Secretion Of Insulin-like Growth Factor 2 Research Articles

TMED10 mediates the trafficking of insulin-like growth factor 2 along the secretory pathway for myoblast differentiation.

The insulin-like growth factor 2 (IGF2) plays critical roles in cell proliferation, migration, differentiation, and survival. Despite its importance, the molecular mechanisms mediating the trafficking of IGF2 along the secretory pathway remain unclear. Here, we utilized a Retention Using Selective Hook system to analyze molecular mechanisms that regulate the secretion of IGF2. We found that a type I transmembrane protein, TMED10, is essential for the secretion of IGF2 and for differentiation of mouse myoblast C2C12 cells. Further analyses indicate that the residues 112-140 in IGF2 are important for the secretion of IGF2 and these residues directly interact with the GOLD domain of TMED10. We then reconstituted the release of IGF2 into COPII vesicles. This assay suggests that TMED10 mediates the packaging of IGF2 into COPII vesicles to be efficiently delivered to the Golgi. Moreover, TMED10 also mediates ER export of TGN-localized cargo receptor, sortilin, which subsequently mediates TGN export of IGF2. These analyses indicate that TMED10 is critical for IGF2 secretion by directly regulating ER export and indirectly regulating TGN export of IGF2, providing insights into trafficking of IGF2 for myoblast differentiation.

Imatinib use in the management of a patient with Doege-Potter syndrome.

Doege-Potter syndromeis a paraneoplastic syndrome characterized by nonislet cell tumor hypoglycemia due to a solitary fibrous tumor, which produces insulin-like growth factor II. In this report, we present the case of a 67-year-old male with recurrent and refractory hypoglycemia due to DPS successfully treated with imatinib. He initially presented with neuroglycopenic symptoms and dyspnea secondary to a giant tumor in the left hemithorax, which was totally resected. During follow-up, 7 years later, he presented with thoracoabdominal tumor recurrence associated with severe hypoglycemia and underwent subtotal tumor resection, with a subsequent improvement of symptoms. The following year, he had a recurrence of his intra-abdominal tumor, which was unresectable, associated with severe hypoglycemia refractory to dextrose infusion and corticosteroids, thus receiving imatinib with a favorable response. The clinical presentation, diagnostic approach, progression of the disease, and response to treatment with imatinib in the management of a patient with large, recurrent, and unresectable mesenchymal tumors with insulin-like growth factor-2 secretion causing hypoglycemiahighlight the importance of this case report. Doege-Potter syndrome (DPS) is a rare cause of tumoral hypoglycemia of non-pancreatic origin. Some malignant or benignant neoplasms have ectopic secretion of insulin-like growth factor-2. Total surgical removal is the principal treatment in patients with DPS. Tyrosine kinase inhibitors management in DPS may contribute to improved tumor control in patients with unresectable tumors and severe hypoglycemia.

PSUN286 A Rare Case of Non-Islet Cell Tumor Hypoglycemia in Young Female With Metastatic Neuroendocrine Tumor

Abstract Background Non-islet cell tumor hypoglycemia (NICTH) is a rare paraneoplastic syndrome caused by a benign or cancerous tumor that over-secretes various hormones that lead to hypoglycemia. Multiple etiologies were described in literature, the most common of them is abnormal secretion of insulin-like growth factor-2 (IGF-2) known as Doege-Potter syndrome (first described in 1929). Other etiologies include oversecretion of proinsulin, IGF-1, and somatostatin. Estimated incidence of NICTH is one per one million person-years. Tumors of mesenchymal origin are most often described in association with NICTH. Laboratory diagnosis of fasting hypoglycemia associated with serum insulin levels below 3 μU/ml confirm the diagnosis of NICTH. In the case of Doege-Potter syndrome, lab findings show low levels of insulin, c-peptide, IGF-1 and a high IGF-2: IGF-1 ratio, which indicate non-insulin-mediated hypoglycemia due to IGF-2 oversecretion. Neuroendocrine tumors presented with NICTH are rare and only described in several case reports. Clinical Case We report a case of a 34 year old female with metastatic neuroendocrine high grade tumor (WHO grade 3, non small cell, diagnosed 3 years prior to presentation) with metastasis to the bone (status post debulking and radiation) and brain (status post radiation therapy). Patient was admitted due to left ankle cellulitis. Endocrinology was consulted due to incidental finding of hypoglycemia serum glucose of 44 mg/dL. Repeat serum levels showed c-peptide 1 ng/dL (normal 1.1 - 4.4) and glucose of 54 mg/dL. She denied a history of diabetes mellitus, alcohol intake, insulin or sulfonylurea use. She had normal renal and liver function. Continuous infusion of D10 started and after 24 hours the patient's hypoglycemia improved. However, hypoglycemia recurred every morning. Following morning hypoglycemia repeat serum labs showed insulin 1.3 (Normal 2.6-24.9), c-peptide 0.9 ng/dL, and serum glucose of 114 mg/dL confirming that her hypoglycemia was not insulin induced. She was not on any medications known to cause hypoglycemia. CT abdomen without contrast showed left upper quadrant heterogeneous hypoattenuating mass with calcifications, measuring approximately 8.5 cm×10.1 cm×2.6 cm. Conclusion This is a case of a patient with a high grade non small cell metastatic neuroendocrine tumor who presented with NICTH. This is the first case of a neuroendocrine tumor with non-small cell pathology causing NICTH that has been described to-date. Based on the patient's clinical presentation and available laboratory findings with recurrent fasting hypoglycemia, low insulin and c-peptide levels we confirmed diagnosis of NICTH. This patient`s hypoglycemia was most likely induced by IGF-2 overexpression (Doege-Potter syndrome) which is the most common cause of NICTH. Presentation: Sunday, June 12, 2022 12:30 p.m. - 2:30 p.m.

ODP187 Doedge-Potter syndrome and papillary thyroid carcinoma: An unusual association

Abstract Hypoglycemia can occur in non-diabetic individuals. Doege-Potter is a rare paraneoplastic syndrome presented as a hypoinsulinemic hypoglycemia caused by ectopic secretion of insulin-like growth factor-2 (IGF-2) from a solitary fibrous tumor. There is recent evidence of the involvement of the IGF system in different malignancies as papillary thyroid carcinoma, being exceptional. A 69-year-old female was brought to Emergency after a routine analysis uncovered severe hypoglycemia. She mentioned a history of morning sickness, dizziness, and confusion for the last week that reverted after breakfast, and dry cough for three months. She had a 25-pack-year history of smoking, no history of alcohol use, no other antecedent of interest, no history ofdiabetes mellitus or weight loss surgery, nor chronic treatments. Physical examinations revealed diminished breath sounds in the left middle lung. A thorax radiography showed a submassive left pleural effusion and toracocentesis revealed a hemorrhagic exudate. Laboratory data showed normal GH, and cortisol levels. Despite marked hypoglycemia, insulin level was 0.4 μIU/mL (2,6-24,9), C-peptide 0.11ng/ml (1,1-4,4) and hydroxy butyrate <0.1 mg/dl. The serum IGF-2 level was 428μg/dL (350-1000), and the IGF-1 level was 55.4 μg/L(161-384). Anti-insulin antibodies and sulfonylureas concentration were negative. CT showed a solid mass concerning parietal pleura with areas of necrosis and calcifications, measuring 14.6×15.4×20cm. Low blood glucose with suppressed insulin, C-peptide and IGF-1 levels, elevated IGF-2/IGF-1 ratio (7,7, normal<3) and CT findings suggested the possibility of non-islet cell tumor hypoglycemia (NICTH). Corticoid treatment was initiated to manage the hypoglycemia. Biopsy of the mass showed a malignant solitary fibrous tumor, positive for CD34 and BCL2. After surgery, glucose and IGF-2 levels returned to normal, the patient didn't experience further episodes of hypoglycemia, and corticoid treatment was gradually suspended. A FDG-PET scan, part of extension study, found a PET-positive thyroid nodule. A thyroid ultrasound confirmed a solid, taller than wide thyroid nodule, measuring 6.7×6.4×9,5mm and fine-needle aspiration biopsy was suggestive of papillary thyroid carcinoma. Hemithyroidectomy was performed. The patient is with no evidence of disease. Doege-Potter syndrome is a rare cause of hypoglycemia, resulting from a tumor producing elevated levels of IGF-2. Diagnosis is suggested by low insulin, C-peptide and IGF-1 levels, normal or elevated IGF-2, and elevated IGF2/IGF1 ratio. High levels of ''big-IGF-2'' are associated with high risk papillary thyroid carcinoma, suggesting the relation between the IGF-system and tumor growth, nevertheless there are few cases reported describing this association. Surgical resection of the tumor is the treatment of choice. Presentation: No date and time listed

IGF-2 mediated hypoglycemia and the paradox of an apparently benign lesion: a case report & review of the literature

BackgroundNon-islet cell tumour hypoglycemia (NICTH) is rarely encountered in clinical practice. Insulin-like growth factor 2 (IGF2) is the most common cause of NICTH observed in the setting of mesenchymal and epithelial neoplasia. This is a paraneoplastic syndrome caused by IGF2 activation of the insulin receptor.Case presentationAn 80 year old female presented with a short history of recurrent episodes of confusion with laboratory confirmed hypoglycemia with a plasma glucose of 2.7 mmol/L on fasting which fulfilled Whipple’s triad. Diagnostic clues to the aetiology at presentation include the fasting pattern of hypoglycemia, hypokalaemia and the absence of weight gain. A 72 hour fast with results showed early hypoglycemia and suppression of serum insulin, c-peptide, and proinsulin. Serum insulin antibody was not detected. Subsequent measurement of the serum IGF2:IGF1 ratio was elevated at 22.3 and consistent with IGF-2 mediated hypoglycemia and imaging studies demonstrated a pelvic mass. Dietary intervention and oral prednisolone abated hypoglycemia prior to surgery. Ultimately, hypoglycemia resolved following operative intervention and steroid therapy was successfully withdrawn. Histopathology was remarkable for dual neoplastic processes with uterine solitary fibrous tumour (SFT) confirmed as the source of IGF2 hypersecretion on IGF-2 immunohistochemistry and a coincidental invasive high grade serous carcinoma involving the fimbria of the right fallopian tube.ConclusionThe paradox in this case is that the benign solitary fibrous tumour accounted for patient morbidity through secretion of IGF2 and without treatment, posed a mortality risk. This is despite the synchronous presence of a highly malignant fallopian tube neoplasm. This case reinforces the need for thorough clinical evaluation of hypoglycemia to allow prompt and definitive management.

Liver Expression of IGF2 and Related Proteins in ZBED6 Gene-Edited Pig by RNA-Seq.

Simple SummaryZinc finger BED-type containing 6 (ZBED6), as a regulatory factor, has different regulatory mechanisms in animal development. The intron of insulin-like growth factor 2 (IGF2) regulates the development of animal muscle and adipose by combining with the binding site of ZBED6. As a member of the insulin-like growth factor family, IGF2 plays an important role in embryonic growth and development, cell proliferation, muscle growth and genome imprinting. In order to further study the regulatory mechanism of ZBED6 on IGF2, we detected the expression of IGF2 and related genes in ZBED6 single allele knockout (ZBED6-SKO) pig tissues and analyzed differently expressed genes of the transcriptome of ZBED6-SKO pig liver. The results showed that the partial knockout of ZBED6 could affect the secretion of IGF2 in pig liver but had no significant difference at the protein level. This research provides a new idea for the interaction between IGF2 and ZBED6.Zinc finger BED-type containing 6 (ZBED6), a highly conservative transcription factor of placental mammals, has conservative interaction of insulin-like growth factor 2 (IGF2) based on the 16 bp binding sites of ZBED6 on the IGF2 sequence. IGF2 is related to embryo growth and cell proliferation. At the same time, its functions in muscle and adipose in mammals have been widely mentioned in recent studies. To further investigate the mechanism of ZBED6 on IGF2, we detected the expression of IGF2 and related genes in ZBED6 single allele knockout (ZBED6-SKO) pig tissues and analyzed the transcriptome of ZBED6-SKO pig liver. Through RNA-seq, we captured nine up-regulated genes and eight down-regulated genes which related to lipid metabolism. The results showed that the mRNA of IGF2 had an upward trend after the partial knockout of ZBED6 in liver and had no significant difference in protein expression of IGF2. In summary, ZBED6-SKO could affect the secretion of IGF2 in pig liver and its own lipid metabolism. Our research has provided basic information for revealing the regulatory mechanism of the interaction between ZBED6 and IGF2 in mammals.

Insulin-Like Growth Factor 2 (IGF2) Signaling in Colorectal Cancer-From Basic Research to Potential Clinical Applications.

Colorectal cancer (CRC) is one of the most common cancers in men and women worldwide as well as is the leading cause of death in the western world. Almost a third of the patients has or will develop liver metastases. While genetic as well as epigenetic mechanisms are important in CRC pathogenesis, the basis of the most cases of cancer is unknown. High spatial and inter-patient variability of the molecular alterations qualifies this cancer in the group of highly heterogeneous tumors, which makes it harder to elucidate the mechanisms underlying CRC progression. Determination of highly sensitive and specific early diagnosis markers and understanding the cellular and molecular mechanism(s) of cancer progression are still a challenge of the current era in oncology of solid tumors. One of the accepted risk factors for CRC development is overexpression of insulin-like growth factor 2 (IGF2), a 7.5-kDa peptide produced by liver and many other tissues. IGF2 is the first gene discovered to be parentally imprinted. Loss of imprinting (LOI) or aberrant imprinting of IGF2 could lead to IGF2 overexpression, increased cell proliferation, and CRC development. IGF2 as a mitogen is associated with increased risk of developing colorectal neoplasia. Higher serum IGF2 concentration as well as its tissue overexpression in CRC compared to control are associated with metastasis. IGF2 protein was one of the three candidates for a selective marker of CRC progression and staging. Recent research indicates dysregulation of different micro- and long non-coding RNAs (miRNAs and lncRNAs, respectively) embedded within the IGF2 gene in CRC carcinogenesis, with some of them indicated as potential diagnostic and prognostic CRC biomarkers. This review systematises the knowledge on the role of genetic and epigenetic instabilities of IGF2 gene, free (active form of IGF2) and IGF-binding protein (IGFBP) bound (inactive form), paracrine/autocrine secretion of IGF2, as well as mechanisms of inducing dysplasia in vitro and tumorigenicity in vivo. We have tried to answer which molecular changes of the IGF2 gene and its regulatory mechanisms have the most significance in initiation, progression (including liver metastasis), prognosis, and potential anti-IGF2 therapy in CRC patients.

SUN-328 Non-Islet Cell Tumor-Induced Hypoglycemia in a Patient with IGF-2 Producing Spindle Cell Neoplasm

Background: Non-islet cell tumor-induced hypoglycemia is a rare paraneoplastic syndrome with approximately 300 reported cases in the literature. Classically these tumors secrete insulin-like growth factor-2 (IGF-2) to induce hypoglycemia, but the etiology and regulation of the hormone secretion remains poorly understood. We present a case of non-islet cell hypoglycemia from a spindle cell neoplasm of the lung, which appeared to show inconsistent secretion of IGF-2. Clinical case: A 72-year-old, non-diabetic male presented with episodes of altered mental status due to hypoglycemia with unintentional weight loss. Blood glucose was 35 mg/dL upon admission. CT chest showed an 18 cm tumor in the left hemithorax. The patient continued to have hypoglycemia throughout his hospital stay. At a time when the serum glucose was 48 mg/dL, a biochemical work up for hypoglycemia showed C-peptide 0.21 ng/mL (0.78-5.19 ng/mL), proinsulin 1.6 pmol/L (0-10 pmol/L), insulin 0.1 U/mL (1.9-23 U/mL), β-hydroxybutyrate 0.1 mmol/L (0-0.4 mmol/L), and a negative sulfonylurea screen. Given the large lung tumor, IGF hormones were measured: IGF-1 59 ng/mL (36-215 ng/mL), IGF-2 670 ng/mL (333-967 ng/mL). While both were within normal ranges, the IGF-2/IGF-1 ratio was 11.35. Biopsy of the lung mass showed benign spindle cell neoplasm, consistent with solitary fibrous tumor. The patient was diagnosed with paraneoplastic non-insulin mediated hypoglycemia and was transferred to our hospital for resection of the lung mass. During the first 96 hours after transfer, the patient had no hypoglycemic episodes and repeat laboratory values showed IGF-1 of 72 ng/mL, IGF-2 of 162 ng/mL, IGF-2/IGF-1 ratio of 2.25 with a glucose of 119 mg/dL, indicating the secretion of IGF-2 may have been intermittent. However, he later developed recurrent hypoglycemia and underwent resection of the tumor. Pathology showed a malignant solitary fibrous tumor. Blood sugars improved within 2 days after tumor resection and patient did not have further hypoglycemia. Discussion: Literature review shows that tumors causing non-islet cell hypoglycemia are large with one-third located in the chest and two-thirds in the retroperitoneum. They can secrete “big” IGF-2, which can activate insulin receptors to inhibit glucose output from the liver and increase uptake in muscle and adipose tissue leading to hypoglycemia. Interestingly, the IGF-2 level is not always outside the normal range, but a ratio of IGF-2/IGF-1 greater than 10 can be another marker of paraneoplastic hypoglycemia. We believe our case is unique in that during a 48-hour period of intense observation, the patient had no hypoglycemia and the IGF-2 levels and IGF-2/IGF-1 ratio decreased to the normal range. Thus, in patients with suspected tumor-mediated hypoglycemia, the IGF levels may need to be drawn during a hypoglycemic event, much like the workup for insulinoma.

Seborrheic keratoses and severe hypoinsulinemic hypoglycemia associated with insulin grow factor 2 secretion by a malignant solitary fibrous tumor

A rare sign of some malignant tumors is a sudden eruption of multiple seborrheic keratoses called Leser-Trélat sign. Overproduction of insulin-like growth factor-2 (IGF2) or its precursor is the main mechanism related to non-islet cell tumor hypoglycemia. Doege-Potter syndrome is the name given to paraneoplastic hypoinsulinemic hypoglycemia in presence of a solitary fibrous tumor. This report describes a case of a patient with hypoinsulinemic hypoglycemia and Leser-Trélat sign associated with a malignant solitary fibrous tumor with IGF2 secretion. Both conditions have improved after tumor excision.

Prenatal ethanol exposure and placental hCG and IGF2 expression

IntroductionFetal alcohol spectrum disorder (FASD) is the main cause of preventable non-genetic mental retardation. Diagnosis of prenatal exposure to ethanol (PEE) is based on questionnaires and biomarkers in perinatal matrices. Early diagnosis of FASD is important to mitigate secondary disabilities that will arise later in life. It is important to identify biomarkers related to cellular damage caused by PEE. The main objective was to identify novel candidate biomarkers from placental tissue using an in vitro model of exposure to ethanol and to support it in placental tissue obtained from pregnancies with PEE assessed by fatty acid esters in meconium samples. MethodsFirst, hormone production was examined using two different human trophoblast cell lines, JEG3 and BeWo. Viable cell count by exclusion method was analyzed and human chorionic gonadotrophin (hCG) and insulin-like growth factor 2 (IGF2) were quantified by Western blot and ELISA. Second, these techniques were used in protein lysates from human placentas from pregnancies with and without exposure to ethanol. ResultsBoth trophoblast cell lines showed a decrease in cell viability accompanied with apoptosis activation after a chronic ethanol treatment. Moreover, we showed an increase in the secretion of hCG and IGF2 in a dose-dependent manner. Interestingly, this increase was also observed in a set of human placenta tissue from fetuses exposed prenatally to ethanol. DiscussionEthanol exposure during pregnancy causes placenta cell damage, so altering its normal function. The specific hCG and IGF2 release pattern is a candidate surrogated biomarker of the damage due to PEE.

Glutamine Stimulates Biosynthesis and Secretion of Insulin-like Growth Factor 2 (IGF2), an Autocrine Regulator of Beta Cell Mass and Function

IGF2 is an autocrine ligand for the beta cell IGF1R receptor and GLP-1 increases the activity of this autocrine loop by enhancing IGF1R expression, a mechanism that mediates the trophic effects of GLP-1 on beta cell mass and function. Here, we investigated the regulation of IGF2 biosynthesis and secretion. We showed that glutamine rapidly and strongly induced IGF2 mRNA translation using reporter constructs transduced in MIN6 cells and primary islet cells. This was followed by rapid secretion of IGF2 via the regulated pathway, as revealed by the presence of mature IGF2 in insulin granule fractions and by inhibition of secretion by nimodipine and diazoxide. When maximally stimulated by glutamine, the amount of secreted IGF2 rapidly exceeded its initial intracellular pool and tolbutamide, and high K(+) increased IGF2 secretion only marginally. This indicates that the intracellular pool of IGF2 is small and that sustained secretion requires de novo synthesis. The stimulatory effect of glutamine necessitates its metabolism but not mTOR activation. Finally, exposure of insulinomas or beta cells to glutamine induced Akt phosphorylation, an effect that was dependent on IGF2 secretion, and reduced cytokine-induced apoptosis. Thus, glutamine controls the activity of the beta cell IGF2/IGF1R autocrine loop by increasing the biosynthesis and secretion of IGF2. This autocrine loop can thus integrate changes in feeding and metabolic state to adapt beta cell mass and function.

Abstract 4361: Role of Id1-induced IGF2 in autocrine/endocrine promotion of esophageal cancer tumorigenesis and metastasis

Abstract Esophageal cancer is the 7th leading cause of cancer death in Hong Kong. The disease is highly lethal (with a 5-year survival rate of ∼14%) because many cases go undetected until the disease is at an advanced stage. Understanding of the underlying mechanisms and the signaling pathways involved in esophageal tumorigenesis is the key to identify new molecular targets for intervention. Id1 (inhibitor of differentiation or DNA binding), a member of the helix-loop-helix (HLH) proteins, is overexpressed in many types of human cancer. Our previous reports provided the first evidence that Id1 activates the phosphatidylinositol-3-kinase (PI3K)/AKT pathway in esophageal squamous cell carcinoma (ESCC) cells, and that there is a positive correlation between Id1 and phosphorylated (p)-AKT expressions in clinical esophageal cancer specimens. We also demonstrated in experimental animal models that Id1 promotes tumor growth and metastasis through activation of PI3K/AKT. Increasing evidence suggests that growth factors secreted from cancer cells can play an important role in cancer progression by modulating the tumor microenvironment and the host systemic environment via autocrine, paracrine, and endocrine mechanisms. In this study, we found that Id1 induced the expression and secretion of IGF2 (insulin-like growth factor 2) in ESCC cells, and that this induction mediated the activation of PI3K/AKT in an autocrine manner. Knockdown of IGF2 in cancer cells or addition of IGF2 neutralizing antibody abrogated the effects of Id1 on proliferation, survival, and invasion of esophageal cancer cells. The mechanisms by which Id1 upregulates IGF2 were studied, and the results indicated that Id1 could protect IGF2 from ubiquitin-proteasomal degradation in ESCC cells. Immunohistochemical analysis of human esophageal cancer tissue microarray showed elevated IGF2 expression in ESCC, which correlated with increased Id1 and p-AKT expressions. Furthermore, our in vivo experiments showed that IGF2 secreted by Id1-overexpressing ESCC xenograft could instigate the growth of distant ESCC tumors, as well as promote metastasis of circulating ESCC cells. Targeting IGF2 with the use of neutralizing antibody had significant antitumor effects on esophageal cancer xenografts in mice. Our findings therefore support that the Id1-IGF2-PI3K/AKT signaling cascade plays an important role in esophageal cancer, and that Id1-overexpressing primary tumors may affect the host systemic environment and contribute to cancer progression. In conclusion, we have uncovered a novel mechanism in the role of Id1 in tumorigenesis which may have significant implications for the development of targeted therapy for esophageal cancer. [This study is supported by the Research Grants Council of the Hong Kong SAR, China (General Research Fund Project no. HKU 762610M)] Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 103rd Annual Meeting of the American Association for Cancer Research; 2012 Mar 31-Apr 4; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2012;72(8 Suppl):Abstract nr 4361. doi:1538-7445.AM2012-4361

Glucagon-like Peptide-1 Increases β-Cell Glucose Competence and Proliferation by Translational Induction of Insulin-like Growth Factor-1 Receptor Expression

Glucagon-like peptide-1 (GLP-1) protects beta-cells against apoptosis, increases their glucose competence, and induces their proliferation. We previously demonstrated that the anti-apoptotic effect was mediated by an increase in insulin-like growth factor-1 receptor (IGF-1R) expression and signaling, which was dependent on autocrine secretion of insulin-like growth factor 2 (IGF-2). Here, we further investigated how GLP-1 induces IGF-1R expression and whether the IGF-2/IGF-1R autocrine loop is also involved in mediating GLP-1-increase in glucose competence and proliferation. We show that GLP-1 up-regulated IGF-1R expression by a protein kinase A-dependent translational control mechanism, whereas isobutylmethylxanthine, which led to higher intracellular accumulation of cAMP than GLP-1, increased both IGF-1R transcription and translation. We then demonstrated, using MIN6 cells and primary islets, that the glucose competence of these cells was dependent on the level of IGF-1R expression and on IGF-2 secretion. We showed that GLP-1-induced primary beta-cell proliferation was suppressed by Igf-1r gene inactivation and by IGF-2 immunoneutralization or knockdown. Together our data show that regulation of beta-cell number and function by GLP-1 depends on the cAMP/protein kinase A mediated-induction of IGF-1R expression and the increased activity of an IGF-2/IGF-1R autocrine loop.

A HIF‐1α‐dependent autocrine feedback loop promotes survival of serum‐deprived prostate cancer cells

We previously reported that normoxic, serum-deprived prostate cancer (PCa) cells upregulate hypoxia-inducible factor-1alpha (HIF-1alpha) protein, which promotes survival during serum deprivation via insulin-like growth factor-2 (IGF-2) upregulation. This study investigated the molecular mechanism of autocrine regulation of HIF-1alpha, IGF-2 and cell survival in serum-deprived PC-3 and LNCaP PCa cells. Cell viability was assessed by trypan blue assay. PI3K activity was inhibited with LY294002, and PTEN overexpression. mRNA was assessed by RT-PCR, and IGF-2 protein by ELISA. Activated insulin-like growth factor-I receptor (IGF-IR) was detected by probing immunoprecipitated IGF-IR for phospho-tyrosine. IGF-IR activity was inhibited with IGF-2 neutralizing antibody and IGF-IR-specific siRNA. HIF-1alpha, phospho-Akt, total-Akt and IGF-IR protein was assessed by immunoblots. HIF-1alpha was suppressed with siRNA. We detected a time-dependent increase in Akt activation during serum deprivation, and inhibition of Akt activation attenuated the serum deprivation-mediated increase in HIF-1alpha and cell survival. Importantly, IGF-2 secretion significantly increased during serum deprivation, and was accompanied by increased activation of its receptor, IGF-IR. Additionally, inhibition of IGF-2 activity markedly attenuated the serum deprivation-mediated increase in IGF-IR and Akt activation, HIF-1alpha expression, and also its own transcription, suggesting autocrine regulation of HIF-1alpha expression via IGF-2. Cross-talk between IGF-2/ IGF-IR system and PI3K-Akt pathway was further demonstrated by findings wherein IGF-IR suppression inhibited Akt activation, and IGF-IR activation was inhibited following PI3K inhibition. Furthermore, HIF-1alpha suppression attenuated the serum deprivation-mediated increase in Akt and IGF-IR activation. Collectively, our study demonstrates existence of a pro-survival HIF-1alpha-dependent autocrine feedback loop in normoxic, serum-deprived PCa cells.

Post-translational Processing of the Insulin-like Growth Factor-2 Precursor: ANALYSIS OF O-GLYCOSYLATION AND ENDOPROTEOLYSIS

Insulin-like growth factor-2 (IGF-2) is expressed in most embryonic tissues and is required for normal development during gestation. After birth IGF-2 expression is extinguished in most tissues, but the gene is often reactivated during tumorigenesis. Tumors secrete high molecular weight forms of IGF-2 that result from aberrant post-translational processing of pro-IGF-2. As a first step toward understanding how high molecular weight IGF-2 peptides might contribute to tumor progression, we have characterized the biosynthesis of IGF-2 in a human embryonic cell line. We have found that pro-IGF-2 can initially form two disulfide isomers that undergo rearrangement to a single conformation in vivo. The addition of N-acetylgalactosamine to Ser71, Thr72, Thr75, and Thr139 likely occurs in the cis- Golgi apparatus. Sialic acid addition begins in the trans- Golgi apparatus, but IGF-2 peptides must reach the trans-Golgi network for oligosaccharide maturation to be completed. Endoproteolysis occurs concomitant to or slightly after oligosaccharide maturation. Cleavage was observed only at Arg104, resulting in the secretion of IGF-2-(1-104) and free E-peptide. Proteolysis required basic residues in the P1 (Arg104) and P4 (Arg101) positions, was completely blocked by a furin inhibitor, and was enhanced by coexpression with furin, PACE4, PC6A, PC6B, and LPC. These data suggest that members of the subtilisin-related proprotein convertase family mediate processing of pro-IGF-2 at Arg104. We did not detect the IGF-2 peptides that are most abundant in normal serum, mature IGF-2, and IGF-2-(1-87), in this expression system, which indicates that novel endoproteases are responsible for generating these products.