Insulin-like Growth Factor-1 Treatment Research Articles

Improvement of Eye Alignment in Adult Strabismic Monkeys by Sustained IGF-1 Treatment.

PurposeThe goal of this study was to determine if continuous application of insulin-like growth factor-1 (IGF-1) could improve eye alignment of adult strabismic nonhuman primates and to assess possible mechanisms of effect.MethodsA continuous release pellet of IGF-1 was placed on one medial rectus muscle in two adult nonhuman primates (M1, M2) rendered exotropic by the alternating monocular occlusion method during the first months of life. Eye alignment and eye movements were recorded for 3 months, after which M1 was euthanized, and the lateral and medial rectus muscles were removed for morphometric analysis of fiber size, nerve, and neuromuscular density.ResultsMonkey 1 showed a 40% reduction in strabismus angle, a reduction of exotropia of approximately 11° to 14° after 3 months. Monkey 2 showed a 15% improvement, with a reduction of its exotropia by approximately 3°. The treated medial rectus muscle of M1 showed increased mean myofiber cross-sectional areas. Increases in myofiber size also were seen in the contralateral medial rectus and lateral rectus muscles. Similarly, nerve density increased in the contralateral medial rectus and yoked lateral rectus.ConclusionsThis study demonstrates that in adult nonhuman primates with a sensory-induced exotropia in infancy, continuous IGF-1 treatment improves eye alignment, resulting in muscle fiber enlargement and altered innervational density that includes the untreated muscles. This supports the view that there is sufficient plasticity in the adult ocular motor system to allow continuous IGF-1 treatment over months to produce improvement in eye alignment in early-onset strabismus.

PREX1 Protein Function Is Negatively Regulated Downstream of Receptor Tyrosine Kinase Activation by p21-activated Kinases (PAKs)

Downstream of receptor tyrosine kinase and G protein-coupled receptor (GPCR) stimulation, the phosphatidylinositol 3,4,5-trisphosphate (PIP3)-dependent Rac exchange factor (PREX) family of guanine nucleotide exchange factors (GEFs) activates Rho GTPases, leading to important roles for PREX proteins in numerous cellular processes and diseases, including cancer. PREX1 and PREX2 GEF activity is activated by the second messengers PIP3 and Gβγ, and further regulation of PREX GEF activity occurs by phosphorylation. Stimulation of receptor tyrosine kinases by neuregulin and insulin-like growth factor 1 (IGF1) leads to the phosphorylation of PREX1; however, the kinases that phosphorylate PREX1 downstream of these ligands are not known. We recently reported that the p21-activated kinases (PAKs), which are activated by GTP-bound Ras-related C3 botulinum toxin substrate 1 (Rac1), mediate the phosphorylation of PREX2 after insulin receptor activation. Here we show that certain phosphorylation events on PREX1 after insulin, neuregulin, and IGF1 treatment are PAK-dependent and lead to a reduction in PREX1 binding to PIP3 Like PREX2, PAK-mediated phosphorylation also negatively regulates PREX1 GEF activity. Furthermore, the onset of PREX1 phosphorylation was delayed compared with the phosphorylation of AKT, supporting a model of negative feedback downstream of PREX1 activation. We also found that the phosphorylation of PREX1 after isoproterenol and prostaglandin E2-mediated GPCR activation is partially PAK-dependent and likely also involves protein kinase A, which is known to reduce PREX1 function. Our data point to multiple mechanisms of PREX1 negative regulation by PAKs within receptor tyrosine kinase and GPCR-stimulated signaling pathways that have important roles in diseases such as diabetes and cancer.

Effects of age and insulin-like growth factor-1 on rat neurotrophin receptor expression after nerve injury.

Neurotrophin receptors, such as p75(NTR) , direct neuronal response to injury. Insulin-like growth factor-1 receptor (IGF-1R) mediates the increase in p75(NTR) during aging. The aim of this study was to examine the effect of aging and insulin-like growth factor-1 (IGF-1) treatment on recovery after peripheral nerve injury. Young and aged rats underwent tibial nerve transection with either local saline or IGF-1 treatment. Neurotrophin receptor mRNA and protein expression were quantified. Aged rats expressed elevated baseline IGF-1R (34% higher, P = 0.01) and p75(NTR) (68% higher, P < 0.01) compared with young rats. Post-injury, aged animals expressed significantly higher p75(NTR) levels (68.5% above baseline at 4 weeks). IGF-1 treatment suppressed p75(NTR) gene expression at 4 weeks (17.2% above baseline, P = 0.002) post-injury. Local IGF-1 treatment reverses age-related declines in recovery after peripheral nerve injuries by suppressing p75(NTR) upregulation and pro-apoptotic complexes. IGF-1 may be considered a viable adjuvant therapy to current treatment modalities. Muscle Nerve 54: 769-775, 2016.

Neuroprotective levels of IGF-1 exacerbate epileptogenesis after brain injury

Exogenous Insulin-Like Growth Factor-1 (IGF-1) is neuroprotective in animal models of brain injury, and has been considered as a potential therapeutic. Akt-mTOR and MAPK are downstream targets of IGF-1 signaling that are activated after brain injury. However, both brain injury and mTOR are linked to epilepsy, raising the possibility that IGF-1 may be epileptogenic. Here, we considered the role of IGF-1 in development of epilepsy after brain injury, using the organotypic hippocampal culture model of post-traumatic epileptogenesis. We found that IGF-1 was neuroprotective within a few days of injury but that long-term IGF-1 treatment was pro-epileptic. Pro-epileptic effects of IGF-1 were mediated by Akt-mTOR signaling. We also found that IGF-1 – mediated increase in epileptic activity led to neurotoxicity. The dualistic nature of effects of IGF-1 treatment demonstrates that anabolic enhancement through IGF-1 activation of mTOR cascade can be beneficial or harmful depending on the stage of the disease. Our findings suggest that epilepsy risk may need to be considered in the design of neuroprotective treatments for brain injury.

Dexras1 links glucocorticoids to insulin-like growth factor-1 signaling in adipogenesis.

Glucocorticoids are associated with obesity, but the underlying mechanism by which they function remains poorly understood. Previously, we showed that small G protein Dexras1 is expressed by glucocorticoids and leads to adipocyte differentiation. In this study, we explored the mechanism by which Dexras1 mediates adipogenesis and show a link to the insulin-like growth factor-1 (IGF-1) signaling pathway. Without Dexras1, the activation of MAPK and subsequent phosphorylation of CCAAT/enhancer binding protein β (C/EBPβ) is abolished, thereby inhibiting mitotic clonal expansion and further adipocyte differentiation. Dexras1 translocates to the plasma membrane upon insulin or IGF-1 treatment, for which the unique C-terminal domain (amino acids 223–276) is essential. Dexras1-dependent MAPK activation is selectively involved in the IGF-1 signaling, because another Ras protein, H-ras localized to the plasma membrane independently of insulin treatment. Moreover, neither epidermal growth factor nor other cell types shows Dexras1-dependent MAPK activation, indicating the importance of Dexras1 in IGF-1 signaling in adipogenesis. Dexras1 interacts with Shc and Raf, indicating that Dexras1-induced activation of MAPK is largely dependent on the Shc-Grb2-Raf complex. These results suggest that Dexras1 is a critical mediator of the IGF-1 signal to activate MAPK, linking glucocorticoid signaling to IGF-1 signaling in adipogenesis.

449. IGF-1 Treatment Enhances the Myogenic Potential of Human Skeletal Muscle-Derived Stem Cells

Human muscle-derived stem cells (hMDSCs) have been shown to promote the regeneration of a variety of tissues, including damaged heart, peripheral nerve, bone, articular cartilage and skeletal muscle1xSee all References, 2xSee all References, 3xSee all References; however, without appropriate stimulation their differentiation potential remains limited.4xSee all References In this study we examined whether insulin-like growth factor 1 (IGF-1) treatment could also enhance the myogenic potential of hMDSC, since it has been shown that IGF-1 treatment increased the myogenic potential of bone marrow-derived stem cells (BMSCs).5xSee all References Methods: HMDSCs were isolated from human adult muscle biopsies via a modified preplate technique, as previously described.6xSee all References Young female hMDSCs were plated in proliferation medium at a density of 1.58 × 104 cells/cm2. After 24h in proliferation medium, the cells were cultured for an additional 7 days in myogenic medium containing various levels of IGF-1: 0ng/mL, 50 ng/mL, 100 ng/mL, or 200 ng/mL. To evaluate the extent of myogenesis, cells were fixed in MeOH and stained for fast myosin heavy chain (fMHC). Activity levels of creatine kinase, which is highly expressed in tissues like skeletal muscle that rapidly consume ATP, were measured according to the company's protocol. Additionally, RNA was extracted from cells, and cDNA was synthesized. Semi-quantitative PCR and qRT-PCR analyses were performed for myogenin, a late myogenic differentiation marker. Results: (1) fMHC staining indicated that IGF-1 treatment increased myotube formation (Fig. 1AFig. 1A). However, the mortality rates of the cells in all IGF-1 treatment groups were increased compared to the non-treated group. (2) hMDSCs treated with 50 ng/ml IGF-1 had significantly higher CK activity compared to all other groups when normalized to total protein. Furthermore, cells treated with 200 ng/ml IGF-1 also had significantly more CK activity than the untreated cells. This suggests that IGF-1 may enhance CK activity at low levels. At higher IGF-1 concentrations, CK activity may be reduced due to increased cell death (Fig. 1BFig. 1B). (3) Semi-quantitative analysis showed that IGF-1 treatment, at all concentrations, increased myogenin expression. The 100ng/mL concentration appeared to have the greatest effect. These results were corroborated by qRT-PCR analysis, which indicated that, compared to the non treated group, the 50ng/mL, 100ng/mL, and 200ng/mL groups had a 16.1 fold, 30.1 fold, and 14.3 fold increase in myogenin expression, respectively. Conclusion: These results indicate that IGF-1 enhances the myogenic potential of hMDSCs. However, we observed more cell death in IGF-1 treatment groups, which may be a result of overgrown cells during differentiation. In the future, we will determine whether IGF-1 treated hMDSCs display a higher regenerative potential in skeletal muscle, after injury and disease, than non-treated hMDSCs.View Large Image | Download PowerPoint SlideView Large Image | Download PowerPoint Slide

Growth Modulation of Diabetic Factors and Antidiabetic Drugs on Prostate Cancer Cell Lines.

Risk factors for prostate cancer (PCa) include age, hormones, race, family history and diet. Recently, epidemiologic evidence has indicated that history of diabetes mellitus (DM) is inversely associated with risk of PCa. However, epidemiological investigations have yielded inconsistent results. Hence, the exact mechanism of DM-induced reduction in the incidence of PCa has yet to be fully elucidated. The aim of this study was to investigate the effects of DM factors, including glucose, insulin and insulin-like growth factor-1 (IGF-1), on the proliferation of PCa cell lines in vitro. Cell proliferation and expression of hormone receptors was examined in MTT assay and Western blot analysis, respectively. The results showed that DM factors did not affect the viability of androgen receptor (AR)-expressing PCa cell lines. However, cell proliferation increased after treatment with DM factors in androgen-independent PCa cell lines. On PCa tissue arrays, intensities of total AR and nuclear IGF-1R were higher in malignant tissues than in normal prostate glands. In terms of hormonal receptors, androgen-dependent LNCaP cells treated with insulin and IGF-1 in a low-serum medium showed decreased expression of insulin receptor beta (IRβ) and elevated expression of IGF-1 receptor beta (IGF-1Rβ). Moreover, expression of AR was upregulated after insulin and IGF-1 treatment in LNCaP cells, but not in the other PCa cell lines. Most of the studied antidiabetic drugs promoted the viability of PCa cells. However, metformin decreased the viability of AR-expressing PCa cells. These results suggest that diabetic factors modify the expression of AR, IR and IGF-1R to increase cancer cell proliferation. Moreover, the growth suppressing effects of metformin on PCa may be via the regulation of the AR signaling pathway.

Zinc has insulin-mimetic properties which enhance spinal fusion in a rat model

Background ContextPrevious studies have found that insulin or insulin-like growth factor treatment can stimulate fracture healing in diabetic and normal animal models, and increase fusion rates in a rat spinal fusion model. Insulin-mimetic agents, such as zinc, have demonstrated antidiabetic effects in animal and human studies, and these agents that mimic the effects of insulin could produce the same beneficial effects on bone regeneration and spinal fusion. PurposeThe purpose of this study was to analyze the effects of locally applied zinc on spinal fusion in a rat model. Study Design/SettingInstitutional Animal Care and Use Committee-approved animal study using Sprague-Dawley rats was used as the study design. MethodsThirty Sprague-Dawley rats (450–500 g) underwent L4–L5 posterolateral lumbar fusion (PLF). After decortication and application of approximately 0.3 g of autograft per side, one of three pellets were added to each site: high-dose zinc calcium sulfate (ZnCaSO4), low-dose ZnCaSO4 (half of the high dose), or a control palmitic acid pellet (no Zn dose). Systemic blood glucose levels were measured 24 hours postoperatively. Rats were sacrificed after 8weeks and the PLFs analyzed qualitatively by manual palpation and radiograph review, and quantitatively by micro-computed tomography (CT) analysis of bone volume and trabecular thickness. Statistical analyses with p-values set at .05 were accomplished with analysis of variance, followed by posthoc tests for quantitative data, or Mann-Whitney rank tests for qualitative assessments. ResultsCompared with controls, the low-dose zinc group demonstrated a significantly higher manual palpation grade (p=.011), radiographic score (p=.045), and bone formation on micro-CT (172.9 mm3 vs. 126.7 mm3 for controls) (p<.01). The high-dose zinc also demonstrated a significantly higher radiographic score (p=.017) and bone formation on micro-CT (172.7 mm3 vs. 126.7 mm3) (p<.01) versus controls, and was trending toward higher manual palpation scores (p=.058). ConclusionsThis study demonstrates the potential benefit of a locally applied insulin-mimetic agent, such as zinc, in a rat lumbar fusion model. Previous studies have demonstrated the benefits of local insulin application in the same model, and it appears that zinc has similar effects.

Insulin-like growth factor-1 improves diabetic cardiomyopathy through antioxidative and anti-inflammatory processes along with modulation of Akt/GSK-3β signaling in rats.

Diabetic cardiomyopathy (DCM), a serious complication of diabetes mellitus, is associated with changes in myocardial structure and function. This study sought to explore the ability of insulin-like growth factor-1 (IGF-1) to modulate DCM and its related mechanisms. Twenty-four male Wistar rats were injected with streptozotocin (STZ, 60 mg/kg) to mimic diabetes mellitus. Myocardial fibrosis and apoptosis were evaluated by histopathologic analyses, and relevant proteins were analyzed by Western blotting. Inflammatory factors were assessed by ELISA. Markers of oxidative stress were tested by colorimetric analysis. Rats with DCM displayed decreased body weight, metabolic abnormalities, elevated apoptosis (as assessed by the bcl-2/bax ratio and TUNEL assays), increased fibrosis, increased markers of oxidative stress (MDA and SOD) and inflammatory factors (TNF-α and IL-1β), and decreased phosphorylation of Akt and glycogen synthase kinase (GSK-3β). IGF-1 treatment, however, attenuated the metabolic abnormalities and myocardial apoptosis, interstitial fibrosis, oxidative stress and inflammation seen in diabetic rats, while also increasing the phosphorylation levels of Akt and GSK-3β. These findings suggest that IGF-1 ameliorates the pathophysiological progress of DCM along with an activation of the Akt/GSK-3β signaling pathway. Our findings suggest that IGF-1 could be a potential therapeutic choice for controlling DCM.

Abstract A89: Istiratumab (MM-141), a bispecific antibody co-targeting IGF-1R and ErbB3, potentiates the activity of immune checkpoint inhibitors

Abstract Purpose: Regulatory T cells (Tregs) help maintain immunological tolerance to cancer cells by suppressing T cell effector function. The depletion of Tregs with the CTLA-4 inhibitor antibody ipilimumab has been proven to be a productive therapeutic strategy in melanoma. Additionally, levels of insulin-like growth factor 1 (IGF-1) are often elevated following chemotherapy treatment and have been associated with poor prognosis in multiple malignances. We demonstrate that IGF-1 can stimulate Treg proliferation. Istiratumab, a tetravalent bispecific antibody that targets both the IGF-1 receptor (IGF-1R) and ErbB3, inhibits IGF-1 signaling by blocking ligand binding and inducing rapid receptor internalization. Based on its mechanism of action, we evaluated the activity of istiratumab on Treg proliferation and in combination with immune checkpoint targeting antibodies in murine models of cancer. Experimental Procedures: Mouse-derived splenocytes were harvested to evaluate the effect of IGF-1 and/or istiratumab treatment on Treg expansion and proliferation in vitro. Flow cytometry studies assessed surface receptor expression of T cell populations isolated from murine splenocytes. Efficacy studies using immunocompetent mice bearing syngeneic murine tumors determined the activity of istiratumab and immune checkpoint targeting antibodies, alone and in combination, on tumor growth alongside isotype-matched controls. Mice whose tumors were eradicated by treatment were maintained and subsequently used in tumor re-challenge studies to assess the development of tumor-specific, immunological, long-term memory. In addition, pharmacodynamic studies analyzed the expression of tumor cell signaling pathway markers, effects on relevant T cell sub-populations and effects on post-treatment cytokine profiles. Data Summary: Our in vitro T cell analyses indicated that sub-populations of murine CD4+ CD25+ FoxP3+ Tregs express IGF-1R, and that istiratumab can reverse IGF-1 driven Treg proliferation in vitro. In addition, istiratumab monotherapy treatment had significant anti-tumor activity in vivo in immunogenic, syngeneic murine tumor models. Moreover, istiratumab potentiated the activity of immune checkpoint targeting antibodies in efficacy studies leading to curative outcomes in a subset of treated animals. These animals developed long-term immunological memory directed against the original tumor cell line. A pharmacodynamic analysis of tumor cell signaling- and immunology-related markers demonstrated that istiratumab impacts tumor cell survival signaling and contributes to overall immunological changes that favor breaking immunological tolerance in relevant syngeneic tumor models, thus enabling the development of an effective and durable anti-tumor immune response. Conclusions: Our in vitro and in vivo pre-clinical studies demonstrate that istiratumab (MM-141) inhibits both tumor cell survival and Treg proliferation in host mice, thereby potentiating the anti-tumor activity of clinically-relevant immune checkpoint inhibitor antibodies. Citation Format: Sharlene Adams, Michael D. Curley, Adam J. Camblin, Sergio Iadevaia, Lin Nie, Gege Tan, Chrystal U. Louis, Alexey A. Lugovskoy. Istiratumab (MM-141), a bispecific antibody co-targeting IGF-1R and ErbB3, potentiates the activity of immune checkpoint inhibitors. [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2015 Nov 5-9; Boston, MA. Philadelphia (PA): AACR; Mol Cancer Ther 2015;14(12 Suppl 2):Abstract nr A89.

D-4F Decreases White Matter Damage After Stroke in Mice.

Stroke-induced neuroinflammation and white matter damage are associated with neurological deficits. Whether D-4F, an apolipoprotein A-I mimetic peptide, treatment of stroke decreases neuroinflammation and white matter damage and improves functional outcome has not been investigated. Adult male C57BL/6 mice were subjected to permanent middle cerebral artery occlusion (MCAo) and were orally administered saline as a vehicle control and different doses of D-4F (2, 4, 8, 16, or 32 mg/kg) starting at 2 h after MCAo and daily until euthanized at 7 days after MCAo. D-4F treatment did not alter the blood levels of high-density lipoprotein, total cholesterol, triglyceride, blood-brain barrier leakage, and infarction volume compared with control group. D-4F (16 mg/kg) treatment of stroke significantly improved functional outcome, increased the white matter density and the number of oligodendrocyte progenitor cells in the ischemic boundary zone of the ipsilateral striatum, and increased myelin basic protein, insulin-like growth factor-1 (IGF1), but decreased inflammatory factor Toll-like receptor-4 and tumor necrosis factor-α expression in the ischemic brain 7 days after MCAo (P<0.05, n=11/group). The neurite/axonal outgrowth in primary cultured neurons was significantly increased when treated with D-4F (100 ng/mL) and IGF1 (100 ng/mL) compared with the nontreatment control. Inhibition of IGF1 significantly attenuated D-4F or IGF1 treatment-induced axonal outgrowth. D-4F-treatment did not increase oligodendrocyte-progenitor cell proliferation but decreased oligodendrocyte-progenitor cell death. D-4F treatment initiated 2 h after MCAo decreases neuroinflammation and white matter damage and improves functional outcome after stroke. D-4F-induced increase in IGF1 may contribute to D-4F-induced neurite/axonal outgrowth after stroke.

Insulin-like growth factor 1 (IGF-1) regulates prolactin, growth hormone, and IGF-1 receptor expression in the pituitary gland of the gilthead sea bream Sparus aurata.

The role of insulin-like growth factor 1 (IGF-1) on regulation of growth hormone (GH) and prolactin (PRL) as well as the possible involvement of IGF-1 receptor subtype a (IGF-1Ra) mRNA was assessed in juvenile specimens of Sparus aurata. IGF-1Ra was successfully cloned, and active receptor domains were localized in its mRNA precursor. Also, phylogenetic analysis of the protein sequence indicated a closer proximity to IGF-1Ra isoform found in zebrafish and other teleosts, than to the isoform IGF-1Rb. The most abundant presence of IGF-1Ra mRNA was detected in white muscle, whereas head kidney showed the lowest gene expression among 24 different studied tissues. Pituitaries of juvenile specimens of S. aurata were incubated in vitro with different doses of IGF-1 (0, 1, 100, and 1000 ng mL(-1)) during a period of 10 h. Total RNA with a high quality could be obtained from these pituitaries. PRL mRNA expression significantly increased with increasing IGF-1 doses. Similarly, IGF-1Ra mRNA increased its expression in response to IGF-1. However, GH mRNA levels decreased in a dose-dependent manner after IGF-1 treatment. The contradictory responses of GH and PRL expressions to IGF-1 in our experiment are possibly mediated by IGF-1Ra presence on the somatotrophs and prolactotrophs. The increase in IGF-1Ra mRNA levels may be related to the proper activation of the PI3-K/Akt signal transduction pathways which are normally involved in GH and PRL regulation.

Insulin-like growth factor 1 prevents diastolic and systolic dysfunction associated with cardiomyopathy and preserves adrenergic sensitivity.

Insulin-like growth factor 1 (IGF-1)-dependent signalling promotes exercise-induced physiological cardiac hypertrophy. However, the in vivo therapeutic potential of IGF-1 for heart disease is not well established. Here, we test the potential therapeutic benefits of IGF-1 on cardiac function using an in vivo model of chronic catecholamine-induced cardiomyopathy. Rats were perfused with isoproterenol via osmotic pump (1 mg kg(-1) per day) and treated with 2 mg kg(-1) IGF-1 (2 mg kg(-1) per day, 6 days a week) for 2 or 4 weeks. Echocardiography, ECG, and blood pressure were assessed. In vivo pressure-volume loop studies were conducted at 4 weeks. Heart sections were analysed for fibrosis and apoptosis, and relevant biochemical signalling cascades were assessed. After 4 weeks, diastolic function (EDPVR, EDP, tau, E/A ratio), systolic function (PRSW, ESPVR, dP/dtmax) and structural remodelling (LV chamber diameter, wall thickness) were all adversely affected in isoproterenol-treated rats. All these detrimental effects were attenuated in rats treated with Iso+IGF-1. Isoproterenol-dependent effects on BP were attenuated by IGF-1 treatment. Adrenergic sensitivity was blunted in isoproterenol-treated rats but was preserved by IGF-1 treatment. Immunoblots indicate that cardioprotective p110α signalling and activated Akt are selectively upregulated in Iso+IGF-1-treated hearts. Expression of iNOS was significantly increased in both the Iso and Iso+IGF-1 groups; however, tetrahydrobiopterin (BH4) levels were decreased in the Iso group and maintained by IGF-1 treatment. IGF-1 treatment attenuates diastolic and systolic dysfunction associated with chronic catecholamine-induced cardiomyopathy while preserving adrenergic sensitivity and promoting BH4 production. These data support the potential use of IGF-1 therapy for clinical applications for cardiomyopathies.

Dendritic Spine Instability in a Mouse Model of CDKL5 Disorder Is Rescued by Insulin-like Growth Factor 1

BackgroundCDKL5 (cyclin-dependent kinase-like 5) is mutated in many severe neurodevelopmental disorders, including atypical wRett syndrome. CDKL5 was shown to interact with synaptic proteins, but an in vivo analysis of the role of CDKL5 in dendritic spine dynamics and synaptic molecular organization is still lacking. MethodsIn vivo two-photon microscopy of the somatosensory cortex of Cdkl5−/y mice was applied to monitor structural dynamics of dendritic spines. Synaptic function and plasticity were measured using electrophysiological recordings of excitatory postsynaptic currents and long-term potentiation in brain slices and assessing the expression of synaptic postsynaptic density protein 95 (PSD-95). Finally, we studied the impact of insulin-like growth factor 1 (IGF-1) treatment on CDKL5 null mice to restore the synaptic deficits. ResultsAdult mutant mice showed a significant reduction in spine density and PSD-95-positive synaptic puncta, a reduction of persistent spines, and impaired long-term potentiation. In juvenile mutants, short-term spine elimination, but not formation, was dramatically increased. Exogenous administration of IGF-1 rescued defective rpS6 phosphorylation, spine density, and PSD-95 expression. Endogenous cortical IGF-1 levels were unaffected by CDKL5 deletion. ConclusionsThese data demonstrate that dendritic spine stabilization is strongly regulated by CDKL5. Moreover, our data suggest that IGF-1 treatment could be a promising candidate for clinical trials in CDKL5 patients.

Insulin-Like Growth Factor-1 Contributes to Mucosal Repair by β-Arrestin2–Mediated Extracellular Signal-Related Kinase Signaling in Experimental Colitis

Insulin-like growth factor-1 (IGF-1) possesses the ability to attenuate intestinal damage and promote mucosal repair of colitis. β-Arrestins, as the scaffolding proteins of G protein-coupled receptors or non-G protein-coupled receptors signaling, can be involved in IGF-1-mediated signaling pathways. However, the interaction of IGF-1 and β-arrestin2 in the mucosal repair of experimental colitis remains unexplored. Ulcerative colitis was induced in β-arrestin2 wild-type mice and β-arrestin2 knockout littermates by using 3% dextran sulfate sodium for 5 days, followed by regular water consumption for 1, 2, 3, and 4 weeks to analyze the mucosal repair from experimental colitis. Disease activity index and histologic score analyses were performed. Apoptosis and proliferation were assessed by terminal deoxynucleotidyl transferase-mediated dUTP nick-end labeling and Ki-67 staining, respectively. The expressions of β-arrestin2, phospho (p)-IGF-1R, and p-extracellular signal-regulated kinase (ERK)1/2 were examined. Furthermore, β-arrestin2 was overexpressed or altered in HCT116 cells by transfection before IGF-1 treatment in vitro. IGF-1 and β-arrestin2 expression was up-regulated in the repairing phase of experimental colitis. Targeted deletion of β-arrestin2 delayed the repair of colitis by inhibiting cell proliferation without affecting the levels of IGF-1 and p-IGF-1R. The β-arrestin2/ERK signaling pathway was involved in IGF-1-mediated mucosal repair through promoting epithelial cell and goblet cell regeneration from experimental colitis. These results indicate that IGF-1 contributes to the mucosal repair by β-arrestin2-mediated ERK signaling in experimental colitis.

Integrative Analysis of MicroRNA and mRNA Data Reveals an Orchestrated Function of MicroRNAs in Skeletal Myocyte Differentiation in Response to TNF-α or IGF1.

IntroductionSkeletal muscle cell differentiation is impaired by elevated levels of the inflammatory cytokine tumor necrosis factor-α (TNF-α) with pathological significance in chronic diseases or inherited muscle disorders. Insulin like growth factor-1 (IGF1) positively regulates muscle cell differentiation. Both, TNF-α and IGF1 affect gene and microRNA (miRNA) expression in this process. However, computational prediction of miRNA-mRNA relations is challenged by false positives and targets which might be irrelevant in the respective cellular transcriptome context. Thus, this study is focused on functional information about miRNA affected target transcripts by integrating miRNA and mRNA expression profiling data.Methodology/Principal FindingsMurine skeletal myocytes PMI28 were differentiated for 24 hours with concomitant TNF-α or IGF1 treatment. Both, mRNA and miRNA expression profiling was performed. The data-driven integration of target prediction and paired mRNA/miRNA expression profiling data revealed that i) the quantity of predicted miRNA-mRNA relations was reduced, ii) miRNA targets with a function in cell cycle and axon guidance were enriched, iii) differential regulation of anti-differentiation miR-155-5p and miR-29b-3p as well as pro-differentiation miR-335-3p, miR-335-5p, miR-322-3p, and miR-322-5p seemed to be of primary importance during skeletal myoblast differentiation compared to the other miRNAs, iv) the abundance of targets and affected biological processes was miRNA specific, and v) subsets of miRNAs may collectively regulate gene expression.ConclusionsJoint analysis of mRNA and miRNA profiling data increased the process-specificity and quality of predicted relations by statistically selecting miRNA-target interactions. Moreover, this study revealed miRNA-specific predominant biological implications in skeletal muscle cell differentiation and in response to TNF-α or IGF1 treatment. Furthermore, myoblast differentiation-associated miRNAs are suggested to collectively regulate gene clusters and targets associated with enriched specific gene ontology terms or pathways. Predicted miRNA functions of this study provide novel insights into defective regulation at the transcriptomic level during myocyte proliferation and differentiation due to inflammatory stimuli.

Astrocyte-elevated gene-1 mediates insulin-like growth factor 1-induced the progression of cardiac myxoma.

Recently, astrocyte-elevated gene-1 (AEG-1) and insulin-like growth factor 1 (IGF-1) have been involved in the regulation of multiple signaling pathways in tumorigenesis. To date, the detailed mechanisms underlying IGF-1-AEG-1 pathway-induced proliferation and apoptosis in cardiac myxoma (CM) was not reported. In the present study, we used immnohistochemistry, immunoblotting, and qRT-PCR to detect the expression profile of IGF-1 and AEG-1 in 90 CM tissues, and then cultured CM cells were subjected to si-AEG-1, in vitro, and in vivo assays. Our findings showed that IGF-1 and AEG-1 were obviously upregulated in CM tissues and markedly associated with tumor size. When CM cells were treated with si-AEG-1, si-AEG-1 attenuated IGF-1-induced CM cell growth and enhanced cell apoptosis. Mechanically, we validated the expression of AEG-1, p-Erk1/2, and p-Akt increased in CM cells in response to IGF-1 treatment in a time-dependent manner. However, si-AEG-1 affected the expression of these proteins. Functionally, we found the knockdown of AEG-1-inhibited G1/S transition and tumor formation of CM cells. In conclusion, AEG-1 regulates IGF-1-induced proliferation and apoptosis via Erk1/2 and Akt signaling in CM development, which suggests IGF-1-AEG-1 signaling could be recommended to be a useful target to exert anti-tumor effects on CM.

Effects of insulin-like growth factor-1 on the relaxation responses of the cavernous smooth muscle from aged rats

Objective. The aim of this study was determine whether intracavernosal injection (ICI) of insulin-like growth factor-1 (IGF-1) protein can improve corpus cavernosal smooth muscle relaxation in aging rats. Materials and methods. Ten young (4-month-old) and 30 old (24-month-old) Sprague–Dawley male rats were enrolled in the study. The old rats were divided into three groups: vehicle-only (n = 10), IGF-1 1 µg/kg (n = 10) and IGF-1 10 µg/kg treatment groups (n = 10). After 4 weeks of single IGF-1 injection treatment, strips of corporal tissue were precontracted with phenylephrine, and dose–response curves were generated to evaluate endothelial-dependent [acetylcholine (ACh)], endothelial-independent [sodium nitroprusside (SNP)] and electrical field stimulation (EFS) vasoreactivity. The changes in percentage of cavernosal smooth muscle and the concentration of nitric oxide (NO) in penile tissue were also evaluated. Results. After IGF-1 treatment, the vasoreactivity was significantly improved in both the 1 µg/kg and the 10 µg/kg treatment groups compared with the vehicle-only group at 4 weeks in response to ACh, SNP and EFS (all p < 0.05). The percentage of cavernosal smooth muscle was increased in the IGF-1 treatment groups. The NO concentrations were increased after IGF-1 treatment. Conclusions. These data demonstrate that ICI of IGF-1 can improve vasoreactivity via endothelium-dependent and endothelial-independent mechanisms in the corpus cavernosum of the aging rat.

Combined effect of insulin-like growth factor-1 and CC chemokine ligand 2 on angiogenic events in endothelial cells.

Therapeutic angiogenesis may be applied in medical conditions to promote stimulation of angiogenesis. Angiogenesis is a multistep process, which includes endothelial cell proliferation, migration, and tube formation, which is mediated by various angiogenic polypeptides. Thus, studies that elucidate the cellular mechanisms involved in these processes are necessary to develop novel therapeutic strategies. This study investigated the in vitro effects of the pro-angiogenic factors, insulin-like growth factor-1 (IGF-1) and/or chemokine (CC motif) ligand 2 (CCL2), on endothelial cells. Flow cytometry analysis showed that IGF-1 and CCL2 treatment did not interfere with IGF-1 receptor (IGF-1R) expression, but CCL2 treatment increased CCL2 receptor (CCR2) expression. Immunofluorescence analysis revealed that the IGF-1/CCL2 combination induced a greater increase in fibronectin deposition, but the treatments did not alter the expression of the fibronectin receptors, CD49e and CD44. The interaction of fibronectin with cytokines demonstrated that IGF-1/CCL2 promoted changes in intermediate F-actin remodeling that may result in increased endothelial cell adhesion and cell migration mediated by fibronectin. Furthermore, IGF-1/CCL2 stimulated endothelial cells, grown on fibronectin, to form capillary-like structures and intercellular lumina with greater luminal area. These data suggest that IGF-1/CCL2 combination and a fibronectin matrix may contribute to the angiogenesis process to stimulate adhesion, migration, and tube formation by endothelial cells as a result of F-actin remodeling.

Proliferation of cultured mouse choroid plexus epithelial cells.

The choroid plexus (ChP) epithelium is a multifunctional tissue found in the ventricles of the brain. The major function of the ChP epithelium is to produce cerebrospinal fluid (CSF) that bathes and nourishes the central nervous system (CNS). In addition to the CSF, ChP epithelial cells (CPECs) produce and secrete numerous neurotrophic factors that support brain homeostasis, such as adult hippocampal neurogenesis. Accordingly, damage and dysfunction to CPECs are thought to accelerate and intensify multiple disease phenotypes, and CPEC regeneration would represent a potential therapeutic approach for these diseases. However, previous reports suggest that CPECs rarely divide, although this has not been extensively studied in response to extrinsic factors. Utilizing a cell-cycle reporter mouse line and live cell imaging, we identified scratch injury and the growth factors insulin-like growth factor 1 (IGF-1) and epidermal growth factor (EGF) as extrinsic cues that promote increased CPEC expansion in vitro. Furthermore, we found that IGF-1 and EGF treatment enhances scratch injury-induced proliferation. Finally, we established whole tissue explant cultures and observed that IGF-1 and EGF promote CPEC division within the intact ChP epithelium. We conclude that although CPECs normally have a slow turnover rate, they expand in response to external stimuli such as injury and/or growth factors, which provides a potential avenue for enhancing ChP function after brain injury or neurodegeneration.