Growth Factor Binding Protein Research Articles

Response to Comment on Pellegrinelli et al. Human Adipocytes Induce Inflammation and Atrophy in Muscle Cells During Obesity. Diabetes 2015;64:3121–3134

We appreciate the comments by Moro and Bourlier (1) regarding our recent study (2), which shows the capacity of visceral fat–derived adipocytes from obese patients to promote inflammation and atrophy of muscle cells. Briefly, we identified interleukin (IL)-6 and IL-1β as drivers of the inflammatory burst of cocultured cells and a substantial amelioration of the atrophic phenotype by IGF-II/insulin growth factor binding protein-5. Given that our data do not support the direct role of IL-6 or IL-1β in promoting atrophy in muscle cells, we agree that further research is needed to …

Evaluation of insulin-like growth factor (IGF-1) and retinol binding protein (RBP-4) concentration levels in patients with newly diagnosed pancreatic adenocarcinoma

Evaluation of insulin-like growth factor (IGF-1) and retinol binding protein (RBP-4) concentration levels in patients with newly diagnosed pancreatic adenocarcinoma

Differential expression of insulin-like growth factor binding protein- 3 in the different grades of glioma and its correlation with malignancy of glial tumors

Objective To explore the differential expression of insulin- like growth factor- binding protein- 3(IGFBP- 3)in normal brain tissue and different grades of gliomas,and its corralation with the differentiation and malignance of gliomas. Methods By real- time quantitative polymerase chain reaction (Real- time PCR)and immunohistochemical staining methods, the expression of IGFBP- 3 was detected in 11 cases of grade-Ⅱglioma tissue samples, 14 cases of grade-Ⅲglioma tissue specimens, 9 cases of grade-Ⅳgrade glioma tissue samples,and 16 specimens of normal brain tissue. Results The average optical density values of IGFBP- 3 in normal brain tissue specimens and the specimens of grade-Ⅱ,-Ⅲ,-Ⅳgliomas were 1.082 2±0.334 7, 1.535 0±0.226 7, 4.383 1±0.521 8, and 5.094 1±0.652 5 respectively.Single factor analysis of variance(One- way ANOVA)and LSD, SNK- t analysis comparison showed there was significant difference in the expression of IGFBP- 3 between human gliomas and normal brain tissue(P< 0.05), and between grade-Ⅱand grade-Ⅲor grade-Ⅳgliomas(P< 0.05).By immunohistochemical staining methods,the positive expression rate of IGFBP- 3 was 20.0% in normal brain tissue specimens, which was significantly lower than in grade-Ⅱ(33.3%),-Ⅲ(57.1%), and-Ⅳ(80.0%)glimas(P<0.05 for all).There was no significant difference among different grades of gliomas. Conclusion The expression of IGFBP- 3 was significantly reduced in gliomas,and closely related with disease progression. Key words: Glioma; Insulin- like growth factor binding protein- 3; Promote apoptosis

Novel anti IGFBP2 single chain variable fragment inhibits glioma cell migration and invasion.

Insulin like growth factor binding protein 2 (IGFBP2) is highly up regulated in glioblastoma (GBM) tissues and has been one of the prognostic indicators. There are compelling evidences suggesting important roles for IGFBP2 in glioma cell proliferation, migration and invasion. Extracellular IGFBP2 through its carboxy terminal arginine glycine aspartate (RGD) motif can bind to cell surface α5β1 integrins and activate pathways downstream to integrin signaling. This IGFBP2 activated integrin signaling is known to play a crucial role in IGFBP2 mediated invasion of glioma cells. Hence a molecular inhibitor of carboxy terminal domain of IGFBP2 which can inhibit IGFBP2-cell surface interaction is of great therapeutic importance. In an attempt to develop molecular inhibitors of IGFBP2, we screened single chain variable fragment (scFv) phage display libraries, Tomlinson I (Library size 1.47 × 10(8)) and Tomlinson J (Library size 1.37 × 10(8)) using human recombinant IGFBP2. After screening we obtained three IGFBP2 specific binders out of which one scFv B7J showed better binding to IGFBP2 at its carboxy terminal domain, blocked IGFBP2-cell surface association, reduced activity of matrix metalloprotease 2 in the conditioned medium of glioma cells and inhibited IGFBP2 induced migration and invasion of glioma cells. We demonstrate for the first time that in vitro inhibition of extracellular IGFBP2 activity by using human scFv results in significant reduction of glioma cell migration and invasion. Therefore, the inhibition of IGFBP2 can serve as a potential therapeutic strategy in the management of GBM.

Higher protein kinase C ζ in fatty rat liver and its effect on insulin actions in primary hepatocytes.

We previously showed the impairment of insulin-regulated gene expression in the primary hepatocytes from Zucker fatty (ZF) rats, and its association with alterations of hepatic glucose and lipid metabolism. However, the molecular mechanism is unknown. A preliminary experiment shows that the expression level of protein kinase C ζ (PKCζ), a member of atypical PKC family, is higher in the liver and hepatocytes of ZF rats than that of Zucker lean (ZL) rats. Herein, we intend to investigate the roles of atypical protein kinase C in the regulation of hepatic gene expression. The insulin-regulated hepatic gene expression was evaluated in ZL primary hepatocytes treated with atypical PKC recombinant adenoviruses. Recombinant adenovirus-mediated overexpression of PKCζ, or the other atypical PKC member PKCι/λ, alters the basal and impairs the insulin-regulated expressions of glucokinase, sterol regulatory element-binding protein 1c, the cytosolic form of phosphoenolpyruvate carboxykinase, the catalytic subunit of glucose 6-phosphatase, and insulin like growth factor-binding protein 1 in ZL primary hepatocytes. PKCζ or PKCι/λ overexpression also reduces the protein level of insulin receptor substrate 1, and the insulin-induced phosphorylation of AKT at Ser473 and Thr308. Additionally, PKCι/λ overexpression impairs the insulin-induced Prckz expression, indicating the crosstalk between PKCζ and PKCι/λ. We conclude that the PKCζ expression is elevated in hepatocytes of insulin resistant ZF rats. Overexpressions of aPKCs in primary hepatocytes impair insulin signal transduction, and in turn, the down-stream insulin-regulated gene expression. These data suggest that elevation of aPKC expression may contribute to the hepatic insulin resistance at gene expression level.

Human Adipocytes Induce Inflammation and Atrophy in Muscle Cells During Obesity.

Inflammation and lipid accumulation are hallmarks of muscular pathologies resulting from metabolic diseases such as obesity and type 2 diabetes. During obesity, the hypertrophy of visceral adipose tissue (VAT) contributes to muscle dysfunction, particularly through the dysregulated production of adipokines. We have investigated the cross talk between human adipocytes and skeletal muscle cells to identify mechanisms linking adiposity and muscular dysfunctions. First, we demonstrated that the secretome of obese adipocytes decreased the expression of contractile proteins in myotubes, consequently inducing atrophy. Using a three-dimensional coculture of human myotubes and VAT adipocytes, we showed the decreased expression of genes corresponding to skeletal muscle contractility complex and myogenesis. We demonstrated an increased secretion by cocultured cells of cytokines and chemokines with interleukin (IL)-6 and IL-1β as key contributors. Moreover, we gathered evidence showing that obese subcutaneous adipocytes were less potent than VAT adipocytes in inducing these myotube dysfunctions. Interestingly, the atrophy induced by visceral adipocytes was corrected by IGF-II/insulin growth factor binding protein-5. Finally, we observed that the skeletal muscle of obese mice displayed decreased expression of muscular markers in correlation with VAT hypertrophy and abnormal distribution of the muscle fiber size. In summary, we show the negative impact of obese adipocytes on muscle phenotype, which could contribute to muscle wasting associated with metabolic disorders.

Insulin like growth factor binding protein 7 (IGFBP7) expression is linked to poor prognosis but may protect from bone disease in multiple myeloma.

BackgroundInsulin like growth factor binding protein 7 (IGFBP7) is a secreted protein binding insulin like growth factor 1 (IGF-1), insulin, vascular endothelial growth factor A (VEGFA), and activin A. It antagonizes bone morphogenetic proteins and is involved in the tumour propagation of solid as well as haematological malignancies. Its role in multiple myeloma (MM) is not defined so far. We therefore aim here to investigate its prognostic and pathophysiological role in MM.MethodsThe clinical significance of IGFBP7 gene expression was investigated by gene expression profiling in two independent cohorts (n = 948) of newly-diagnosed MM patients. Methylation of the IGFBP7 promoter was analysed by pyrosequencing and treatment of MM cell lines with 5-aza-2-deoxycytidine. The impact of IGFBP7 on MM cells was studied by CCK-8 assay, BrdU assay and flow cytometry, respectively. IGFBP7 expression in bone marrow stromal cells (BMSCs) was studied by quantitative RT-PCR. For osteoblast development, immortalized and primary human BMSCs were cultured in osteogenic differentiation medium for 7–14 days in the presence of recombinant human IGFBP7 and/or activin A.ResultsMedian IGFBP7 expression is significantly lower in CD138-purified plasma cells from individuals with MGUS and MM, compared to normal bone marrow plasma cells. IGFBP7 gene expression in MM cells is regulated by methylation, shown by pyrosequencing and exposure to demethylating agents (5-aza-2-deoxycytidine). High expression of IGFBP7 in MM cells is associated with adverse survival in two independent cohorts of 247 and 701 newly-diagnosed MM patients treated with high-dose therapy and autologous stem cell transplantation. IGFBP7 is associated with prognostically adverse chromosomal aberrations (t(4;14) and gain of 1q21), MMSET expression, and higher myeloma cell proliferation. In vitro, IGFBP7 overcomes activin A induced osteoblast suppression and promotes osteogenesis. MM cells downregulate IGFBP7 in stromal cells, possibly contributing to the osteoblast suppression found in MM. Conversely, higher IGFBP7 expression is associated with a lower probability of myeloma bone disease.ConclusionsOur data indicate that IGFBP7 expression is a marker for a specific methylation pattern in myeloma, linked to translocation t(4;14) associated MMSET expression, showing clinical features of adverse prognosis with absence of myeloma bone disease.Electronic supplementary materialThe online version of this article (doi:10.1186/s13045-014-0105-1) contains supplementary material, which is available to authorized users.

Using induced pluripotent stem cell-derived conditional medium to attenuate the light-induced photodamaged retina of rats

BackgroundLight injury to photoreceptor cells and retinal pigment epithelium may lead to oxidative stress and irreversible degeneration of retina, especially degeneration of the high energy-demanded macula. The model of retinal photodamage could be applied to age-related macular degeneration and other degenerative retinal diseases for exploring new treatments. Based on broadly investigated induced pluripotent stem cells (iPSC) in the field of retinal degeneration, we aimed to clarify further how the interaction progresses between iPSC-conditional medium (CM) and light-damaged retina. MethodsiPSCs were generated from murine embryonic fibroblasts of C57/B6 mice by retroviral transfection of three factors: Oct4, Sox2, and Klf4. Cytokine array was performed to analyze the components of CM. Sprague–Dawley rats receiving white light exposure to retina were viewed as an animal model of light injury. The rats were divided into four subgroups: light-injured rats receiving intravitreal injection of iPSC-CM, apoptotic iPSC-CM, or sodium phosphate buffer (PBS); and a control group without light damage. The electroretinography and thickness of outer nuclear layer were measured to document the therapeutic effects in each condition. Apoptosis arrays for detecting annexin V and caspase 3 were performed in the retinal tissues from each group. ResultsMurine embryonic fibroblasts were induced into iPSCs and expressed the marker genes similar to embryonic stem cells. These iPSCs can differentiate into Embryoid bodies (EBs), three germ layers in vitro and develop teratoma in severe combined immunodeficiency mice. The quantitative polymerase chain reaction of our iPSC-CM showed significantly elevated fibroblast growth factor-2, glial cell-derived neurotrophic factor, and insulin-like growth factor-binding proteins-1, -2, and -3. Compared to rats without photodamage, the light-injured rats receiving iPSC-CM had less reduction of outer nuclear layer thickness on Day 21 than other groups treated with either PBS or apoptotic iPSC-CM. In the same animal model, both a- and b-waves of electroretinography measurement in the group treated with iPSC-CM were significantly maintained compared to the control group and others with apoptotic iPSC-CM or PBS treatment. The apoptosis assay also demonstrated lower levels of annexin V and caspase 3 in the group with iPSC-CM treatment than in other groups presenting increasing apoptotic markers. ConclusionThe conditional medium of iPSCs contains plenty of cytoprotective, immune-modulative and rescue chemicals, contributing to the maintenance of neuronal function and retinal layers in light-damaged retina compared with apoptotic iPSC-CM and PBS. The antiapoptotic effect of iPSC-CM also shows promise in restoring damaged neurons. This result demonstrates that iPSC-CM may serve as an alternative to cell therapy alone to treat retinal light damage and maintain functional and structural integrity of the retina.

Proteomics analysis reveals IGFBP2 as a candidate diagnostic biomarker for heart failure

BackgroundDiagnostic biomarkers for heart failure (HF) such as the natriuretic peptides (NPs) are widely used but have limitations. Innovative biomarkers could provide improved diagnostic performance. MethodsWe launched a prospective case–control proteomic study and investigated for polypeptides specific to HF through a capillary electrophoresis-mass spectrometry (CE-MS) proteomic analysis. The putative biomarker was identified by Orbitrap liquid chromatography-MS, validated by western blot, then by ELISA using plasmas from multicentric international cohorts. A rat model of HF was tested for biomarker expression levels. ResultsWe identified insulin like growth factor binding protein 2 (IGFBP2) as a new diagnostic biomarker for HF with a high sensitivity and specificity (AUC=0.93; 95% CI, 0.89-0.96; p<0.0001) in the local cohort and IGFBP2 levels provided an AUC of 0.943 (95% CI, 0.860-1.026) which gave a 87 % sensitivity in AHF and 90 % specificity at the cut off value previously determined in the discovery cohort, i.e. 556ng/ml. ROC curve analysis of IGFBP2 and NTproBNP showed an AUC of 0.784 (95% CI, 0.744-0.820) for IGFBP2 and a significantly higher AUC of 0.927 (95% CI, 0.900-0.949) for NT-proBNP, p<0.0001 in a Dutch cohort. In this cohort, the optimal cut off value for IGFBP2 gave a sensibility of 71% (95% CI, 66–76) and a specificity of 75% (95% CI, 65–83). ConclusionIGFBP2 is a new biomarker to diagnose HF which could be used to provide additional information to the NPs. Animals models will help in the evaluation of the putative IGFBP2 regulated mechanisms in HF.

Insulin-like growth factor-1 binding protein 3 (IGFBP-3) promotes recovery from trauma-induced expression of inflammatory and apoptotic factors in retina

Ocular trauma affects 20% of Americans in their lifetime and can cause permanent visual system damage. We have used a mouse model of ocular trauma (exposure to an air blast from a paintball gun) to examine pathways that trigger the resulting retinal damage and to develop treatment strategies that might ameliorate the deleterious effects of trauma on retinal tissue. Our previous studies have shown that ocular blast causes an increase in protein levels of inflammatory mediators and apoptotic factors, including tumor necrosis factor alpha (TNFα) and interleukin-1-beta (IL-1β), as well as the apoptotic markers, Bax, cytochrome C, and cleaved caspase 3. Furthermore, topical treatment by eye drop application of a β-adrenergic receptor agonist, Compound 49b, was shown to decrease these inflammation/apoptosis markers and thus ameliorate the effects of blast trauma. We postulate that the protective effect of Compound 49b may be linked to its demonstrated ability to activate the β-adrenergic receptor and in turn trigger production of insulin-like growth factor binding protein 3 (IGFBP-3). In the current study, we tested this hypothesis using mice with minimal IGFBP-3 activity (IGFBP-3 knockdown mouse) vs. wildtype mice. We found that ocular blast alone did not affect IGFBP-3 levels in retinas of wild type or knockdown mice and surprisingly, the lower levels of IGFBP-3 in knockdown animals did not exacerbate the blast-induced increase in protein levels of inflammation/apoptosis markers. Nevertheless, the levels of IGFBP-3 were significantly increased in knockdown mouse retina by treatment with Compound 49b 24h post-trauma and as expected, the increase in IGFBP-3 was linked to a decrease in inflammation/apoptosis markers. We conclude that while lowered IGFBP-3 may not make the retina more vulnerable to blast injury, an increase in IGFBP-3 post-trauma may play an important role in limiting trauma-induced inflammatory and apoptotic pathways leading to retinal damage. Eye drop application of the β-adrenergic receptor agonist, Compound 49b, provides a promising treatment strategy for increasing IGFBP-3 levels to promote recovery from retinal inflammation and apoptosis after ocular blast.

IGF-1 predicts survival in chronic heart failure. Insights from the T.O.S.CA. (Trattamento Ormonale Nello Scompenso CArdiaco) registry

Recent evidence suggests that anabolic deficiencies play a pivotal role in chronic heart failure (CHF) progression [ [1] Jankowska E.A. Biel B. Majda J. et al. Anabolic deficiency in men with chronic heart failure: prevalence and detrimental impact on survival. Circulation. 2006; 114: 1829-1837 Crossref PubMed Scopus (320) Google Scholar ]. However, studies dwelling upon their prognostic value are few and, with particular regard to the Growth Hormone/Insulin-like Growth Factor-1 (GH/IGF-1) axis, controversial [ 1 Jankowska E.A. Biel B. Majda J. et al. Anabolic deficiency in men with chronic heart failure: prevalence and detrimental impact on survival. Circulation. 2006; 114: 1829-1837 Crossref PubMed Scopus (320) Google Scholar , 2 Watanabe S. Tamura T. Ono K. et al. Insulin-like growth factor axis (insulin-like growth factor-I/insulin-like growth factor-binding protein-3) as a prognostic predictor of heart failure: association with adiponectin. Eur J Heart Fail. 2010; 12: 1214-1222 Crossref PubMed Scopus (57) Google Scholar ]. It is well established that GH and IGF-1 are essential regulators of myocardial architecture and function in normal and diseased states [ [3] Sacca L. Cittadini A. Fazio S. Growth hormone and the heart. Endocr Rev. 1994; 15: 555-573 Crossref PubMed Scopus (493) Google Scholar ]. To address the predictive role of IGF-1 in CHF, we prospectively studied a population of 207 consecutive patients with CHF NYHA classes I–III. Patients with LV ejection fraction of 40% or below were recruited in the outpatient clinic after at least a 3-month period of clinical stability. Unstable patients and those with significant comorbidity (active cancer, liver cirrhosis Child C, and end-stage renal failure) were excluded. After obtaining an informed consent (ClinicalTrials.gov NCT00511927), circulating levels of IGF-1 and serum IGF Binding Protein-3 (IGFBP-3) were determined at baseline. All patients also underwent a physical examination as well as an echocardiographic study and a Cardiopulmonary Exercise Test (CPET). Patients were regularly followed-up in the outpatient clinic for a median period of 3.5 years (range 1–72 months).

1α,25(OH)₂D₃ inhibits FGF-2 release from oral squamous cell carcinoma cells through down-regulation of HBp17/FGFBP-1.

Heparin-binding protein 17/fibroblast growth factor binding protein-1 (HBp17/FGFBP-1, GenBank accession no. NP-005121) is prominent for its role as the chaperone for fibroblast growth factor-2 (FGF-2), which plays a crucial role in angiogenesis as well as promoting tumor growth. HBp17/FGFBP-1 has been proposed as a candidate biomarker for a number of cancers since it is frequently found to be elevated in many cancer types including in the tissue and cell lines of oral squamous cell carcinomas (OSCC). Previously, we reported that 1α,25(OH)2D3 suppressed the HBp17/FGFBP-1 expression in OSCC by inhibiting nuclear factor-kappaB (NF-κB) expression via vitamin D3 receptor (VDR). In this paper, to further characterize the inhibitory effect of 1α,25(OH)2D3 on HBp17/FGFBP-1, we examined the cellular localization of HBp17/FGFBP-1 protein and FGF-2 protein in the UE OSCC cell line. We found that the treatment of OSCC cells with 40-nM 1α,25(OH)2D3 suppressed HBp17/FGFBP-1 expression both in the nucleus and cytosol and reduced FGF-2 release into the culture medium. The expression of HBp17/FGFBP-1 and FGF-2 was analyzed by immunofluorescence and enzyme-linked immunosorbent assay (ELISA). In summary, the ability of 1α,25(OH)2D3 to suppress the expression of HBp17/FGFBP-1 and FGF-2 strongly suggests a therapeutic potential as a molecular-targeted anticancer drug for FGF-dependent cancers.

197 Alpha-5 Beta-1 Integrin: a Promising Therapeutic Target in the Field of Insulin Resistance and Cardiovascular Disease

Obesity is a key factor in the development of insulin resistance. Resistance to the vascular effects of insulin plays a central role in the initiation and progression of cardiovascular disease....

Intravitreal Injection of IGFBP-3 Restores Normal Insulin Signaling in Diabetic Rat Retina

Diabetes-induced changes in growth factor binding protein 3 (IGFBP-3) and tumor necrosis factor alpha (TNFα) have been linked to decreased insulin receptor signaling in diabetic retinopathy. Our previous studies in retinas of diabetic rats have shown that Compound 49b, a novel β-adrenergic receptor agonist, prevented diabetic changes by increasing IGFBP-3 and decreasing TNFα, thus restoring insulin signaling and protection against diabetic retinopathy. The current study was designed to determine whether boosted expression of IGFBP-3 NB (a non-IGF-1 binding form of IGFBP-3) alone is sufficient to mimic the full actions of Compound 49b in protecting against diabetic retinopathy, as well as testing whether IGFBP-3 NB is linked to a restoration of normal insulin signal transduction. Two months after initiation of streptozotocin-induced diabetes, rats received a single intravitreal injection of IGFBP-3 NB plasmid in the right eye. Four days after injection, electroretinogram (ERG) analyses were performed prior to sacrifice. Whole retinal lysates from control, diabetic, diabetic + control plasmid, and diabetic+ IGFBP-3 NB were analyzed for IGFBP-3, TNFα, suppressor of cytokine signaling 3 (SOCS3), and insulin receptor signaling partners using Western blotting or ELISA. Data show that a single intraocular injection of IGFBP-3 NB in diabetic animals significantly reduced TNFα levels, concomitant with reductions in IRS-1Ser307, SOCS3, and pro-apoptotic markers, while restoring insulin receptor phosphorylation and increasing anti-apoptotic marker levels. These cellular changes were linked to restoration of retinal function. Our findings establish IGFBP-3 as a pivotal regulator of the insulin receptor/TNFα pathway and a potential therapeutic target for diabetic retinopathy.

Association of serum free IGF-1 and IGFBP-1 with insulin sensitivity and insulin secretory defects in Bangladeshi type 2 diabetes mellitus

Abstract Objectives Serum free insulin like growth factors-1 (IGF-1) and insulin like growth factor binding protein-1 (IGFBP-1) may have an association with insulin resistance and B-cell secretory dysfunction. Present study is undertaken to explore the independent association of IGF-1 and IGFBP-1with two basic defects in type 2 diabetic patients in a Bangladeshi population. Methods Sixty eight type 2 diabetes mellitus (DM) subjects are studied along with age-, sex- and BMI-matched 61 healthy controls without family history of diabetes or prediabetes. Insulin, free IGF-1 and IGFBP-1 are measured by the standard ELISA method. Insulin secretory capacity (HOMA B) and insulin sensitivity (HOMA S) are calculated using fasting glucose and fasting insulin by HOMA-CIGMA software. Results Fasting free IGF-1 and IGFBP-1 levels are not significantly different between the study groups. But the level of free IGF-1 is significantly higher ( p = 0.03) in type 2 DM subjects compared to controls in the low BMI group (BMI ⩽ 23). In stepwise multiple regression analysis, positive association of HOMA B with free IGF-1 is evident. Similarly positive association of HOMA B and HOMA S with IGFBP-1 is shown in different models. In the same analysis, both free IGF-1 and IGFBP-1 are inversely associated with fasting insulin in different models of stepwise multiple regression analysis. Conclusion Both IGF-1 and IGFBP-1 are negatively associated with fasting insulin levels. Moreover B-cell secretory dysfunction is positively associated with IGF-1 and both B-cell secretory dysfunction and insulin sensitivity are also positively associated with IGFBP-1 in type 2 diabetic patients.

Insulin like growth factor binding protein 4 promotes GBM progression and regulates key factors involved in EMT and invasion

Insulin like growth factor binding protein 4 (IGFBP4) regulates growth and development of tissues and organs by negatively regulating IGF signaling. Among most cancers, IGFBP4 has growth inhibitory role and reported as a down-regulated gene, except for renal cell carcinoma, wherein IGFBP4 promotes tumor progression. IGFBP4 expression has been shown to be higher in increasing grades of astrocytoma. However, the functional role of IGFBP4 in gliomas has not been explored. Surgical biopsies of 20 normal brain and 198 astrocytoma samples were analyzed for IGFBP4 expression by qRT-PCR. Highest expression of IGFBP4 mRNA was seen in GBM tumors compared to control brain tissues (median log2 of 2.035, p<0.0001). Immunohistochemical analysis of 53 tissue samples revealed predominant nuclear staining of IGFBP4, seen maximally in GBMs when compared to DA and AA tumors (median LI=29.12±16.943, p<0.001). Over expression of IGFBP4 in U343 glioma cells resulted in up-regulation of molecules involved in tumor growth, EMT and invasion such as pAkt, pErk, Vimentin, and N-cadherin and down-regulation of E-cadherin. Functionally, IGFBP4 over expression in these cells resulted in increased proliferation, migration and invasion as assessed by MTT, transwell migration, and Matrigel invasion assays. These findings were confirmed upon IGFBP4 knockdown in U251 glioma cells. Our data suggest a pro-tumorigenic role for IGFBP4 in glioma.

Kininogen 1 and insulin‐like growth factor binding protein 6: candidate serum biomarkers of proliferative vitreoretinopathy

BackgroundThe aim was to validate whether kininogen 1 (KNG1) or insulin‐like growth factor binding protein 6 (IGFBP‐6) are serum biomarkers of proliferative vitreoretinopathy (PVR).MethodsSamples from vitreous and corresponding serum samples were collected from patients with PVR. The donor vitreous samples and serum samples from healthy volunteers and volunteers who had undergone vitrectomies for other conditions were used as controls. The samples were subsequently analysed using Western blotting (WB) and enzyme‐linked immunosorbent assay.ResultsThe Western blotting outcomes indicated both IGFBP‐6 and KNG1 could be specifically detected in the vitreous and serum samples of patients with PVR. The concentrations of KNG1 and IGFBP‐6 were significantly higher in both vitreous and serum samples from patients with severe PVR than in the samples from patients with moderate PVR. The serum concentrations of KNG1 or IGFBP‐6 had decreased by the post‐vitrectomy examinations. The receiver operating characteristic (ROC) analyses when the concentrations of IGFBP‐6 or KNG1 were greater than 181.4-pg/ml or 441.75-ng/ml, respectively, predicted severe PVR with both a sensitivity and specificity of over 70 per cent. When the concentrations of IGFBP‐6 or KNG1 were greater than 98.5-pg/ml or 88.5-ng/ml, respectively, they predicted the PVR prognosis with both a sensitivity and specificity of 80 per cent.ConclusionsKNG1 and IGFBP‐6 may be candidate serum biomarkers of PVR.

IL-6-STAT3 signaling and premature senescence

Cytokines play several roles in developing and/or reinforcing premature cellular senescence of young cells. One such cytokine, interleukin-6 (IL-6), regulates senescence in some systems in addition to its known functions of immune regulation and promotion of tumorigenesis. In this review, we describe recent advances in studies on the roles of IL-6 and its downstream signal transducer and activator of transcription 3 (STAT3) in regulating premature cellular senescence. IL-6/sIL-6Rα stimulation forms a senescence-inducing circuit involving the STAT3-insulin-like growth factor-binding protein 5 (IGFBP5) as a key axis triggering and reinforcing component in human fibroblasts. We describe how cytokines regulate the process of senescence by activating STAT3 in one system and anti-senescence or tumorigenesis in other systems. The roles of other STAT members in premature senescence also will be discussed to show the multiple mechanisms leading to cytokine-induced senescence.

Cancer Stemness in Apc- vs. Apc/KRAS-Driven Intestinal Tumorigenesis

Constitutive activation of the Wnt pathway leads to adenoma formation, an obligatory step towards intestinal cancer. In view of the established role of Wnt in regulating stemness, we attempted the isolation of cancer stem cells (CSCs) from Apc- and Apc/KRAS-mutant intestinal tumours. Whereas CSCs are present in Apc/KRAS tumours, they appear to be very rare (<10−6) in the Apc–mutant adenomas. In contrast, the Lin−CD24hiCD29+ subpopulation of adenocarcinoma cells appear to be enriched in CSCs with increased levels of active β-catenin. Expression profiling analysis of the CSC-enriched subpopulation confirmed their enhanced Wnt activity and revealed additional differential expression of other signalling pathways, growth factor binding proteins, and extracellular matrix components. As expected, genes characteristic of the Paneth cell lineage (e.g. defensins) are co-expressed together with stem cell genes (e.g. Lgr5) within the CSC-enriched subpopulation. This is of interest as it may indicate a cancer stem cell niche role for tumor-derived Paneth-like cells, similar to their role in supporting Lgr5+ stem cells in the normal intestinal crypt. Overall, our results indicate that oncogenic KRAS activation in Apc-driven tumours results in the expansion of the CSCs compartment by increasing ®-catenin intracellular stabilization.

Expression of insulin‐like growth factor binding proteins during mouse cochlear development

Insulin-like growth factor (IGF) signaling plays important roles in growth and cellular differentiation in the cochlear sensory epithelium. However, the roles of IGF binding proteins (IGFBPs), a family of IGF modulators, remain to be elucidated in this system. To begin to examine the role of IGFBPs, we used reverse transcription polymerase chain reaction (RT-PCR) and in situ hybridization to determine the temporal and spatial patterns of expression for Igfbps within the developing mouse cochlea. RT-PCR analysis indicates that Igfpb2-5 are expressed in the cochlea between embryonic day (E) 13.5 and postnatal day (P) 0. In addition, the expression of each Igfbp significantly increased between E13.5 and P0. In situ hybridization indicates that Igfbp2, 3, 4, and 5 have distinct and complementary expression patterns in the developing cochlea. Moreover, expression patterns of Igfbp3 and 5 demonstrate contrasting gradients along the basal-to-apical axis of the cochlea. Igfbp2-5 are expressed in distinct and complementary patterns during cochlear development. These data suggest that IGFBPs may act to precisely regulate activation of IGF signaling in the developing cochlea in a cell type-specific manner, contributing to cellular patterning and differentiation in the cochlea.