Differential production of insulin-like growth factor-binding proteins in liver fibrosis progression.

Noninvasive methods for liver disease diagnoses offer great advantages over biopsy, but they cannot be utilized in all cases. Therefore, specific indicators for chronic liver disease management are necessary. The aim was to assess the production of insulin-like growth factor-binding proteins (IGFBPs) 1-7 and their correlation with the different stages of fibrosis in chronic hepatitis C (CHC). A prospective, cross-sectional, multicenter study was conducted. CHC patients were categorized by FibroTest® and/or FibroScan®. Serum concentrations of IGFBPs 1-7 were determined through multiple suspension arrangement array technology. Significant differences were validated by the Kruskal-Wallis and Mann-Whitney U tests. Logistic regression models were performed to assess the association between the IGFBPs and fibrosis stages. The association was determined utilizing odds ratios (ORs), and receiver operating characteristic (ROC) curves were constructed to distinguish the IGFBPs in relation to the diagnosis of fibrosis. IGFBP-1 and IGFBP-7 concentrations were higher in CHC than in the healthy individuals, whereas IGFBP-3, IGFBP-5, and IGFBP-6 were downregulated in the patients. An apparent increase of all the IGFBPs was found at fibrosis stage F4, but with different regulations. IGFBP-2, -4, -6, and -7 had the best OR, showing the relation to fibrosis progression. The ROC curves showed that IGFBP-7 was the only protein that distinguished F1 from F3 and F2 from F3. IGFBPs participate in liver fibrosis progression and could be employed as circulating novel protein panels for diagnosis and as possible therapeutic targets in liver fibrosis progression.

Extensive serum biomarker analysis in patients with non-small-cell lung carcinoma.

Lung cancer is a common malignant disease, nearly 2.09 million new patients occurred last year. Approximately 85% of the patients are classified as non-small-cell lung cancer (NSCLC). It is therefore important to identify new diagnostic and prognostic biomarkers for the early detection of this disease. The presented study identifies biomarkers in the serum of NSCLC patients. The expression of 274 cytokines was measured by a novel antibody array methodology and ELISA was applied to validate the array results. The levels of MIP-1 α, IL-8, MIP-1 ß, Resistin, GDF-15, HGF, CA125, FLRG, VCAM-1, DKK-3, sTNF-R1, CTACK, Acrp30, CXCL-16 and LYVE-1 were significantly higher in serum from NSCLC patients, while the level of TIMP-2 and IGFBP-6 were lower. More importantly, the validation supported the result of the antibody array. The result of the antibody array indicates that these cytokines might be novel auxiliary biomarkers in the diagnosis and prognosis of NSCLC.

Protein microarray analysis identifies key cytokines associated with malignant middle cerebral artery infarction.

INTRODUCTION: We aimed to explore potential cytokines involved in the malignant middle cerebral artery infarction (MMI) and elucidate their underlying regulatory mechanisms. METHODS: We first developed a cytokine profile by Quantibody® Human Cytokine Antibody Array7000 using serum samples from eight patients with MMI and eight patients with non-acute cerebral infarction (NACI). The differentially expressed cytokines were then identified in patients with MMI using two-tailed Student's t-test and Fisher's Exact Test compared with patients with NACI. Gene Ontology and pathway enrichment analyses were performed using DAVID. Protein-protein interaction (PPI) network was constructed based on STRING database. RESULTS: A total of 10 differentially expressed cytokines were identified from 320 unique inflammatory cytokines in serums. Among them, four cytokines, like NCAM1 (neural cell adhesion molecule 1), IGFBP-6 (insulin-like growth factor binding protein 6), LYVE1 (lymphatic vessel endothelial hyaluronan receptor 1), and LCN2 (Lipocalin2), were up-regulated, while another six cytokines, such as TGFB1 (transforming growth factor, beta 1, also known as LAP), EGF (epidermal growth factor), PDGFA (platelet-derived growth factor alpha polypeptide), MMP-10 (matrix metallopeptidase 10), IL-27 (interleukin 27), and CCL2 (chemokine (C-C motif) receptor 2), were down-regulated. Moreover, cytokine-cytokine receptor interaction pathway was significantly enriched. CONCLUSIONS: Our findings indicate that 10 differentially expressed cytokines, such as NCAM1, LCN2, IGFBP-6, LYVE1, MMP-10, IL-27, PDGFA, EGF, CCL2, and TGFB1 may participate in the development of MMI. Moreover, cytokine-cytokine receptor interaction pathway may be an important mechanism involved in this disease. These differentially expressed cytokines may serve as diagnostic biomarkers or drug targets for MMI.

Correlation Between IGFs-Related Proteins Expression and Incidence of Colorectal Cancer in Diabetic Patients and Related Mechanisms.

BACKGROUND: Diabetes mellitus a common metabolic disorder with hyperglycemia, is caused by the interaction of genetic and environmental factors. Approximately 12~20% of diabetic patients have risk of colorectal cancer. Recent studies revealed that the insulin-like growth factor system (IGFs) plays an important role in tumor occurrence. This study thus investigated the relationship between IGFs-related proteins in diabetic patients and the incidence of colorectal carcinoma. MATERIAL/METHODS: A retrospective study was performed in a total of 206 individuals, including 85 diagnosed with diabetes. The incidence of colorectal cancer was tracked, along with the detection of IGFs expression in serum. During the surgical resection, tumor tissues and adjacent tissues were collected and quantified for IGFs expression level. RESULTS: We found no significant difference in age or sex between the diabetic and control groups. Diabetic patients, however, had elevated body weight and higher incidence of colorectal cancer compared to non-diabetic controls (p<0.05). The diabetic group also had higher IGF-I and IGF-IR mRNA levels in serum, while IGFBP-6 expression was down-regulated. In comparison to adjacent healthy tissues, tumor tissue had higher levels of IGF-I and IGF-IR but lower levels of IGFBP-6 (p<0.05). CONCLUSIONS: Our study showed higher incidence of colorectal cancer in diabetics compared to non-diabetics. The occurrence of colorectal cancer in diabetic patients may be associated with elevated IGFs-related protein expression level.

Quantitative analysis of the concentrations of IGFs and several IGF-binding proteins in a large fibrous abdominal tumor and the circulation of a patient with hypoglycemia.

The syndrome of nonislet cell tumor induced hypoglycemia (NICTH) represent extreme cases of excessive expression and production of incompletely processed high-molecular-mass pro-IGF-II forms (big IGF-II) by an often large tumor. Tumor-derived big IGF-II is responsible for enhanced insulin-like effects in the body through complicated mechanisms, leading to hypoglycemia. Case studies on NICTH usually focus on measurements of diagnostic parameters in the circulation of patients. Some studies have also reported on qualitative immunohistochemical analysis of tumor tissue, in particular with respect to the expression of IGF-II at the mRNA or protein level. However, quantitative data on the concentrations of IGFs and IGFBPs in tumor specimen causing NICTH, in relation to their corresponding plasma levels are lacking. Such an analysis would provide an estimate of the total potential of (big) IGF-II retained by the tumor and more insight in the relative levels of different IGFBPs and their origin in the circulation, that is, systemically induced by tumor related factors or directly tumor-derived. Here we investigated quantitatively the levels of IGFs and IGFBPs in a large, 1.76 kg weighing, solitary fibrous tumor from a typical case of NICTH using highly specific immunometric assays. Besides a high level of big IGF-II, patient's plasma also contained increased levels of both IGFBP-2 and -6 which declined after removal of the tumor. These IGFBPs have a higher affinity for (pro-) IGF-II than IGF-I and exhibit intrinsic IGF-independent bioactivities. Tumor tissue contained high amounts of big IGF-II and IGFBP-6, exceeding that in patient's circulation many-fold. A relatively low tumor content of IGFBP-2 was found suggesting that the preoperative high levels in plasma were attributable to systemic mechanisms. The background literature and possible implications of these findings are briefly discussed. Based on the present results we postulate that tumor tissue is not the source of the elevated levels of IGFBP-2 often seen in NICTH patients. Large tumors that cause NICTH can produce IGFBP-6 leading to enhanced levels of this IGFBP in the circulation. Hence, the measurement of IGFBP-6 in plasma may serve as an additional marker of this disease pattern.

Circulating Proteomic Signatures of Chronological Age.

To elucidate the proteomic features of aging in plasma, the subproteome targeted by the SOMAscan assay was profiled in blood samples from 202 females from the TwinsUK cohort. Findings were replicated in 677 independent individuals from the AddNeuroMed, Alzheimer's Research UK, and Dementia Case Registry cohorts. Results were further validated using RNAseq data from whole blood in TwinsUK and the most significant proteins were tested for association with aging-related phenotypes after adjustment for age. Eleven proteins were associated with chronological age and were replicated at protein level in an independent population. These were further investigated at gene expression level in 384 females from the TwinsUK cohort. The two most strongly associated proteins were chordin-like protein 1 (meta-analysis ß [SE] = 0.013 [0.001], p = 3.66 × 10(-46)) and pleiotrophin (0.012 [0.005], p = 3.88 × 10(-41)). Chordin-like protein 1 was also significantly correlated with birthweight (0.06 [0.02], p = 0.005) and with the individual Framingham 10-years cardiovascular risk scores in TwinsUK (0.71 [0.18], p = 9.9 × 10(-5)). Pleiotrophin is a secreted growth factor with a plethora of functions in multiple tissues and known to be a marker for cardiovascular risk and osteoporosis. Our study highlights the importance of proteomics to identify some molecular mechanisms involved in human health and aging.

CC chemokine ligand 18 and IGF-binding protein 6 as potential serum biomarkers for prostate cancer.

Prostate cancer (PCa) is the second leading cause of cancer-related death in men globally. However, there are few sensitive biomarkers for PCa, especially those which can distinguish PCa and benign prostate hyperplasia (BPH). Antibody microarrays allow for high-throughput and high-sensitivity detection of multiple proteins simultaneously, providing a powerful tool for biomarker screening. Here, we selected 46 patients with PCa and 42 controls with BPH, and compared the serum levels of different cytokines in PCa and BPH patients using antibody microarrays. The results indicated that serum levels of macrophage colony-stimulating factor (M-CSF) and CC chemokine ligand 18 (CCL-18) were remarkably higher in PCa patients than those in BPH patients, while serum levels of insulin-like growth factor-binding protein 6 (IGFBP-6) and Fas receptor (Fas), also called tumor necrosis factor receptor superfamily member 6 (TNFRSF6), were significantly lower. M-CSF and Fas/TNFRSF6 have been reported to be associated with PCa pathogenesis, and thus were used as positive controls in the present study. CCL-18 is a chemokine primarily involved in recruitment of the adaptive immune system, while IGFBP-6 has been reported to inhibit proliferation of PCa cells. Serum levels of these four cytokines could distinguish PCa from BPH with high sensitivity and high specificity. Furthermore, the area under the ROC curve (AUC) was above 0.925 and 0.835 for CCL-18 and IGFBP-6, respectively, implying their high diagnostic value. In conclusion, we have identified CCL-18 and IGFBP-6 as new potential serum biomarkers for PCa.

Expression of IGFBP­6 in proliferative vitreoretinopathy rat models and its effects on retinal pigment epithelial­J cells.

Proliferative vitreoretinopathy (PVR) is one of the most common causes for failed retinal detachment surgeries. The aim of the present study was to investigate the role of insulin­like growth factor­binding protein­6 (IGFBP­6) in PVR using rat models and its effects on retinal pigment epithelial­J (RPE­J) cells. PVR Wistar rat models were administered intravitreal injection of RPE­J cells (1x106/5 µl) combined with platelet­rich plasma (1x107/5 µl). The concentration of IGFBP­6 in the vitreous and serum of rats was tested by an enzyme­linked immunosorbent assay and the expression of IGFBP­6 mRNA in the liver and retina of rats was determined by quantitative polymerase chain reaction (qPCR). The expression of IGFBP­6 mRNA in the RPE­J cells stimulated by vitreous or serum from PVR patients or normal volunteers was also determined by qPCR. The proliferation of RPE­J cells was evaluated by the 3­(4,5­dimethylthiazol­2­yl)­5­(3­carboxymethoxyphenyl)­2­(4­sulfophenyl)­2H­tetrazolium, inner salt (MTS) method. The success rate of PVR rat model induction at the 8th week was 89.5% (34/38). The concentration of IGFBP­6 in the vitreous and serum of PVR rats was significantly higher than that of the control group (P<0.05). The expression of IGFBP­6 mRNA in the retina of PVR rats was also significantly higher compared with the control group (P<0.05). The vitreous from PVR patients and donors significantly stimulated the expression of IGFBP­6 mRNA in the RPE­J cells (P<0.05). IGFBP­6 only inhibited IGF­II­stimulated proliferation but not the basal level of proliferation or the PDGF/VEGF­stimulated RPE­J cell proliferation. Thus, the trends and effects of IGFBP­6 provide the possibility of PVR therapeutic targets, with the vitreous representing a significant environmental factor in the progression of PVR.

Kininogen 1 and insulin-like growth factor binding protein 6: candidate serum biomarkers of proliferative vitreoretinopathy.

BACKGROUND: The aim was to validate whether kininogen 1 (KNG1) or insulin-like growth factor binding protein 6 (IGFBP-6) are serum biomarkers of proliferative vitreoretinopathy (PVR). METHODS: Samples from vitreous and corresponding serum samples were collected from patients with PVR. The donor vitreous samples and serum samples from healthy volunteers and volunteers who had undergone vitrectomies for other conditions were used as controls. The samples were subsequently analysed using Western blotting (WB) and enzyme-linked immunosorbent assay. RESULTS: The Western blotting outcomes indicated both IGFBP-6 and KNG1 could be specifically detected in the vitreous and serum samples of patients with PVR. The concentrations of KNG1 and IGFBP-6 were significantly higher in both vitreous and serum samples from patients with severe PVR than in the samples from patients with moderate PVR. The serum concentrations of KNG1 or IGFBP-6 had decreased by the post-vitrectomy examinations. The receiver operating characteristic (ROC) analyses when the concentrations of IGFBP-6 or KNG1 were greater than 181.4 pg/ml or 441.75 ng/ml, respectively, predicted severe PVR with both a sensitivity and specificity of over 70 per cent. When the concentrations of IGFBP-6 or KNG1 were greater than 98.5 pg/ml or 88.5 ng/ml, respectively, they predicted the PVR prognosis with both a sensitivity and specificity of 80 per cent. CONCLUSIONS: KNG1 and IGFBP-6 may be candidate serum biomarkers of PVR.

Female recruits sustaining stress fractures during military basic training demonstrate differential concentrations of circulating IGF-I system components: a preliminary study.

OBJECTIVE: Stress fracture injuries sustained during military basic combat training (BT) are a significant problem and occur at a higher rate in female recruits than male recruits. Insulin-like growth factor-I (IGF-I) is an easily measured biomarker that is involved in bone formation and positively correlated with bone mineral density, especially in women. This study examined the response of the IGF-I system between female soldiers that sustained a stress fracture (SFX, n=13) during BT and female soldiers who did not (NSFX, n=49). DESIGN: Female soldiers (n=62, 18.8 ± 0.6 yr) from 2 companies of a gender-integrated combat battalion in the Israeli Defense Forces participated in this study. Height, weight and blood draws were taken upon entry to BT (preBT) and after a four-month BT program (postBT). Stress fractures were diagnosed by bone scan. Serum was analyzed for total IGF-I, free IGF-I, IGF binding proteins (IGFBP)1-6, BAP, calcium, CTx, IL1ß, IL6, PINP, PTH, TNFα, TRAP, and 25(OH)D. Statistical differences between SFX and NSFX groups and time points were assessed by RM ANOVA with Fisher post-hoc (p≤0.05). RESULTS: The SFX group was significantly taller and had lower BMI than NSFX (p≤0.05). Serum concentrations of total IGF-I, bioavailable IGF-I, other bone biomarkers, and cytokines were not significantly different between SFX and NSFX preBT. Serum IGFBP-2 and IGFBP-5 were significantly higher in the SFX compared to the NSFX preBT (p≤0.05). In both groups, total IGF-I increased pre to postBT (p≤0.05). Additionally, a significant difference was observed in the bioavailable IGF-I response pre to postBT for both groups. The SFX group demonstrated a significant decrease in bioavailable IGF-I pre to postBT (preBT: 0.58 ± 0.58 ng/mL; postBT 0.39 ± 0.48; p≤0.05) whereas the NSFX group demonstrated a significant increase in bioavailable IGF-I pre to postBT (preBT: 0.53 ± 0.37 ng/mL; postBT: 0.63 ± 0.45; p≤0.05). CONCLUSIONS: Our study demonstrated that serum IGF-I changes during basic training and that women sustaining stress fractures during BT significantly decreased bioavailable IGF-I, whereas their uninjured counter parts increased bioavailable IGF-I. These results suggest that stress fracture susceptibility may be related to differential IGF-I system concentrations and response to physical training.

[Kininogen-1 and insulin-like growth factor binding protein-6 as serum biomarkers for proliferative vitreoretinopathy].

OBJECTIVE: To evaluated the predictive potential of kininogen-1 and insulin-like growth factor binding protein-6 (IGFBP-6) for PVR. METHODS: Vitreous and serum samples were obtained from 24 PVR patients. Vitreous from 8 donated normal eyes, and serum samples from 20 healthy volunteers served as control. Patients who underwent vitrectomy with C(3)F(8) gas tamponade (n = 15) and silicone tamponade (n = 8) and patients who experienced recurrent retinal detachment after scleral buckling surgery (n = 8) were recruited for serum tests as well. Western blot analysis was employed to detect the presence of kininogen-1 and IGFBP-6. The protein concentration was measured by using enzyme linked immunosorbent assay (ELISA) analysis. All date were analyzed with the SPSS 3.0 for Windows (only-way analysis of variance and t test). RESULTS: Western blot analysis displayed that except that IGFBP-6 was absent in 2 PVR vitreous, both kininogen-1 and IGFBP-6 were otherwise found in all PVR vitreous and serum samples. Neither kininogen-1 nor IGFBP-6 can be detected in normal vitreous or serum samples. Protein expression was more intensive in severe PVR vitreous than in mild PVR vitreous, which was confirmed by a significantly higher concentration of each protein in sever PVR vitreous. The ELISA outcomes documented that kininogen-1 concentration in vitreous were significantly higher in severe PVR patients than those in mild PVR (281.0 ± 63.0 & 237.5 ± 32.1) µg/L (t = 5.44, P < 0.05). Kininogen-1 was about 2 times higher in serum than in vitreous (443.3 ± 190.1) µg/L (t = 5.27, P < 0.05). At 6 months after vitrectomy with gas tamponade in 15 patients, their kininogen-1 level in serum was significantly lower than that of preoperation (81.9 ± 18.6 & 443.3 ± 190.1) µg/L (t = 5.26, P < 0.05) and encircling failure group was (116.8 ± 45.1) µg/L, it was higher than that of normal and silicone tamponade groups (t = 3.95, 4.34;P < 0.05). Similarly, IGFBP-6 concentration in vitreous were significantly higher in severe PVR patients than those in mild PVR (352.9 ± 64.4 & 283.9 ± 69.9) ng/L (t = 5.08, P < 0.05) and its level in serum was (185.3 ± 34.9) ng/L and lower than that of in vitreous(t = 7.95, P < 0.05). At 6 months after vitrectomy with gas tamponade in 15 patients, their IGFBP-6 level in serum decreased comparing that of preoperation (65.4 ± 31.8) ng/L (t = 11.10, P < 0.05) and encircling failure group was (109.2 ± 6.6) ng/L, it was higher than that of normal and silicone tamponade groups (t = 3.16, 2.77; P < 0.05). CONCLUSIONS: Kininogen-1 and IGFBP-6 are presented in serum and vitreous in PVR patients. The strength of protein expression is related to the severity of PVR. These results suggested that kininogen-1 and IGFBP-6 can be biomarkers for severe PVR.

Growth hormone-binding protein is directly and IGFBP-3 is inversely associated with risk of female breast cancer.

OBJECTIVE: Several components of the GH and IGF systems have been implicated in the development of malignancies. All components of these hormonal systems have never been jointly evaluated in female breast cancer, and previous studies have not examined the role of IGF-binding proteins (IGFBP-4, IGFBP-6) or GH-binding protein (GHBP). DESIGN: Hospital-based case-control study. METHODS: In this sample of primarily postmenopausal women, we obtained serum measures of IGF-I, IGF-II, and binding proteins IGFBP-1, IGFBP-3, IGFBP-4, IGFBP-6, as well as GHBP, insulin, and leptin from 74 breast cancer cases and 76 control subjects. RESULTS: In crude analyses, we found lower age-standardized mean IGF-I, IGFBP-3, IGFBP-4, IGFBP-6, and higher IGFBP-1 and GHBP in breast cancer cases when compared with controls. Multivariate models mutually adjusted for other GH-IGF system components and classical breast cancer risk factors demonstrated an inverse association between IGFBP-3 and risk of breast cancer (odds ratio (OR) = 0.2, P < 0.01) and a direct association between GHBP and disease risk (OR = 3.3, P < 0.01). No significant associations were detected in multivariate analyses among IGF-I, IGF-II or IGFBP-1, IGFBP-4, IGFBP-6 with risk of breast cancer, indicating that these factors may not have effects independent of and/or comparable with IGFBP-3 and GHBP. CONCLUSIONS: These results support a protective role of IGFBP-3 and demonstrate for the first time an increased risk of breast cancer with higher GHBP, after accounting for variation in IGFs, IGFBPs, and classical breast cancer risk factors.

The IGF system in a case of Costello syndrome.

Costello syndrome is characterized by facial dysmorphia, hyperpigmented skin, palmar and plantar hyperkeratosis, curly hair, perioral and nasal papillomata (more rarely localized anally and on vocal cords), short stature, mental retardation and sociable personality. Although growth retardation is typical of Costello syndrome, its cause is not defined. We report on a 10-yr-old Caucasian girl affected by Costello syndrome with fasting hypoglycemia and short stature, associated low circulating levels of acid-labile subunit (ALS), relatively low levels of IGF-I and IGFBP-3, and normal IGF-II, mostly circulating in a binary complex with IGFBP-2 and -6 instead of in a 150 kDa ternary complex. The reduced ALS concentration and the consequent impaired formation of the circulating 150 kDa ternary complex can induce an accelerated clearance rate of IGF peptides and of IGFBP-3, contributing to the decreased IGF-I growth promoting activity in our patient. Moreover, the presence of IGF-II in the binary complex, which has been postulated to increase the insulin-like effects of these peptides, can explain, at least in part, the patient's asymptomatic fasting hypoglycemia.

Ethinylestradiol and testosterone have divergent effects on circulating IGF system components in adolescents with constitutional tall stature.

OBJECTIVE: Pharmacological doses of estrogens or testosterone are used to limit the final height of girls or boys with constitutional tall stature but the mechanism behind this growth inhibition is still debated. We therefore studied the changes in the circulating components of the insulin-like growth factor (IGF) system during high dose sex steroid therapy. DESIGN AND METHODS: Twenty three girls and twenty boys with constitutional tall stature were treated with 100 microg ethinylestradiol per day or 250 mg testosterone ester every 14 days respectively. In 19 girls and 18 boys, the levels of IGF-I, free IGF-I, IGF-II, acid-labile subunit (ALS) and IGF binding proteins (IGFBP)-2 to -6 were measured before and 3-6 months after the start of therapy (group 1). In 18 girls and 11 boys, samples were collected at the end of therapy and 3 to 6 months afterwards (group 2). Fourteen girls and nine boys belonged to both groups. All parameters were measured by radioimmunoassay or ELISA. RESULTS: Levels of IGF-I were decreased significantly by estrogen treatment but remained unchanged during testosterone treatment. Free IGF-I decreased during estrogen treatment but increased during testosterone therapy. Estrogens increased IGF-II and testosterone reduced it. The important reduction of IGFBP-2 during estrogen therapy is not reproduced by androgen therapy, neither is the stimulation by estrogens of IGFBP-4. IGFBP-3 is not modulated by either sex steroid. We found that IGFBP-6 is up-regulated by testosterone but not by estrogens; the reverse is true for ALS, which increased during estrogen treatment but remained unchanged during testosterone treatment. CONCLUSIONS: Our findings demonstrate that androgens and estrogens exert differential effects on the circulating levels of several IGF components.

Mutations in the NSD1 gene in patients with Sotos syndrome associate with endocrine and paracrine alterations in the IGF system.

OBJECTIVE: To investigate the effect of nuclear receptor Su-var, 3-9, enhancer of zeste, trithorax (SET) domain-containing protein 1 (NSD1) gene alteration in patients with Sotos syndrome on plasma IGFs and IGF-binding proteins (IGFBPs), as well as on the IGF/IGFBP system activity at the tissue level. DESIGN: Twenty-nine patients suspected of Sotos syndrome were divided into two groups: patients with heterozygous deletions or mutations in the NSD1 gene (NSD1(+/-)) (n=11) and subjects without (NSD1(+/+)) (n=18). Plasma samples (n=29) and skin fibroblasts (n=23) were obtained. The results of both groups were compared and related to reference values. METHODS: IGF-I, IGF-II, IGFBP-2, IGFBP-3, IGFBP-4 and IGFBP-6 levels were determined by RIAs. The mitogenic response of fibroblasts to IGFs was investigated by [methyl-(3)H]thymidine incorporation. IGFBP-3 levels in the culture media were measured by RIA. IGFBP-3 mRNA expression was determined by real time RT-PCR. RESULTS: NSD1(+/-) patients showed significantly altered levels of IGF-I (mean-1.2 SDS), IGF-II (-1.2), IGFBP-3 (-1.7), IGFBP-4 (-0.4), IGFBP-2 (+0.8) and IGFBP-6 (+1.5). The NSD1(+/+) patients did not differ from the reference, with the exception of the mean IGFBP-3 level (-1.3). Basal proliferation and mitogenic response to IGFs was diminished in NSD1(+/-) fibroblasts compared with NSD1(+/+) (basal, P=0.02; IGF-I, P<0.001; IGF-II, P=0.02). Compared with control fibroblasts, only the mitogenic response was diminished (basal, P=0.07; IGF-I, P=0.04; IGF-II, P=0.04). A trend of higher IGFBP-3 secretion after IGF-I stimulation (P=0.09) and 3.5-5 times higher mRNA expression of IGFBP-3 in basal conditions was found in NSD1(+/-) fibroblasts in comparison to controls. CONCLUSIONS: NSD1(+/-) patients show endocrine and paracrine changes in the IGF system. These changes may contribute to the abnormal growth pattern.

Negative energy balance plays a major role in the IGF-I response to exercise training.

Circulating IGF-I is correlated with fitness, but results of prospective exercise training studies have been inconsistent, showing both increases and decreases in IGF-I. We hypothesized that energy balance, often not accounted for, is a regulating variable such that training plus an energy intake deficit would cause a reduction in IGF-I, whereas training plus energy intake excess would lead to an increased IGF-I. To test this, 19 young, healthy men completed a 7-day strenuous exercise program in which they were randomly assigned to either a positive energy balance [overfed (OF), n = 10] or negative energy balance [underfed (UF), n = 9] group. IGF-I (free and total), insulin, and IGF-binding protein-1 were measured before, during, and 1 wk after the training. Weight decreased in the UF subjects and increased in the OF subjects. Free and total IGF-I decreased substantially in the UF group (P < 0.0005 for both), but, in the OF group, IGF-I remained unchanged. The UF group also demonstrated an increase in IGF-binding protein-1 (P < 0.027), whereas glucose levels decreased (P < 0.0005). In contrast, insulin was reduced in both the OF and UF exercise-training groups (P < 0.044). Finally, within 7 days of the cessation of the diet and training regimen, IGF-I and IGF-binding protein-1 in the UF group returned to preintervention levels. We conclude that energy balance during periods of exercise training influences circulating IGF-I and related growth mediators. Exercise-associated mechanisms may inhibit increases in IGF-I early in the course of a training protocol, even in overfed subjects.

The relationship between serum levels of insulin-like growth factor-I and its binding proteins and microvascular function in acromegalic patients.

The system of insulin-like growth factor-I (IGF-I) and its binding proteins is thought to be involved in the pathogenesis of vascular damage under different pathological circumstances. The results of various studies are rather controversial. This study considers the relationship between the activity of this system and the function of microcirculation in acromegalic patients. Thirteen patients with hormonally active acromegaly and 15 healthy controls were included in the study. The growth hormone, free IGF-I, IGF-I, IGF binding protein (IGFBP) -1, -2, -3 and -6 serum levels and parameters of lipid metabolism were determined. The function of microcirculation was determined by laser Doppler fluxmetry and the intima media thickness of the common carotid artery was measured by ultrasound. We noted significant reduction in postocclusive reactive hyperaemia (PORH(max)) (P < 0.01), in thermal hyperaemia (TH(max)) (P < 0.05) and in the velocity of reaction in both tests in the group of acromegalic patients. A significant negative correlation between free IGF-I serum levels and maximal perfusion during thermal hyperaemia TH(max) (P < 0.02) was found in the control group. Statistically significant positive correlation between free IGF-I serum levels and the time to maximal perfusion in postocclusive reactive hyperaemia PORH(max) (P < 0.05) was found in the group with hormonally active acromegaly. Moreover, a positive relationship between IGFBP-1 serum levels and serum levels of total (P < 0.01) and low density lipoprotein (LDL) (P < 0.05) cholesterol was found in the group of patients with acromegaly. We conclude that the function of microcirculation is impaired in patients with acromegaly and that free IGF-I serum levels may affect the microvascular function as measured by laser Doppler fluxmetry. In addition, we found a significant relationship between the serum levels of IGFBP-1 and those of total and LDL cholesterol in the group of patients with hormonally active acromegaly.

Insulin-like growth factors and insulin-like growth factor binding proteins in adult patients with severe liver disease before and after orthotopic liver transplantation.

INTRODUCTION: The liver is the main source of serum insulin-like growth factors (IGFs) and IGF-binding proteins (IGFBPs) and the concentration of these proteins might reflect liver function. METHODS: In a retrospective longitudinal study we examined serum levels of total and free IGF-I, IGF-II, IGFBP-1, IGFBP-2, IGFBP-3 and IGFBP-6 in 21 adult patients with end-stage liver disease before and after orthotopic liver transplantation (LTX) by sensitive and specific RIAs. In each patient, the mean value of at least three measurements before and after LTX was calculated. RESULTS: Before LTX, serum levels of total and free IGF-I, IGF-II, IGFBP-3 were low and showed a rapid and significant increase in almost all patients after successful LTX (total IGF-I: 30 +/- 7 vs. 256 +/- 30 ng/ml, p < 0.001; free IGF-I: 1.3 +/- 0.3 vs. 3.5 +/- 0.6 ng/ml, p < 0.01; IGF-II: 177 +/- 28 vs. 618 +/- 30 ng/ml, p < 0.001; IGFBP-3: 1,230 +/- 136 vs. 3,665 +/- 264 ng/ml, p < 0.001). In contrast, IGFBP-1 was found to be high immediately before LTX, and declined to normal levels after LTX (210 +/- 40 vs. 90 +/- 15 ng/ml, p < 0.01), while IGFBP-2 did not show any significant changes (1,154 +/- 296 vs. 1,303 +/- 192 ng/ ml). Positive correlations were found between IGF-I, IGF-II or IGFBP-3, and serum pseudocholinesterase (R = 0.50, 0.72 and 0.61 respectively, p < 0.001). Negative correlations were found between IGF-I, IGF-II or IGFBP-3, and prothrombin time (R = 0.50, 0.59 and 0.51 respectively, p < 0.001). CONCLUSION: Patients with severe liver disease show decreased levels of total and free IGF-I, IGF-II and IGFBP-3, and increased levels of IGFBP-1. These abnormalities are promptly normalized after successful LTX. Thus, serum levels of IGF-I, IGF-II and IGFBP-3 might be useful parameters for the assessment of liver function.

Gender differences in regional body composition and somatotrophic influences of IGF-I and leptin.

This study evaluated the arm, trunk, and leg for fat mass, lean soft tissue mass, and bone mineral content (BMC) assessed via dual-energy X-ray absorptiometry in a group of age-matched (approximately 29 yr) men (n = 57) and women (n = 63) and determined their relationship to insulin-like growth factor I (IGF-I) and leptin. After analysis of covariance adjustment to control for differences in body mass between genders, the differences that persisted (P < or = 0.05) were for lean soft tissue mass of the arm (men: 7.1 kg vs. women: 6.4 kg) and fat mass of the leg (men: 5.3 kg vs. women: 6.8 kg). Men and women had similar (P > or = 0.05) values for fat mass of the arms and trunk and lean soft tissue mass of the legs and trunk. Serum IGF-I and insulin-like growth factor binding protein-3 correlated (P < or = 0.05) with all measures of BMC (r values ranged from 0.31 to 0.39) and some measures of lean soft tissue mass for women (r = 0.30) but not men. Leptin correlated (P < or = 0.05) similarly for measures of fat mass for both genders (r values ranging from 0.74 to 0.85) and for lean soft tissue mass of the trunk (r = 0.40) and total body (r = 0.32) for men and for the arms in women (r = 0.56). These data demonstrate that 1) the main phenotypic gender differences in body composition are that men have more of their muscle mass in their arms and women have more of their fat mass in their legs and 2) gender differences exist in the relationship between somatotrophic hormones and lean soft tissue mass.

No evidence of insulin-like growth factor-binding protein 3 proteolysis during a maximal exercise test in elite athletes.

The aim of the present study was to examine the GH/insulin-like growth factor (IGF) axis, post exercise, with emphasis on IGF-binding protein (IGFBP)-3 proteolysis. Sixteen elite rowers (8 female/8 male) performed a stepwise submaximal rowing test followed by a 6- to 7-min-long maximal test. Blood samples were drawn at baseline, t = 0 (end of exercise) and t = 15, 30, 60, 90, and 120 min. GH and IGFBP-1 levels increased post exercise (P < 0.0005). Total IGF-I and IGF-II increased significantly post exercise (P < 0.0005) but not after albumin correction. Free IGF-I decreased after exercise with nadir coincidently with the IGFBP-1 peak, and free IGF-II decreased post exercise coincidently with the IGFBP-6 peak. IGFBP-3, measured by immunoradiometric assay, increased after exercise (P < 0.0005) but not after albumin adjustment. IGFBP-3 proteolysis (%) (measured by a specific in vitro proteolytic activity assay) and IGFBP-3 (measured by Western ligand blotting) were unchanged post exercise. Albumin-adjusted levels of IGFBP-6 increased by 18% (P < 0.0005), whereas IGFBP-2 and IGFBP-4 did not change significantly post exercise. Our findings do not support the hypothesis that short-term strenuous exercise induces major acute changes in the GH/IGF axis. To what degree the protein anabolic effects of regular exercise are associated with acute alterations in the GH/IGF axis remains unclear.