Prevalence of thrombophilia-associated genetic risk factors in blood donors of a regional hospital in southern Brazil

ABSTRACT Introduction: Thromboembolic events occur due to an imbalance in the hemostasis and some factors associated with this condition can be inherited. In order to evaluate the frequency of genotypes considered to be common hereditary risk factors for thrombophilia associated with venous thrombosis (g.1691G>A and g.20210G>A) and hyperhomocysteinemia (g.677C>T and g.1298A>C), samples from voluntary healthy blood donors at the Hospital de Clínicas de Porto Alegre were tested. Methods: We examined 325 blood samples from blood donors collected from October 2017 to July 2018. Blood was collected on filter paper and the DNA was extracted for single nucleotide polymorphisms (SNPs) analysis using the qualitative real time polymerase chain reaction. Results: The calculated frequencies of each genetic variant in heterozygosity were 4% for the FV gene (g.1691G> A), 4% for the F2 gene (g.20210G> A) and 42% and 39% for methylenetetrahydrofolate reductase (MTHFR), g.677C>T and g.1298A>C, respectively. Only the genetic variants of MTHFR were found in homozygosity, with frequencies of 14% and 6% (g.677C>T and g.1298A>C), respectively. Discussion: Altogether, these results describe the frequencies of genetic variants associated with venous thrombosis and hyperhomocysteinemia in the analyzed group and are important to enhance our current knowledge about the genetic profiles of Brazilian blood donors.

Association between metabolic syndrome and euthyroid nodular goiter: a case-control study / Asociación entre el síndrome metabólico y el bocio nodular eutiroideo: un estudio de casos y controles

Abstract Background: Metabolic syndrome is a cluster of metabolic abnormalities and abdominal obesity; its pathophysiologic basis, insulin resistance, has been shown to act as agent in thyroid cell proliferation. Few studies analyze the relationship between metabolic syndrome and thyroid nodular disease, with a substantial knowledge gap. Objective: Determine the association between metabolic syndrome and nodular thyroid disease in a region with adequate iodine intake. Methods: Case-control study. A total of 182 patients referred to radiology to undergo thyroid ultrasonography due to suspicion of thyroid disease. Cases had at least one thyroid nodule greater than 3 mm (n= 91). Controls did not have evidence of thyroid nodules (n= 91). Results: Bivariate analysis showed a significant association between metabolic syndrome and the presence of thyroid nodule (OR 2.56, 95% CI: 1.41-4.66, p <0.05). Low levels of HDL (OR 2.81, 95% CI: 1.54-5.12, p <0.05) and impaired fasting glucose (OR 2.05, 95%CI 1.10 to 3.78, p <0.05) were significantly associated with the presence of thyroid nodule, independent of the presence of metabolic syndrome. Multivariate analysis maintained the association between metabolic syndrome and thyroid nodule with an OR of 2.96 (95%CI 1.47 to 5.95, p <0.05); similarly, the associations of low levels of HDL (OR 2.77, 95%CI 1.44 to 5.3, p <0.05) and impaired fasting glucose (OR 2.23, 95%CI 1.14 to 4.34, p<0.05) with thyroid nodule remained significant. Conclusion: The thyroid nodular disease is associated with increased risk of metabolic syndrome, specifically decreased HDL and impaired fasting glucose levels were the factors that increased association was found.

Resumen Antecedentes: el síndrome metabólico es un conjunto de anormalidades metabólicas y obesidad abdominal; Se ha demostrado que su base fisiopatológica, la resistencia a la insulina, actúa como agente en la proliferación de las células tiroideas. Pocos estudios analizan la relación entre el síndrome metabólico y la enfermedad nodular tiroidea, con una brecha de conocimiento sustancial. Objetivo: determinar la asociación entre el síndrome metabólico y la enfermedad tiroidea nodular en una región con una ingesta adecuada de yodo. Métodos: estudio de casos y controles. Un total de 182 pacientes remitidos a radiología para someterse a una ecografía tiroidea debido a la sospecha de enfermedad tiroidea. Los casos tenían al menos un nódulo tiroideo mayor de 3 mm (n = 91). Los controles no tenían evidencia de nódulos tiroideos (n = 91). Resultados: El análisis bivariado mostró una asociación significativa entre el síndrome metabólico y la presencia de nódulo tiroideo (OR 2.56, IC 95%: 1.41-4.66, p <0.05). Los niveles bajos de HDL (OR 2.81, IC 95%: 1.54-5.12, p <0.05) y glucosa en ayunas alterada (OR 2.05, IC 95% 1.10 a 3.78, p <0.05) se asociaron significativamente con la presencia de nódulo tiroideo, independiente de la presencia de síndrome metabólico. El análisis multivariado mantuvo la asociación entre el síndrome metabólico y el nódulo tiroideo con un OR de 2.96 (IC 95% 1.47 a 5.95, p <0.05); de manera similar, las asociaciones de niveles bajos de HDL (OR 2.77, IC 95% 1.44 a 5.3, p <0.05) y glucosa en ayunas alterada (OR 2.23, IC 95% 1.14 a 4.34, p <0.05) con nódulo tiroideo permanecieron significativas. Conclusión: la enfermedad nodular tiroidea se asocia con un mayor riesgo de síndrome metabólico, específicamente la disminución de HDL y los niveles de glucosa en ayunas alterados fueron los factores que aumentaron la asociación.

Expression of long non-coding RNA NR2F2-AS1 in human epithelial ovarian cancer / 肿瘤

Objective: To analyze the expression of long non-coding RNA (lncRNA) nuclear receptor subfamily 2 group F member 2 (NR2F2)-antisense RNA 1 (AS1) in human epithelial ovarian cancer. Methods: Agilent human lncRNA microarray was used to detect the expressions of lncRNA and mRNA in 3 pairs of ovarian cancer tissues and para-cancerous tissues. The abnormally expressions of mRNAs and lncRNAs were screened. Real-time fuorescent quantitative PCR was performed to detect the expression levels of lncRNA NR2F2-AS1, small nucleolar RNA host gene 4(SNHG4), LOC101927905 and OVE5-21006 in 22 ovarian cancer tissues and 10 benign ovarian cyst tissues. Moreover, the expression of NR2F2 (an antisense gene of NR2F2-AS1) in ovarian cancer SKOV3 cells after transfection with the specific siRNA targeting NR2F2-AS1 was detected by real-time fuorescent quantitative PCR. Results: Based on microarray analysis, a total of 33 403 lncRNAs and 20 351 mRNAs with different expression levels were screened from 3 pairs of epithelial ovarian cancer and paracancerous tissues. The expression level of NR2F2-AS1 in ovarian cancer tissues was lower than that in benign ovarian cyst tissues (P = 0.003 5). The expression level of NR2F2-AS1 in patients with ovarian cancer of stage I-II was lower than that of stage III-IV (P = 0.042 2). After the transfection with NR2F2-AS1 siRNA, the expression level of NR2F2 mRNA was downregulated in SKOV3 cells (P = 0.049 5). Conclusion: The expression profiles of lncRNAs in ovarian cancer tissues are significantly different from those in the corresponding para-cancerous tissues. NR2F2-AS1 is downregulated in ovarian cancer tissues, and is related to the pathological stage of ovarian cancer. After inhibiting the expression of NR2F2-AS1 in ovarian cancer SKOV3 cells, the expression level of its antisense gene NR2F2 is decreased.

Non-islet-cell tumor induced hypoglycemia:one case report and literature review / 中华内分泌代谢杂志

A patient with severe hypoglycemia due to insulin-like growth factor ( IGF)-IIsecreted by a giant solitary fibrous tumor of the pleura ( SFTP) was investigated through comprehensively reviewing his medical history and clinical records. The patient had severe hypoglycemia accompanied with significantly decreased serum insulin level. A solitary fibrous tumor of the pleura was found, and right pneumonectomy removed this giant tumor. Two years after the operation, the patient was fit and well with no further hypoglycemia episodes. Non-islet-cell tumor hypoglycemia should be considered in patients who have hypoglycemia episodes accompanied with significantly decreased serum insulin level.

Hipoglucemia inducida por carcinoma adrenal / Adrenal carcinoma induced hypoglycemia

El carcinoma suprarrenal es una neoplasia maligna infrecuente y de mal pronóstico. La presentación clínica más común es originada por la producción hormonal excesiva, mientras que el desarrollo de hipoglucemia sintomática es excepcional. Presentamos el caso de una mujer de 37 años que ingresó al hospital por síntomas de hipoglucemias graves, hipertensión arterial, hipopotasemia y amenorrea secundaria. En el laboratorio se halló hipoglucemia con insulina inhibida y niveles de andrógenos en rango tumoral. La tomografía computarizada (TC) de abdomen y pelvis mostró voluminosa formación heterogénea de aspecto sólido sin plano de clivaje con respecto al parénquima hepático e intenso realce con contraste. Luego de la extirpación de la masa retroperitoneal, evolucionó con valores de glucemia y potasemia normales, estabilizó la presión arterial y recuperó los ciclos menstruales.

Adrenal carcinoma is a rare malignancy of poor prognosis. The most common clinical presentation is secondary to hormone production, while the development of symptomatic hypoglycemia is exceptional. We report the case of a 37 year old-woman admitted to hospital with severe hypoglycemia, hypertension, hypokalemia and amenorrhea. In the laboratory we found hypoglycemia, with low insulin levels, and androgen levels in tumor range. CT of abdomen and pelvis showed a heterogeneous lesion of solid appearance without a cleavage plane relative to liver parenchyma, and intense contrast enhancement. Retroperitoneal mass was removed, and the patient evolved without complications, blood glucose and potassium were normalized, blood pressure stabilized and menstrual cycles recovered.

The role of HIF-1α, β-catenin, IMP3 and PCNA in proliferation of gastric cancer SGC-7901 cells / 肿瘤

Objective: To explore the effects of HIF-1α (hypoxia-inducible factor-1α), β-catenin, IMP3 (insulin-like growth factor II mRNA-binding protein 3) and PCNA (proliferating cell nuclear antigen) on proliferation of gastric cancer SGC-7901 cells. Methods: The pcDNA™ 6.2-GW/EmGFP-miR- β-catenin plasmid was transfected into SGC-7901 cells to establish stably transfected cell line miR- β-catenin-7901. In this experiment, four groups were designed: control group (SGC-7901 cells were cultured under normoxic conditions), hypoxia group (SGC-7901 cells were cultured under hypoxic conditions), transfection group (miR-β-catenin-7901 cells were cultured under normoxic conditions), and combination of transfection and hypoxia group (miR-β-catenin-7901 cells were cultured under hypoxic conditions). The proliferation of SGC-7901 cells in four groups was detected by colony formation assay and cell doubling time assay. The expression levels of HIF-1α, β-catenin, IMP3 and PCNA proteins of SGC-7901 cells in four groups were detected by Western blotting. Results: As compared with the control group, the number of colonies was increased and the cell doubling time was shortened in hypoxia group (P < 0.05). As compared with the hypoxia group, the number of colonies was reduced and the cell doubling time was prolonged in the combination of transfection and hypoxia group (P < 0.05). The expression levels of HIF-1α, β-catenin, IMP3 and PCNA proteins in hypoxia group were higher than those in the control group (P < 0.05). The expression levels of HIF-1α, β-catenin, IMP3 and PCNA proteins in the transfection group were lower than those in the control group (P < 0.05). The expression levels of HIF-1α, β-catenin, IMP3 and PCNA proteins in the combination of transfection and hypoxia group were lower than those in the hypoxia group (P < 0.05). Conclusion: The proliferation of gastric cancer SGC-7901 cells may be related to the interaction of HIF-1α and β-catenin, which resulted in the increased expression levels of IMP3 and PCNA. Copyright © 2013 by TUMOR.

Role of insulin-like growth factor II receptor in transdifferentiation of free silica-induced primary rat lung fibroblasts / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To study the role of insulin-like growth factor II receptor in free silica-induced transdifferentiation of primary rat lung fibroblasts.</p><p><b>METHODS</b>Rat lung fibroblasts and rat alveolar macrophages were cultured. A transdifferentiation model of primary rat lung fibroblasts was induced by free silica. Levels of α-SMA protein, IGF-IIR protein and mRNA were measured by immunocytochemistry, Western blot and RT-PCR, respectively. Lung fibroblasts were treated with Wortmannin.</p><p><b>RESULTS</b>The expression levels of α-SMA and IGF-IIR increased with the increasing free silica concentration and decreased after Wortmannin was used.</p><p><b>CONCLUSION</b>The IGF-IIR plays an important role in free silica-induced transdifferentiation of primary rat lung fibroblasts.</p>

Large-scale meta-analysis of genetic studies in ischemic stroke: Five genes involving 152,797 individuals.

BACKGROUND: Ischemic stroke descent has a genetic basis. Stroke represents a complex trait, which is assumed to be polygenic. On this topic, the role of a wide number of candidate genes has been investigated in stroke through association studies. MATERIALS AND METHODS: We performed a literature-based systematic review of genetic association studies in stroke abound several populations. Odds ratios (ORs) and 95% confidence intervals (CIs) were determined for each gene-disease association. Following a review of 300 manuscripts, five candidate gene variants were analyzed among 152,797 individuals (45,433 cases and 107,364 controls). RESULTS: For these five candidate genes studied, the prothrombin OR is 1,57 (1,23-2,89), the factor V Leiden OR is 1,43 (0,67-6,24), the mean OR of angiotensin I converting enzyme (ACE) insertion/deletion (I/D) polymorphism is 1,11 (1,02-1,25), the summary OR for the C677T variant of 5,10-methylenetetrahydrofolate reductase (MTHFR) is 1,23 (0,61-1,47) and the pooled OR for the apolipoprotein E (APOE) gene is 0,95 (0,77-1,14) . CONCLUSION: These data suggest the genetic associations of some genes with ischemic stroke and it is necessary to compete with other genes. Our findings could represent an epidemiological base and a useful tool to address further molecular investigations and to realize more detailed meta-analyses.

Clinical significance of serum IGF-2 and IGFBP-3 in perioperative period patients with ovarian cancer / 中国医师杂志

Objective To investigate the changes of serum insulin-like growth factor-2 (IGF-2) and insulin-link growth factor binding protein-3(IGFBP-3) in the patients with ovarian cancer before and after operation,and evaluate their clinical significance.MethodsThe contents of serum IGF-2 and IGFBP-3 in 82 patients with ovarian cancer were detected by ELISA before and after operation and compared with that in health controls.ResultsThe contents of IGF-2 in the patients before operation were significantly higher than that in control group[(101.5±22.2)ng/ml,(49.3±15.6)ng/ml vs (69.6±17.7)ng/ml,(23.9±11.3)ng/ml,t＝3.74,2.85,P＜0.05].The contents of IGFBP-3[(39.8±11.1)ng/ml]in the patients before operation were significantly lower than that in control group[(55.8±19.2)ng/ml](t′＝4.49,P＜0.05).There was significant correlation between the contents with lymph node metastasis and clinical stage[(107.5±24.0)ng/ml,(41.7±16.9)ng/ml vs (91.6±17.7)ng/ml,(56.9±19.1)ng/ml;(103.4±27.2)ng/ml,(50.2±16.6)ng/ml vs (86.6±12.3)ng/ml,(41.1±17.1)ng/ml,t＝2.83,2.37,2.48,3.32,P＜0.05).The contents of IGF-2 was significantly decreased,and IGFBP-3 was significantly increased in patients after radical operation [(86.6±12.3)ng/ml,(41.1±17.1)ng/ml vs (103.2±26.0)ng/ml,(45.3±14.9)ng/ml,t′＝3.46,t＝2.67,P＜0.05].But there was no significant difference on the level of IGF-2 and IGFBP-3 before and after palliative resection(P＞0.05).ConclusionsThe contents of serum IGF-2 and IGFBP-3 are closely related to tumor invasion,metastasis and clinical stage.Dynamic determination of the contents of serum IGF-2 and IGFBP-3 may be an important index for evaluation of invasion,metastasis,efficacy and prognosis for the patients with ovarian cancer.

Insulin-like growth factor- II induces mTOR pathway change in Rh1 sarcoma cells / 第二军医大学学报

Objective To observe the effect of insuiin-iike growth factor-II (IGF-2) on the growth and the mTOR pathway of Rh1 sarcoma cells. Methods Rh1 cells were cultured routinely, and were treated with IGF-2 at a final concentration of 10 ng/ml after starving with pure RPMI 1640 medium. The growth of cells was analyzed by flow cytometry 72 h after IGF-2 treatment. The phosphorylation of S6 and Akt (s473) proteins were examined by Western blotting analysis at 5, 10, 20, 30, and 60 min after IGF-2 treatment. Results IGF-2 treatment promoted the survival and inhibited the apoptosis of Rh1 cells compared with the control group. IGF-2 also increased the phosphorylation of S6 in a time-dependent manner. However, the phosphorylation of Akt(s473) was relatively stable in Rh1 cells. Conclusion IGF-2 can gradually increase the function of S6 in the mTOR pathway, and the function of Akt (s473) is kept relatively stable.

A Case of Gastrointestinal Stromal Tumor with Recurrent Hypoglycemia

Non-islet cell tumor induced hypoglycemia (NICTH) is attributable to overproduction of insulin-like growth factor-II (IGF-II) by solid tumors, and these tumors usually originate from mesenchymal or epithelial cells. Gastrointestinal stromal tumor (GIST) is a rare mesenchymal tumor and most commonly find in the gastrointestinal tract. It is usually expresses the CD117 (stem cell factor receptor, c-kit) detected by immunohistochemistry. Hypoglycemia associated with GIST is very rare and this has not yet been reported in Korea. A 72-year-old man was hospitalized due to frequent episodes of confusion. It was observed that non-hyperinsulinemic hypoglycemia, an elevated serum IGF-II level and a huge liver mass. The histology of liver mass showed c-kit (CD117) positivity, which was consistent with GIST, but it was surgically unresectable. He was treated with imatinib mesylate. Although he recieved palliative treatment, he still experienced intermittent fasting hypoglycemia. After 2 months, the serum IGF-II level was even higher than before. We changed imatinib mesylate to sunitinib malate and performed radiotherapy on the liver mass. Although the change of the liver mass was not significant, he did not suffer from hypoglycemia for three months afterwards.

High frequency of loss of allelic integrity at Wilms' tumor suppressor gene-1 locus in advanced breast tumors associated with aggressiveness of the tumor.

Background: The product of Wilms' tumor suppressor gene (WT1), a nuclear transcription factor, regulates the expression of the insulin-like growth factor (IGF) and transforming growth factor (TGF) systems, both of which are implicated in breast tumorigenesis and are known to facilitate angiogenesis. In the present study, WT1 allelic integrity was examined by Loss of Heterozygosity (LOH) studies in infiltrating breast carcinoma (n=60), ductal carcinoma in situ (DCIS) (n=10) and benign breast disease (n=5) patients, to determine its possible association with tumor progression. Methods: LOH at the WT1 locus (11p13) as determined by PCR-RFLP for Hinf1 restriction site and was subsequently examined for its association with intratumoral expression of various growth factors i.e. TGF-β1, IGF-II, IGF-1R and angiogenesis (VEGF and Intratumoral micro-vessel density) in breast carcinoma. Results: Six of 22 (27.2%) genetically heterozygous of infiltrating breast carcinoma and 1 of 4 DCIS cases showed loss of one allele at WT1 locus. Histologically, the tumors with LOH at WT1 were Intraductal carcinoma (IDC) and were of grade II and III. There was no correlation in the appearance of LOH at WT1 locus with age, tumor stage, menopausal status, chemotherapy status and lymph node metastasis. The expression of factor IGF-II and its receptor, IGF-1R was significantly higher in carcinoma having LOH at WT1 locus. A positive correlation was observed between the TGF-β1, VEGF expression and IMD scores in infiltrating carcinoma. Conclusions: The current study indicates that the high frequency of loss of allelic integrity at Wilms' tumor suppressor gene-1 locus in high-graded breast tumors is associated with aggressiveness of the tumor.

Correlation between preoperative serum levels of five biomarkers and relationships between these biomarkers and cancer stage in epithelial overian cancer / 부인종양

OBJECTIVE: To examine the correlation among the preoperative serum levels of five biomarkers presumed to be useful for early detection of epithelial ovarian cancer and evaluate the relationships between serum levels of these five biomarkers and epithelial ovarian cancer stage. METHODS: We analyzed 56 newly diagnosed epithelial ovarian cancer patients. Preoperative serum levels of leptin, prolactin, osteopontin (OPN), insulin-like growth factor-II, and CA-125 were determined by ELISA. We also examined the correlation between the serum levels of the biomarkers and ovarian cancer stage. Significant differences in the mean serum levels of two proteins, leptin and CA-125, were observed between stage subsets. RESULTS: There was a significant negative correlation between prolactin and leptin and a significant positive correlation between prolactin and OPN. Of the five biomarkers, only the mean serum CA-125 level showed a significant positive correlation with cancer stage (Spearman rho=0.24, p<0.01). OPN showed a marginally significant positive correlation with stage (Spearman rho=0.14, p=0.07). CONCLUSION: We demonstrated the relationship between five biomarkers in epithelial ovarian cancer. These tumor markers may be useful in screening for ovarian cancer, in characterizing disease states, and in developing therapeutic interventions targeting these marker proteins. Large-scale studies that include potential confounding factors and modifiers are necessary to more accurately define the value of these novel biomarkers in ovarian cancer.

PTEN/MMAC1 enhances the growth inhibition by anticancer drugs with downregulation of IGF-II expression in gastric cancer cells

PTEN/MMAC1 is a tumor suppressor gene that is mutated in a variety of advanced and metastatic cancers. Its major function is likely to be the phosphatase activity that regulates the phosphotidylinositol (PI)3-kinase/ Akt pathway. On the other hand, IGF system plays an important role in cell proliferation and cell survival via PI3-kinase/Akt and mitogen-activated protein kinase pathways in many cancer cells. To evaluate effect of PTEN on cell growth and IGF system in gastric cancer, human gastric adenocarcinoma cells (SNU-5 & -216) were transfected with human PTEN cDNA. Those PTEN- transfected gastric cancer cells had a lower proliferation rate than the pcDNA3-transfected cells. PTEN overexpression induced a profound decrease in the IGF-II and IGF-IR expression levels, and downregulation of IGF-II expression by PTEN was mediated through the regulation of the IGF-II promoter. In addition, a PI3-kinase inhibitor, LY294002, induced the downregulation of IGF-II expression. The PTEN-overexpressing SUN-5 and -216 cells were more sensitive to death induced by etoposide and adriamycin that induce DNA damage than the pcDNA3-transfected cells. These findings suggest that PTEN suppresses the cell growth through modulation of IGF system and sensitizing cancer cells to cell death by anticancer drugs.

The Tumor Suppressor Function of PTEN/MMAC1 through the Regulation of IGFs and IGFBPs / 소아과

PURPOSE: PTEN/MMAC1, a novel tumor suppressor gene, is mutated in a variety of advanced and metastatic cancers. It acts as a phosphatase, and thereby, regulates the PI-3 kinase/Akt pathway. In this study, we examined to evaluate the new function of anti-tumor effects of PTEN/MMAC1 through the regulation of the IGFs-IGFBPs in gastric cancer cells. METHODS: PTEN/MMAC1 was expressed in an adenovirus-mediated gene delivery system and introduced into gastric cancer cells(SNU-484 & SNU-668) in vitro. The effect of cell growth and the expression of IGFs and IGFBPs after Ad/PTEN infection was analyzed by MTT assay, RT-PCR and Western immunoblot. RESULTS: Ad/PTEN infected cells were inhibited in cell growth compared with moak cells and Ad/ LacZ infected cells. Overexpression of PTEN/MMAC1 induced decrease in expression of IGF-I, -II and IGF-I receptors which are known as growth prompt molecules in a variety of cancers. Of the six IGFBPs, the expressions of IGFBP-4 and IGFBP-6 were decreased in Ad/PTEN infected cells. In contrast, IGFBP-3 expression was markedly increased by up to 3-fold in Ad/PTEN infected cells. Overexpression of PTEN/MMAC1 inhibited the activation of Akt/PKB pathway, but had no effect on the MAPK pathway. CONCLUSION: These findings suggest that the tumor suppressor function of PTEN/MMAC1 is, at least in part, mediated through the down-regulation of IGF-I abd IGF-II, and up-regulation of IGFBP-3 in gastric cancer cells by the inhibition of PI-3 kinase pathway.

Study of the Mechanism for the Growth Inhibitory Effects of Conjugated Linoleic Acid on Caco-2 Colon Cancer Cells

Conjugated linoleic acid (CLA) is a group of positional and geometric isomers of linoleic acid (LA) and exhibits anticarcinogenic activity in a variety of animal models. We have previously observed that CLA inhibited the growth of Caco-2 cells, a human colon adenocarcinoma cell line. The present study was performed to determine whether the growth inhibitory effect of CLA is related to change in secretion of IGF- II and/or IGF-binding proteins (IGFBPs) that have been shown to regulate Caco-2 cell proliferation by an autocrine mechanism. Cells were incubated in serum-free medium with various concentrations of CLA or linoleic acid (LA). Immunoblot analysis of 24-hours, serum-free, conditioned medium using a monoclonal anti-IGF-IIantibody revealed that Caco-2 cells secreted both mature 6,500 Mr and higher Mr forms of pro IGF-II. The levels of pro IGF-II and mature IGF-IIwere decreased by 43+/-2% and 53+/-6%, respectively by treatment with 50 micrometer CLA. LA slightly increased pro IGF- II levels. Results from Northern blot analysis showed that CLA decreased IGF-II mRNA levels at 50 micrometer concentration suggesting that CLA regulation of IGF-II protein expression occurs partly at the transcriptional level. Ligand blot analysis of conditioned media using 1251-IGF-II revealed that CLA slightly decreased IGFBP-2 levels and increased IGFBP-4 levels. We confirmed our previous results that CLA inhibited cell growth in a dose-dependent manner but LA slightly increased cell growth. Exogenous IGF-II mitigated the growth inhibitory effect of CLA. These results indicate that the growth inhibitory effect of CLA may be at least in part mediated by decreasing IGF-II and IGFBP-2 secretion and increasing IGFBP-4 secretion in Caco-2 cells.

Clinical Efficacy of Serum Insulin-like Growth Factor- I (IGF-I), IGF-II, and IGF Binding Protein-3 (IGFBP-3) in Predicting the Prognosis of In Vitro Fertilization and Embryo Transfer / 대한산부인과학회잡지

OBJECTIVE: To evaluate the clinical efficacy of serum insulin-like growth factor-I (IGF-I), IGF-II, and IGF binding protein-3 (IGFBP-3) levels in predicting the prognosis of in vitro fertilization and embryo transfer (IVF-ET). MATERIALS AND METHODS: In 84 patients undergoing IVF-ET, serum levels of IGF-I , IGF-II, and IGFBP-3 were measured using immunoradiometric assay (IRMA) before the gonadotropin administration and on the hCG day of controlled ovarian hyperstimulation (COH). Serum levels of IGFs and IGFBP-3, and the outcomes of IVF-ET were retrospectively analyzed and compared between the pregnant (n=18) and nonpregnant (n=66) groups. RESULTS: There were no significant differences in the outcomes of COH such as total dosage of gonadotropins used, duration of COH, serum estradiol (E2) level on the hCG day, numbers of oocytes retrieved and fertilized, and number of embryos transferred between the pregnant and nonpregnant groups. No differences were found in serum levels of IGF- I , IGF-II, and IGFBP-3, and their ratios before the gonadotropin administration and on the hCG day of COH. Basal serum level of IGF-II was lower with the borderline significance in the pregnant group (796.9+/-159.6 vs. 908.9+/-338.9 ng/ml, p=0.056). The ratio of change in IGF-I to that of IGF-II was significantly higher in the pregnant group (0.066+/-0.489 vs. -0.582+/-2.091, p=0.045). CONCLUSION: Even though basal serum level of IGF-II was lower and the ratio of changes in IGF-I to IGF-II was higher in the pregnant group, serum levels of IGF-I , IGF-II, and IGFBP-3 do not seem to predict the prognosis of IVF-ET. Further investigations are necessary in a larger group of patients to elucidate the clinical efficacy of serum IGFs and IGFBPs levels in predicting the prognosis of IVF-ET.

Transforming Growth Factor-alpha Regulation of Insulin-like Growth Factor-II, Insulin-like Growth Factor Binding Protein-1 and 3 secretion and Epidermal Growth Factor receptor expression in human luteinized granulosa cells / 대한산부인과학회잡지

OBJECTIVE: To investigate the effects of transforming growth factor (TGF)-alpha on insulin-like growth factor (IGF)-II, insulin-like growth factor binding protein (IGFBP)-1, and 3 secretion and epidermal growth factor (EGF) receptor expression in cultured human luteinized granulosa cells. MATERIALS AND METHODS: Human luteinized granulosa cells were obtained from follicular fluid by transvaginal oocyte aspiration from infertile patients undergoing in vitro fertilization and cultured for 72 hours with TGF-alpha at concentration of 1.0, 10.0, 100.0 ng/ml. The luteinized granulosa cells not treated with TGF-alpha served as control. The secretion of IGF-II, IGFBP-1 and 3 were determined in conditioned media by immunoradiometric assay (IRMA) and reverse transcription-polymerase chain reaction (RT-PCR) was performed for EGF receptor mRNA expression. RESULTS: The cell numbers of 1.0 and 10.0 ng/ml supplement groups were significantly decreased compared to control (p<0.05, p<0.05, respectively), although the cell viabilities were similar in all groups. IGF-II levels were significantly higher in TGF-alpha treatment group at 1.0 and 10.0 ng/ml (p<0.01, p<0.01, respectively), but lower in 100.0 ng/ml (p<0.01). However, the concentrations of IGFBP-1, and 3 per one granulosa cell in each group were no statistically significant differences among the groups. The mRNA concentration of EGF receptor in cultured human luteinized granulosa cells were not significantly different among the groups. CONCLUSION: This study suggests that TGF-alpha regulate intrafollicular bioavailable IGF-II levels, by which TGF-alpha might involved luteinizations. However, TGF-alpha may not directly regulate EGF receptor mRNA expression in cultured human luteinized granulosa cells.

Expression of insulin-like growth factor I and II in various regionally important solid tumors.

Insulin-like growth factor I (IGF-I) may thwart a T cell response to several cancers, since suppression of IGF-I expression using an episome transcribing an antisense RNA into rat glioma and murine teratocarcinoma, which are known to actively express IGF-I, resulted in tumor regression and CD8+ T cell infiltration.1,2,3,4 This finding led to the gene therapy protocol for human glioblastoma multiforme, a known IGF-I over-expressing tumor, using the same strategy in 1993. The hypothesis that cancers which over-express IGF-I can be cured by this episome has led us to search for other cancers which actively produce IGF-I. Many cancers were reported to express IGF-I4, but most of the geographically important tumors in Thailand had not been studied for the expression of IGF-I. Our immediate goal was to survey the leading human cancers in the Tumor Registry of Siriraj Hospital (i.e., non-small cell lung cancer, cholangiocarcinoma, hepatocellular carcinoma, papillary thyroid carcinoma, nasopharyngeal carcinoma) including all stages of astrocytomas for the expression of IGF-I and IGF-II (a homologue of IGF-I). Tumor tissue collected from paraffin blocks deposited at the Department of Pathology, Siriraj Hospital was screened for the presence of the predominant form of IGF's (IGF-I or -II). Tumor tissues that express any specific form of IGF's more than adjacent wild-type cells are potential candidates for an antisense gene therapy against the expression of that specific form of IGF. Paraffin sections containing these cancer cells and adjacent wild-type cells were examined by immunohistochemical technique using mouse IgG-1 antihuman IGF-I, or antihuman IGF-II as primary antibodies. Normal liver tissue, known as the largest producer of both forms of IGF's, was used as a positive control. The staining pattern of both IGF-I and IGF-II in the liver was highest in the cytoplasm of normal hepatocytes. The majority of the tumors except for papillary thyroid carcinoma and nasopharyngeal carcinoma strongly expressed IGF-I. Only a few samples of the tissues studied expressed a low level of IGF-II, if any. This raises a further working proposal that the blockade of IGF-I synthesis in these tumors may help to delay the tumor progression so that an effective immune response can be established and destroy the tumor.

Expression of cancer gene products of IGF-Ⅱ, IGF-Ⅱ receptors, CSF-1 receptors in primary hepatic cancer and non-cancerous liver tissue / 第三军医大学学报

The expression of cancer gene products of insuline-like growth factor Ⅱ (IGF-Ⅱ),IGF-Ⅱ receptors (IGF-Ⅱ-R),and colony-stimulating factor 1 receptors (CSF-1-R) /c-fms in 17 cases of human primary hepatic cancer,the non-cancerous liver tissue adjacent to the cancer,and normal liver tissue was studied with immunocytochemistry (ABC),Western blot and Northern blot techniques.It was found that the expression of IGF-Ⅱ,IGF-Ⅱ-R and CSF-1-R was significantly higher in the cancers than in normal liver tissues,and the expression of IGF-Ⅱ and IGF-Ⅱ-R was higher in the non-cancerous liver tissues than in the cancers.It was noteworthy that the expression of IGF-Ⅱ in both the cancerous and non-cancerous hepatic tissues was characterized by its fetal type.However the expression of CSF-1-R was distinctly higher in the cancers than in the non-cancerous hepatic tissue.These findings,the authors believe,imply that the aberrant overexpression of IGF-Ⅱ,IGF-Ⅱ-R and CSF-l-R might be related to the mechanism of auto-crine-stimulated growtth of the cells of human primary hepatic cancer.