Compromised IGF signaling causes caspase-6 activation in Huntington disease.

Huntington disease (HD) is an autosomal dominant neurodegenerative disorder caused by an expansion of a polyglutamine repeat in the huntingtin (HTT) protein. Aberrant activation of caspase-6 and cleavage of mutant HTT generating the toxic N-terminal 586 HTT fragment are important steps in the pathogenesis of HD. Similarly, alterations in the insulin-like growth factor 1 (IGF-1) signaling pathway have been implicated in the disease as a result of decreased plasma IGF-1 levels in HD patients. In addition, two recent studies have demonstrated therapeutic benefit of IGF-1 treatment in mouse models of HD. Since IGF-1 promotes pro-survival pathways, we examined the relationship between IGF-1 signaling and aberrant caspase-6 activation in HD. Using immortalized mouse striatal cells expressing wild-type (STHdhQ7) or mutant HTT (STHdhQ111), we show that reduced levels of IGF-1 are associated with enhanced activation of caspase-6, increased cell death, and mutant HTT cleavage in a cellular stress paradigm. We demonstrate that IGF-1 supplementation reverses these effects and lowers the level of the toxic 586 HTT fragment. In addition, transcriptional analysis in the R6/2 HD transgenic mouse model demonstrated that the IGF-1 signaling system is dysregulated at multiple levels in several tissues including liver, muscle, and brain. Among these changes, we found increased expression of IGF-1 binding protein 3 (IGFBP-3), which may further reduce the bioavailability of IGF-1 as a consequence of increased IGF-1 binding. Our findings thus suggest that the therapeutic benefit of IGF-1 supplementation in HD may be significantly improved if other defects in the IGF-1 signaling pathway are corrected concurrently.

Suppression of tumor growth via IGFBP3 depletion as a potential treatment in glioma.

OBJECTIVE: Despite intensive medical treatment, patients with glioblastoma (grade IV glioma [GBM]) have a low 5-year survival rate of 5.5%. In this study, the authors tried to improve currently used therapies by identification of a therapeutic target, IGFBP3, for glioma treatment. METHODS: IGFBP3 RNA expression in 135 patients newly diagnosed with glioma was correlated with clinicopathological factors. Immunohistochemical analysis was performed to determine IGFBP3 protein expression in glioma specimens. The effect of IGFBP3 depletion on cell proliferation was examined using IGFBP3 knockdown glioma cells. Intracranial infusion of IGFBP3 siRNAs was performed to evaluate the effect of IGFBP3 depletion in mouse intracranial xenograft models. RESULTS: We demonstrated higher IGFBP3 expression in GBM than in tumor margin and grade II glioma. IGFBP3 expression was not only positively correlated with tumor grades but also associated with tumor histology and IDH1/2 mutation status. Additionally, higher IGFBP3 expression predicted shorter overall survival in glioma and GBM proneural subgroup patients. In vitro cell culture studies suggested IGFBP3 knockdown suppressed cell proliferation and induced cell cycle G2/M arrest as well as apoptosis in glioma cells. Also, accumulation of DNA double-strand breaks and γH2AX was observed in IGFBP3 knockdown cells. IGFBP3 knockdown delayed in vivo tumor growth in mouse subcutaneous xenograft models. Furthermore, convection-enhanced delivery of IGFBP3 siRNA to mouse brain suppressed intracranial tumor growth and prolonged survival of tumor-bearing mice. CONCLUSIONS: Our findings suggest IGFBP3 predicts poor outcome of glioma patients and is a potential therapeutic target for which depletion of its expression suppresses tumor growth through inducing apoptosis and accumulation of DNA damage in glioma cells.

MiR-592 functions as a tumor suppressor in glioma by targeting IGFBP2.

A growing body of evidence suggests that microRNA-592 is involved in tumor initiation and development in several types of human cancers. However, the biological functions and molecular mechanism of microRNA-592 in glioma remain unclear. In this study, we explored the potential role of microRNA-592 in glioma as well as the possible molecular mechanisms. Our results proved that microRNA-592 expression was significantly downregulated in glioma tissues and cell lines (p < 0.01). Functional assays revealed that overexpression of microRNA-592 dramatically reduced the cell proliferation, migration, and invasion and induced cell arrest at G1/G0 phase in vitro. Mechanistic investigations defined insulin-like growth factor binding protein 2 as a direct and functional downstream target of microRNA-592, which was involved in the microRNA-592-mediated tumor-suppressive effects in glioma cells. Moreover, the in vivo study showed that microRNA-592 overexpression produced the smaller tumor volume and weight in nude mice. In summary, these results elucidated the function of microRNA-592 in glioma progression and suggested a promising application of it in glioma treatment.

Metastasis-associated protein 1 is an upstream regulator of DNMT3a and stimulator of insulin-growth factor binding protein-3 in breast cancer.

Despite a recognized role of DNA methyltransferase 3a (DNMT3a) in human cancer, the nature of its upstream regulator(s) and relationship with the master chromatin remodeling factor MTA1, continues to be poorly understood. Here, we found an inverse relationship between the levels of MTA1 and DNMT3a in human cancer and that high levels of MTA1 in combination of low DNMT3a status correlates well with poor survival of breast cancer patients. We discovered that MTA1 represses DNMT3a expression via HDAC1/YY1 transcription factor complex. Because IGFBP3 is an established target of DNMT3a, we investigated the effect of MTA1 upon IGFBP3 expression, and found a coactivator role of MTA1/c-Jun/Pol II coactivator complex upon the IGFBP3 transcription. In addition, MTA1 overexpression correlates well with low levels of DNMT3a which, in turn also correlates with a high IGFBP3 status in breast cancer patients and predicts a poor clinical outcome for breast cancer patients. These findings suggest that MTA1 could regulate the expression of IGFBP3 in both DNMT3a-dependent and -independent manner. Together findings presented here recognize an inherent role of MTA1 as a modifier of DNMT3a and IGFBP3 expression, and consequently, the role of MTA1-DNMT3a-IGFBP3 axis in breast cancer progression.

IGFBP-2 and -3 co-ordinately regulate IGF1 induced matrix mineralisation of differentiating human dental pulp cells.

Human dental pulp cells (DPCs), which are known to contain a subset of stem cells capable of reforming a dentin and pulp-like complex upon in vivo transplantation, were isolated from third molars of three healthy donors and differentiated to a matrix mineralisation phenotype using by culture in dexamethasone and l-ascorbic acid. qRT-PCR analysis of insulin-like growth factor ( IGF) axis gene expression indicated that all genes, except insulin-like growth factor 1 (IGF1) and insulin-like growth factor binding protein-1 ( IGFBP-1), were expressed in DPCs. During differentiation upregulation of insulin-like growth factor binding protein-2 (IGFBP-2) and downregulation of insulin-like growth factor binding protein-3 (IGFBP-3) expression was observed. Changes in IGFBP-2 and IGFBP-3 mRNA expression were confirmed at the protein level by ELISA of DPC conditioned medium functional analysis indicated that IGF1 stimulated the differentiation of DPCs and that the activity of the growth factor was enhanced by pre-complexation with IGFBP-2 but inhibited by pre-complexation with IGFBP-3. Therefore changes in IGFBP-2 and -3 expression during differentiation form part of a co-ordinated functional response to enhance the pro-differentiative action of IGF1 and represent a novel mechanism for the regulation of DPC differentiation.

Compound 49b Restores Retinal Thickness and Reduces Degenerate Capillaries in the Rat Retina following Ischemia/Reperfusion.

We have recently reported that Compound 49b, a novel ß-adrenergic receptor agonist, can significantly reduce VEGF levels in retinal endothelial cells (REC) grown in diabetic-like conditions. In this study, we investigated whether Compound 49b could protect the retina under hypoxic conditions using the ischemia-reperfusion (I/R)-induced model in rats, as well REC cultured in hypoxic conditions. Some rats received 1mM topical Compound 49b for the 2 (5 rats each group) or 10 (4 rats in each group) days post-I/R. Analyses for retinal thickness and cell loss in the ganglion cell layer was done at 2 days post-I/R, while numbers of degenerate capillaries and pericyte ghosts were measured at 10 days post-I/R. Additionally, REC were cultured in normal oxygen or hypoxia (5% O2) only or treated with 50 nM Compound 49b for 12 hours. Twelve hours after Compound 49b exposure, cells were collected and analyzed for protein levels of insulin-like growth factor binding protein 3 (IGFBP-3), vascular endothelial cell growth factor (VEGF) and its receptor (KDR), angiopoietin 1 and its receptor Tie2 for Western blotting. Data indicate that exposure to I/R significantly decreased retinal thickness, with increasing numbers of degenerate capillaries and pericyte ghosts. Compound 49b treatment inhibited these retinal changes. In REC cultured in hypoxia, levels of IGFBP-3 were reduced, which were significantly increased by Compound 49b. Hypoxia significantly increased protein levels of VEGF, KDR, Angiopoiein 1, and Tie2, which were reduced following Compound 49b treatment. These data strongly suggested that Compound 49b protected the retina against I/R-induced injury. This provides additional support for a role of ß-adrenergic receptor actions in the retina.

MicroRNA-197 Mediates the Overgrowth and Anti-Apoptotic Effects by Downregulating Insulin-Like Growth Factor-Binding Protein-3 During Nephroblastoma Tumorigenesis.

MiR-197 is frequently upregulated to induce a series of oncogenic effects, which is closely associated with poor survival and prognosis of multiple malignancies. However, the roles of miR-197 in tumorigenesis and the detailed molecular mechanism in Wilms tumor (WT) have rarely been reported. This study aimed to evaluate the expression of miR-197 in WT in vivo and the potential effects of miR-197 on the proliferation and apoptosis in SK-NEP-1 cells. A total of 15 patients with a pathologically confirmed diagnosis of WT and 15 paraneoplastic controls were enrolled. Real-time quantitative PCR (RT-qPCR) identified the upregulation of miR-197 and downregulation of insulin-like growth factors binding protein 3 (IGFBP3) in WT tissues in comparison with adjacent normal tissue (p < 0.001). CCK-8 and flow cytometry assay found that inhibition of miR-197 caused a significantly reduced proliferation along with a dramatically enhanced apoptosis of SK-NEP-1 cells (p < 0.01). IGFBP3 was overexpressed in SK-NEP-1 cells by pEGFP-C1-IGFBP3 plasmid transfection. Overexpression of IGFBP3 suppressed the proliferation and induced the apoptosis of SK-NEP-1 cells (p < 0.01). Further study detected the decreased IGFBP3 expression with miR-197 mimics SK-NEP-1 cells and increased IGFBP3 expression with miR-197 inhibitor SK-NEP-1 cells compared with mock (p < 0.01). Dual luciferase reporter assay revealed a direct interaction between miR-197 and 3'-UTR site of IGFBP3. Overall, the above results indicated that miR-197 targeted IGFBP3 to induce the overgrowth and anti-apoptotic effects of WT cells, which could promote nephroblastoma tumorigenesis. Therefore, miR-197 may be further assessed as a potential target for the treatment of WT.

Effects of Phlomis umbrosa Root on Longitudinal Bone Growth Rate in Adolescent Female Rats.

This study aimed to investigate the effects of Phlomis umbrosa root on bone growth and growth mediators in rats. Female adolescent rats were administered P. umbrosa extract, recombinant human growth hormone or vehicle for 10 days. Tetracycline was injected intraperitoneally to produce a glowing fluorescence band on the newly formed bone on day 8, and 5-bromo-2'-deoxyuridine was injected to label proliferating chondrocytes on days 8-10. To assess possible endocrine or autocrine/paracrine mechanisms, we evaluated insulin-like growth factor-1 (IGF-1), insulin-like growth factor binding protein-3 (IGFBP-3) or bone morphogenetic protein-2 (BMP-2) in response to P. umbrosa administration in either growth plate or serum. Oral administration of P. umbrosa significantly increased longitudinal bone growth rate, height of hypertrophic zone and chondrocyte proliferation of the proximal tibial growth plate. P. umbrosa also increased serum IGFBP-3 levels and upregulated the expressions of IGF-1 and BMP-2 in growth plate. In conclusion, P. umbrosa increases longitudinal bone growth rate by stimulating proliferation and hypertrophy of chondrocyte with the increment of circulating IGFBP-3. Regarding the immunohistochemical study, the effect of P. umbrosa may also be attributable to upregulation of local IGF-1 and BMP-2 expressions in the growth plate, which can be considered as a GH dependent autocrine/paracrine pathway.

Gene Expression Profiling Identifies Interferon Signalling Molecules and IGFBP3 in Human Degenerative Annulus Fibrosus.

Low back pain is a major cause of disability especially for people between 20 and 50 years of age. As a costly healthcare problem, it imposes a serious socio-economic burden. Current surgical therapies fail to replace the normal disc in facilitating spinal movements and absorbing load. The focus of regenerative medicine is on identifying biomarkers and signalling pathways to improve our understanding about cascades of disc degeneration and allow for the design of specific therapies. We hypothesized that comparing microarray profiles from degenerative and non-degenerative discs will lead to the identification of dysregulated signalling and pathophysiological targets. Microarray data sets were generated from human annulus fibrosus cells and analysed using IPA ingenuity pathway analysis. Gene expression values were validated by qRT-PCR, and respective proteins were identified by immunohistochemistry. Microarray analysis revealed 238 differentially expressed genes in the degenerative annulus fibrosus. Seventeen of the dysregulated molecular markers showed log2-fold changes greater than ±1.5. Various dysregulated cellular functions, including cell proliferation and inflammatory response, were identified. The most significant canonical pathway induced in degenerative annulus fibrosus was found to be the interferon pathway. This study indicates interferon-alpha signalling pathway activation with IFIT3 and IGFBP3 up-regulation, which may affect cellular function in human degenerative disc.

A pleiotropic effect of the single clustered hepatic metastamiRs miR-96-5p and miR-182-5p on insulin-like growth factor II, insulin-like growth factor-1 receptor and insulin-like growth factor-binding protein-3 in hepatocellular carcinoma.

MicroRNAs (miRs) have a major role in the pathogenesis of hepatocellular carcinoma (HCC). As the insulin-like growth factor (IGF) axis is a highly tumorigenic pathway in HCC, the present study attempted to target it with miRs. Potential targeting of crucial members of the IGF axis by miRNAs at the 3'-untranslated region (3'-UTR) was predicted using bioinformatic tools, such as microrna.org, Diana lab and Targetscan, while 5'-UTR targeting was predicted using bibiserv software. Expression profiling of obtained miRNAs was performed using quantitative polymerase chain reaction (qPCR) in 22 non-metastatic HCC biopsy samples and 10 healthy tissues. To investigate the impact of miRNAs on their potential downstream targets, transfection of miRNAs was performed in HuH-7 cells and the targets' expression was quantified using qPCR. Transcripts of insulin-like growth factor-1 receptor (IGF-1R), insulin-like growth factor binding protein-3 (IGFBP-3) and IGF-II were found to be potentially targeted at the 5'-UTR and 3'-UTR regions by the single clustered hepatic metastamiRs miR-96-5p and miR-182-5p. The two miRNAs showed a similar expression pattern in HCC tissues compared to those in healthy tissues. Forced expression of miR-96-5p and miR-182-5p in the HCC cell line HuH-7 had inducing effects on IGFBP-3 and IGF-II transcripts. Of note, the two miRs had differential effects on IGF-1R, where miR-96-5p induced IGF-1R mRNA expression and miR-182-5p inhibited its expression. The present study revealed the pleiotropic impact of the single clustered hepatic metastamiRs miR-96-5p and miR-182-5p on IGF-1R, and an inducing effect on IGF-II and IGFBP-3 in hepatocellular carcinoma.

Pioglitazone restores IGFBP-3 levels through DNA PK in retinal endothelial cells cultured in hyperglycemic conditions.

PURPOSE: Previously, we reported that pioglitazone prevented insulin resistance and cell death in type 2 diabetic retina by reducing TNFα and suppressor of cytokine signaling 3 (SOCS3) levels. Numerous reports suggest prominent vasoprotective effects of insulin growth factor binding protein-3 (IGFBP-3) in diabetic retinopathy. We hypothesized that pioglitazone protects against retinal cell apoptosis by regulating IGFBP-3 levels, in addition to reducing TNFα. The current study explored potential IGFBP-3 regulatory pathways by pioglitazone in retinal endothelial cells cultured in high glucose. METHODS: Primary human retinal endothelial cells (REC) were grown in normal (5 mM) and high glucose (25 mM) and treated with pioglitazone for 24 hours. Cell lysates were processed for Western blotting and ELISA analysis to evaluate IGFBP-3, TNFα, and cleaved caspase 3 protein levels. RESULTS: Our results show that treatment with pioglitazone restored the high glucose-induced decrease in IGFBP-3 levels. This regulation was independent of TNFα actions, as reducing TNFα levels with siRNA did not prevent pioglitazone from increasing IGFBP-3 levels. Pioglitazone required protein kinase A (PKA) and DNA-dependent protein kinase (DNA PK) activity to regulate IGFBP-3, as specific inhibitors for each protein prevented pioglitazone-mediated normalization of IGFBP-3 in high glucose. Insulin growth factor binding protein-3 activity was increased and apoptosis decreased by pioglitazone, which was eliminated when serine site 156 of IGFBP-3 was mutated suggesting a key role of this phosphorylation site in pioglitazone actions. CONCLUSIONS: Our findings suggest that pioglitazone mediates regulation of IGFBP-3 via activation of PKA/DNA PK pathway in hyperglycemic retinal endothelial cells.

Expression and underlying roles of IGFBP-3 in paclitaxel-treated gastric cancer SGC-7901 cells.

PURPOSE: To study the expression of insulin-like growth factor binding proteins (IGFBPs) in paclitaxel-treated gastric cancer SGC-7901 cells, and to further investigate underlying mechanisms. MATERIALS AND METHODS: Real time PCR and Western blot assays were applied to detect the mRNA and protein expression of IGFBP-2, -3 and -5 after paclitaxel (10 nM) treatment of SGC-7901 cells. In addition IGFBP-3 expression was silenced by RNA interference to determine effects. Cell viability was determined by MTT assay. Cell cycling and apoptosis were assessed by flow cytometry. RESULTS: Compared to the control group, only IGFBP-3 expression was elevated significantly after paclitaxel (10 nM) treatment (p<0.05). Paclitaxel treatment caused cell cycle arrest and apoptosis via downregulating Bcl-2 expression. However, the effect could be abrogated by IGFBP-3 silencing. CONCLUSIONS: IGFBP-3 exhibits anti-apoptotic effects on paclitaxel-treated SGC-7901 cells via elevating Bcl-2 expression.

Sulforaphane inhibits the proliferation of the BIU87 bladder cancer cell line via IGFBP-3 elevation.

AIM: To investigate effects of sulforaphane on the BIU87 cell line and underlying mechanisms involving IGFBP-3. METHODS: Both BIU87 and IGFBP-3-silenced BIU87 cells were treated with sulforaphane. Cell proliferation was detected by MTT assay. Cell cycle and apoptosis were determined via flow cytometry. Quantitative polymerase chain reaction and Western blotting were applied to analyze the expression of IGFBP-3 and NF-κB at both mRNA and protein levels. RESULTS: Sulforaphane (80 µM) treatment could inhibit cell proliferation, inducing apoptosis and cell cycle arrest at G2/M phase. All these effects could be antagonized by IGFBP-3 silencing. Furthermore, sulforaphane (80 µM) could down-regulate NF-κB expression while elevating that of IGFBP-3. CONCLUSIONS: Sulforaphane could suppress the proliferation of BIU87 cells via enhancing IGFBP-3 expression, which negatively regulating the NF-κB signaling pathway.

ß-D-glucan inhibits endocrine-resistant breast cancer cell proliferation and alters gene expression.

Endocrine therapies have been successfully used for breast cancer patients with estrogen receptor α (ERα) positive tumors, but ~40% of patients relapse due to endocrine resistance. ß-glucans are components of plant cell walls that have immunomodulatory and anticancer activity. The objective of this study was to examine the activity of ß-D-glucan, purified from barley, in endocrine-sensitive MCF-7 versus endocrine-resistant LCC9 and LY2 breast cancer cells. ß-D-glucan dissolved in DMSO but not water inhibited MCF-7 cell proliferation in a concentration-dependent manner as measured by BrdU incorporation with an IC50 of ~164 ± 12 µg/ml. ß-D-glucan dissolved in DMSO inhibited tamoxifen/endocrine-resistant LCC9 and LY2 cell proliferation with IC50 values of 4.6 ± 0.3 and 24.2 ± 1.4 µg/ml, respectively. MCF-10A normal breast epithelial cells showed a higher IC50 ~464 µg/ml and the proliferation of MDA-MB-231 triple negative breast cancer cells was not inhibited by ß-D-glucan. Concentration-dependent increases in the BAX/BCL2 ratio and cell death with ß-D-glucan were observed in MCF-7 and LCC9 cells. PCR array analysis revealed changes in gene expression in response to 24-h treatment with 10 or 50 µg/ml ß-D-glucan that were different between MCF-7 and LCC9 cells as well as differences in basal gene expression between the two cell lines. Select results were confirmed by quantitative real-time PCR demonstrating that ß-D-glucan increased RASSF1 expression in MCF-7 cells and IGFBP3, CTNNB1 and ERß transcript expression in LCC9 cells. Our data indicate that ß-D-glucan regulates breast cancer-relevant gene expression and may be useful for inhibiting endocrine-resistant breast cancer cell proliferation.

Inhibition of human MCF-7 breast cancer cells and HT-29 colon cancer cells by rice-produced recombinant human insulin-like growth binding protein-3 (rhIGFBP-3).

BACKGROUND: Insulin-like growth factor binding protein-3 (IGFBP-3) is a multifunctional molecule which is closely related to cell growth, apoptosis, angiogenesis, metabolism and senescence. It combines with insulin-like growth factor-I (IGF-I) to form a complex (IGF-I/IGFBP-3) that can treat growth hormone insensitivity syndrome (GHIS) and reduce insulin requirement in patients with diabetes. IGFBP-3 alone has been shown to have anti-proliferation effect on numerous cancer cells. METHODOLOGY/PRINCIPAL FINDINGS: We reported here an expression method to produce functional recombinant human IGFBP-3 (rhIGFBP-3) in transgenic rice grains. Protein sorting sequences, signal peptide and endoplasmic reticulum retention tetrapeptide (KDEL) were included in constructs for enhancing rhIGFBP-3 expression. Western blot analysis showed that only the constructs with signal peptide were successfully expressed in transgenic rice grains. Both rhIGFBP-3 proteins, with or without KDEL sorting sequence inhibited the growth of MCF-7 human breast cancer cells (65.76 ± 1.72% vs 45.00 ± 0.86%, p < 0.05; 50.84 ± 1.97% vs 45.00 ± 0.86%, p < 0.01 respectively) and HT-29 colon cancer cells (65.14 ± 3.84% vs 18.01 ± 13.81%, p < 0.05 and 54.7 ± 9.44% vs 18.01 ± 13.81%, p < 0.05 respectively) when compared with wild type rice. CONCLUSION/SIGNIFICANCE: These findings demonstrated the feasibility of producing biological active rhIGFBP-3 in rice using a transgenic approach, which will definitely encourage more research on the therapeutic use of hIGFBP-3 in future.

Comparison of growth hormone receptor, igf-1r and igfbp-3 between tumoral and non-tumoral areas in non-melanoma skin cancers.

OBJECTIVE: A relationship between the pathogenesis of some cancers and growth hormone, insulin-like growth factor-1 and insulin-like growth factors binding protein-3 has been shown. Our aim was to evaluate the expression of growth hormone receptor, insulin-like growth factor-1 receptor and insulin-like growth factors binding protein-3 in actinic keratosis, basal cell carcinoma and squamous cell carcinoma, and to compare the expression patterns of tumoral areas with normal epidermis and skin appendages. MATERIAL AND METHOD: The formalin-fixed, paraff in-embedded tissues of 40 patients which were diagnosed as 15 actinic keratosis, 15 basal cell carcinoma and 15 squamous cell carcinoma were analyzed for growth hormone receptor, insulin-like growth factor-1 receptor and insulin-like growth factors binding protein-3 with the immunohistochemical method using the streptavidin-biotin-peroxidase technique. RESULTS: There was no difference between tumoral areas of actinic keratosis, basal cell carcinoma and squamous cell carcinoma in expression of growth hormone receptor and insulin-like growth factors binding protein-3 (P > 0.05). However, a significantly higher expression of insulin-like growth factor-1 receptor was observed in tumoral areas of squamous cell carcinoma (P < 0.01). In basal cell carcinoma, a significantly lower intensity of immunostaining with growth hormone receptor, insulin-like growth factor-1 receptor and insulin-like growth factors binding protein-3 in tumoral areas than skin appendages was seen (P < 0.01). In squamous cell carcinoma, higher expressions of growth hormone receptor, insulin-like growth factor-1 receptor and insulin-like growth factors binding protein-3 in tumoral areas than peritumoral epidermis was found (P =0.06, P < 0.01 and P =0.02, respectively). CONCLUSION: Our study points out that, growth hormone receptor, insulin-like growth factor-1 receptor and insulin-like growth factors binding protein-3 have a role in the pathogenesis of non-melanoma skin cancers, especially squamous cell carcinoma.

Interactions between vitamin D and IGF-I: from physiology to clinical practice.

The interplay between vitamin D and IGF-I is complex and occurs at both endocrine and paracrine/autocrine levels. Vitamin D has been shown to increase circulating IGF-I and IGFBP-3, with the consistent finding of a positive correlation between vitamin D and IGF-I serum values in population-based cohorts of healthy subjects. The modulation of IGF-I and IGFBP-3 concentrations by vitamin D may impact recombinant human (rh) GH dosing for the treatment of GHD. It might also underlie some of the extra-skeletal beneficial effects ascribed to vitamin D. On the other hand, IGF-I stimulates renal production of 1,25-dihydroxyvitamin D, which increases calcium and phosphate availability in the body and suppresses PTH secretion. This effect is responsible for an altered calcium-phosphate balance in uncontrolled acromegaly and might also account for the improvement in bone metabolism associated with rhGH treatment in patients with GHD. Data on the paracrine/autocrine vitamin D-IGF-I interactions are abundant, but mostly not linked to one another. As a result, it is not possible to draw a comprehensive picture of the physiological and/or pathological interrelations between vitamin D, IGF-I and IGF-binding proteins (IGFBP) in different tissues. A potential role of vitamin D action is related to its association with carcinogenesis, a paradigm being breast cancer. Current evidence indicates that, in breast tumours, vitamin D modulates the IGF-I/IGFBP ratio to decrease proliferation and increase apoptosis.

CD146 and insulin-like growth factor 2 mRNA-binding protein 3 predict prognosis of asbestos-induced rat mesothelioma.

Malignant mesothelioma (MM), which is associated with asbestos exposure, is one of the most deadly tumors in humans. Early MM is concealed in the serosal cavities and lacks specific clinical symptoms. For better treatment, early detection and prognostic markers are necessary. Recently, CD146 and insulin-like growth factor 2 mRNA-binding protein 3 (IMP3) were reported as possible positive markers of MM to distinguish from reactive mesothelia in humans. However, their application on MM of different species and its impact on survival remain to be elucidated. To disclose the utility of these molecules as early detection and prognostic markers of MM, we injected chrysotile or crocidolite intraperitoneally to rats, thus obtaining 26 peritoneal MM and establishing 11 cell lines. We immunostained CD146 and IMP3 using paraffin-embedded tissues and cell blocks and found CD146 and IMP3 expression in 58% (15/26) and 65% (17/26) of MM, respectively, but not in reactive mesothelia. There was no significant difference in both immunostainings for overexpression among the three histological subtypes of MM and the expression of CD146 and IMP3 was proportionally associated. Furthermore, the overexpression of CD146 and/or IMP3 was proportionally correlated with shortened survival. These results suggest that CD146 and IMP3 are useful diagnostic and prognostic markers of MM.

Intramammary infusion of Panax ginseng extract in the bovine mammary gland at cessation of milking modifies components of the insulin-like growth factor system during involution.

The objective of this study was to evaluate the effects of a single intramammary infusion of Panax ginseng extract (GS) on insulin-like growth factors (IGF) in bovine mammary gland during early involution. Eight mammary quarters from six nonpregnant cows in late lactation were infused with 10 mL of ginseng extract solution (3 mg/mL), six quarters were treated with 10 mL of placebo (vehicle alone) and six quarters were maintained as uninoculated controls. Milking was interrupted after infusion. Concentrations of IGF1 in mammary secretions were higher in GS-treated quarters than in placebo and uninoculated control quarters at 24, 48 and 72 h post-treatment (p<0.05). Treatment with GS did not affect mammary secretion of IGF2 (p=0.942). At 7 d of post-lactational involution, a decrease of immunostained area and mRNA expression for IGF1 was observed in mammary tissue of GS-treated quarters compared with placebo-treated quarters and uninoculated controls (p<0.05). The IGF2 immunostained area and mRNA expression for this growth factor were not affected by GS treatment (p=0.216 and p=0.785, respectively). An increase in protein levels and mRNA expression in mammary tissue of IGFBP3, IGFBP4 and IGFBP5 was observed in GS-treated quarters compared with placebo-treated quarters and uninoculated controls (p<0.05). These results provide evidence that intramammary inoculation of GS extract at cessation of milking may promote early mammary involution through the inhibition of IGF1 local production and bioavailability.

Analysis of oxygen-dependent cytokine expression in human mesenchymal stem cells derived from umbilical cord.

Efficient cell expansion is a basic requirement for obtaining clinically relevant numbers of mesenchymal stem cells designed for cell-based therapies or tissue-engineering application. Previous studies have demonstrated that mesenchymal stem cells (MSC) cultivated under reduced atmospheric oxygen concentrations (2.5% O2) possess enhanced proliferation potential and can maintain their differentiation properties. We have analyzed the oxygen-dependent cytokine expression of human MSC derived from umbilical cord and attempted to link the results to the proliferation and differentiation capacities of these cells. By quantitative reverse transcription plus the polymerase chain reaction and by protein microarray, we measured the gene expression and intracellular protein concentration of several growth factors and growth factor receptors. Fibroblast growth factor-7, two growth factor receptors (vascular endothelial growth factor receptor 2 and stem cell factor receptor), and two growth-factor-binding proteins (insulin-like growth-factor-binding proteins 3 and 6) were over-expressed under hypoxic conditions, indicating that their signaling pathways participate in cell proliferation. On the other hand, typical differentiation factors such as bone morphogenetic protein-4, endothelial growth factor, and tissue growth factor-ß1 were absent in cells cultivated under hypoxic and normoxic conditions. The absolute concentration of some intracellular cytokines was also measured for the first time under hypoxia and normoxia. Our results in combination with previous findings indicate that enhanced proliferation potential and a maintained undifferentiated cell state can be ascribed to the oxygen-dependent expression of a set of cytokines. This knowledge might help in the understanding of MSC physiology and in the achievement of directed cell fate of MSC for clinical application.