, we performed a second experiment in which OVX mice were treated with placebo or estrogen implants. In addition to observing similar results compared with our first experiment, estrogen treatment partially rescued the increased body weight and bone size and completely rescued body fat and IGF-I levels. The increased bone accretion in OVX mice was due to increased bone formation rate (as determined by bone histomorphometry) and increased serum procollagen peptide. In conclusion, contrary to the known estrogen effect as an initiator of GH/IGF surge and thereby pubertal growth spurt, our findings demonstrate that loss of estrogen and/or other hormones during the prepubertal growth period effect leads to an increase in IGF-I production and bone accretion in mice. [ABSTRACT FROM AUTHOR]

IGF-I action, is essential for the regulation of tissue formation and remodeling, bone growth, prenatal growth, brain development, and muscle metabolism. Cellular effects of IGF-I are mediated through the IGF-I receptor, a transmembrane tyrosine kinase that phosphorylates intracellular substrates, resulting in the activation of multiple intracellular signaling cascades. Dysregulation of IGF-I actions due to impairment in the postreceptor signaling machinery may contribute to multiple diseases in humans. This article will review current information on IGF-I signaling and illustrate recent results demonstrating how impaired IGF-I signaling and action may contribute to the pathogenesis of human diseases, including osteoporosis, neuro-degenerative disorders, and reduced fetal growth in utero. [ABSTRACT FROM AUTHOR]

Chronic resistive airway loading (CAL) impairs growth in juvenile rats. The effects of CAL on epiphyseal growth plate (EGP) structure and insulin-like growth factor (IGF)-I gene expression have not been explored. Little is known about whether stimulants of endogenous growth hormone (GH) secretion can normalize this growth impairment. This study explored the effect of CAL on circulating and EGP GH/IGF-I pathway GH and the effect of ritanserin (endogenous GH stimulant) on somatic growth and the GH/IGF-I axis. We hypothesized that CAL would lead to a decrease in body temperature (T[subb]) and alterations of GH/IGF-I pathways, consequently leading to growth retardation. The tracheae of 22-day-old male rats were obstructed by tracheal banding (38 sham-operated control, 42 CAL). Tibial EGP morphometry, liver and EGP IGF mRNA, and serum GH and IGF-I levels were analyzed with quantitative real-time PCR and ELISA. T[subb] and locomotion activity (MA) were measured with telemetric transmitters inserted into the abdominal cavity. CAL animals had lower T[subb] and MA despite preserved food consumption. CAL impaired both tibial and tail length gains. Tail and tibial length gains inversely correlated with tracheal resistance. Circulating GH and IGF-I, liver and EGP IGF-I mRNA, and EGP width were decreased in the CAL group. Ritanserin administration to CAL animals normalized circulating and local EGP GH and IGF-I levels and minimized the longitudinal growth impairment. We conclude that CAL causes growth delay associated with alterations in the GH/IGF-I axis. Stimulation of GH release by ritanserin restored both global and local GH/IGF-I pathways, yet growth parameters were only partially restored. [ABSTRACT FROM AUTHOR]

Low bone mineral density (BMD) is a phenotype associated with osteoporosis and increased risk of fracture. Since 60–80% of variation in BMD is associated with genetic factors, we used the novel approach of chromosome substitution strains (CSS) to identify chromosomes that harbor genes that regulate BMD. Specifically, we evaluated 24 wk old C57BL/6J-Chr #A/J/NaJ CSS (n = 7 to 18) in which each chromosome in the host C57BL/6J strain is replaced by a corresponding chromosome from the donor A/J strain (19 autosomes, X, Y). We determined several A/J chromosomes contribute to body weight (BW), percent body fat (BF), whole body areal BMD (aBMD), and serum insulin-like growth factor (IGF)-I in both a positive and negative manner when compared with C57BL/6J. Specifically, C57BL/6J-Chr 5A/J/NaJ (B.A-5) (males) and B.A-13 (females) contributed to increased BW, whereas B.A-3, 4, 8, 9, 12, and 18 (males) and B.A-3, 4, and 11 (females) contributed to reduced BW. B.A-5 (males) and B.A-13 (females) contributed to increased BF, whereas B.A-12 (males) and B.A-3, 4, 10, and 11 (females) contributed to reduced BF. B.A-14 (females) contributed to increased aBMD and B.A-1, 2, 6, 9, 10, and 18 (males) and B.A-8, 9, and 10 (females) contributed to reduced aBMD. To determine if similar chromosomes regulate aBMD and IGF-I, we determined serum concentrations of IGF-I. B.A-14 and Y (males) and B.A-6 (females) contributed to increased IGF-I and male B.A-3 and female B.A-8 contributed to reduced IGF-I. Overall, we identified several sex-dependent and -independent chromosomes that regulate aBMD and IGF-I in adult mice. [ABSTRACT FROM AUTHOR]

Hammers DW, Merritt EK, Matheny W, Adamo ML, Walters TJ, Estep JS, Farrar RP. Functional deficits and insulin-like growth factor-I gene expression following tourniquet-induced injury of skeletal muscle in young and old rats. JAppi Physiol 105: 1274-1281,2008. First published July 31, 2008; doi: 10.11 52/japplphysiol.9041 8.2008.-This study investigated the effect of age on recovery of skeletal muscle from an ischemia-reperfusion (LIR)-induced injury. Young (6 mo old) and old (24-27 mo old) Sprague-Dawley rats underwent a 2-h bout of hindlimb ischemia induced by a pneumatic tourniquet (TK). The TK was released to allow reperfusion of the affected limb, and animals were divided into 7and 14-day recovery groups. Maximum plantar flexor force production was assessed in both 7and 14-day recovery groups of both ages, followed by histological evaluation. Subsequent analysis of IGF-I gene expression and intracellular signaling in 7-day recovery muscles was performed by RT-PCR and Western blotting, respectively. Old rats had significantly greater deficits in force production and exhibited more evidence of histological pathology than young at both 7 and 14 days postinjury. In addition, old rats demonstrated an attenuated upregulation of IGF-I mRNA and induction of proanabolic signaling compared with young in response to injury. We conclude that aged skeletal muscle exhibits more damage and/or defective regeneration following hR and identify an age-associated decrease in local IGF-I responsiveness as a potential mechanism for this phenomenon. [ABSTRACT FROM AUTHOR]

Chronic treatment with insulin-like growth factor I (IGF-I) improves contractile function in congestive heart failure and ischemic cardiomyopathy. The present study investigated the effect of chronic treatment with IGF-I on intrinsic myocyte function and the role of the phosphatidylinositol (PI)3-kinase-Akt-sarco(endo)plasmic reticulum Ca2+-ATPase (SERCA)2a signaling cascade in these responses. Myocytes were isolated from 23 adult rats and cultured with and without IGF-I (10-6 M). After 48 h of treatment, myocyte function was evaluated. IGF-I increased contractile function (percent contraction, 7.7 ± 0.3% vs. 4.5 ± 0.3%; P < 0.01) and accelerated relaxation time (time for 70% relengthening, 81 ± 4 vs. 106 ± 5 ms; P < 0.05) compared with untreated myocytes [control (Con)]. The enhanced function was associated with an in- crease in Ca2+ transients assessed by fura-2 (340/380 nm; IGF-I, 0.42 ± 0.02 vs. Con, 0.25 ± 0.01; P <0.01). The PI3-kinase inhibitor LY-249002 (10-9 M) abolished the enhanced function caused by IGF-I. IGF-I increased both Akt and SERCA2a protein levels 2.5- and 4.8-fold, respectively, compared with those of Con (P < 0.01); neither phospholamban nor calsequestrin was affected. To evaluate whether the SERCA2a protein was directly mediated by Akt-SERCA2a signaling, IGF-I-induced changes in the SERCA2a protein were compared in myocytes transfected with adenovirus harboring either constitutively active Akt [multiplicity of infection (MOI), 15] or dominant negative Akt (dnAkt; MOI, 15). The ability of IGF-I to upregulate the SERCA2a protein in myocytes transected With active Akt was absent in dnAkt myocytes. Taken together, our findings indicate that chronic treatment with IGF-I enhances intrinsic myocyte function and that this effect is due to an enhancement in intracellular Ca2+ handling, secondary to the activation of the PI3-kinase-Akt-SERCA2a signaling cascade. [ABSTRACT FROM AUTHOR]

Energy restriction coupled with high energy expenditure from arduous work is associated with an altered insulin-like growth factor-I (IGF-I) system and androgens that are coincident with losses of fat-free mass. The aim of this study was to determine the effects of two levels of dietary protein content and its effects on IGF-I, androgens, and losses of fat-free mass accompanying energy deficit. We hypothesized that higher dietary protein content would attenuate the decline of anabolic hormones and, thus, prevent losses of fat-free mass. Thirty-four men [24 (SD 0.3) yr, 180.1 (SD 1.1) cm, and 83.0 (SD 1.4) kg] participated in an 8-day military exercise characterized by high energy expenditure (16.5 Mi/day), low energy intake (6.5 MJ/day), and sleep deprivation (4 h/24 h) and were randomly divided into two dietary groups: 0.9 and 0.5 g/kg dietary protein intake. IGF-I system analytes, androgens, and body composition were assessed before and on days 4 and 8 of the intervention. Total, free, and nonternary IGF-I and testosterone declined 50%, 64%, 55%, and 45%, respectively, with similar reductions in both groups. There was, however, a diet × time interaction on day 8 for total IGF-I and sex hormone-binding globulin. Decreases in body mass (3.2 kg), fat-free mass (1.2 kg), fat mass (2.0 kg), and percent body fat (1.5%) were similar in both groups (P = 0.01). Dietary protein content of 0.5 and 0.9 g/kg minimally attenuated the decline of IGF-I, the androgenic system, and fat-free mass during 8 days of negative energy balance associated with high energy expenditure and low energy intake. [ABSTRACT FROM AUTHOR]

The main disorders of human pregnancy are rooted in defective placentation. Normal placental development depends on proliferation, differentiation, and fusion of cytotrophoblasts to form and maintain an overlying syncytiotrophoblast. There is indirect evidence that the insulin-like growth factors (IGFs), which are aberrant in pregnancy disorders, are involved in regulating trophoblast turnover, but the processes that control human placental growth are poorly understood. Using an explant model of human first-trimester placental villus in which the spatial and ontological relationships between cell populations are maintained, we demonstrate that cytotrophoblast proliferation is enhanced by IGF-I/IGF-II and that both factors can rescue cytotrophoblast from apoptosis. Baseline cytotrophoblast proliferation ceases in the absence of syncytiotrophoblast, although denuded cytotrophoblasts can proliferate when exposed to IGF and the rate of cytotrophoblast differentiation/fusion and, consequently, syncytial regeneration, increases. Use of signaling inhibitors suggests that IGFs mediate their effect on cytotrophoblast proliferation/syncytial formation through the MAPK pathway, whereas effects on survival are regulated by the phosphoinositide 3-kinase pathway. These results show that directional contact between cytotrophoblast and syncytium is important in regulating the relative amounts of the two cell populations. However, IGFs can exert an exogenous regulatory influence on placental growth/development, suggesting that manipulation of the placental IGF axis may offer a potential therapeutic route to the correction of inadequate placental growth. [ABSTRACT FROM AUTHOR]

Luminal nutrients stimulate structural and functional regeneration in the intestine through mechanisms thought to involve insulin-like growth factor I (IGF-I) and glucagon-like peptide-2 (GLP-2). We investigated the relationship between IGF-I and GLP-2 responses and mucosal growth in rats fasted for 48 h and then refed for 2 or 4 days by continuous intravenous or intragastric infusion or ad libitum feeding. Fasting induced significant decreases in body weight, plasma concentrations of IGF-I and bioactive GLP-2, jejunal mucosal cellularity (mass, protein, DNA, and villus height), IGF-I mRNA, and ileal proglucagon mRNA. Plasma IGF-I concentration was restored to fed levels with 2 days of ad libitum refeeding but not with 4 days of intravenous or intragastric refeeding. Administration of an inhibitor of endogenous GLP-2 (rat GLP-23-33) during ad libitum refeeding partially attenuated mucosal growth and prevented the increase in plasma IGF-I to fed levels; however, plasma GLP-2 and jejunal IGF-I mRNA were restored to fed levels. Intragastric refeeding restored intestinal cellularity and functional capacity (sucrase activity and sodium-glucose transporter-1 expression) to fed levels, whereas intravenous refeeding had no effect. Intestinal regeneration after 4 days of intragastric or 2 days of ad libitum refeeding was positively associated with increases in plasma concentrations of GLP-2 and jejunal IGF-I mRNA. These data suggest that luminal nutrients stimulate intestinal growth, in part, by increased expression of both GLP-2 and IGF-I. [ABSTRACT FROM AUTHOR]

IGF-I rescues diabetic heart defects and oxidative stress, although the underlying mechanism of action remains poorly understood. This study was designed to delineate the beneficial effects of IGF-I with a focus on RhoA, Akt, and eNOS coupling. Echocardiography was performed in normal or diabetic Friend Virus-B type (FVB) and IGF-I transgenic mice. Cardiomyocyte contractile properties were evaluated using peak shortening (PS), time-to-90% relengthening (TR90), and intracellular Ca2+ rise and decay. Diabetes reduced fraction shortening, PS, and intracellular Ca2+ it increased chamber size, prolonged TR90, and intracellular Ca2+ decay. Levels of RhoA mRNA, active RhoA, and O2- were elevated, whereas nitric oxide (NO) levels were reduced in diabetes. Diabetes-induced O2- accumulation was ablated by the NO synthase (NOS) inhibitor nitro-L-arginine methyl ester (L-NAME), indicating endothelial NOS (eNOS) uncoupling, all of which except heart size were negated by IGF-I. The IGF-I-elicited beneficial effects were mimicked by the Rho kinase inhibitor Y27632 and BH4. Diabetes depressed expression of Kv1.2 and dihydrofolate reductase (DHFR), increased β-myosin heavy-chain expression, stimulated p38 MAPK, and reduced levels of total Akt and phosphorylated Akt/eNOS, all of which with the exception of myosin heavy chain were attenuated by IGF-I. In addition, Y27632 and the eNOS coupler folate abrogated glucose toxicity-induced PS decline, TR90 prolongation, while it increased O2- and decreased NO and Kv1.2 levels. The DHFR inhibitor methotrexate impaired myocyte function, NO/O2- balance, and rescued Y27632-induced cardiac protection. These results revealed that IGF-I benefits diabetic hearts via Rho inhibition and antagonism of diabetes-induced decrease in pAkt, eNOS uncoupling, and K+ channel expression. [ABSTRACT FROM AUTHOR]

Osteoblasts are key players in bone remodeling. The accessibility of human primary osteoblast-like cells (HObs) from bone explants makes them a lucrative model for studying molecular physiology of bone turnover, for discovering novel anabolic therapeutics, and for mesenchymal cell biology in general. Relatively little is known about resting and dynamic expression profiles of HObs, and to date no studies have been conducted to systematically assess the osteoblast transcriptome. The aim of this study was to characterize HObs and investigate signaling cascades and gene networks with genomewide expression profiling in resting and bone morphogenic protein (BMP)-2- and dexamethasone-induced cells. In addition, we compared HOb gene expression with publicly available samples from the Gene Expression Omnibus. Our data show a vast number of genes and networks expressed predominantly in HObs compared with closely related cells such as fibroblasts or chondrocytes. For instance, genes in the insulin-like growth factor (IGF) signaling pathway were enriched in HObs (P = 0.003) and included the binding proteins (IGFBP-1, -2, -5) and IGF-II and its receptor. Another HOb-specific expression pattern included leptin and its receptor (P < 10–8). Furthermore, after stimulation of HObs with BMP-2 or dexamethasone, the expression of several interesting genes and pathways was observed. For instance, our data support the role of peripheral leptin signaling in bone cell function. In conclusion, we provide the landscape of tissue-specific and dynamic gene expression in HObs. This resource will allow utilization of osteoblasts as a model to study specific gene networks and gene families related to human bone physiology and diseases. [ABSTRACT FROM AUTHOR]

Laboratory inbred mouse strains show a broad range of variation in phenotypes, such as body composition, bone mineral density (BMD), plasma leptin, and insulin-like growth factor I (IGF-I), and thus provide a basis for the study of associations among them. We analyzed these phenotypes in male and female mice from 43 inbred strains fed on a high-fat (30% caloric content) diet and from 30 inbred strains fed on a low-fat (6%) diet. Structural equation modeling of these data reveals that the relationship of body fat content and areal BMD is altered by dietary factors and genotypes. Sex has no net effect on areal BMD, but after accounting for body mass difference females have higher areal BMD. Leptin is affected by relative fat mass and has no net effect on areal BMD. IGF-I has a direct effect on areal BMD. [ABSTRACT FROM AUTHOR]

The insulin-like growth factors (IGF-I and IGF-II), working through the type I IGF receptor (IGF-1R), are key mediators of skeletal muscle fiber growth and hypertrophy. These processes are largely dependent on stimulation of proliferation and differentiation of muscle precursor cells, termed myoblasts. It has not been rigorously determined whether the IGFs can also mediate skeletal muscle hypertrophy in a myoblast-independent fashion. Similarly, although the phosphatidyl-inositol 3-kinase (PI3K) and calcineurin signaling pathways have been implicated in skeletal muscle hypertrophy, these pathways are also involved in skeletal myoblast differentiation. To determine whether the IGFs can stimulate skeletal muscle hypertrophy in a myoblast-independent fashion, we developed and validated a retroviral expression vector that mediated overexpression of the human IGF-1R in rat L6 skeletal myotubes (immature muscle fibers), but not in myoblasts. L6 myotubes transduced with this vector accumulated significantly higher amounts of myofibrillar proteins, in a ligand- and receptor-dependent manner, than controls and demonstrated significantly increased rates of protein synthesis. Stimulation of myotube hypertrophy was independent of myoblast contributions, inasmuch as these cultures did not exhibit increased levels of myoblast proliferation or differentiation. Experiments with PI3K and calcineurin inhibitors indicated that myoblast-independent myotube hypertrophy was mediated by PI3K, but not calcineurin, signaling. This study demonstrates that IGF can mediate skeletal muscle hypertrophy in a myoblast-independent fashion and suggests that muscle-specific overexpression of the IGF-1R or stimulation of its signaling pathways could be used to develop strategies to ameliorate muscle wasting without stimulating proliferative pathways leading to carcinogenesis or other pathological sequelae. [ABSTRACT FROM AUTHOR]

Fibroblast growth factors (FGFs) play critical roles in development, maintenance, and repair following injury or disease in the lung. Their activity is modulated by a variety of factors, including FGF-binding protein (FGF-BP; HBp-17) and N-deacetylase/N-sulfotransferase-1 (NDST-1). Functionally, FGF-BP shuttles FGFs from binding sites in ECMs to cell surfaces and enhances FGF binding and signaling, whereas NDST-1 adds sulfate groups to FGF coreceptor proteoglycans and modulates alveolar type II (ATII) cell maturation and differentiation. Since the sulfated nature of ECMs is a critical determinant of their relationship with FGFs, we predicted that ECMs and their sulfation would modulate the expression of FGF-BP and NDST-1. To examine this question, selected culture conditions of rat ATII cells were manipulated [with and without coculture with rat lung fibroblasts (RLFs)] by treatment with heparin or sodium chlorate (inhibitor of sulfation) for 24-96 h. In addition, ECMs biosynthesized by RLFs for up to 10 days before coculture were used as model intervening barriers to communication between alveolar cells and fibroblasts. FGF-BP expression was enhanced in ATII cells by coculture with RLF cells and least suppressed by desulfated heparin. NDST-1 expression in ATII cells was most sensitive to the amount of sulfation in medium and ECM and enhanced by fully sulfated heparin. Preformed ECM appears to supply factors that modify subsequent treatment effects. These results demonstrate a potentially important modulatory influence of sulfated ECMs and fibroblasts on FGF-BP and NDST-I at the gene expression level. [ABSTRACT FROM AUTHOR]

Estrogen receptors (ERs) are members of the nuclear receptor superfamily and are involved in regulation of fallopian tube functions (i.e., enhancement of protein secretion, formation of tubal fluid, and regulation of gamete transport). However, the ER subtype-mediated mechanisms underlying these processes have not been completely clarified. Recently, we identified ERβ expression and localization in rat fallopian tubes, suggesting a potential biological function of ERβ related to calcium-dependent ciliated beating. Here we provide for the first time insight into the less studied ERα isoforms, which mediate estrogen-dependent production and secretion of IGFs in vivo. First, Western blot studies revealed that three ERα isoforms were expressed in mouse fallopian tubes. Subsequent immunohistochemical analysis showed that ERa was detected in all cell types, whereas ERβ was mainly localized in ciliated epithelial cells. Second, ERrs isoform levels were dramatically down-regulated in mouse fallopian tubes by treatment with E2 or PPT, an ERα agonist, in a time-dependent manner. Third, the presence of ICI 182,780, an ER antagonist, blocked the E2- or PPT-induced down-regulation of tubal ERα isoform expression in mice. However, alteration of ERα immunoreactivity following ICI 182,780 treatment was only detected in epithelial cells of the ampullary region. Fourth, changes in ERa isoform expression were found to be coupled to multiple E2 effects on tubal growth, protein synthesis, and secretion in mouse fallopian tube tissues and fluid. In particular, E2 exhibited positive regulation of IGF-I and IGF-II protein levels. Finally, using growth hormone receptor (GHR) gene-disrupted mice, we showed that regulation by E2 of IGF production was independent of GH- induced GHR signaling in mouse fallopian tubes in vivo. These data, together with previous studies from our laboratory, suggest that the long-term effects of estrogen agonist promote IGF synthesis and secretion in mouse tubal epithelial cells and fallopian tube fluid via stimulation of ERα. [ABSTRACT FROM AUTHOR]