The receptor tyrosine kinase IGF1R and its associated GPCRs are co-regulated by the noncoding RNA NEAT1 in Alzheimer's disease.

The study is based on the complexity of Insulin like growth factor receptor (IGF1R) signaling and its regulation by noncoding RNAs (ncRNAs). IGF1R signaling is an important cascade in Alzheimer's disease (AD); however, its regulation and roles are poorly understood. Due to the presence of ß-arrestin and GPCR Receptor Kinase binding sites, this protein has been termed a 'functional hybrid', as it can take part in both kinase and GPCR signaling pathways, further adding to its complexity. The objective of this study is to understand the underlying ncRNA regulation controlling IGF1R and GPCRs in AD to find commonalities in the network. We found through data mining that 45 GPCRs were reportedly deregulated in AD and built clusters based on GO/KEGG pathways to show shared functionality with IGF1R. Eight miRs were further discovered that could coregulate IGF1R and GPCRs. We validated their expression in an AD cell model and probed for common lncRNAs downstream that could regulate these miRs. Seven such candidates were identified and further validated. A combined network comprising IGF1R with nine GPCRs, eight miRs, and seven lncRNAs was created to visualize the interconnectivity within pathways. Betweenness centrality analysis showed a cluster of NEAT1, hsa-miR-15a-5p, hsa-miR-16-5p, and IGF1R to be crucial form a competitive endogenous RNA-based (ceRNA) tetrad that could relay information within the network, which was further validated by cell-based studies. NEAT1 emerged as a master regulator that could alter the levels of IGF1R and associated GPCRs. This combined bioinformatics and experimental study for the first time explored the regulation of IGF1R through ncRNAs from the perspective of neurodegeneration.

Insulin promotes growth in breast cancer cells through the type I IGF receptor in insulin receptor deficient cells.

Breast cancer is the most common cancer in women. The upregulation of insulin-like growth factor (IGF) system observed in certain types of breast cancers was linked to growth, metastasis, and survival resulting in multiple strategies designed to target the type I IGF receptor (IGF-1R) in breast cancer. These attempts failed to prove beneficial and it has been suggested that insulin receptor (IR) could also play an important role in breast cancer biology. To better understand the IR's role in breast cancer cells, the receptor was deleted from MCF-7L cells using CRISPR technology, and fluorescence-assisted cell sorting was used to obtain clone 35 (CL35). It was found that CL35 activated signaling pathways upon insulin stimulation despite the absence of IR expression. We hypothesized that CL35 used a surrogate receptor for sustained growth and development. IGF-1R was able to activate insulin signaling and growth in CL35. Thus, insulin may play a central role in regulating breast cancer growth due to its ability to activate all the receptors of the IGF family. These findings argue that dual targeting of IR and IGF-IR may be required to inhibit breast cancer growth.

Cell cycle control by the insulin-like growth factor signal: at the crossroad between cell growth and mitotic regulation.

In proliferating cells and tissues a number of checkpoints (G1/S and G2/M) preceding cell division (M-phase) require the signal provided by growth factors present in serum. IGFs (I and II) have been demonstrated to constitute key intrinsic components of the peptidic active fraction of mammalian serum. In vivo genetic ablation studies have shown that the cellular signal triggered by the IGFs through their cellular receptors represents a non-replaceable requirement for cell growth and cell cycle progression. Retroactive and current evaluation of published literature sheds light on the intracellular circuitry activated by these factors providing us with a better picture of the pleiotropic mechanistic actions by which IGFs regulate both cell size and mitogenesis under developmental growth as well as in malignant proliferation. The present work aims to summarize the cumulative knowledge learned from the IGF ligands/receptors and their intracellular signaling transducers towards control of cell size and cell-cycle with particular focus to their actionable circuits in human cancer. Furthermore, we bring novel perspectives on key functional discriminants of the IGF growth-mitogenic pathway allowing re-evaluation on some of its signal components based upon established evidences.

The CX3CL1 intracellular domain exhibits neuroprotection via insulin receptor/insulin-like growth factor receptor signaling.

CX3CL1, also known as fractalkine, is best known for its signaling activity through interactions with its cognate receptor CX3CR1. However, its intrinsic function that is independent of interaction with CX3CR1 remains to be fully understood. We demonstrate that the intracellular domain of CX3CL1 (CX3CL1-ICD), generated upon sequential cleavages by α-/ß-secretase and γ-secretase, initiates a back signaling activity, which mediates direct signal transmission to gene expression in the nucleus. To study this, we fused a synthetic peptide derived from CX3CL1-ICD, named Tet34, with a 13-amino acid tetanus sequence at the N terminus to facilitate translocation into neuronal cells. We show that treatment of mouse neuroblastoma Neuro-2A cells with Tet34, but not its scrambled control (Tet34s), induced cell proliferation, as manifested by changes in protein levels of transcription factors and progrowth molecules cyclin D1, PCNA, Sox5, and Cdk2. Further biochemical assays reveal elevation of phosphorylated insulin receptor ß subunit, insulin-like growth factor-1 receptor ß subunit, and insulin receptor substrates as well as activation of proliferation-linked kinase AKT. In addition, transgenic mice overexpressing membrane-anchored C-terminal CX3CL1 also exhibited activation of insulin/insulin-like growth factor-1 receptor signaling. Remarkably, we found that this Tet34 peptide, but not Tet34s, protected against endoplasmic reticulum stress and cellular apoptosis when Neuro-2A cells were challenged with toxic oligomers of ß-amyloid peptide or hydrogen peroxide. Taken together, our results suggest that CX3CL1-ICD may have translational potential for neuroprotection in Alzheimer's disease and for disorders resulting from insulin resistance.

Determinants of IGF-II influencing stability, receptor binding and activation.

Insulin like growth factor II (IGF-II) is involved in metabolic and mitogenic signalling in mammalian cells and plays important roles in normal fetal development and postnatal growth. It is structurally similar to insulin and binds not only with high affinity to the type 1 insulin-like growth factor receptor (IGF-1R) but also to the insulin receptor isoform A (IR-A). As IGF-II expression is commonly upregulated in cancer and its signalling promotes cancer cell survival, an antagonist that blocks IGF-II action without perturbing insulin signalling would be invaluable. The high degree of structural homology between the IR and IGF-1R makes selectively targeting either receptor in the treatment of IGF-II-dependent cancers very challenging. However, there are sequence differences between insulin and IGF-II that convey receptor selectivity and influence binding affinity and signalling outcome. Insulin residue YB16 is a key residue involved in maintaining insulin stability, dimer formation and IR binding. Mutation of this residue to glutamine (as found in IGF-II) results in reduced binding affinity. In this study we sought to determine if the equivalent residue Q18 in IGF-II plays a similar role. We show through site-directed mutagenesis of Q18 that this residue contributes to IGF-II structural integrity, selectivity of IGF-1R/IR binding, but surprisingly does not influence IR-A signalling activation. These findings provide insights into a unique IGF-II residue that can influence receptor binding specificity whilst having little influence on signalling outcome.

SBGN Bricks Ontology as a tool to describe recurring concepts in molecular networks.

A comprehensible representation of a molecular network is key to communicating and understanding scientific results in systems biology. The Systems Biology Graphical Notation (SBGN) has emerged as the main standard to represent such networks graphically. It has been implemented by different software tools, and is now largely used to communicate maps in scientific publications. However, learning the standard, and using it to build large maps, can be tedious. Moreover, SBGN maps are not grounded on a formal semantic layer and therefore do not enable formal analysis. Here, we introduce a new set of patterns representing recurring concepts encountered in molecular networks, called SBGN bricks. The bricks are structured in a new ontology, the Bricks Ontology (BKO), to define clear semantics for each of the biological concepts they represent. We show the usefulness of the bricks and BKO for both the template-based construction and the semantic annotation of molecular networks. The SBGN bricks and BKO can be freely explored and downloaded at sbgnbricks.org.

Identification and characteristics of insulin-like growth factor system in the brain, liver, and gonad during development of a seasonal breeding teleost, Pampus argenteus.

Reproductive activity is closely related to the development and function of the brain and liver in teleosts, particularly in seasonal breeding teleosts. This study measured the involvement of the insulin-like growth factor (IGF) system in controlling the reproduction of the silver pomfret Pampus argenteus, a seasonal breeding tropical to temperate commercial fish. We cloned and characterized the cDNAs of igfs (igf2 and igf3) and igfrs (igf1ra, igf1rb, and igf2r) and examined their transcript levels in relation to seasonal reproduction. Phylogenetic analyses revealed that two types of IGFs (IGF-1 and IGF-2) and three types of IGFRs (IGF1RA, IGF1RB, and IGF2R) of the silver pomfret were clustered with those of teleosts; however, IGF-3 was a transmembrane protein different with the IGF-3 of other teleosts. The expression of IGF-3 was gonad-specific in the silver pomfret. The transcript levels of igf1 in the female brain were the highest, and the levels of igfrs in both sexes' brains increased during gametogenesis. Meanwhile, igfs and igfrs maintained high transcript levels in both sexes' liver and gonad during vitellogenesis and spermatogonia proliferation. We concluded that the development and activities of brain, liver, and gonad were related to the IGF system (IGFs and IGFRs). And the IGFs were mainly expressed in the liver. Nevertheless, gonadal development, especially vitellogenesis and spermatogonia proliferation, were related with IGFs in this species.

Disorders of IGFs and IGF-1R signaling pathways.

The insulin-like growth factor (IGF) system comprises two ligands, IGF-I and IGF-II, that regulate multiple physiological processes, including mammalian development, metabolism and growth, through the type 1 IGF receptor (IGF-1R). The growth hormone (GH)-IGF-I axis is the major regulator of longitudinal growth. IGF-II is expressed in many tissues, notably the placenta, to regulate human pre- and post-natal growth and development. This review provides a brief introduction to the IGF system and summarizes findings from reports arising from recent larger genomic sequencing studies of human genetic mutations in IGF1 and IGF2 and genes of proteins regulating IGF action, namely the IGF-1R, IGF-1R signaling pathway components and the IGF binding proteins (IGFBPs). A perspective on the effect of homozygous mutations on structure and function of the IGFs and IGF-1R is also given and this is related to the effects on growth.

Short-Term Diet Induced Changes in the Central and Circulating IGF Systems Are Sex Specific.

Insulin-like growth factor (IGF) 1 exerts a wide range of functions in mammalians participating not only in the control of growth and metabolism, but also in other actions such as neuroprotection. Nutritional status modifies the IGF system, although little is known regarding how diet affects the newest members of this system including pregnancy-associated plasma protein-A (PAPP-A) and PAPP-A2, proteases that liberate IGF from the IGF-binding proteins (IGFBPs), and stanniocalcins (STCs) that inhibit PAPP-A and PAPP-A2 activity. Here we explored if a 1-week dietary change to either a high-fat diet (HFD) or a low-fat diet (LFD) modifies the central and peripheral IGF systems in both male and female Wistar rats. The circulating IGF system showed sex differences in most of its members at baseline. Males had higher levels of both free (p < 0.001) and total IGF1 (p < 0.001), as well as IGFBP3 (p < 0.001), IGFBP5 (p < 0.001), and insulin (p < 0.01). In contrast, females had higher serum levels of PAPP-A2 (p < 0.05) and IGFBP2 (p < 0.001). The responses to a short-term dietary change were both diet and sex specific. Circulating levels of IGF2 increased in response to LFD intake in females (p < 0.001) and decreased in response to HFD intake in males (p < 0.001). In females, LFD intake also decreased circulating IGFBP2 levels (p < 0.001). In the hypothalamus LFD intake increased IGF2 (p < 0.01) and IGFBP2 mRNA (p < 0.001) levels, as well as the expression of NPY (p < 0.001) and AgRP (p < 0.01), but only in males. In conclusion, short-term LFD intake induced more changes in the peripheral and central IGF system than did short-term HFD intake. Moreover, these changes were sex-specific, with IGF2 and IGFBP2 being more highly affected than the other members of the IGF system. One of the main differences between the commercial LFD employed and the HFD or normal rodent chow is that the LFD has a significantly higher sucrose content, suggesting that this nutrient could be involved in the observed responses.

Expression of the Insulin-like Growth Factor System in First- and Second-Trimester Human Embryonic and Fetal Gonads.

CONTEXT: Insulin-like growth factor (IGF) signaling is crucial for sex differentiation and development of Leydig and Sertoli cells in fetal mice testes. No such information is available for human embryonic and fetal testes and ovaries. OBJECTIVE: To investigate presence and activity of the IGF signaling system during human embryonic and fetal ovarian and testicular development. DESIGN: Human embryonic and fetal gonads were obtained following legal terminations of pregnancies. Gene expression was assessed by microarray and qPCR transcript analyses. Proteins of the IGF system components were detected with immunohistochemistry and immunofluorescence analyses. Specimens were included from 2010 to 2017. SETTING: University Hospital. PATIENTS/PARTICIPANTS: Ovaries and testes from a total of 124 human embryos and fetuses aged 5 to 17 postconception weeks were obtained from healthy women aged 16 to 47 years resident in Denmark or Scotland. MAIN OUTCOME MEASURES: Gene expression analysis using microarray was performed in 46 specimens and qPCR analysis in 56 specimens, both sexes included. Protein analysis included 22 specimens (11 ovaries, 11 testes). RESULTS: IGF system members were detected in embryonic and fetal testes and ovaries, both at gene transcript and protein level. A higher expression of IGF regulators was detected in testes than ovaries, with a preferred localization to Leydig cells. CONCLUSIONS: These data indicate that the IGF system is active during very early gestation, when it may have a regulatory role in Leydig cells.

Increasing knowledge in IGF1R defects: lessons from 35 new patients.

BACKGROUND: The type 1 insulin-like growth factor receptor (IGF1R) is a keystone of fetal growth regulation by mediating the effects of IGF-I and IGF-II. Recently, a cohort of patients carrying an IGF1R defect was described, from which a clinical score was established for diagnosis. We assessed this score in a large cohort of patients with identified IGF1R defects, as no external validation was available. Furthermore, we aimed to develop a functional test to allow the classification of variants of unknown significance (VUS) in vitro. METHODS: DNA was tested for either deletions or single nucleotide variant (SNV) and the phosphorylation of downstream pathways studied after stimulation with IGF-I by western blot analysis of fibroblast of nine patients. RESULTS: We detected 21 IGF1R defects in 35 patients, including 8 deletions and 10 heterozygous, 1 homozygous and 1 compound-heterozygous SNVs. The main clinical characteristics of these patients were being born small for gestational age (90.9%), short stature (88.2%) and microcephaly (74.1%). Feeding difficulties and varying degrees of developmental delay were highly prevalent (54.5%). There were no differences in phenotypes between patients with deletions and SNVs of IGF1R. Functional studies showed that the SNVs tested were associated with decreased AKT phosphorylation. CONCLUSION: We report eight new pathogenic variants of IGF1R and an original case with a homozygous SNV. We found the recently proposed clinical score to be accurate for the diagnosis of IGF1R defects with a sensitivity of 95.2%. We developed an efficient functional test to assess the pathogenicity of SNVs, which is useful, especially for VUS.

[Effect of GPX4 on proliferation and metastasis of renal clear cell carcinoma and its relationship with expression of IGF-1R and COX-2].

Objective: To investigate the effect of human glutathione peroxidase 4 (GPX4) on the proliferation and metastasis of renal clear cell carcinoma and its relationship with the expression of IGF-1R and COX-2. Methods: Culture of human normal tubular cell line HK-2 and human renal clear cell carcinoma Caki-1, A498, Caki-2, 786-o in vitro. Detection of GPX4 mRNA and protein expression in different cell lines by quantitative real-time PCR (RT-PCR) and Western blot assay. Overexpression of GPX4 cell lines, including blank carrier (Vector) and overexpress GPX4 (oeGPX4) group, and interference with GPX4 renal clear cell carcinoma cell lines, including random sequence (shControl), interference GPX4#1 (shGPX4#1) and interference GPX4#2 (shGPX4#2) group by lentiviral transfection. RT-PCR technology and Western blot were used to detect the expression of GPX4, IGF-1R and COX-2 mRNA and protein. CCK-8 assay was used to detect the relative proliferation of cells at 0, 24, 48, 72 and 96 h in each group. Transwell invasion and migration assay to detect the invasion and migration ability of cells of each group. Results: GPX4 is highly expressed in renal clear cell carcinoma cell lines compared to human normal tubular cell lines; The expression of GPX4, IGF-1R and COX-2 mRNA was significantly increased in oeGPX4 cells compared with Vector cells, the expression of GPX4,IGF-1R and COX-2 mRNA was significantly decreased in shGPX4#1 and shGPX4#2 compared with shControl cells; oeGPX4 cells significantly increased proliferative capacity compared to Vector cells at 72 and 96 h, the proliferation of shGPX4#1 and shGPX4#2 cells was significantly lower than that of shControl cells at 72 and 96 h; The number of invading and migrating cells of oeGPX4 cells was significantly higher than that of Vector cells, the number of invasive and migrating cells in shGPX4#1 and shGPX4#2 cells was significantly lower than that in shControl cells. Conclusion: GPX4 is highly expressed in renal clear cell carcinoma cells, which is positively correlated with the expression of IGF-1R and COX-2, and can promote cell proliferation and metastasis in vitro.

A specific type of insulin-like peptide regulates the conditional growth of a beetle weapon.

Evolutionarily conserved insulin/insulin-like growth factor (IGF) signaling (IIS) has been identified as a major physiological mechanism underlying the nutrient-dependent regulation of sexually selected weapon growth in animals. However, the molecular mechanisms that couple nutritional state with weapon growth remain largely unknown. Here, we show that one specific subtype of insulin-like peptide (ILP) responds to nutrient status and thereby regulates weapon size in the broad-horned flour beetle Gnatocerus cornutus. By using transcriptome information, we identified five G. cornutus ILP (GcorILP1-5) and two G. cornutus insulin-like receptor (GcorInR1, -2) genes in the G. cornutus genome. RNA interference (RNAi)-mediated gene silencing revealed that a certain subtype of ILP, GcorILP2, specifically regulated weapon size. Importantly, GcorILP2 was highly and specifically expressed in the fat body in a condition-dependent manner. We further found that GcorInR1 and GcorInR2 are functionally redundant but that the latter is partially specialized for regulating weapon growth. These results strongly suggest that GcorILP2 is an important component of the developmental mechanism that couples nutritional state to weapon growth in G. cornutus. We propose that the duplication and subsequent diversification of IIS genes played a pivotal role in the evolution of the complex growth regulation of secondary sexual traits.

A homozygous mutation in the highly conserved Tyr60 of the mature IGF1 peptide broadens the spectrum of IGF1 deficiency.

BACKGROUND: IGF1 is a key factor in fetal and postnatal growth. To date, only three homozygous IGF1 gene defects leading to complete or partial loss of IGF1 activity have been reported in three short patients born small for gestational age. We describe the fourth patient with severe short stature presenting a novel homozygous IGF1 gene mutation. RESULTS: We report a boy born from consanguineous parents at 40 weeks of gestational age with intrauterine growth restriction and severe postnatal growth failure. Physical examination revealed proportionate short stature, microcephaly, facial dysmorphism, bilateral sensorineural deafness and mild global developmental delay. Basal growth hormone (GH) fluctuated from 0.2 to 29 ng/mL, while IGF1 levels ranged from -1.15 to 2.95 SDS. IGFBP3 was normal-high. SNP array delimited chromosomal regions of homozygosity, including 12q23.2 where IGF1 is located. IGF1 screening by HRM revealed a homozygous missense variant NM_000618.4(IGF1):c.322T>C, p.(Tyr108His). The change of the highly conserved Tyr60 in the mature IGF1 peptide was consistently predicted as pathogenic by multiple bioinformatic tools. Tyr60 has been described to be critical for IGF1 interaction with type 1 IGF receptor (IGF1R). In vitro, HEK293T cells showed a marked reduction of IGF1R phosphorylation after stimulation with serum from the patient as compared to sera from age-matched controls. Mutant IGF1 was also less efficient in inducing cell growth. CONCLUSION: The present report broadens the spectrum of clinical and biochemical presentation of homozygous IGF1 defects and underscores the variability these patients may present depending on the IGF/IGF1R pathway activity.

Insulin-like growth factor axis in pregnancy and gestational diabetes mellitus.

The insulin-like growth factor (IGF) is involved in the regulation of growth and metabolism. The aim of this study was to determine selected parameters of IGF system at systemic and local levels [subcutaneous (SAT) and visceral adipose tissue (VAT)] to assess its possible role in gestational diabetes mellitus (GDM). 37 pregnant women (21 with GDM and 16 without GDM) and 15 age-matched non-pregnant females were included in the study. Blood samples were taken in 28-32 and 36-38 weeks of gestation and 6-12 months after delivery. SAT and VAT samples were obtained during delivery or surgery. Compared with non-pregnant women, serum IGF-1 and IGFBP-3 were increased in both groups of pregnant women. IGF-2 was elevated only in GDM women from 36 weeks of gestation culminating 6 months after delivery (p=0.003). Serum IGFBP-3 was increased and IGFBP-4 decreased in GDM women vs. pregnant women without GDM during the whole study (IGFBP-3: p?0.001 for GDM vs. non-GDM; IGFBP-4: p=0.004 for GDM vs. non-GDM). Pregnant women with GDM had decreased mRNA expression of IGF-1, IGF-1R and IGF-2R and IGFBP-4 in VAT and IGF-1R in SAT compared to pregnant women without GDM. Changes in local activity of IGF are associated with the development of GDM.

Myeloid Zinc Finger 1 (MZF1) Maintains the Mesenchymal Phenotype by Down-regulating IGF1R/p38 MAPK/ERα Signaling Pathway in High-level MZF1-expressing TNBC cells.

BACKGROUND/AIM: Signaling regulation of myeloid zinc finger 1 (MZF1) has been implicated in the progression of many human malignancies; however, the mechanistic action of MZF1 in triple-negative breast cancer (TNBC) progression remains elusive. In this study, the aim was to investigate the molecular mechanisms of MZF1 and its functional role in TNBC cellular migration and invasion. MATERIALS AND METHODS: Hs578T and MDA-MB-231 cells were transfected to stably express the acidic domain of MZF1 (MZF160-72), or were transfected with MZF1-specific or ELK1-specific short hairpin RNA (shRNA). Changes in cell morphology and distributions of cellular proteins were observed and subsequently migration and invasion were measured by wound healing and transwell assays. Expression levels of epithelial-mesenchymal transition (EMT)-related genes were carried out using immunoblotting and quantitative reverse transcription-polymerase chain reaction (RT-PCR) assays. Data of transcriptional regulation were obtained from promoter-luciferase reporter and chromatin immunoprecipitation (ChIP) assays. RESULTS: Herein, we found that MZF1 in high-level MZF1-expressing TNBC cells is associated with cell migration, invasion, and mesenchymal phenotype. MZF1 interacted with the promoter region of insulin-like growth factor 1 receptor (IGF1R) to drive invasion and metastasis of high-level MZF1-expressing TNBC cells. Exogenous expression of the acidic domain of MZF1 repressed the binding of endogenous MZF1 to IGF1R promoter via blocking the interaction with ETS-like gene 1 (ELK1). This blockage not only caused MZF1 protein degradation, but also restrained ELK1 nuclear localization in high-level MZF1-expressing TNBC cells. MZF1, but not ELK1, was necessary for the retention of mesenchymal phenotype by repressing IGF1R promoter activity in TNBC cells expressing high levels of MZF1. Activation of the IGF1R-driven p38MAPK-ERα-slug-E-cadherin signaling axis mediated the conversion of mesenchymal cell to epithelial phenotype, caused by MZF1 destabilization. These results suggest that MZF1 is an oncogenic inducer. CONCLUSION: Blocking of the MZF1/ELK1 interaction to reduce MZF1 protein stability by saturating the endogenous MZF1/ELK1 binding domains might be a promising therapeutic strategy for the treatment of high-level MZF1-expressing TNBC.

Insulin signaling mediates previtellogenic development and enhances juvenile hormone-mediated vitellogenesis in a lepidopteran insect, Maruca vitrata.

BACKGROUND: Insulin/insulin-like growth peptide signaling (IIS) down-regulates hemolymph sugar level and facilitates larval growth in the soybean pod borer, Maruca vitrata. The objective of this study is to determine whether IIS of M. vitrata can mediate ovarian development of adult females. RESULTS: A pair of ovaries consists of 8 ovarioles, each of which is separated into distal germarium and proximal vitellarium in M. vitrata. In the germarium, oocyte development occurred with active mitotic activity which was visible by incorporating bromodeoxyribose uridine. Previtellogenic development and subsequent vitellogenesis began soon after adult emergence. They continued with increase of female age. Oocyte development was facilitated by up-regulation of vitellogenin (Vg) and Vg receptor (VgR) gene expression. Larval diets significantly influenced on ovarian development of M. vitrata because oocyte development varied with pupal size derived from larvae treated with different nutritional diets. Its ovarian development was dependent on endocrine signal(s) from the head because decapitation soon after adult emergence prevented oogenesis and subsequent vitellogenesis along with marked reduction of Vg and VgR expression. Topical application of juvenile hormone (JH) significantly recovered its ovarian development whereas farnesoic acid (a precursor of JH biosynthesis) or 20-hydroxyecdysone treatment did not. JH stimulated vitellogenesis and choriogenesis, but not previtellogenic development. In contrast, insulin injection to decapitated females stimulated oocyte differentiation and vitellogenesis along with increase of Vg and VgR expression. To further analyze the effect of insulin on ovarian development, expression of four IIS components (InR, FOXO, Akt, and TOR) genes was manipulated by RNA interference. Hemocoelic injection of gene-specific double stranded RNAs significantly reduced their target gene mRNA levels and interfered with ovarian development. An addition of insulin to JH treatment against decapitated females enhanced the gonadotropic effect of JH by stimulating oogenesis. CONCLUSIONS: IIS plays crucial role in mediating previtellogenic development of M. vitrata in response to nutrient signal. It also enhances the gonadotropic effect of JH II on vitellogenesis.

IGF1R Is a Potential New Therapeutic Target for HGNET-BCOR Brain Tumor Patients.

(1) Background: The high-grade neuroepithelial tumor of the central nervous system with BCOR alteration (HGNET-BCOR) is a highly malignant tumor. Preclinical models and molecular targets are urgently required for this cancer. Previous data suggest a potential role of insulin-like growth factor (IGF) signaling in HGNET-BCOR. (2) Methods: The primary HGNET-BCOR cells PhKh1 were characterized by western blot, copy number variation, and methylation analysis and by electron microscopy. The expression of IGF2 and IGF1R was assessed by qRT-PCR. The effect of chemotherapeutics and IGF1R inhibitors on PhKh1 proliferation was tested. The phosphorylation of IGF1R and downstream molecules was assessed by western blot. (3) Results: Phkh1 cells showed a DNA methylation profile compatible with the DNA methylation class "HGNET-BCOR" and morphologic features of cellular cannibalism. IGF2 and IGF1R were highly expressed by three HGNET-BCOR tumor samples and PhKh1 cells. PhKh1 cells were particularly sensitive to vincristine, vinblastine, actinomycin D (IC50 < 10 nM for all drugs), and ceritinib (IC50 = 310 nM). Ceritinib was able to abrogate the proliferation of PhKh1 cells and blocked the phosphorylation of IGF1R and AKT. (4) Conclusion: IGF1R is as an attractive target for the development of new therapy protocols for HGNET-BCOR patients, which may include ceritinib and vinblastine.

Downregulation of lncRNA CCDC26 contributes to imatinib resistance in human gastrointestinal stromal tumors through IGF-1R upregulation.

Imatinib is the first line of therapy for patients with metastatic or gastrointestinal stromal tumors (GIST). However, drug resistance limits the long-term effect of imatinib. Long non-coding RNAs (lncRNAs) are emerging as key players in regulating drug resistance in cancer. In this study, we investigated the association between lncRNA CCDC26 and IGF-1R in GIST and their involvement in drug resistance. Considering the key role of lncRNAs in drug resistance in cancer, we hypothesized that IGF-1R is regulated by lncRNAs. The expression of a series of reported drug resistance-related lncRNAs, including CCDC26, ARF, H19, NBR2, NEAT1, and HOTAIR, in GIST cells treated with imatinib H19 was examined at various time-points by qRT-PCR. Based on our results and published literature, CCDC26, a strongly down-regulated lncRNA following imatinib treatment, was chosen as our research target. GIST cells with high expression of CCDC26 were sensitive to imatinib treatment while knockdown of CCDC26 significantly increased the resistance to imatinib. Furthermore, we found that CCDC26 interacted with c-KIT by RNA pull down, and that CCDC26 knockdown up-regulated the expression of IGF-1R. Moreover, IGF-1R inhibition reversed CCDC26 knockdown-mediated imatinib resistance in GIST. These results indicated that treatments targeting CCDC26-IGF-1R axis would be useful in increasing sensitivity to imatinib in GIST.

miR-140-5p could suppress tumor proliferation and progression by targeting TGFBRI/SMAD2/3 and IGF-1R/AKT signaling pathways in Wilms' tumor.

BACKGROUND: Wilms' tumor is also called nephroblastoma and is the most common pediatric renal cancer. Several genetic and epigenetic factors have been found to account for the development of Wilms' tumor. MiRNAs play important roles in this tumorigenic process. In the present study, we aimed to investigate the role of miR-140-5p in nephroblastoma by identifying its targets, as well as its underlying molecular mechanism of action. METHODS: The miRNA expression profile of nephroblastoma samples was investigated and the targets of miR-140-5p were predicted and validated using the miRNA luciferase reporter method. Moreover, the roles of miR-140-5p in regulating nephroblastoma cell proliferation, migration and cell cycle were analyzed by the CCK8, migration and flow cytometry assays, respectively. The downstream protein of the direct target of miR-140-5p was also identified. RESULTS: miR-140-5p was downregulated in Wilms' tumor tissues, whereas in the nephroblastoma cell lines G401 and WT-CLS1 that exhibited high levels of miRNA-140-5p, inhibition of cellular proliferation and metastasis were noted as well as cell cycle arrest at the G1/S phase. TGFBRI and IGF1R were identified as direct target genes for miRNA-140-5p. In addition, SMAD2/3 and p-AKT were regulated by TGFBRI and IGF1R separately and participated in the miRNA-140-5p regulatory network. Ectopic expression of TGFBR1 and IGF-1R could abrogate the inhibitory effect of miR-140-5p. CONCLUSION: We demonstrated that miRNA-140-5p participates in the progression of Wilms' tumor by targeting the TGFBRI/SMAD2/3 and the IGF-1R/AKT signaling pathways.