IMP3 Immunohistochemical Expression Is Related with Progression and Metastases in Xenografted and Cutaneous Melanomas.

INTRODUCTION: Insulin-like growth factor-II messenger RNA-binding protein-3 (IMP3) over-expression is a predictor of tumor recurrence and metastases in some types of human melanoma. Our objective was to evaluate the immunohistochemical expression of IMP3 and other molecules related to tumor prognosis in melanoma-xeno-tumors undergoing treatment. We test the effect of radiotherapy (RT) and mesenchymal stromal cells (MSCs) treatment, analyzing the tumorigenic and metastatsizing capacity in a mice melanoma xenograft model. MATERIALS AND METHODS: We inoculated A375 and G361 human melanoma cell lines into NOD/SCID gamma mice (n = 64). We established a control group, a group treated with MSCs, a group treated with MSCs plus RT, and a group treated with RT. We assessed the immunohistochemical expression of IMP3, E-cadherin, N-cadherin, PARP1, HIF-1α, and the proliferation marker Ki-67. Additionally, we performed a retrospective study including 114 histological samples of patients diagnosed with malignant cutaneous superficial spreading melanoma (n = 104) and nodular melanoma (n = 10) with at least 5 years of follow-up. RESULTS: Most morphological and immunohistochemical features show statistically significant differences between the 2 cell lines. The A375 cell line induced the formation of metastases, while the G361 cell line provoked tumor formation but not metastases. All three treatments reduced the cell proliferation evaluated by the Ki-67 nuclear antigen (p = 0.000, one-way ANOVA test) and reduced the number of metastases (p = 0.004, one-way ANOVA test). In addition, the tumor volumes reduced in comparison with the control groups, 31.74% for RT + MSCs in the A357 tumor cell line, and 89.84% RT + MSCs in the G361 tumor cell line. We also found that IMP3 expression is associated with greater tumor aggressiveness and was significantly correlated with cell proliferation (measured by the expression of Ki-67), the number of metastases, and reduced expression of adhesion molecules. CONCLUSIONS: The combined treatment of RT and MSCs on xenografted melanomas reduces tumor size, metastases frequency, and the epithelial to mesenchymal transition/PARP1 metastatic phenotype. This treatment also reduces the expression of molecules related to cellular proliferation (Ki-67), molecules that facilitate the metastatic process (E-cadherin), and molecules related with prognosis (IMP3).

The Role of SOX9 in IGF-II-Mediated Pulmonary Fibrosis.

Pulmonary fibrosis (PF) associated with systemic sclerosis (SSc) results in significant morbidity and mortality. We previously reported that insulin-like growth factor-II (IGF-II) is overexpressed in lung tissues and fibroblasts from SSc patients, and IGF-II fosters fibrosis by upregulating collagen type I, fibronectin, and TGFß. We now show that IGF-II augments mRNA levels of profibrotic signaling molecules TGFß2 (p ≤ 0.01) and TGFß3 (p ≤ 0.05), collagen type III (p ≤ 0.01), and the collagen posttranslational modification enzymes P4HA2 (p ≤ 0.05), P3H2 (p ≤ 0.05), LOX (p = 0.065), LOXL2 (p ≤ 0.05), LOXL4 (p ≤ 0.05) in primary human lung fibroblasts. IGF-II increases protein levels of TGFß2 (p ≤ 0.01), as well as COL3A1, P4HA2, P4Hß, and LOXL4 (p ≤ 0.05). In contrast, IGF-II decreases mRNA levels of the collagen degradation enzymes cathepsin (CTS) K, CTSB, and CTSL and protein levels of CTSK (p ≤ 0.05). The SRY-box transcription factor 9 (SOX9) is overexpressed in SSc lung tissues at the mRNA (p ≤ 0.05) and protein (p ≤ 0.01) levels compared to healthy controls. IGF-II induces SOX9 in lung fibroblasts (p ≤ 0.05) via the IGF1R/IR hybrid receptor, and SOX9 regulates TGFß2 (p ≤ 0.05), TGFß3 (p ≤ 0.05), COL3A1 (p ≤ 0.01), and P4HA2 (p ≤ 0.001) downstream of IGF-II. Our results identify a novel IGF-II signaling axis and downstream targets that are regulated in a SOX9-dependent and -independent manner. Our findings provide novel insights on the role of IGF-II in promoting pulmonary fibrosis.

Novel Isoform DTX3c Associates with UBE2N-UBA1 and Cdc48/p97 as Part of the EphB4 Degradation Complex Regulated by the Autocrine IGF-II/IRA Signal in Malignant Mesothelioma.

EphB4 angiogenic kinase over-expression in Mesothelioma cells relies upon a degradation rescue signal provided by autocrine IGF-II activation of Insulin Receptor A. However, the identity of the molecular machinery involved in EphB4 rapid degradation upon IGF-II signal deprivation are unknown. Using targeted proteomics, protein-protein interaction methods, PCR cloning, and 3D modeling approaches, we identified a novel ubiquitin E3 ligase complex recruited by the EphB4 C tail upon autocrine IGF-II signal deprivation. We show this complex to contain a previously unknown N-Terminal isoform of Deltex3 E3-Ub ligase (referred as "DTX3c"), along with UBA1(E1) and UBE2N(E2) ubiquitin ligases and the ATPase/unfoldase Cdc48/p97. Upon autocrine IGF-II neutralization in cultured MSTO211H (a Malignant Mesothelioma cell line that is highly responsive to the EphB4 degradation rescue IGF-II signal), the inter-molecular interactions between these factors were enhanced and their association with the EphB4 C-tail increased consistently with the previously described EphB4 degradation pattern. The ATPase/unfoldase activity of Cdc48/p97 was required for EphB4 recruitment. As compared to the previously known isoforms DTX3a and DTX3b, a 3D modeling analysis of the DTX3c Nt domain showed a unique 3D folding supporting isoform-specific biological function(s). We shed light on the molecular machinery associated with autocrine IGF-II regulation of oncogenic EphB4 kinase expression in a previously characterized IGF-II+/EphB4+ Mesothelioma cell line. The study provides early evidence for DTX3 Ub-E3 ligase involvement beyond the Notch signaling pathway.

Genetic ablation of the preptin-coding portion of Igf2 impairs pancreatic function in female mice.

Preptin is a 34-amino acid peptide derived from the E-peptide of pro-insulin-like growth factor 2 and is co-secreted with insulin from ß-cells. Little is understood about the effects of endogenous preptin on whole body glucose metabolism. We developed a novel mouse model in which the preptin portion of Igf2 was genetically ablated in all tissues, hereafter referred to as preptin knockout (KO), and tested the hypothesis that the removal of preptin will lead to a decreased insulin response to a metabolic challenge. Preptin KO and wild-type (WT) mice underwent weekly fasting blood glucose measurements, intraperitoneal insulin tolerance tests (ITT) at 9, 29, and 44 wk of age, and an oral glucose tolerance test (GTT) at 45 wk of age. Preptin KO mice of both sexes had similar Igf2 exon 2-3 mRNA expression in the liver and kidney compared with WT mice, but Igf2 exon 3-4 (preptin) expression was not detectable. Western blot analysis of neonatal serum indicated that processing of pro-IGF2 translated from the KO allele may be altered. Preptin KO mice had similar body weight, body composition, ß-cell area, and fasted glucose concentrations compared with WT mice in both sexes up to 47 wk of age. Female KO mice had a diminished ability to mount an insulin response following glucose stimulation in vivo. This effect was absent in male KO mice. Although preptin is not essential for glucose homeostasis, when combined with previous in vitro and ex vivo findings, these data show that preptin positively impacts ß-cell function.NEW & NOTEWORTHY This is the first study to describe a model in which the preptin-coding portion of the Igf2 gene has been genetically ablated in mice. The mice do not show reduced size at birth associated with Igf2 knockout suggesting that IGF2 functionality is maintained, yet we demonstrate a change in the processing of mature Igf2. Female knockout mice have diminished glucose-stimulated insulin secretion, whereas the insulin response in males is not different to wild type.

Insulin-like growth factor II-producing colonic carcinoma presenting with non-islet cell tumor hypoglycemia: An autopsy report revealing neuroendocrine differentiation in the metastatic foci and literature review.

Non-islet cell tumor hypoglycemia (NICTH) is a very rare symptom of severe hypoglycemia associated with extrapancreatic tumors. It is considered to be caused by insulin-like growth factor (IGF)-II. There have been no autopsy cases of colorectal carcinoma with NICTH confirmed with both serum high molecular weight and tumoral IGF-II. We report the case of a 46-year-old woman with advanced sigmoid colon cancer and liver metastases. She underwent open sigmoidectomy, and histologically, the lesion was a differentiated-type tubular adenocarcinoma. Postoperative chemotherapy was initiated. However, she experienced repeated hypoglycemia attacks 10 months after the operation, while the liver metastases increased. We examined the cause of hypoglycemia, and finally diagnosed her with NICTH associated with high molecular weight IGF-II production, which was proven by Western immunoblot of the serum. She died 12 months after surgery and was examined by autopsy. Liver metastases showed a transition from adenocarcinoma to carcinoma with neuroendocrine differentiation. Immunohistochemistry showed that both metastatic carcinoma of the liver and primary colonic adenocarcinoma were positive for IGF-II. Neuroendocrine differentiation in liver metastases proven by an autopsy may have contributed to tumor growth, which may have exacerbated the symptoms.

Tanshinone IIA inhibits Leu27IGF-II-induced insulin-like growth factor receptor II signaling and myocardial apoptosis via estrogen receptor-mediated Akt activation.

Different stress condition stimulates the expression level of insulin-like growth factor receptor II (IGF-IIR) in cardiomyoblasts that lead to apoptosis. Tanshinone IIA (TSN), a pharmacologically active component from Danshen, has been shown cardioprotective effects against cardiac apoptosis induced by several stress conditions. Therefore, this study was conducted to assess the cardioprotective effects of TSN IIA mediated through the estrogen receptor (ER) in order to inhibit the Leu27IGF-II-enhanced IGF-IIR-mediated cardiac apoptosis. The estrogenic activity of TSN IIA was examined after myocardial cells were pretreated with the ER antagonist, and inhibited the phospho-inositide-3 kinase (PI3K). Here, we found that TSN IIA significantly induced ER that phosphorylated Akt. Further, Akt activation considerably suppressed the Leu27IGF-II induced IGF-IIR expression level and the downstream effectors, including Gαq and calcineurin as well as mitochondrial dependent apoptosis proteins including Bad, cytochrome c, and active caspase-3 that result in cardiac apoptosis resistance. However, the western blot analysis, JC-1 staining, and terminal deoxynucleotide transferase-mediated dUTP nick end labeling assay revealed that TSN IIA attenuated Leu27IGF-II-induced IGF-IIR mediated cardiac apoptosis was reversed by an ER antagonist such as ICI 182780, and PI3K inhibition. All these findings demonstrate that TSN IIA exerts estrogenic activity, which can activate PI3K-Akt pathway, and thereby inhibits Leu27IGFII induced IGF-IIR mediated cardiac apoptosis. Thus, TSN IIA can be considered as an effective therapeutic strategy against IGF-IIR signaling cascade to suppress cardiac apoptosis.

Association of Insulin-like Growth Factor-II Apa1 and MspI Polymorphisms with Intrauterine Growth Restriction Risk.

BackgroundInsulin-like growth factor-II (IGF-II) has a prominent role in fetal growth and development. The aim of this study was to investigate the association of IGF-II Apa1 and MspI polymorphisms with intrauterine growth restriction (IUGR) risk. Methods: A total of 45 infants with IUGR and 45 infants appropriate for gestational (AGA) were enrolled. Genotyping of Apa1 and MspI polymorphisms was assayed by PCR-RFLP approach. Results: The heterozygote genotype (AG) of IGF-II Apa1 CT was associated with an increased risk of IUGR. Genotypes and alleles of IGF-II MspI polymorphism had no significant association with IUGR susceptibility (P > 0.05). Conclusions: The current study suggests that IGF-II Apa1 polymorphism is associated with an increased risk of IUGR, while IGF-II MspI showed no association with IUGR. Thus, IGF-II Apa1 polymorphism could be used as a relevant molecular marker to identify the fetus at risk of developing IUGR.

Insulin-Like Growth Factor II Targets the mTOR Pathway to Reverse Autism-Like Phenotypes in Mice.

Autism spectrum disorder (ASD) is a developmental disability characterized by impairments in social interaction and repetitive behavior, and is also associated with cognitive deficits. There is no current treatment that can ameliorate most of the ASD symptomatology; thus, identifying novel therapies is urgently needed. We used male BTBR T+Itpr3tf /J (BTBR) mice, a model that reproduces most of the core behavioral phenotypes of ASD, to test the effects of systemic administration of insulin-like growth factor II (IGF-II), a polypeptide that crosses the blood-brain barrier and acts as a cognitive enhancer. We show that systemic IGF-II treatments reverse the typical defects in social interaction, cognitive/executive functions, and repetitive behaviors reflective of ASD-like phenotypes. In BTBR mice, IGF-II, via IGF-II receptor, but not via IGF-I receptor, reverses the abnormal levels of the AMPK-mTOR-S6K pathway and of active translation at synapses. Thus, IGF-II may represent a novel potential therapy for ASD.SIGNIFICANCE STATEMENT Currently, there is no effective treatment for autism spectrum disorder (ASD), a developmental disability affecting a high number of children. Using a mouse model that expresses most of the key core as well as associated behavioral deficits of ASD, that are, social, cognitive, and repetitive behaviors, we report that a systemic administration of the polypeptide insulin-like growth factor II (IGF-II) reverses all these deficits. The effects of IGF-II occur via IGF-II receptors, and not IGF-I receptors, and target both basal and learning-dependent molecular abnormalities found in several ASD mice models, including those of identified genetic mutations. We suggest that IGF-II represents a potential novel therapeutic target for ASD.

The complementary role of insulin-like growth factor II mRNA-binding protein 3 (IMP3) in diagnosis of Hodgkin's lymphoma.

BACKGROUND: Hodgkin's Lymphoma (HL) is a peculiar subtype of lymphoid malignancies. The etiology of HL is still unknown. Insulin-like growth factor II mRNA-binding Protein 3 (IMP3) is a member of IMP family. In HL, IMP3 is expressed in the cytoplasmic compartment of the tumor cells (in both HRS cells & LP cells) against completely negative background of non-tumor cells except for residual germinal centers. MATERIALS & METHODS: Of 51 cases of Hodgkin's lymphoma (HL) referred to surgical pathology laboratory at oncology center, Mansoura University (OCMU), Egypt. All cases were stained for CD20, CD3, CD15, CD30 and IMP3. Regarding IMP3 antibody, The slides were then incubated with the Anti-IMP3, mouse monoclonal antibody (1:200, clone sc-365640, concentrated, California) that was used as primary antibody for IMP3 detection for 1 hour at room temperature, followed by incubation with the secondary antibody, poly HRP (horseradish peroxidase), conjugate for mouse/rabbit, for 20 minutes. RESULTS: IMP3 showed cytoplasmic immunoreactivity in 43 (83%) cases while 8 cases were negative. The sensitivity of combined CD30 & CD15, combined CD30 & IMP3, combined CD15 & IMP3 were 96%, 98% and 94% respectively. On the other hand, the sensitivity of CD30, CD15 and IMP3 alone were 92.2%, 68.6% and 84.3% respectively. All 23 studied cases of NSCHL, all the 17 cases of MCCHL, 7 out of 8 cases of LRCHL and the only case of LDCHL had predominant T-lymphocytes in their background. On the other hand, the 2 cases of NLPHL and only case of LRCHL had predominant B-lymphocytes in their background. CONCLUSION: IMP3 is a novel marker that is expressed in large proportion of both types of HL against nearly negative background. It has no significant increase in sensitivity in detection of the tumor cells when combined with CD30. There are insignificant relations between IMP3 expression and different clinicopathological parameters. Further studies about IMP3 on a large scale of cases are required to confirm its mechanistic role in generation of HL.

Alteration of oncogenic IGF-II gene methylation status associates with hepatocyte malignant transformation.

BACKGROUND: Oncogenic insulin-like growth factor-II (IGF-II) is overexpressed in hepatocellular carcinoma (HCC). The present study aimed to analyze the dynamic alteration of IGF-II CpG site methylation status and its molecular mechanism in HCC progression. METHODS: IGF-II alterations were observed in rat hepatocarcinogenesis models induced by 2-acetylaminofluorene. Liver IGF-II expression was compared by immunohistochemistry or tissue IGF-II specific concentration (nmol/mg protein). Status of human IGF-II promoter 3 (P3) or rat IGF-II P2 CpG site methylation was amplified by methylation-specific polymerase chain reaction (MSP). Serum IGF-II levels were quantitatively detected by an enzyme-linked immunosorbent assay. RESULTS: The levels of hepatic IGF-II expression were significantly elevated in the HCC group (P < 0.001). The unmethylation rate of IGF-II P3 CpG sites was 100% in the HCC-, 52.5% in the paracancerous-, and none (0%) in the distal noncancerous-tissues. Abnormal IGF-II expression was related to differentiation degree, tumor invasion, and positive HBV-DNA (all P < 0.001), with a negative correlation between P3 methylation degree and IGF-II expression. There was a positive correlation between liver IGF-II specific concentration and circulating IGF-II level (r = 0.97, P < 0.001). Significantly negative correlation was found between IGF-II P2 CpG site methylation and circulating IGF-II (rs = -0.89, P < 0.001) or liver IGF-II level (rs = -0.84, P < 0.001). CONCLUSIONS: The increase of serum IGF-II and the alteration of oncogenic gene IGF-II methylation may be biomarkers for HCC diagnosis and DNA methylation may be the therapeutic target of HCC.

Increased motor neuron resilience by small molecule compounds that regulate IGF-II expression.

The selective vulnerability of motor neurons in amyotrophic lateral sclerosis (ALS) is evident by sparing of a few subpopulations during this fast progressing and debilitating degenerative disease. By studying the gene expression profile of resilient vs. vulnerable motor neuron populations we can gain insight in what biomolecules and pathways may contribute to the resilience and vulnerability. Several genes have been found to be differentially expressed in the vulnerable motor neurons of the cervical spinal cord as compared to the spared motor neurons in CNIII/IV. One gene that is differentially expressed and present at higher levels in less vulnerable motor neurons is insulin-like growth factor II (IGF-II). The motor neuron protective effect of IGF-II has been demonstrated both in vitro and in SOD1 transgenic mice. Here, we have screened a library of small molecule compounds and identified inducers of IGF-II mRNA and protein expression. Several identified compounds significantly protected motor neurons from glutamate excitotoxicity in vitro. One of the compounds, vardenafil, resulted in a complete motor neuron protection, an effect that was reversed by blocking receptors of IGF-II. When administered to naïve rats vardenafil was present in the cerebrospinal fluid and increased IGF-II mRNA expression in the spinal cord. When administered to SOD1 transgenic mice, there was a significant delay in motor symptom onset and prolonged survival. Vardenafil also increased IGF-II mRNA and protein levels in motor neurons derived from healthy subject and ALS patient iPSCs, activated a human IGF-II promoter and improved survival of ALS-patient derived motor neurons in culture. Our findings suggest that modulation of genes differentially expressed in vulnerable and resilient motor neurons may be a useful therapeutic approach for motor neuron disease.

Serum from Chronic Hepatitis B Patients Promotes Growth and Proliferation via the IGF-II/IGF-IR/MEK/ERK Signaling Pathway in Hepatocellular Carcinoma Cells.

BACKGROUND/AIMS: Chronic hepatitis B virus (HBV) infection (CHB) plays a central role in the etiology of hepatocellular carcinoma (HCC). Emerging evidence implicates insulin-like growth factor (IGF)-II as a major risk factor for the growth and development of HCC. However, the relationship between HBV infection and IGF-II functions remains to be elucidated. METHODS: Levels of circulating IGF-II and IGF-I receptor (IGF-IR) in healthy donors (HDs) and CHB patients were tested by ELISA. Human HCC cell lines (HepG-2, SMMC-7721, MHCC97-H) were incubated with serum from HDs and CHB patients at various concentrations for 24, 48, and 72 h. MTT and plate colony formation assays, BrdU ELISA, ELISA, small-interfering RNA (siRNA) transfection, quantitative real-time PCR, and western blot were applied to assess the functional and molecular mechanisms in HCC cell lines. RESULTS: Serum levels of IGF-II and IGF-IR were significantly higher in CHB patients than in HDs. Additionally, serum from CHB patients directly induced cell growth, proliferation, IGF-II secretion, and HDGF-related protein-2 (HRP-2) and nuclear protein 1 (NUPR1) mRNA and protein expression in HCC cells. Moreover, serum from CHB patients increased IGF-II-induced cell growth, proliferation, and HRP-2 and NUPR1 mRNA and protein expression in HCC cells. Blockade of IGF-IR clearly inhibited the above effects. Most importantly, interference with IGF-II function markedly repressed the cell proliferation and HRP-2 and NUPR1 mRNA and protein expression induced by serum from CHB patients. Furthermore, serum from CHB patients induced ERK phosphorylation via IGF-IR, with the MEK inhibitor PD98059 significantly decreasing CHB patient serum-induced IGF-II secretion, cell proliferation, and HRP-2 and NUPR1 mRNA and protein expression. CONCLUSION: Serum from CHB patients increases cell growth and proliferation and enhances HRP-2 and NUPR1 expression in HCC cells via the IGF-II/IGF-IR/MEK/ERK signaling pathway. These findings help to explain the molecular mechanisms underlying HBV-related HCC and may lead to the development of effective therapies.

Regulatory Role for Growth Hormone in Statural Growth: IGF-Dependent and IGF-Independent Effects on Growth Plate Chondrogenesis and Longitudinal Bone Growth.

It was initially thought that the growth-promoting effects of GH were exclusively mediated by liver-derived Insulin-like Growth Factor-I (IGF-I). Subsequent studies demonstrated that GH promotes IGF-I synthesis and activity in other organs and in the growth plate. GH has also IGF-I-independent growth-promoting effects. In Igf1 null mice, high circulating GH levels may be responsible for normal chondrocyte proliferation. Furthermore, tibial growth is reduced more in Ghr null mice than in Igf1 null mice, while the body of mice lacking both Ghr and Igf1 is smaller than that of mice lacking Igf1 or Ghr. The increased IGF-II expression in the growth plate in Igf1 null mice suggests that the IGF-I-independent effects of GH may be mediated by IGF-II. The effects of Igf1 receptor (Igf1r) gene deletion in chondrocytes indicate that GH may promote growth directly at the growth plate even when the local effects of IGF-I and IGF-II are abrogated.

Pediatric Growth Hormone Deficiency (GHD) in the Recombinant Human GH (rhGH) Era.

During the phase of using hGH extracted from pituitaries (pit hGH) - 1958-1985 - fundamental experience related to the diagnosis and treatment was accumulated. However, since recombinant hGH (rhGH) had become available diagnosis and treatment of GHD were conducted world-wide in a more standardized way. Treatment with rhGH was also accompanied by documentations in large international pharmaco-epidemiological surveys, which provided new insight. Despite of this development the treatment of children and adolescents with GHD raises still issues related to the most effective and efficacious as well as safe use of rhGH. This brief review attempts to discuss a few aspects related to these topics as they have developed during the rhGH era.

Silencing IGF-II impairs C-myc and N-ras expressions of SMMC-7721 cells via suppressing FAK/PI3K/Akt signaling pathway.

Emerging evidence confirms that insulin-like growth factor -II (IGF-II), oncogenes C-myc and N-ras are an essential regulator for development and growth in hepatocellular carcinoma (HCC). Although our previous study also indicated that IGF-II might upregulate levels of oncogenes C-myc and N-ras in hepatoma carcinoma cells, the molecular mechanism had not been fully elucidated. Herein, we successfully silenced IGF-II expression in SMCC-7721 cells by small RNA interference. Functional analysis showed that knockdown of IGF-II significantly suppressed growth and proliferation of SMMC-7721 cells and decreased C-myc and N-ras mRNA and protein levels. And this function was mediated by the FAK/PI3K/Akt signaling pathway. Taken together, IGF-II siRNA inactivates the FAK/PI3K/Akt signaling pathway, and further reduces cell proliferation, N-ras and C-myc levels in SMMC-7721 cells. Especially, understanding the relationship between IGF-II and oncogenes N-ras and C-myc in cancer cells will provide novel clues for clinic HCC treatment in the future.

Levels of glucose-regulatory hormones in patients with non-islet cell tumor hypoglycemia: including a review of the literature.

Non-islet cell tumor hypoglycemia (NICTH) is one of the causes of spontaneous hypoglycemia. The pathogenesis of NICTH is thought to be an excessive production by tumors of big insulin-like growth factor (IGF)-II. This study investigated the levels of glucose-regulatory hormones in patients with NICTH with high serum levels of big IGF-II (big IGF-II group) and compared these with profiles of patients with spontaneous hypoglycemia with normal IGF-II (normal IGF-II group). Circulating IRI, CPR, ACTH, cortisol, GH, and IGF-I levels measured during hypoglycemic episodes were examined retrospectively in 37 patients with big IGF-II producing NICTH and 6 hypoglycemic patients with normal IGF-II. The hormone profile data of 15 patients with NICTH from published case reports were reviewed and included in the analyses. Mean plasma glucose levels (36 vs. 29 mg/dL), serum IRI (0.53 vs. 0.37 µIU/mL), CPR (0.15 vs. 0.20 ng/mL), IGF-I SDS (-3.55 vs. -3.18 SD) and ACTH levels (27.3 vs. 33.8 pg/mL) were not significantly different between the big and normal IGF-II groups. However, mean serum GH (0.85 vs. 9.62 ng/mL) and plasma cortisol levels (16.2 vs. 34.5 µg/dL) were significantly lower in the big IGF-II group than in the normal IGF-II group (both p<0.05). In conclusion, although the magnitude of the decrease in insulin and IGF-I levels did not differ between spontaneous hypoglycemic patients caused by other etiologies, patients with NICTH tended to have low basal GH levels during hypoglycemic episodes. These differences in hormone profile may be helpful for selecting patients who require analysis of IGF-II.

The impact of a human IGF-II analog ([Leu27]IGF-II) on fetal growth in a mouse model of fetal growth restriction.

Enhancing placental insulin-like growth factor (IGF) availability appears to be an attractive strategy for improving outcomes in fetal growth restriction (FGR). Our approach was the novel use of [Leu(27)]IGF-II, a human IGF-II analog that binds the IGF-II clearance receptor IGF-IIR in fetal growth-restricted (FGR) mice. We hypothesized that the impact of [Leu(27)]IGF-II infusion in C57BL/6J (wild-type) and endothelial nitric oxide synthase knockout (eNOS(-/-); FGR) mice would be to enhance fetal growth and investigated this from mid- to late gestation; 1 mg·kg(-1)·day(-1) [Leu(27)]IGF-II was delivered via a subcutaneous miniosmotic pump from E12.5 to E18.5. Fetal and placental weights recorded at E18.5 were used to generate frequency distribution curves; fetuses <5th centile were deemed growth restricted. Placentas were harvested for immunohistochemical analysis of the IGF system, and maternal serum was collected for measurement of exogenously administered IGF-II. In WT pregnancies, [Leu(27)]IGF-II treatment halved the number of FGR fetuses, reduced fetal(P = 0.028) and placental weight variations (P = 0.0032), and increased the numbers of pups close to the mean fetal weight (131 vs. 112 pups within 1 SD). Mixed-model analysis confirmed litter size to be negatively correlated with fetal and placental weight and showed that [Leu(27)]IGF-II preferentially improved fetal weight in the largest litters, as defined by number. Unidirectional (14C)MeAIB transfer per gram placenta (System A amino acid transporter activity) was inversely correlated with fetal weight in [Leu(27)]IGF-II-treated WT animals (P < 0.01). In eNOS(-/-) mice, [Leu(27)]IGF-II reduced the number of FGR fetuses(1 vs. 5 in the untreated group). The observed reduction in FGR pup numbers in both C57 and eNOS(-/-) litters suggests the use of this analog as a means of standardizing and rescuing fetal growth, preferentially in the smallest offspring.

Pro-Insulin-Like Growth Factor-II Ameliorates Age-Related Inefficient Regenerative Response by Orchestrating Self-Reinforcement Mechanism of Muscle Regeneration.

Sarcopenia, age-related muscle weakness, increases the frequency of falls and fractures in elderly people, which can trigger severe muscle injury. Rapid and successful recovery from muscle injury is essential not to cause further frailty and loss of independence. In fact, we showed insufficient muscle regeneration in aged mice. Although the number of satellite cells, muscle stem cells, decreases with age, the remaining satellite cells maintain the myogenic capacity equivalent to young mice. Transplantation of young green fluorescent protein (GFP)-Tg mice-derived satellite cells into young and aged mice revealed that age-related deterioration of the muscle environment contributes to the decline in regenerative capacity of satellite cells. Thus, extrinsic changes rather than intrinsic changes in satellite cells appear to be a major determinant of inefficient muscle regeneration with age. Comprehensive protein expression analysis identified a decrease in insulin-like growth factor-II (IGF-II) level in regenerating muscle of aged mice. We found that pro- and big-IGF-II but not mature IGF-II specifically express during muscle regeneration and the expressions are not only delayed but also decreased in absolute quantity with age. Supplementation of pro-IGF-II in aged mice ameliorated the inefficient regenerative response by promoting proliferation of satellite cells, angiogenesis, and suppressing adipogenic differentiation of platelet derived growth factor receptor (PDGFR)α(+) mesenchymal progenitors. We further revealed that pro-IGF-II but not mature IGF-II specifically inhibits the pathological adipogenesis of PDGFRα(+) cells. Together, these results uncovered a distinctive pro-IGF-II-mediated self-reinforcement mechanism of muscle regeneration and suggest that supplementation of pro-IGF-II could be one of the most effective therapeutic approaches for muscle injury in elderly people.

Involvement of IGF-II receptors in the antioxidant and neuroprotective effects of IGF-II on adult cortical neuronal cultures.

Insulin-like growth factor-II (IGF-II) is a naturally occurring peptide that exerts known pleiotropic effects ranging from metabolic modulation to cellular development, growth and survival. IGF-II triggers its actions by binding to and activating IGF (IGF-I and IGF-II) receptors. In this study, we assessed the neuroprotective effect of IGF-II on corticosterone-induced oxidative damage in adult cortical neuronal cultures and the role of IGF-II receptors in this effect. We provide evidence that treatment with IGF-II alleviates the glucocorticoid-induced toxicity to neuronal cultures, and this neuroprotective effect occurred due to a decrease in reactive oxygen species (ROS) production and a return of the antioxidant status to normal levels. IGF-II acts via not only the regulation of synthesis and/or activity of antioxidant enzymes, especially manganese superoxide dismutase, but also the restoration of mitochondrial cytochrome c oxidase activity and mitochondrial membrane potential. Although the antioxidant effect of IGF-I receptor activation has been widely reported, the involvement of the IGF-II receptor in these processes has not been clearly defined. The present report is the first evidence describing the involvement of IGF-II receptors in redox homeostasis. IGF-II may therefore contribute to the mechanisms of neuroprotection by acting as an antioxidant, reducing the neurodegeneration induced by oxidative insults. These results open the field to new pharmacological approaches to the treatment of diseases involving imbalanced redox homeostasis. In this study, we demonstrated that the antioxidant effect of IGF-II is at least partially mediated by IGF-II receptors.