Expression Level Of IGF2 mRNA Research Articles

The effect of laser-assisted hatching on the methylation and expression pattern of imprinted gene IGF2/H19 in mouse blastocysts and offspring

This study aimed to determine the effects of drilling and thinning treatment of laser-assisted hatching on the expression and methylation of imprinted gene IGF2/H19 in embryos and offspring. The prehatching blastocysts with treatment of drilling or thinning, or control prehatching blastocysts, were transplanted in surrogate uteri. The DNA methylation of IGF2/H19 imprinting control region (ICR) and the expression of IGF2 and H19 were respectively evaluated using bisulfite conversion-mediated sequencing and real-time PCR. The drilling group showed a significant increase in the development rate of hatched blastocysts in comparison with the control and thinning group. DNA methylation level of IGF2/H19 ICR of hatched blastocysts in the thinning group was 27.33% in comparison with the 38.67% and 36% observed in the control and drilling group. The thinning treatment increased the DNA methylation level of IGF2/H19 ICR in the placenta in comparison with the control and drilling group. The drilling and thinning treatment decreased the expression level of H19 mRNA in prehatching and hatched blastocysts as well as placenta, while a significant increase in the expression level of H19 mRNA of offspring was observed in the thinning group. The thinning treatment increased the expression level of IGF2 mRNA of prehatching blastocysts and offspring and a significant decrease in placenta, while the drilling treatment resulted in a significant increase in the expression level of IGF2 mRNA of hatched blastocysts and placenta. These observations suggested that drilling used for hatching of in vitro cultured mouseblastocysts may improvethe production of offspring.

High IGF1R protein expression correlates with disease-free survival of patients with stage III colon cancer

The aim of this study was to investigate the association between expression of insulin-like growth factor-1 receptor (IGF1R) and its ligand, IGF-II, and disease-free survival (DFS) in patients with stage III colon cancer (CC). In this retrospective study we included consecutive patients who underwent curative surgery for stage III CC. IGF1R and IGF-II/IGF2 status were evaluated in tumour samples by immunohistochemistry and quantitative real-time PCR (qRT-PCR). Associations of markers with DFS were analysed using Cox proportional hazards models. Hundred and fifty-one CC patients were included (median age, 66.6years; female, 54.3%). Low levels of IGF1R and IGF-II protein expression were observed in 16.1% and 10.7% of the cases, respectively. No significant differences in clinicopathological characteristics between patients with tumours expressing low IGF1R or IGF-II protein levels and those with high levels were observed. A low IGF1R protein expression was found to be significantly associated with a shorter DFS (HR 3.32; 95% CI, 1.7-6.31; p= 0.0003), while no association was observed between IGF-II protein expression and DFS (HR 0.91; 95% CI, 0.28-2.96; p= 0.87). In a multivariate analysis, IGF1R protein status remained an independent prognostic factor for DFS (HR 2.73; 95% CI, 1.40-5.31; p= 0.003). Furthermore, we found that neither IGF1R nor IGF2 mRNA expression levels as measured by qRT-PCR correlated with the respective protein expression levels as assessed by immunohistochemistry. Neither of the mRNA expression levels was significantly associated with DFS. From our data we conclude that low IGF1R protein expression represents a poor prognostic biomarker in stage III colon cancer.

Association of body weight gain with muscle, fat, and liver expression levels of growth hormone receptor, insulin-like growth factor I, and beta-adrenergic receptor mRNAs in steers

The physiological basis of feed efficiency is unclear. Administration of GH or beta-adrenergic agonists improves feed efficiency in various animals. The objective of this study was to test the hypothesis that more efficient cattle have greater expression of GH receptor (GHR) or beta-adrenergic receptor (ADRB) mRNA in skeletal muscle, fat, and liver, the major target tissues of GH and beta-adrenergic agonists. Fifty Angus steers were fed a finishing diet for 75 d to determine residual feed intake (RFI). Carcass measures, skeletal muscle, subcutaneous fat, and liver samples were collected from the top 10 high-RFI steers and top 10 low-RFI steers at slaughter. Abundances of GHR, insulin-like growth factor I (IGF1), IGF1 receptor (IGF1R), beta-1 adrenergic receptor (ADRB1), ADRB2, and ADRB3 mRNAs were quantified by real-time reverse transcription-PCR. Low-RFI steers consumed 11% less dry matter intake than high-RFI steers (P = 0.004). Low- and high-RFI steers, however, did not differ in ADG or other growth or carcass measures. Low-RFI steers had a tendency to have smaller birth weights than high-RFI steers (P = 0.089). The expression levels of GHR, IGF1, IGF1R, ADRB1, ADRB2, and ADRB3 mRNAs in muscle, fat, and liver were neither different (P > 0.1) between high- and low-RFI steers nor correlated (P > 0.1) with RFI. These results do not support our original hypothesis. However, the expression levels of GHR, IGF1, and IGF1R mRNAs in muscle and fat were positively correlated with ADG (r = 0.52 to 0.65, P = 0.002 to 0.02), whereas the expression levels of GHR mRNA (r = −0.50, P = 0.03) and IGF1 mRNA (r = −0.47, P = 0.04) in the liver were negatively correlated with ADG. These results suggest that the GHR, IGF1, and IGF1R mRNA expression levels in the muscle and fat have a positive effect, whereas the GHR and IGF1 mRNA expression levels in the liver have a negative effect on postweaning body weight gain in cattle.

Regulation of compensatory growth by molecular mechanism in Labeo rohita juveniles under different feeding regimes

A study was carried out to assess the regulation of compensatory growth under different restriction feeding regimes in Labeo rohita juveniles by the interaction of various feed intake and growth regulating genes. A 60 day feeding trial was conducted with five treatment groups, Control (3% body weight, bw), T1 (alternate days), T2 (0.5% bw), T3 (1% bw) and T4 (2% bw) and feeding was done for first 30 days of the trial. For next 30 days, all the treatment groups were fed at a rate of 3% bw as in the control group. There was significant (p < 0.05) difference in the weight gain among the treatment groups with lowest FCR and highest PER was found in T2 group. Ghrelin gene mRNA levels were upregulated during first 30th days of the trial with highest expression levels in the T2 group. The expression levels of leptin gene mRNA were found significantly different (p < 0.05) among the treatments, which was down-regulated during initial 30 days and upregulated as the experiment progress towards 60th day. The IGF-1 mRNA expression levels were upregulated more in liver compared to the muscle tissue. The results of the study suggest that increased ghrelin levels and decreased leptin levels lead to hyperphagia during the onset of refeeding, which further triggers the compensatory growth in L. rohita. The present study describes the molecular mechanism behind the compensatory growth following a different feed restriction regime in L. rohita which is regulated due to the interaction of different energy homeostasis and growth regulating genes.

Molecular endocrine changes of Gh/Igf1 axis in gilthead sea bream (Sparus aurata L.) exposed to different environmental salinities during larvae to post-larvae stages.

The influence of acclimation of the euryhaline gilthead sea bream (Sparus aurata) larvae/post-larvae to brackish water on growth, energetic contents, and mRNA levels of selected hormones and growth-regulating hypothalamic neurohormones was assessed. Specimens from 49days post-hatching were acclimated during 28days to two different environmental salinities: 38 and 20psu (as brackish water). Both groups were then transferred to 38psu and acclimated for an additional week. Early juveniles were sampled after 28days of acclimation to both salinities and one week after transfer to 38psu. Pituitary adenylate cyclase-activating peptide (adcyap1; pacap), somatostatin-I (sst1), growth hormone (gh1), insulin-like growth factor-I (igf1), and prolactin (prl) mRNA expression were all studied by QPCR. Post-larvae acclimated to 20psu showed better growth performance and body energetic content than post-larvae maintained at 38psu. prl, adcyap1, and igf1 mRNA expression levels increased in 20-psu-acclimated post-larvae but decreased upon transfer to 38psu. GH1 expression did not show significant changes under both experimental conditions. Our results suggested an enhanced general performance for post-larvae in brackish water, supported by the actions of adcyap1, igf1, and prl.

Abstract 5131: Establishment of uterine carcinosarcoma primary cell lines for chemosensitivity testing and evaluation of targeted therapy

Abstract To establish primary cell lines from uterine carcinosarcoma (CS) patient samples and to evaluate combination treatment with standard chemotherapy with or without the addition of an anti-IGF1R antibody. CS tumor tissue was obtained under IRB approval at the time of primary surgery. We reported the establishment of patient-derived xenografts elsewhere. Tissue pieces were minced extensively and digested by collagenase treatment. The single cell solution was then seeded in F-media containing EGF, insulin, hydrocortisone, adenine, cholera toxin and the Rock inhibitor Y-27632. A previously established carcinosarcoma cell line CS99 (Schulten et al, 2008) was used for comparison. To verify origin, Short Tandem Repeat (STR) profiles of the primary cell lines CS13, CS18, CS19, CS21 and CS22 were compared with the patient's tumor samples. Drug sensitivity to Taxol, cisplatin and carboplatin was determined individually or in combination using the sulforhodamine B (SRB) proliferation assay. The effect of IMC-A12 (anti-IGF1R antibody, Eli Lily / Imclone) alone or with chemotherapy was also determined with the SRB assay. Total and promoter-specific IGF2 mRNA levels were determined by reverse transcriptase quantitative PCR. All STR profiles of the primary cell lines matched their patient sample counterparts. IGF2 mRNA expression levels of the cell lines were similar to the patient samples, and over 10,000-fold higher than in CS99. IGF2 promoter-specific primers showed that the IGF2 mRNA transcripts of the primary cell lines were initiated at the oncofetal IGF2 promoters P3 and P4. For Taxol, the IC50 (inhibitory concentration of 50%) ranged from 4.7 nM for CS21 to 9.8 nM for CS22. For cisplatin, the IC50 ranged from 1.0 μM for CS99 to 4.8 μM for CS19. For carboplatin, CS99 cells had the lowest IC50 of 12.5 μM while CS19 had the highest IC50 of 58.1 μM. In the presence of 10 μg/ml IMC-A12, the primary cell lines CS19, CS21 and CS22 showed significant sensitization to Taxol (∼50% decrease in IC50), to carboplatin (∼65% decrease in IC50), and to combination Taxol/carboplatin treatment (∼50% decrease in IC50). We conclude that establishing primary cell lines of this rare cancer is a promising approach for testing sensitivity to current and novel chemotherapies and combination treatment strategies. We also found that high oncofetal promoter-driven IGF2 mRNA expression was observed in all CS samples tested and that IGF1R blockade sensitized the primary cell lines to standard chemotherapy. Citation Format: Jurriaan Brouwer-Visser, Eirwen Scott, Shijun Mi, Maria J. Cossio, Tiffany Hebert, Gloria S. Huang. Establishment of uterine carcinosarcoma primary cell lines for chemosensitivity testing and evaluation of targeted therapy. [abstract]. In: Proceedings of the 106th Annual Meeting of the American Association for Cancer Research; 2015 Apr 18-22; Philadelphia, PA. Philadelphia (PA): AACR; Cancer Res 2015;75(15 Suppl):Abstract nr 5131. doi:10.1158/1538-7445.AM2015-5131

Pre and postprandial regulation of ghrelin, amino acids and IGF1 in Atlantic salmon (Salmo salar L.) at optimal and elevated seawater temperatures

Atlantic salmon farmed in seacage facilities are exposed to changing environmental conditions. While growth decline under lower water temperatures are reasonably well understood, the mechanisms behind production decline under high water temperatures are yet to be well elucidated and have been hampered by a lack of data describing pre- and postprandial patterns of endocrine fluctuation. The present research therefore aims to determine whether peak nutrient flux into the blood plasma is the most appropriate time point to investigate the endocrine regulation of growth and appetite under conditions of normal and high temperature, and to investigate the interrelationship between appetite and growth on a pre- and post-prandial time scale. Two experiments are presented which examine ghrelin (GHRL) as an indicator of appetite stimulation, the GH–IGF (growth hormone–insulin-like growth factor) system to describe growth-regulating processes and free amino acids (FAA) to indicate postprandial nutrient influx and link appetite and anabolic processes. Postprandial sampling of plasma and white muscle tissue from short-term adapted postsmolt was conducted at 13 °C and 19 °C at 4, 8, 12, 16, 20 and 24 hours (h). The same samples were taken from long-term adapted big salmon at 12 °C, − 4, − 2, − 1, 0 h pre-prandially and 2, 3, 4 and 6 h post-prandially. While limited relationship between plasma ghrelin concentration and meal times was found for short term adapted postsmolt, clear ghrelin peaks were described for long term adapted salmon prior to the timing of anticipated meals. Possible explanations and consequences to experimental design in this area will be discussed. Postprandial FAA in plasma and white muscle from postsmolt was reduced at 19 °C compared to 13 °C and plasma levels peaked 8 h post-prandially. Muscle Igf1 mRNA expression levels were consistently higher at 13 °C than 19 °C, with no clear postprandial patters. In contrast, plasma IGF-1 concentration was relatively constant over time at 12 °C and 13 °C, but significantly declined from 20 h postprandially at 19 °C. GH receptor (ghr1) mRNA expression in muscle was unaffected by temperature, peaking 4 h post-prandially at both temperatures. This paper describes growth and appetite-regulating processes under conditions of normal and elevated temperature for Atlantic salmon, which is fundamental to our understanding of growth limitations inherent to high water temperature situations.

Quality improvement of fish feed by using insulin-like growth factor 1 as a potential indicator of amino acid deficiency in yellowtail

Abstract We developed a new method for improving the quality of fish feed, by using insulin-like growth factor 1 (IGF1) as a potential indicator of dietary amino acid deficiency in yellowtail ( Seriola quinqueradiata ). First, 7 dispensable amino acid solutions (alanine, asparagine, cysteine, glutamine, glycine, serine and proline) and 0.9% NaCl (control) solution were injected into the dorsal muscle of fish using a syringe. Twenty-four hours post-administration, white muscle tissue was collected from the fish opposite to the injection site and assayed for igf1 mRNA expression. Only fish injected with proline showed a significant increase in igf1 mRNA expression levels. Subsequently, a feeding trial was conducted with 3 different diets—the control diet; the control diet supplemented with 1% proline; and the control diets supplemented with 3% proline. Juvenile yellowtails were fed exclusively with 1 of the 3 diets until apparent satiation. The body weight and feed efficiency were significantly higher in fish fed with the 3% proline-supplemented diet than fish fed with the control diet. Further, after 3 weeks, the expression levels of igf1 in white muscle and liver were significantly higher in fish fed with the 3% proline-supplemented diet than in fish fed with the control diet. Our results suggest that igf1 mRNA expression in the white muscle tissue of yellowtails serves as a reliable indicator of amino acid deficiency. Additionally, the results of our feeding trial suggest that proline is required for juvenile yellowtail growth. Moreover, white muscle igf1 may represent a valuable tool for improving the quality of fish feed.

Expression of genes involved in the embryo–maternal interaction in the early-pregnant canine uterus

Although there is no acute luteolytic mechanism in the absence of pregnancy in the bitch, a precise and well-timed embryo-maternal interaction seems to be required for the initiation and maintenance of gestation. As only limited information is available about these processes in dogs, in this study, the uterine expression of possible decidualization markers was investigated during the pre-implantation stage (days 10-12) of pregnancy and in the corresponding nonpregnant controls. In addition, the expression of selected genes associated with blastocyst development and/or implantation was investigated in embryos flushed from the uteri of bitches used for this study (unhatched and hatched blastocysts). There was an upregulated expression of prolactin receptor (PRLR) and IGF2 observed pre-implantation. The expression of PRL and of IGF1 was unaffected, and neither was the expression of progesterone- or estrogen receptor β (ESR2). In contrast, (ESR1) levels were elevated during early pregnancy. Prostaglandin (PG)-system revealed upregulated expression of PGE2-synthase and its receptors, PTGER2 and PTGER4, and of the PG-transporter. Elevated levels of AKR1C3 mRNA, but not the protein itself, were noted. Expression of prostaglandin-endoperoxide synthase 2 (PTGS2) remained unaffected. Most of the transcripts were predominantly localized to the uterine epithelial cells, myometrium and, to a lesser extent, to the uterine stroma. PGES (PTGES) mRNA was abundantly expressed in both groups of embryos and appeared higher in the hatched ones. The expression level of IGF2 mRNA appeared higher than that of IGF1 mRNA in hatched embryos. In unhatched embryos IGF1, IGF2, and PTGS2 mRNA levels were below the detection limit.

Caffeine-induced fetal rat over-exposure to maternal glucocorticoid and histone methylation of liver IGF-1 might cause skeletal growth retardation

Several epidemiological investigations, including previous work by our laboratory, indicate that maternal caffeine consumption is associated with intrauterine growth retardation and impaired fetal length growth. Skeletal development is critical for length growth. In the present study, our goals were to determine the effects of prenatal caffeine exposures on fetal skeletal growth and to investigate the mechanisms associated with such effects. Pregnant Wistar rats were injected intragastrically with 120mg/kg of caffeine intragastrically each day from gestational days 11–20. Maternal prenatal caffeine exposure was associated with decreased fetal femur lengths and inhibited of synthesis of extracellular matrices in fetal growth plates Moreover, caffeine exposure significantly increased the levels of fetal blood corticosterone and decreased IGF-1mRNA expression levels in the liver and growth plate. The expression levels of IGF-1 signaling pathway components (IGF-1R, IRS-1, AKT1/2 and Col2A1) were also reduced. In addition, the results of chromatin immunoprecipitation assays indicated that caffeine exposure down-regulated histone methylation of fetal IGF-1 in the liver. These results suggest that prenatal caffeine exposure may inhibit fetal skeletal growth through a mechanism that is associated with increased fetal exposure to maternal glucocorticoids and results in lower IGF-1 signaling pathway activity. Taken together, these results raise important concerns regarding the skeletal growth toxicity of caffeine and potentially indicate the intrauterine origins of adult osteoporosis and osteoarthritis.

Abstract A48: Effect of constitutive IGF2 overexpression on tumorigenicity of human ovarian carcinoma cells

Abstract Introduction: We previously described the association of high tumoral IGF2 expression with reduced disease-free survival in epithelial ovarian tumors. The purpose of this study was to determine the effect of IGF2 overexpression on the tumorigenicity of human ovarian cancer cells. Methods: Expression constructs containing the full-length coding sequence of IGF2 or no insert (empty vector; EV) were made. HEY cells were stably transfected. Quantitative real-time PCR (of CDNA made from cell and xenograft RNA) and ELISA (of conditioned media) were performed to measure the IGF2 mRNA and protein expression levels, respectively. The relative IGF2 mRNA expression level was calculated by the 2−δδCt method with internal normalization to the cyclophilin B mRNA level. Tumorigenicity was evaluated in athymic nude mice by subcutaneous injection of 1×106 cells, with or without matrigel, followed by serial monitoring of in vivo tumor growth by digital caliper measurements. The two-tailed t-test was used to evaluate the significance of comparisons. Results: The IGF2 mRNA level was 5604 ±617 fold-change higher in HEY-IGF2 cells relative to HEY-EV cells (P&amp;lt;0.05). The IGF2 secreted protein level was 1852 ng/ml per 100,000 HEY-IGF2 cells, and was not detectable in the conditioned media of HEY-EV cells. After 4 weeks, the mean tumor volume of subcutaneous xenografts following cell injection without matrigel was 181 ±53 mm3 versus 52 ±15 mm3 for HEY-IGF2 and HEY-EV, respectively (P&amp;lt;0.05, N=10 per group); and for cells injected with matrigel, the mean tumor volume was 1608 ±330 mm3 versus 705 ±95 mm3 for HEY-IGF2 and HEY-EV xenografts, respectively (P&amp;lt;0.05, N=6 per group). The tumor growth rate differed significantly between the HEY-IGF2 and HEY-EV xenografts (P&amp;lt;0.01). In the HEY-IGF2 xenografts, overexpression of IGF2 relative to HEY-EV xenografts was maintained. Conclusions: Stable overexpression of IGF2 in HEY ovarian cancer cells and xenografts was achieved, and resulted in an increase in IGF2 protein secretion. IGF2 overexpression enhanced in vivo tumorigenicity of HEY ovarian cells, accompanied by an increased xenograft growth rate. Further evaluation is underway to fully characterize the mechanisms by which IGF2 overexpression promotes tumor growth. Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the Second AACR International Conference on Frontiers in Basic Cancer Research; 2011 Sep 14-18; San Francisco, CA. Philadelphia (PA): AACR; Cancer Res 2011;71(18 Suppl):Abstract nr A48.