Overexpression Of Insulin-like Growth factor-II Research Articles

Insulin-like growth factor-II overexpression accelerates parthenogenetic stem cell differentiation into cardiomyocytes and improves cardiac function after acute myocardial infarction in mice

BackgroundParthenogenetic stem cells (PSCs) are a promising source of regenerated cardiomyocytes; however, their application may be limited without a paternal genome. Insulin-like growth factor-II (IGF-II), a paternally expressed growth hormone, is critical in embryonic differentiation. This study investigated whether forced expression of IGF-II in PSCs can accelerate their differentiation.MethodsOverexpression and re-knockdown of IGF-II in PSCs were performed to investigate the role of IGF-II in PSC differentiation. The derivatives of PSCs with different IGF-II manipulations were transplanted into infarcted murine hearts to investigate the role of IGF-II in cardiomyocyte differentiation in vivo.ResultsData showed that the expression of cardiac troponin T and troponin I in IGF-II-PSC outgrowths preceded that of parental PSC outgrowths, suggesting that IGF-II can accelerate PSC differentiation into cardiac lineage. Overexpression of IGF-II accelerated PSC differentiation towards cardiomyocytes while inhibiting PSC proliferation via the IGF-II/IGF1R signaling. Similar to that observed in cardiac marker expression, on differentiation day 24, IGF-II-PSCs showed PCNA and cyclin D2 expression comparable to juvenile mouse cardiomyocytes, showing that IGF-II-PSCs at this stage possess differential and proliferative properties similar to those of juvenile cardiomyocytes. Moreover, the expression pattern of cardiac markers in IGF-II-overexpressing PSC derivatives resembled that of juvenile mouse cardiomyocytes. After transplantation into the infarcted mouse hearts, IGF-II-PSC-derived cardiomyocytes displayed significant characteristics of mature cardiomyocytes, and IGF-II-depletion by shRNA significantly reversed these effects, suggesting the critical role of IGF-II in promoting cardiomyocyte maturation in vivo. Furthermore, IGF-II-overexpressing PSC derivatives reduced collagen deposition and mitochondrial damage in the infarcted areas and improved cardiac function. The re-knockdown of IGF-II could counteract these favorable effects of IGF-II.ConclusionsThese findings suggest that the ectopic expression of IGF-II accelerates PSC differentiation into the cardiac lineage and promotes cardiomyocyte maturation. The underlying process includes the IGF-II/IGF1R signaling, which is involved in the suppressive effect of IGF-II on PSC proliferation. Moreover, transplanting IGF-II-overexpressing PSC derivatives into the infarcted heart could reduce collagen deposition and improve mitochondria biogenesis and measurements of cardiac function, highlighting the importance of IGF-II in the application of PSCs in cardiac regeneration.

Antisense oligonucleotides directed against insulin-like growth factor-II messenger ribonucleic acids delay the progress of rat hepatocarcinogenesis

Background:Hepatocellular carcinoma (HCC) is a multistep complex process, caused by many of genetic alteration. Insulin-like growth factors and their receptor have been widely implicated to HCC. Insulin-like growth factor-II (IGF-II) is a mitogenic polypeptide, found in various fetal and neonatal tissues of humans and rats and expresses in HCC. Here we investigated anticancer potential of phosphorothioate antisense oligonucleotides (ASOs) against three coding exons (exon-1/exon-2/exon-3) of IGF-II messenger ribonucleic acid in rat hepatocarcinogenesis model.Materials and Methods:During diethylnitrosamine and 2-acetylaminofluorene induced hepatocarcinogenesis, rats were treated with ASOs. Various biochemical and histological studies were conducted.Results:About 40% of carcinogen treated rats, which received two oligomers (against exon-1 or-3) did not show any hepatic lesion, hyperplastic nodule or tumor and remaining 60% of those rats showed lesion incidence and had about 59% and 55% reductions in the numbers of hepatic altered foci, respectively. Reductions in the total lesion-area when compared with carcinogen control rats were 64% and 53%, respectively for the animals treated with carcinogen and received the ASOs against exon-1/-3. Fluorescein isothiocyanate-labeled ASO reached in the hepatocytes in 2 h. No predominant IGF-II overexpression was observed in case of rats treated with the two ASOs. Treatment of the antisense IGF-II oligomers in carcinogen treated rats show better hepatocellular integrity along with several preneoplastic/neoplastic marker isoenzyme/enzyme modulations.Conclusions:Two of the three antisense oligomer-types effectively controlled IGF-II overexpression, causing the delay of the development and/or progress of hepatic cancer in rats.

Increased Intragenic IGF2 Methylation is Associated with Repression of Insulator Activity and Elevated Expression in Serous Ovarian Carcinoma

Overexpression of insulin-like growth factor-II (IGF2) is a prominent characteristic of many epithelial ovarian malignancies. IGF2 imprinting and transcription are regulated in part through DNA methylation, which in turn regulates binding of the insulator protein CTCF within the IGF2/H19 imprint center. We have shown that IGF2 overexpression in ovarian cancer is associated with hypermethylation of CTCF binding sites within the IGF2/H19 imprint center. The aim of this study was to investigate the methylation and binding capacity of a novel putative CTCF binding motif located intragenic to IGF2 and determine how this relates to IGF2 expression. Among 35 primary serous epithelial ovarian cancer specimens, methylation of two CpGs, including one within the core binding motif and another adjacent to this motif, was higher in the 18 cancers with elevated IGF2 expression versus 10 with low expression (average 68.2 versus 38.5%; p < 0.0001). We also found that the CpG site within the CTCF binding motif is hypermethylated in male gametes (>92%; average 93.2%; N = 16). We confirmed binding of CTCF to this region in ovarian cancer cells, as well as the paralog of CTCF, Brother Of the Regulator of Imprinted Sites (BORIS), which is frequently overexpressed in cancers. The unmethylated CTCF binding motif has insulator activity in cells that express CTCF or BORIS, but not in cells that express both CTCF and BORIS. These intragenic CpG dinucleotides therefore comprise a novel paternal germline imprint mark and are located in a binding motif for the insulator protein CTCF. Methylation of the CpG dinucleotides is positively correlated with IGF2 transcription, indicating that increased methylation represses insulator function. These combined results suggest that methylation and CTCF binding at this region play important roles in regulating the level of IGF2 transcription. Our data have revealed a novel epigenetic regulatory element within the IGF2/H19 imprinted domain that is highly relevant to aberrant IGF2 expression in ovarian malignancies.

Correlation between epigenetic alterations in the insulin growth factor-II gene and hepatocellular carcinoma

To investigate whether epigenetic alterations in the insulin-like growth factor-II (IGF-II) gene that cause differential transcription or expression are correlated with onset and severity of hepatocellular carcinoma (HCC). Patient-matched specimens of HCC, paracancerous, and non-cancerous tissues were collected from 40 primary liver cancer patients. Epigenetic alterations in the promoter (P3) sequence of the IGF-II gene were analyzed by methylation-specific PCR (MSP) and IGF-II transcription was measured by RT-PCR. IGF-II protein expression and clinicopathological features were assessed by immunohistochemistry and microscopic observation. The rate of IGF-II P3 methylation was significantly lower in HCC tissues (0%) than in paracancerous tissues (vs. 47.5%; x2 = 24.918, P less than 0.001) and non-cancerous tissues (vs. 100%; x2 = 80.000, P less than 0.001). IGF-II mRNA expression was significantly higher in HCC tissues (100%) than in paracancerous tissues (vs. 52.5%; x2 = 24.918, P less than 0.001) and non-cancerous tissues (vs. 0%; x2 = 80.000, P less than 0.001). IGF-II protein expression was significantly higher in HCC tissues (82.5%) than in paracancerous tissues (vs. 45.0%; x2 = 12.170, P less than 0.001) and non-cancerous tissues (vs. 0%; x2 = 56.170, P less than 0.001). IGF-II overexpression in HCC was significantly associated with degree of differentiation, extent of infiltrated serosa, size of tumor, and HBV-positive infection status. Epigenetic alterations in the IGF-II gene regulate its transcription and expression and are closely associated with HCC development and progression.

Hypomethylated P4 Promoter Induces Expression of the Insulin-Like Growth Factor-II Gene in Hepatocellular Carcinoma in a Chinese Population

The expression of human insulin-like growth factor-II (IGF-II) is regulated by the activation of four promoters (P1-P4) acting in a development-dependent, tissue-specific manner. IGF-II overexpression associated with P3 and P4 activation is observed in animal and human hepatocarcinogenesis. We correlated P4 epigenetic alteration with P4 transcript activation and clinicopathologic features. We analyzed P4 epigenetic alteration using methylation-specific PCR in 34 hepatocellular carcinoma (HCC) specimens, 34 matched adjacent nontumor specimens, and 8 normal adult liver specimens. The data were correlated with activation of P4 transcription by using reverse transcription-PCR. Epigenetic alteration was compared with patients' clinicopathologic features. Compared with normal liver tissue, hypomethylation of P4 CpG islands was significantly more frequent in HCC (P = 0.03) and matched tissues (P = 0.047). P4 mRNA levels in HCC with unmethylated alleles were significantly higher than in HCC without unmethylated alleles (P = 0.001); P4 mRNA levels in matched nontumor tissues with unmethylated alleles were significantly higher than in matched nontumor tissues without unmethylated alleles (P = 0.005). P4 hypomethylation in HCC was associated with portal vein tumor embolus (P = 0.017) and poorer tumor differentiation (P = 0.025). These findings suggest that IGF-II P4 hypomethylation may be an early and frequent event and that it may contribute to P4 transcription expression activation during the transformation of a premalignant liver lesion to HCC. Furthermore, aberrant hypomethylation of P4 CpG islands not only may play an important role during hepatocarcinogenesis but might also be a useful biomarker for poor prognosis of patients with HCC.

A case of colorectal carcinoma in adenoma analyzed by a cDNA array

A 68-year-old woman presenting with bloody stools and anemia was referred to our hospital. Colonoscopy showed a Is-type tumor of 45 mm in diameter in the cecum and three Is-type tumors in the ascending colon. Ileocecal resection with regional lymph node dissection was performed. Microscopically, the large tumor consisted of a well-differentiated adenocarcinoma with a tubulovillous adenoma (TVA) component (carcinoma in adenoma). Some carcinoma (CA) cells had invaded the submucosal layer, but the lymph nodes were negative for malignancy. The other three polyps were diagnosed as TVAs. Because her family history fulfilled the Amsterdam criteria II for hereditary non-polyposis colorectal cancer (HNPCC), genetic analysis was performed. All of the four tumor tissues were classified as microsatellite stable (MSS) according to the National Cancer Institute guideline for analysis of microsatellite instability (MSI). K-ras mutation was detected in both CA and TVA lesions of the carcinoma in adenoma. To clarify relevant alterations of gene expression associated with adenoma-carcinoma progression, the gene expression profiles of these tumor tissues were analyzed by a cDNA array. Although the gene expression profiles were similar, insulin-like growth factor-II (IGF-II) was expressed most differentially in CA and TVA tissues. The results were further substantiated by comparison of the gene expression profiles of CA and TVA lesions of the carcinoma in adenoma. The results suggest that overexpression of IGF-II played an important role in the progression of adenoma to carcinoma in this patient.

Adrenal incidentalomas.

The term adrenal incidentaloma refers to an adrenal mass occasionally and unexpectedly discovered by an abdominal imaging procedure performed for reasons a priori unrelated to adrenal dysfunction. The prevalence of adrenal incidentalomas as discovered by computed tomographic scan examination is estimated to be between 1% and 4%. The vast majority of these lesions are of adrenocortical origin, most often adenomas. Identification of steroid or catecholamine-secreting tumors is important but usually solved with appropriate endocrinologic investigations. A difficult problem, however, is to distinguish between benign and malignant primary or secondary tumors. Size less than 4 cm and an unenhanced computed tomographic attenuation under 10 Hounsfield Units (HU) are findings in favor of a benign adrenocortical adenoma, as is a positive NP 59 scintigraphic examination. The pathogenesis of adrenal tumors is not well understood. However, alterations of the cyclic AMP signalling pathway have recently been observed in benign adrenocortical lesions and molecular defects associated with insulin-like growth factor-II overexpression in malignant adrenocortical tumors.

Overexpression of Insulin-like Growth Factor-II in Mouse Embryonic Stem Cells Promotes Myogenic Differentiation

Embryonic stem (ES) cells derived from androgenetic or parthenogenetic mouse embryos are important tools for studying the roles of imprinted genes in early development. Androgenetic ES cells have been shown to preferentially differentiate into the myogenic lineage both in vitro and after formation of teratocarcinomas in vivo. To clarify if the maternally imprinted Igf2 gene which is expected to be overexpressed in androgenetic ES cells is sufficient to induce myogenic differentiation, R1 ES cells were transfected with human IGF-II expression vectors. Stable ES cell clones exhibiting human IGF-II mRNA and protein expression were studied vs ES cell clones without IGF-II overexpression in a standard in vitro differentiation system involving culture in “hanging drops” and observation of differentiation of the recovered embryoid bodies (EBs). EBs derived from IGF-II overexpressing ES cells showed stimulated myogenic differentiation evident by the appearance of myoblasts already 3 days after plating and by higher levels of skeletal muscle-specific transcripts (myf5, myoD, myogenin) at earlier stages. Our study demonstrates for the first time that overexpression of IGF-II enhances and accelerates myogenic differentiation of ES cells, which has implications for ES cell-derived tissue engineering.

Transgenic mice overexpressing insulin-like growth factor-II in beta cells develop type 2 diabetes.

During embryonic development, insulin-like growth factor-II (IGF-II) participates in the regulation of islet growth and differentiation. We generated transgenic mice (C57BL6/SJL) expressing IGF-II in beta cells under control of the rat Insulin I promoter in order to study the role of islet hyperplasia and hyperinsulinemia in the development of type 2 diabetes. In contrast to islets from control mice, islets from transgenic mice displayed high levels of IGF-II mRNA and protein. Pancreases from transgenic mice showed an increase in beta-cell mass (about 3-fold) and in insulin mRNA levels. However, the organization of cells within transgenic islets was disrupted, with glucagon-producing cells randomly distributed throughout the core. We also observed enhanced glucose-stimulated insulin secretion and glucose utilization in islets from transgenic mice. These mice displayed hyperinsulinemia, mild hyperglycemia, and altered glucose and insulin tolerance tests, and about 30% of these animals developed overt diabetes when fed a high-fat diet. Furthermore, transgenic mice obtained from the N1 backcross to C57KsJ mice showed high islet hyperplasia and insulin resistance, but they also developed fatty liver and obesity. These results indicate that local overexpression of IGF-II in islets might lead to type 2 diabetes and that islet hyperplasia and hypersecretion of insulin might occur early in the pathogenesis of this disease.

Overexpression of insulin-like growth factor-II in transgenic mice is associated with pancreatic islet cell hyperplasia.

We have used an insulin-like growth factor (IGF)-II transgenic mouse model in which mouse IGF-II is widely overexpressed, resulting in increased fetal size and selective organ overgrowth, to investigate the effects on the development of the endocrine pancreas. Fetuses examined on day 19.5-20 of gestation had significantly elevated circulating levels of IGF-II, compared with control mice. The pancreatic islets in transgenic animals were of irregular shape and had a mean area five times greater than in controls, whereas the mean number of islets per tissue section was not altered. The size of individual endocrine cells was not altered. Although the islets in animals expressing the IGF-II transgene were considerably larger, immunohistochemistry for insulin and glucagon showed that the relative proportion of beta-cells was significantly less, and that of alpha-cells was higher. Normal islet morphology was disrupted, with alpha-cells appearing in small groups within the islets, as well as on the periphery, whereas beta-cells were often seen at the edge of the islets. Twice as many islet cells (21.9% vs. 11.4%) were involved in cell replication, detected by the presence of immunoreactive proliferating cell nuclear antigen, in pancreata from transgenic mice vs. controls, whereas the number of cells undergoing apoptosis was significantly reduced. Abundant IGF-II messenger RNAwas found within the islets of transgenic animals by in situ hybridization, and the relative area of islets demonstrating immunoreactive IGF-II was significantly greater. Immunoreactive IGF-I was much less abundant and was further reduced in islets of transgenic animals. The area of islets immunopositive for IGF binding protein-2 was unaltered. Despite the presence of islet hyperplasia, circulating insulin levels and serum glucose levels were not significantly different between transgenic and control mice. These results show that an overexpression of IGF-II in fetal life has a profound effect on islet morphology and causes islet hyperplasia while reducing the attrition of islet cells by apoptosis.

Overexpression of Insulin-Like Growth Factor-II in Transgenic Mice Is Associated with Pancreatic Islet Cell Hyperplasia

We have used an insulin-like growth factor (IGF)-II transgenic mouse model in which mouse IGF-II is widely overexpressed, resulting in increased fetal size and selective organ overgrowth, to investigate the effects on the development of the endocrine pancreas. Fetuses examined on day 19.5–20 of gestation had significantly elevated circulating levels of IGF-II, compared with control mice. The pancreatic islets in transgenic animals were of irregular shape and had a mean area five times greater than in controls, whereas the mean number of islets per tissue section was not altered. The size of individual endocrine cells was not altered. Although the islets in animals expressing the IGF-II transgene were considerably larger, immunohistochemistry for insulin and glucagon showed that the relative proportion of β-cells was significantly less, and that of α-cells was higher. Normal islet morphology was disrupted, with α-cells appearing in small groups within the islets, as well as on the periphery, whereas β-cells were often seen at the edge of the islets. Twice as many islet cells (21.9% vs. 11.4%) were involved in cell replication, detected by the presence of immunoreactive proliferating cell nuclear antigen, in pancreata from transgenic mice vs. controls, whereas the number of cells undergoing apoptosis was significantly reduced. Abundant IGF-II messenger RNA was found within the islets of transgenic animals by in situ hybridization, and the relative area of islets demonstrating immunoreactive IGF-II was significantly greater. Immunoreactive IGF-I was much less abundant and was further reduced in islets of transgenic animals. The area of islets immunopositive for IGF binding protein-2 was unaltered. Despite the presence of islet hyperplasia, circulating insulin levels and serum glucose levels were not significantly different between transgenic and control mice. These results show that an overexpression of IGF-II in fetal life has a profound effect on islet morphology and causes islet hyperplasia while reducing the attrition of islet cells by apoptosis.

Overexpression of insulin-like growth factor-II induces accelerated myoblast differentiation.

Previous studies have shown that exogenous insulin-like growth factors (IGFs) can stimulate the terminal differentiation of skeletal myoblasts in culture and have established a correlation between the rate and the extent of IGF-II secretion by muscle cell lines and the rate of biochemical and morphological differentiation. To investigate the hypothesis that autocrine secretion of IGF-II plays a critical role in stimulating spontaneous myogenic differentiation in vitro, we have established C2 muscle cell lines that stably express a mouse IGF-II cDNA under control of the strong, constitutively active Moloney sarcoma virus promoter, enabling us to study directly the effects of IGF-II overproduction. Similar to observations with other muscle cell lines, IGF-II overexpressing myoblasts proliferated normally in growth medium containing 20% fetal serum, but they underwent enhanced differentiation compared with controls when incubated in low-serum differentiation medium. Accelerated differentiation of IGF-II overexpressing C2 cells was preceded by the rapid induction of myogenin mRNA and protein expression (within 1 h, compared with 24-48 h in controls) and was accompanied by an enhanced proportion of the retinoblastoma protein in an underphosphrylated and potentially active form, by a marked increase in activity of the muscle-specific enzyme, creatine phosphokinase, by extensive myotube formation by 48 h, and by elevated secretion of IGF binding protein-5 when compared with controls. These results confirm a role for IGF-II as an autocrine/paracrine differentiation factor for skeletal myoblasts, and they define a model cell system that will be useful in determining the biochemical mechanisms of IGF action in cellular differentiation.

Hepatic Overexpression of Insulin-like Growth Factor-II in Adulthood Increases Basal and Insulin-stimulated Glucose Disposal in Conscious Mice

The physiological role of circulating insulin-like growth factor-II (IGF-II) in adult humans is poorly understood. We recently generated an IGF-II transgenic murine model of persistent IGF-II production (plasma IGF-II approximately 30-fold increased above normal) through over-expression of the transgene driven by the major urinary protein promoter (Rinderknecht, E., and Humbel, R. E. (1978) J. Biol. Chem. 269, 13779-13784). To determine whether in vivo insulin action is improved in these transgenic mice, we performed euglycemic insulin (18 milliunits/kg.min) clamp studies in conscious IGF-II transgenic and in age- and weight-matched control mice. Plasma glucose and insulin concentrations were significantly lower in the IGF-II transgenic compared with both control grouoff Despite decreased plasma glucose concentration, basal hepatic glucose production (HGP) and glucose clearance were increased. During the insulin clamp studies in IGF-II transgenic mice compared with control mice (a) the rates of glucose infusion and glucose uptake were increased by approximately by 65 and approximately 55%, respectively; (b) glycolysis was increased by approximately 12% while glycogen synthesis was approximately 2-fold higher; (c) while the suppression of plasma free fatty acid was similar, the increment in plasma lactate concentration was significantly higher; (d) although HGP was similarly inhibited by insulin, phosphoenolpyruvate gluconeogenesis was enhanced and accounted for a larger portion of HGP (64% versus approximately 40% in control mice). Our data suggest that the persistence of circulating IGF-II in adult mice to levels commonly observed in adult humans (50-70 nM) causes a marked improvement in peripheral (skeletal muscle) insulin action, which is not due to changes in body composition. These results suggest that circulating IGF-II may exert a regulatory role on insulin sensitivity and body composition in humans.

Hepatitis C virus replication is associated with expression of transforming growth factor-α and insulin-like growth factor-II in cirrhotic livers

The molecular role of hepatitis C virus (HCV) in liver disease has yet to be clarified. In this study, we analyzed the relationship of HCV replication with mRNA expression of growth factors and mutation of tumor suppressor gene, ie, transforming growth factor-beta 1 (TGF-beta 1), which promotes cirrhotic changes; TGF-alpha, insulin-like growth factor-II (IGF-II), which are both related to hepatocyte transformation; and tumor suppressor gene p53, which is associated with HCC progression. A semiquantitative RNA polymerase chain reaction (RNA-PCR) was used to analyze genetic expression in 31 cirrhotic liver specimens from patients with HCV. In order to detect HCV replication, the minus-strand RNA of HCV, which serves as a template for the synthesis of genomic plus-strand RNA, was examined. The expression of the growth factors was semiquantified by RNA-PCR, and the mutation of p53 was detected using PCR-single-strand conformation polymorphism. According to the semiquantitative analysis, HCV replication was not associated with the expression of TGF-beta 1 but was significantly so with the overexpression of TGF-alpha (r = 0.74) and IGF-II (r = 0.65) in the HCV-positive cirrhotic livers. No mutation of p53 was recognized in any of the samples. Our investigation thus suggested that the replication of HCV might mediate the coexpression of TGF-alpha and IGF-II and act as a possible initiating factor for hepatocarcinogenesis.

Rearrangements at the 11p15 locus and overexpression of insulin-like growth factor-II gene in sporadic adrenocortical tumors.

Little is known about the pathophysiology of sporadic adrenocortical tumors in adults. Because loss of heterozygosity at the 11p15 locus has been described in childhood tumors, particularly, in adrenocortical tumors, associated with the Beckwith-Wiedemann syndrome and because insulin-like growth factor-II (IGF-II) is a crucial regulator of fetal adrenal growth, we looked for structural analysis at the 11p15 locus and IGF-II gene expression in 23 sporadic adrenocortical adult tumors: 6 carcinomas (5 with Cushing's syndrome and 1 nonsecreting) and 17 benign adenomas (13 with Cushing's syndrome, 1 pure androgen secreting, and 3 nonsecreting). Twenty-one patients were informative at the 11p15 locus, and six (four carcinomas and two adenomas) of them (28.5%) exhibited 11p15 structural abnormalities in tumor DNA (five, an uniparental disomy and one, a mosaicism). In a single case that could be further studied, a paternal isodisomy was observed. Very high IGF-II mRNA contents were detected in seven tumors (30%; 5 of the 6 carcinomas and 2 of the 17 adenomas). They were particularly found in tumors with uniparental disomy at the 11p15 locus. Overall, a strong correlation existed between IGF-II mRNA contents and DNA demethylation at the IGF-II locus. These data show that genetic alterations involving the 11p15 locus were highly frequent in malignant tumors, but found only in rare adenomas. These results in combination with evidence for overexpression of IGF-II from the 11p15.5 locus suggest that abnormalities in structure and/or expression of the IGF-II gene play a role as a late event of a multistep process of tumorigenesis.

Simultaneous over-expression of insulin-like growth factor-II (IGF II) and IGF-II receptor (IGF-II R) genes in human primary cancer-implication of autocrine and paracrine mechanism in autonomous growth of hepatic cancer

This is first report about the simultaneous over-expression of both Insulin-like growth factor (IGF-II) and its receptor (IGF-II R) at mRNA level in human primary hepatic Cancer (PHC). In 10 PHC samples from China, IGF-II and IGF-II R were both over-expressed, whereas only a background signal was detected in normal liver. In 5 pairs of PHC and its non-tumorous adjacent liver tissues from South Africa, IGF-II and IGF-II R were also over-expressed in PHC. mRNA expression of IGF-II in all 5 cases and IGF-II R in 4 of 5 cases were higher in cancer than non-tumorous adjacent liver tissues. These results strongly implicate that an autocrine and/or paracrine mechanism might be involved in formation and progression of PHC.

Insulin-like growth factor-II overexpression in MCF-7 cells induces phenotypic changes associated with malignant progression

Insulin-like growth factor-II overexpression in MCF-7 cells induces phenotypic changes associated with malignant progression

Insulin-like growth factor-II overexpression in MCF-7 cells induces phenotypic changes associated with malignant progression.

It has been proposed that the insulin-like growth factors (IGFs) can act as autocrine and/or paracrine growth promoters in breast cancer. To investigate this hypothesis, we infected early passage MCF-7 cells with a retroviral vector containing the coding sequence for the IGF-II preprohormone along with a constitutive cytomegalovirus promoter sequence. These cells do not normally express IGF-I or IGF-II. After infection with the retroviral vector, several single cell clones were analyzed. Seven of nine isolated clones expressed very high levels of IGF-II mRNA. Biologically active IGF-II protein was easily detectable in the medium conditioned by the IGF-II-expressing clones, and IGF receptors were down-regulated in these. All IGF-II-expressing clones showed marked morphological changes in anchorage-dependent culture, growing in large clumps and as free-floating colonies. The cells also cloned in soft agar in the absence of estrogen, while the wild-type MCF-7 cells and control cells infected with an irrelevant DNA sequence showed none of these properties. alpha IR-3, an antibody that blocks the type I IGF receptor, inhibited the growth of IGF-II-expressing clones in serum-free medium. This model demonstrates that IGF-II can serve as an autocrine growth stimulant in breast cancer epithelial cells and that IGF-II overexpression may be capable of mediating malignant progression in human breast cancer.