IGF-1 rs6218 polymorphisms modulate the susceptibility to age-related cataract.

Background: Single nucleotide polymorphisms (SNPs), as the most abundant form of DNA variation in the human genome, contribute to age-related cataracts (ARC) development. Apoptosis of lens epithelial cells (LECs) is closely related to ARC formation. Insulin-like growth factor 1 (IGF1) contributes to cell apoptosis regulation. Moreover, IGF1 was indicated to exhibit a close association with cataract formation. Afterward, an investigation was conducted to examine the correlation between polymorphisms in IGF1 and the susceptibility to ARC. Methods: The present investigation was a case-control study. Venous blood draws were collected from the participants for DNA genotyping. Lens capsule samples were collected to detect mRNA and apoptosis. TaqMan RT-PCR was used to detect IGF1 polymorphism genotypes and qRT PCR was used to detect IGF1 mRNA levels in LECs. LEC apoptosis was evaluated through flow cytometry. The chi-square test was used to compare differences between ARCs and controls of each SNP. Results: We found that the G allele frequency in the IGF1-rs6218 was higher in the ARCs than in the controls. Furthermore, it was observed that the rs6218 GG genotype exhibited a positive correlation to elevated levels of IGF1 mRNA in LECs. The IGF1 mRNA in the LECs and the apoptosis of LECs in nuclear type of ARCs (ARNC) was higher than the controls. Conclusion: The susceptibility to ARC was related to IGF1-rs6218 polymorphism, and this polymorphism is associated with IGF1 expression at the mRNA level. Moreover, apoptosis in LECs of ARNCs was found to be increased.

Exploring the Integrated Role of miRNAs and lncRNAs in Regulating the Transcriptional Response to Amino Acids and Insulin-like Growth Factor 1 in Gilthead Sea Bream (Sparus aurata) Myoblasts.

In this study, gilthead sea bream (Sparus aurata) fast muscle myoblasts were stimulated with two pro-growth treatments, amino acids (AA) and insulin-like growth factor 1 (Igf-1), to analyze the transcriptional response of mRNAs, microRNAs (miRNAs) and long non-coding RNAs (lncRNAs) and to explore their possible regulatory network using bioinformatic approaches. AA had a higher impact on transcription (1795 mRNAs changed) compared to Igf-1 (385 mRNAs changed). Both treatments stimulated the transcription of mRNAs related to muscle differentiation (GO:0042692) and sarcomere (GO:0030017), while AA strongly stimulated DNA replication and cell division (GO:0007049). Both pro-growth treatments altered the transcription of over 100 miRNAs, including muscle-specific miRNAs (myomiRs), such as miR-133a/b, miR-206, miR-499, miR-1, and miR-27a. Among 111 detected lncRNAs (>1 FPKM), only 30 were significantly changed by AA and 11 by Igf-1. Eight lncRNAs exhibited strong negative correlations with several mRNAs, suggesting a possible regulation, while 30 lncRNAs showed strong correlations and interactions with several miRNAs, suggesting a role as sponges. This work is the first step in the identification of the ncRNAs network controlling muscle development and growth in gilthead sea bream, pointing out potential regulatory mechanisms in response to pro-growth signals.

[Regulation of Aging Processes: A Perspective of Dietary Restriction Models].

The moderate restriction of dietary energy intake (dietary restriction: DR) extends the lifespan and health span of various laboratory animals, suggesting that it delays the aging process inherent in many animal species. Attenuated growth hormone and insulin-like growth factor-1 (IGF-1) signaling caused by mutations also increases the lifespan of mice, even those allowed to feed freely. In nematodes, the Daf16, mammalian Forkhead box O (FoxO) transcription factor, was shown to be required for lifespan extension in response to reduced IGF-1 signaling. Because DR also decreases the plasma concentration of IGF-1 in mammals, the IGF-1-FoxO axis may play a central role in the lifespan extension effect of DR and, thus, retardation of aging. Studies using knockout mice under DR conditions revealed the importance of FoxO1 and nuclear factor erythroid-derived 2-like 2 (Nrf2) in tumor suppression, and FoxO3 in lifespan extension. Human genomic studies also identified a strong association between a FOXO3 single nucleotide polymorphism and longevity. The aging mechanism is the most important risk factor for disease and frailty in aging humans. Therefore, further research on the application of DR to humans, the development of compounds and drugs that mimic the effects of DR, and mechanisms underlying FOXO3 polymorphisms for longevity is highly relevant to extending the human health span.

Locally Directed Recombinant Adeno- Associated Virus-Mediated IGF-1 Gene Therapy Enhances Osteochondral Repair and Counteracts Early Osteoarthritis In Vivo.

BACKGROUND: Restoration of osteochondral defects is critical, because osteoarthritis (OA) can arise. HYPOTHESIS: Overexpression of insulin-like growth factor 1 (IGF-1) via recombinant adeno-associated viral (rAAV) vectors (rAAV-IGF-1) would improve osteochondral repair and reduce parameters of early perifocal OA in sheep after 6 months in vivo. STUDY DESIGN: Controlled laboratory study. METHODS: Osteochondral defects were created in the femoral trochlea of adult sheep and treated with rAAV-IGF-1 or rAAV-lacZ (control) (24 defects in 6 knees per group). After 6 months in vivo, osteochondral repair and perifocal OA were assessed by well-established macroscopic, histological, and immunohistochemical scoring systems as well as biochemical and micro-computed tomography evaluations. RESULTS: Application of rAAV-IGF-1 led to prolonged (6 months) IGF-1 overexpression without adverse effects, maintaining a significantly superior overall cartilage repair, together with significantly improved defect filling, extracellular matrix staining, cellular morphology, and surface architecture compared with rAAV-lacZ. Expression of type II collagen significantly increased and that of type I collagen significantly decreased. Subchondral bone repair and tidemark formation were significantly improved, and subchondral bone plate thickness and subarticular spongiosa mineral density returned to normal. The OA parameters of perifocal structure, cell cloning, and matrix staining were significantly better preserved upon rAAV-IGF-1 compared with rAAV-lacZ. Novel mechanistic associations between parameters of osteochondral repair and OA were identified. CONCLUSION: Local rAAV-mediated IGF-1 overexpression enhanced osteochondral repair and ameliorated parameters of perifocal early OA. CLINICAL RELEVANCE: IGF-1 gene therapy may be beneficial in repair of focal osteochondral defects and prevention of perifocal OA.

Early nutritional programming in sterlet sturgeon (Acipenser ruthenus) with dietary soybean meal: Assessment of growth performance, body composition, and expression of GH, IGF-I, and Ghrelin genes.

This study was performed to assess the impacts of introducing diets containing different levels of soybean meal (SBM) to sterlet sturgeon (Acipenser ruthenus) larvae on growth performance, body composition, and molecular responses in the juvenile stage. The sterlet larvae (57.68 ± 0.66 mg) were weaned onto the formulated diets as follows: a control diet containing 60% fishmeal (FM), and three experimental diets with replacement levels of 15% (SBM15), 30% (SBM30), and 45% (SBM45) of FM with SBM. Then, a total of 260 fish (initial weight: 323.33 ± 11.76 mg) were fed the four different diets for 28 days in triplicates (phase 1, nutritional programming, NP). All treatments were then fed with the FM diet in phase 2 (common phase), and in phase 3 (challenge phase), all experimental groups (6.14 ± 0.08 g) were transitioned to SBM45 for 28 days. At the end of phases 1 and 2, growth performance showed no significant differences among the groups (P > 0.05), while significantly improved in SBM45 than the control at the end of phase 3 (P < 0.05). No significant differences were found among the groups in any phases for whole body composition (P > 0.05). Additionally, the total saturated fatty acids were significantly higher in SBM-based diets than FM at the end of phase 3 (P < 0.05). The mRNA of GH, IGF-I was significantly affected by variation of FM replacement level (P < 0.05). The expression level of Ghrelin was up-regulated in fish fed SBM at the end of phase 3 (P < 0.05). Our findings revealed that NP can positively enhance the adaptation of juvenile sterlet sturgeon to 45% SBM when exposed to the same diets at the larval stage. Further research is being carried out to provide valuable insights into the underlying mechanisms of digestive performance for this species.

Silencing of secreted phosphoprotein 1 attenuates sciatic nerve injury-induced neuropathic pain: Regulating extracellular signal-regulated kinase and neuroinflammatory signaling pathways.

BACKGROUND: Neuropathic pain (NP) is a chronic pathological pain that affects the quality of life and is a huge medical burden for affected patients. In this study, we aimed to explore the effects of secreted phosphoprotein 1 (SPP1) on NP. METHODS: We established a chronic constriction injury (CCI) rat model, knocked down SPP1 via an intrathecal injection, and/or activated the extracellular signal-regulated kinase (ERK) pathway with insulin-like growth factor 1 (IGF-1) treatment. Pain behaviors, including paw withdrawal threshold (PWT), paw withdrawal latency (PWL), lifting number, and frequency, were assessed. After sacrificing rats, the L4-L5 dorsal root ganglion was collected. Then, SPP1 levels were determined using quantitative polymerase chain reaction (qPCR) and western blot analysis. The levels of interleukin (IL)-1ß, tumor necrosis factor (TNF)-α, IL-6, IL-10, epidermal growth factor (EGF), vascular endothelial growth factor (VEGF), and transforming growth factor (TGF)-ß were determined using qPCR and enzyme-linked immunosorbent assay. The levels of ERK pathway factors were determined via western blot analysis. RESULTS: We found that CCI decreased PWT and PWL, increased the lifting number and frequency, and upregulated SPP1 levels. The loss of SPP1 reversed these CCI-induced effects. Additionally, CCI upregulated IL-1ß, TNF-α, IL-6, EGF, and VEGF levels, downregulated TGF-ß levels, and activated the ERK pathway, while silencing of SPP1 abrogated these CCI-induced effects. Moreover, IGF-1 treatment reversed the effects of SPP1 loss. CONCLUSIONS: The data indicate that silencing SPP1 attenuates NP via inactivation of the ERK pathway, suggesting that SPP1 may be a promising target for NP treatment.

GWAS-identified hyperuricemia-associated IGF1R variant rs6598541 has a limited role in urate mediated inflammation in human mononuclear cells.

Gout is a common autoinflammatory joint diseases characterized by deposition of monosodium urate (MSU) crystals which trigger an innate immune response mediated by inflammatory cytokines. IGF1R is one of the loci associated with both urate levels and gout susceptibility in GWAS to date, and IGF-1-IGF-1R signaling is implicated in urate control. We investigate the role of IGF-1/IGF1R signaling in the context of gouty inflammation. Also, we test the gout and urate-associated IGF1R rs6598541 polymorphism for association with the inflammatory capacity of mononuclear cells. For this, freshly isolated human peripheral blood mononuclear cells (PBMCs) were exposed to recombinant IGF-1 or anti-IGF1R neutralizing antibody in the presence or absence of solubilized urate, stimulated with LPS/MSU crystals. Also, the association of rs6598541 with IGF1R and protein expression and with ex vivo cytokine production levels after stimulation with gout specific stimuli was tested. Urate exposure was not associated with IGF1R expression in vitro or in vivo. Modulation of IGF1R did not alter urate-induced inflammation. Developing urate-induced trained immunity in vitro was not influenced in cells challenged with IGF-1 recombinant protein. Moreover, the IGF1R rs6598541 SNP was not associated with cytokine production. Our results indicate that urate-induced inflammatory priming is not regulated by IGF-1/IGF1R signaling in vitro. IGF1R rs6598541 status was not asociated with IGF1R expression or cytokine production in primary human PBMCs. This study suggests that the role of IGF1R in gout is tissue-specific and may be more relevant in the control of urate levels rather than in inflammatory signaling in gout.

Effect of Gonadotropin-Releasing Hormone Antagonist on the Expression Patterns of Insulin-Like Growth Factor 1, Androgen Receptor, and Luteinizing Hormone Receptor in an Experimental Rat Model of Functional Ovarian Cysts.

We studied the expression of insulin-like growth factor 1 (IGF-1), androgen receptor (AR) and luteinizing hormone receptor (LHR) in the ovaries under the conditions of the modeling and subsequent treatment of functional ovarian cysts with gonadotropin-releasing hormone antagonist (ant-GnRH). The intensity of IGF-1, LHR, and AR expression in the generative elements of rat ovaries changed under conditions of functional ovarian cysts simulation, as well as during treatment with ant-GnRH. In both experimental groups, the expression levels of the studied markers in preantral follicles and epithelial lining of cysts were found to be related to the number of growing follicles and cysts. A divergence of LHR and AR expression indices and a more pronounced decrease in the number of cystic cavities were observed in the group receiving ant-GnRH. These changes demonstrate a positive effect of ant-GnRH on intra-ovarian regulatory factors and a therapeutic effect in functional ovarian cysts.

Combination of blockade of endothelin signalling and compensation of IGF1 expression protects the retina from degeneration.

Retinitis pigmentosa (RP) and macular dystrophy (MD) cause severe retinal dysfunction, affecting 1 in 4000 people worldwide. This disease is currently assumed to be intractable, because effective therapeutic methods have not been established, regardless of genetic or sporadic traits. Here, we examined a RP mouse model in which the Prominin-1 (Prom1) gene was deficient and investigated the molecular events occurring at the outset of retinal dysfunction. We extracted the Prom1-deficient retina subjected to light exposure for a short time, conducted single-cell expression profiling, and compared the gene expression with and without stimuli. We identified the cells and genes whose expression levels change directly in response to light stimuli. Among the genes altered by light stimulation, Igf1 was decreased in rod photoreceptor cells and astrocytes under the light-stimulated condition. Consistently, the insulin-like growth factor (IGF) signal was weakened in light-stimulated photoreceptor cells. The recovery of Igf1 expression with the adeno-associated virus (AAV) prevented photoreceptor cell death, and its treatment in combination with the endothelin receptor antagonist led to the blockade of abnormal glial activation and the promotion of glycolysis, thereby resulting in the improvement of retinal functions, as assayed by electroretinography. We additionally demonstrated that the attenuation of mammalian/mechanistic target of rapamycin (mTOR), which mediates IGF signalling, leads to complications in maintaining retinal homeostasis. Together, we propose that combinatorial manipulation of distinct mechanisms is useful for the maintenance of the retinal condition.

The Lnc-ENST00000602558/IGF1 axis as a predictor of response to treatment with tripterygium glycosides in rheumatoid arthritis patients.

AIMS: Growing clinical evidence suggests that not all patients with rheumatoid arthritis (RA) benefit to the same extent by treatment with tripterygium glycoside (TG), which highlights the need to identify RA-related genes that can be used to predict drug responses. In addition, single genes as markers of RA are not sufficiently accurate for use as predictors. Therefore, there is a need to identify paired expression genes that can serve as biomarkers for predicting the therapeutic effects of TG tablets in RA. METHODS: A total of 17 pairs of co-expressed genes were identified as candidates for predicting an RA patient's response to TG therapy, and genes involved in the Lnc-ENST00000602558/GF1 axis were selected for that purpose. A partial-least-squares (PLS)-based model was constructed based on the expression levels of Lnc-ENST00000602558/IGF1 in peripheral blood. The model showed high efficiency for predicting an RA patient's response to TG tablets. RESULTS: Our data confirmed that genes co-expressed in the Lnc-ENST00000602558/IGF1 axis mediate the efficacy of TG in RA treatment, reduce tumor necrosis factor-α induced IGF1 expression, and decrease the inflammatory response of MH7a cells. CONCLUSION: We found that genes expressed in the Lnc-ENST00000602558/IGF1 axis may be useful for identifying RA patients who will not respond to TG treatment. Our findings provide a rationale for the individualized treatment of RA in clinical settings.

Tmem263 deletion disrupts the GH/IGF-1 axis and causes dwarfism and impairs skeletal acquisition.

Genome-wide association studies (GWAS) have identified a large number of candidate genes believed to affect longitudinal bone growth and bone mass. One of these candidate genes, TMEM263, encodes a poorly characterized plasma membrane protein. Single nucleotide polymorphisms in TMEM263 are associated with bone mineral density in humans and mutations are associated with dwarfism in chicken and severe skeletal dysplasia in at least one human fetus. Whether this genotype-phenotype relationship is causal, however, remains unclear. Here, we determine whether and how TMEM263 is required for postnatal growth. Deletion of the Tmem263 gene in mice causes severe postnatal growth failure, proportional dwarfism, and impaired skeletal acquisition. Mice lacking Tmem263 show no differences in body weight within the first 2 weeks of postnatal life. However, by P21 there is a dramatic growth deficit due to a disrupted growth hormone (GH)/insulin-like growth factor 1 (IGF-1) axis, which is critical for longitudinal bone growth. Tmem263-null mice have low circulating IGF-1 levels and pronounced reductions in bone mass and growth plate length. The low serum IGF-1 in Tmem263-null mice is associated with reduced hepatic GH receptor (GHR) expression and GH-induced JAK2/STAT5 signaling. A deficit in GH signaling dramatically alters GH-regulated genes and feminizes the liver transcriptome of Tmem263-null male mice, with their expression profile resembling wild-type female, hypophysectomized male, and Stat5b-null male mice. Collectively, our data validates the causal role for Tmem263 in regulating postnatal growth and raises the possibility that rare mutations or variants of TMEM263 may potentially cause GH insensitivity and impair linear growth.

A viral insulin-like peptide inhibits IGF-1 receptor phosphorylation and regulates IGF1R gene expression.

OBJECTIVE: The insulin/IGF superfamily is conserved across vertebrates and invertebrates. Our team has identified five viruses containing genes encoding viral insulin/IGF-1 like peptides (VILPs) closely resembling human insulin and IGF-1. This study aims to characterize the impact of Mandarin fish ranavirus (MFRV) and Lymphocystis disease virus-Sa (LCDV-Sa) VILPs on the insulin/IGF system for the first time. METHODS: We chemically synthesized single chain (sc, IGF-1 like) and double chain (dc, insulin like) forms of MFRV and LCDV-Sa VILPs. Using cell lines overexpressing either human insulin receptor isoform A (IR-A), isoform B (IR-B) or IGF-1 receptor (IGF1R), and AML12 murine hepatocytes, we characterized receptor binding, insulin/IGF signaling. We further characterized the VILPs' effects of proliferation and IGF1R and IR gene expression, and compared them to native ligands. Additionally, we performed insulin tolerance test in CB57BL/6 J mice to examine in vivo effects of VILPs on blood glucose levels. Finally, we employed cryo-electron microscopy (cryoEM) to analyze the structure of scMFRV-VILP in complex with the IGF1R ectodomain. RESULTS: VILPs can bind to human IR and IGF1R, stimulate receptor autophosphorylation and downstream signaling pathways. Notably, scMFRV-VILP exhibited a particularly strong affinity for IGF1R, with a mere 10-fold decrease compared to human IGF-1. At high concentrations, scMFRV-VILP selectively reduced IGF-1 stimulated IGF1R autophosphorylation and Erk phosphorylation (Ras/MAPK pathway), while leaving Akt phosphorylation (PI3K/Akt pathway) unaffected, indicating a potential biased inhibitory function. Prolonged exposure to MFRV-VILP led to a significant decrease in IGF1R gene expression in IGF1R overexpressing cells and AML12 hepatocytes. Furthermore, insulin tolerance test revealed scMFRV-VILP's sustained glucose-lowering effect compared to insulin and IGF-1. Finally, cryo-EM analysis revealed that scMFRV-VILP engages with IGF1R in a manner closely resembling IGF-1 binding, resulting in a highly analogous structure. CONCLUSIONS: This study introduces MFRV and LCDV-Sa VILPs as novel members of the insulin/IGF superfamily. Particularly, scMFRV-VILP exhibits a biased inhibitory effect on IGF1R signaling at high concentrations, selectively inhibiting IGF-1 stimulated IGF1R autophosphorylation and Erk phosphorylation, without affecting Akt phosphorylation. In addition, MFRV-VILP specifically regulates IGF-1R gene expression and IGF1R protein levels without affecting IR. CryoEM analysis confirms that scMFRV-VILP' binding to IGF1R is mirroring the interaction pattern observed with IGF-1. These findings offer valuable insights into IGF1R action and inhibition, suggesting potential applications in development of IGF1R specific inhibitors and advancing long-lasting insulins.

Identification of molecular mediators of renal sarcopenia risk: a mendelian randomization analysis.

BACKGROUND: Observational studies have shown an association between reduced renal function and the risk of sarcopenia. However, the causal relationship and the underlying biological mechanisms remain uncertain. Using a Mendelian randomization (MR) framework, we investigated the causal role of 27 hypothetical risk mediators, including metabolites, hormones, inflammation, and stress traits, on the risk of sarcopenia. METHODS: Instrumental variables (IVs) to proxy renal function were identified by selecting single nucleotide polymorphisms (SNPs) reliably associated with creatinine and cystatin C-based glomerular filtration rate (GFR) in CKDGen summary data. IVs for putative risk traits and sarcopenia traits were constructed from relevant genome-wide association studies (GWAS). MR estimated effects were obtained using an inverse-variance weighted effects model, and various sensitivity analyses were performed. The mediating role of hypothetical risk factors in the relationship between GFR and sarcopenia was assessed through multivariate MR. RESULTS: Genetically predicted reduced GFRcrea was associated with higher odds of appendicular lean mass (ALM) (odds ratio (OR): 0.64, 95% confidence interval (CI) 0.37 to 0.68) and grip strength (OR: 0.67; 95% CI 0.58 to 0.78). Likewise, GFRcys highlighted a causal relationship with ALM (OR: 0.52; 95% CI 0.42 to 0.65) and grip strength (OR: 0.66; 95% CI 0.59 to 0.74). Both estimated GFR (eGFR) were negatively associated with IGF-1, IL-16, 25(OH)D, triglycerides (range of effect size per standard deviation: -0.81 to -0.30), and positively correlated with HDL cholesterol (0.62, 0.31). There was a positive correlation between IGF-1, fasting insulin and ALM as well as grip strength (OR range: 1.04-1.67) and a negative correlation between serum CRP and ALM (OR: 0.95) as well as grip strength (OR: 0.98). Additionally, genetically predicted IL-1ß (OR: 0.95) and total cholesterol (OR: 0.96) were negatively associated with ALM. We found evidence that IGF-1 mediates the relationship between eGFR and risk for muscle mass and strength. CONCLUSIONS: This MR study provides insight into the potential causal mechanisms between renal function and the risk of sarcopenia and proposes IGF-1 as a potential target for the prevention of renal sarcopenia.

Erectile function and androgen and estrogen beta receptor gene polymorphisms in acromegalic men.

PURPOSE: Sexual dysfunctions are often experienced by male patients with acromegaly, due to a combination of hypogonadism and other comorbidities, but are a scarcely investigated complication. Erectile dysfunction is also closely related to cardiovascular diseases through endothelial dysfunction. Therefore, this project aimed to assess the prevalence of erectile dysfunction in a population of acromegalic men and evaluate its association with cardio-metabolic disorders, also exploring associations with androgen and estrogen receptor gene polymorphisms. METHODS: Sexually active men aged 18-65 with previous diagnosis of acromegaly were recruited. Clinical and laboratory data were retrospectively collected. Each patient also provided a blood sample for AR and ERß gene polymorphisms analyses and filled out the IIEF-15 questionnaire. RESULTS: Twenty men with previous diagnosis of acromegaly (mean age 48.4 ± 10.0 years) were recruited. 13/20 subjects (65%) had erectile dysfunction, but only four had a concurrent biochemical hypogonadism, with no significant correlation with IIEF-15 scores. Total testosterone negatively correlated with sexual intercourse satisfaction domain (ρ = - 0.595; p = 0.019) and general satisfaction domain (ρ = - 0.651; p = 0.009). IGF-1 levels negatively correlated with biochemical hypogonadism (ρ = - 0.585; p = 0.028). The number of CAG and CA repeats in AR and ERß receptors genes was not significantly associated with IIEF-15 scores or with GH/IGF-1 levels, but a negative correlation between CA repeats and the presence of cardiomyopathy (ρ = - 0.846; p = 0.002) was present. CONCLUSIONS: Men with acromegaly have a high prevalence of erectile dysfunction, but it does not appear to be correlated with treatments, testosterone levels and AR/ER-beta signaling. Nonetheless, a shorter CA polymorphic trait (ERbeta) is associated with the presence of cardiomyopathy. If confirmed, these data may suggest an association between an incorrect hormonal balance and increased cardiovascular risk in acromegaly subjects.

Food deprivation reduces sensitivity of liver Igf1 synthesis pathways to growth hormone in juvenile gopher rockfish (Sebastes carnatus).

Growth hormone (Gh) regulates growth in part by stimulating the liver to synthesize and release insulin-like growth factor-1 (Igf1), which then promotes somatic growth. However, for fish experiencing food limitation, elevated blood Gh can occur even with low circulating Igf1 and slow growth, suggesting that nutritional stress can alter the sensitivity of liver Igf1 synthesis pathways to Gh. Here, we examined how recent feeding experience affected Gh regulation of liver Igf1 synthesis pathways in juvenile gopher rockfish (Sebastes carnatus) to illuminate mechanisms underlying the nutritional modulation of Igf1 production. Juvenile gopher rockfish were maintained under conditions of feeding or complete food deprivation (fasting) for 14 d and then treated with recombinant sea bream (Sparus aurata) Gh or saline control. Gh upregulated hepatic igf1 mRNA levels in fed fish but not in fasted fish. The liver of fasted rockfish also showed a lower relative abundance of gene transcripts encoding teleost Gh receptors 1 (ghr1) and 2 (ghr2), as well as reduced protein levels of phosphorylated janus tyrosine kinase 2 (pJak2) and signal transducer and activator of transcription 5 (pStat5), which function to induce igf1 gene transcription following Gh binding to Gh receptors. Relative hepatic mRNA levels for suppressors of cytokine signaling (Socs) genes socs2, socs3a, and socs3b were also lower in fasted rockfish. Socs2 can suppress Gh activation of Jak2/Stat5, and fasting-related variation in socs expression may reflect modulated inhibitory control of igf1 gene transcription. Fasted rockfish also had elevated liver mRNA abundances for lipolytic hormone-sensitive lipase 1 (hsl1) and Igf binding proteins igfbp1a, -1b and -3a, reduced liver mRNAs encoding igfbp2b and an Igfbp acid labile subunit-like (igfals) gene, and higher transcript abundances for Igf1 receptors igf1ra and igf1rb in skeletal muscle. Together, these findings suggest that food deprivation impacts liver Igf1 responsiveness to Gh via multiple mechanisms that include a downregulation of hepatic Gh receptors, modulation of the intracellular Jak2/Stat5 transduction pathway, and possible shifts in Socs-inhibitory control of igf1 gene transcription, while also demonstrating that these changes occur in concert with shifts in liver Igfbp expression and muscle Gh/Igf1 signaling pathway components.

Anti-Gene IGF-I Vaccines in Cancer Gene Therapy: A Review of a Case of Glioblastoma.

OBJECTIVE: Vaccines for the deadliest brain tumor - glioblastoma (GBM) - are generally based on targeting growth factors or their receptors, often using antibodies. The vaccines described in the review were prepared to suppress the principal cancer growth factor - IGF-I, using anti-gene approaches either of antisense (AS) or of triple helix (TH) type. Our objective was to increase the median survival of patients treated with AS and TH cell vaccines. METHODOLOGY: The cells were transfected in vitro by both constructed IGF-I AS and IGF-I TH expression episomal vectors; part of these cells was co-cultured with plant phytochemicals, modulating IGF-I expression. Both AS and TH approaches completely suppressed IGF-I expression and induced MHC-1 / B7 immunogenicity related to the IGF-I receptor signal. RESULTS: This immunogenicity proved to be stronger in IGF-I TH than in IGF-I AS-prepared cell vaccines, especially in TH / phytochemical cells. The AS and TH vaccines generated an important TCD8+ and TCD8+CD11b- immune response in treated GBM patients and increased the median survival of patients up to 17-18 months, particularly using TH vaccines; in some cases, 2- and 3-year survival was reported. These clinical results were compared with those obtained in therapies targeting other growth factors. CONCLUSION: The anti-gene IGF-I vaccines continue to be applied in current GBM personalized medicine. Technical improvements in the preparation of AS and TH vaccines to increase MHC-1 and B7 immunogenicity have, in parallel, allowed to increase in the median survival of patients.

Detection rate of IGF-1 variants and their implication to protein binding: study of over 240,000 patients.

OBJECTIVES: To determine the detection rate of IGF-1 variants in a clinical population and assess their implications. METHODS: IGF-1 variants were detected based on their predicted mass-to-charge ratios. Most variants were distinguished by their isotopic distribution and relative retention times. A67T and A70T were distinguished with MS/MS. Patient specimens with a detected variant were de-identified for DNA sequencing to confirm the polymorphism. RESULTS: Of the 243,808 patients screened, 1,099 patients containing IGF-1 variants were identified (0.45 %, or 4,508 occurrences per million). Seven patients were identified as homozygous or double heterozygous. Majority of variants (98 %) had amino acid substitutions located at the C-terminus (A62T, P66A, A67S, A67V, A67T, A70T). Isobaric variants A38V and A67V were detected more frequently in children than in adults. Six previously unreported variants were identified: Y31H, S33P, T41I, R50Q, R56K, and A62T. Compared with the overall population, z-score distribution of patients with IGF-1 variants was shifted toward negative levels (median z-score -1.4); however, it resembled the overall population when corrected for heterozygosity. Chromatographic peak area of some variants differed from that of the WT IGF-1 present in the same patient. CONCLUSIONS: In the IGF-1 test reports by LC-MS, the concentrations only account for half the total IGF-1 for patients with heterozygous IGF-1 variants. An IGF-1 variant may change the binding to its receptor and/or its binding proteins, affecting its activity and half-life in circulation. Variants located in or close to the C-domain may be pathogenic. Cross-species sequence comparison indicates that A38V and A70T may have some degree of pathogenicity.

Cloning, phylogenetic and expression analysis of two MyoDs in yellowtail kingfish (Seriola lalandi).

Yellowtail kingfish (Seriola lalandi) is a pelagic piscivore distributed circumglobally. Owing to its great market value, the growth mechanism of S. lalandi, including muscle development and growth, is a hot research topic. The myoblast determination protein (MyoD) gene has been shown to play an important role in formation of myoblasts and the function of somites in fish. The open reading frame (ORF) sequences of MyoD1 and MyoD2 in S. lalandi encoded 298 and 263 amino acids possessing three common characteristic domains, respectively, containing a myogenic basic domain, a bHLH domain, and a ser-rich region (helix III). S. lalandi MyoDs shared the highest identity with the MyoDs of S. dumerili. MyoDs are highly expressed in white muscle (P < 0.05) in S. lalandi. The expression level of MyoD1 mRNA was higher than that of MyoD2 mRNA during embryonic and early developmental stages, indicating that the two MyoD isoforms may have different roles in muscle formation. Moreover, the mRNA expression of MyoDs in the brain, pituitary, liver and muscle of endocrine growth axis were analyzed in the various sizes and ages stages. The expression levels of MyoDs in the different sizes and ages of S. lalandi showed that expression of both these genes was particularly high in 400-g fish and 2-year-old fish (P < 0.05). Moreover, the increases in the mRNA expression and plasma levels of growth hormone (GH) and insulin-like growth factor (IGF-I) were accompanied by an increase in mRNA expression of MyoDs, indicating the roles of GH and IGF-I in muscle development and growth of S. lalandi. Overall, the expression profiles of genes associated with muscle development are the first step taken towards deciphering fast growth mechanism in this important Seriola fish.

Endothelial deficiency of insulin-like growth factor-1 receptor leads to blood-brain barrier disruption and accelerated endothelial senescence in mice, mimicking aspects of the brain aging phenotype.

INTRODUCTION: Age-related blood-brain barrier (BBB) disruption, cerebromicrovascular senescence, and microvascular rarefaction substantially contribute to the pathogenesis of vascular cognitive impairment (VCI) and Alzheimer's disease (AD). Previous studies established a causal link between age-related decline in circulating levels of insulin-like growth factor-1 (IGF-1), cerebromicrovascular dysfunction, and cognitive decline. The aim of our study was to determine the effect of IGF-1 signaling on senescence, BBB permeability, and vascular density in middle-age and old brains. METHODS: Accelerated endothelial senescence was assessed in senescence reporter mice (VE-Cadherin-CreERT2 /Igf1rfl/fl × p16-3MR) using flow cytometry. To determine the functional consequences of impaired IGF-1 input to cerebromicrovascular endothelial cells, BBB integrity and capillary density were studied in mice with endothelium-specific knockout of IGF1R (VE-Cadherin-CreERT2 /Igf1rfl/fl ) using intravital two-photon microscopy. RESULTS: In VE-Cadherin-CreERT2 /Igf1rfl/fl mice: (1) there was an increased presence of senescent endothelial cells; (2) cumulative permeability of the microvessels to fluorescent tracers of different molecular weights (0.3-40 kDa) is significantly increased, as compared to that of control mice, whereas decline in cortical capillary density does not reach statistical significance. CONCLUSIONS: These findings support the notion that IGF-1 signaling plays a crucial role in preserving a youthful cerebromicrovascular endothelial phenotype and maintaining the integrity of the BBB.

Establishment and Characterization of Amitrole-Induced Mouse Thyroid Adenomatous Nodule-Derived Cell Lines.

Background: Thyroid cancer cell lines have been of great value for the study of thyroid cancer. However, the availability of benign thyroid adenoma cell lines is limited. Methods: Cell lines were established from thyroid adenomatous nodules that developed in mice treated with the goitrogen amitrole. Expression of epithelial, mesenchymal, and thyroid markers of these established cell lines was determined, and the effect of lentivirus-transduced overexpression of NKX2-1, a master regulator of thyroid development, on the thyroid marker expression was examined. Signal transduction and cell proliferation were evaluated after treatment with insulin-like growth factor-I (IGF-I) and the selective IGF-I receptor (IGF-IR) inhibitor NVP-ADW742. Xenograft studies were performed to examine tumorigenicity of the cells in mice. Whole-genome sequencing (WGS) was used to comprehensively determine the genetic mutations in the established two cell lines. Results: Five mouse thyroid adenomatous nodules-derived cell lines named CAT (cells from amitrole-treated thyroids) were established. Among these, two cell lines, CAT458/458s (CAT458s: a subline of CAT458) and CAT459, were found to be positive for epithelial markers and negative for a mesenchymal marker. NKX2-1-positive CAT459 cells showed higher messenger RNA (mRNA) expression of some thyroid differentiation markers than NKX2-1-negative CAT458s cells, and NKX2-1 overexpression increased and/or induced their expression. IGF-I signaling was transduced in thyrotropin receptor (Tshr)-negative CAT458s and 459 cells, and NVP-ADW742 suppressed their proliferation. No tumors developed in mice after subcutaneous injection of CAT458s or 459 cells. The WGS analysis revealed the presence of missense mutations in the tumor suppressor genes such as Polk (encoding DNA polymerase kappa) and Tgfb1 (encoding transforming growth factor beta 1), while no mutations were found in the prominent thyroid cancer-related genes Braf, Trp53 (encoding p53), and Tert (encoding telomerase reverse transcriptase). Conclusions: Two mouse thyroid adenomatous nodule-derived cell lines with different thyroid differentiation marker expression were established. NKX2-1 induced partial differentiation of these cell lines. They lacked tumorigenicity and prominent gene mutations involved in thyroid cancer development, while missense mutations were found in some tumor suppressors as revealed by WGS. The CAT458s and 459 provide a new tool to further clarify the process of thyroid multistep carcinogenesis and differentiation.