Joop A. M. J. L. Janssen

Identification of the BclI polymorphism in the glucocorticoid receptor gene: association with sensitivity to glucocorticoids in vivo and body mass index

objective  Sensitivity to glucocorticoids differs between individuals, partially due to genetic variation in the glucocorticoid receptor (GR) gene. We studied the sequence alteration of a previously described intronic BclI polymorphism of the GR gene, and investigated whether there was an association with sensitivity to glucocorticoids and anthropometric parameters in a group of healthy elderly individuals.

Insulin-like Growth Factors, Their Binding Proteins, and Prostate Cancer Risk: Analysis of Individual Patient Data from 12 Prospective Studies

Context Insulin-like growth factors (IGFs) and IGF binding proteins may be associated with some cancers. Contribution This reanalysis of individual patient data from 12 studies of the association between IGFs and IGF binding proteins and prostate cancer suggests that higher levels of serum IGF-I are associated with higher risk for prostate cancer. Caution The 12 studies varied in the types of patients they studied and in how they measured IGFs. Implication High IGF-I levels seem to be a risk factor for prostate cancer. The Editors Prostate cancer is one of the most common types of cancer in men, yet few risk factors for the disease, other than age, race, and a family history, have been established (1, 2). Insulin-like growth factors (IGFs) and their associated binding proteins (IGFBPs) have been the subject of many epidemiologic investigations of prostate cancer because they are known to help regulate cell proliferation, differentiation, and apoptosis (3). Although results from some, but not all, studies suggest an association between IGFs and IGFBPs and prostate cancer risk, there has been much uncertainty about its consistency and magnitude. A previous meta-analysis that included only 3 prospective studies suggested that high levels could be associated with more than a 2-fold increase in risk (4), although recent studies have suggested the risk is lower. Furthermore, given that these peptides are correlated with each other, uncertainty remains about any observed relationships. The individual studies are rarely large enough to allow proper mutual adjustment for these correlated factors, and they are insufficiently powered to investigate the consistency of their findings in key subgroups (for example, stage and grade of disease). Such analyses are important because studies have suggested that IGF-I might be more associated with advanced than with localized disease (5, 6). The Endogenous Hormones and Prostate Cancer Collaborative Group was established to conduct collaborative reanalyses of individual data from prospective studies on the relationships between circulating levels of sex hormones and IGFs and subsequent prostate cancer risk. Results for the sex hormones have been reported elsewhere and show no statistically significant relation between androgen or estrogen levels in men and the subsequent risk for prostate cancer (7). We report results for concentrations of IGFs and IGFBPs. Methods Participants The Endogenous Hormones and Prostate Cancer Collaborative Group is described in detail elsewhere (7). In brief, the group invited principal investigators of all studies, found by searching PubMed, Web of Science, and CancerLit, that provided data on circulating concentrations of sex steroids, IGFs or IGFBPs, and prostate cancer risk by using prospectively collected blood samples to join the collaboration. Thirteen studies collected data on circulating IGF concentrations and the subsequent risk for prostate cancer (5, 6, 820), of which 1 contributed only data on sex hormones (20). Eleven of the studies used a matched casecontrol design nested within a prospective cohort study (5, 6, 812, 16, 19) or a randomized trial (1315, 17). One study used a casecohort design (18) and was converted into a matched casecontrol design by randomly matching up to 3 control participants to each case patient by age at recruitment, time between blood collection and diagnosis, time of blood draw, and race. (Table 1 provides a full description of the studies and matching criteria used.) Most of the prospective studies were population-based, with the exception of 1 based on health plan members (9), 1 that recruited male health professionals (16), and 1 that was a combination of an intervention study and a monitoring study for cardiovascular disease (6, 10). Two of the randomized trials did not have prostate cancer as a primary end point (5, 8, 15); the other 2 were based within a screening trial (13) or were about treatment of prostate-specific antigen (PSA)detected prostate cancer (14). Table 1. Study Characteristics Individual participant data were available for age; height; weight; smoking status; alcohol consumption; marital status; socioeconomic status (assessed by educational achievement); race; concentrations of IGFs, IGFBPs, and endogenous sex steroids; and PSA level. Information sought about prostate cancer included date of diagnosis, stage and grade of disease, and method of case patient ascertainment. Some studies (5, 6, 8, 10, 16) published more than 1 article or performed assays at different times on the association between IGFs and prostate cancer risk, sometimes with different matched casecontrol sets, laboratory measurements, and durations of follow-up. For each study, we created a single data set in which each participant appeared only once. In our analysis, we treated any participant who appeared in a study as both a control participant and a case patient as a case patient only. We removed matched set identifiers, and we generated a series of strata (equivalent to matched sets) in which participants in each study were grouped according to age at recruitment (2-year age bands) and date of recruitment (by year), because these matching criteria were common to most studies (Table 1). The number of strata used in the collaborative analysis was slightly less than that of matched sets used in the original analyses. To ensure that this process did not introduce any bias, we checked that the results for each study, using the original matched sets, were the same as those using the strata described above. Tumors were classified as advanced if the tumor was described as extending beyond the prostate capsule (T3/T4), and/or there was lymph node involvement (N1/N2/N3), and/or there were distant metastases (M1); tumors were classified as localized if they were T0/T1/T2 and N0/NX and M0. We classified tumors as high-grade if they had a Gleason score of 7 or more or were moderately poorly or poorly differentiated; otherwise, they were classified as low-grade. Statistical Analysis We calculated partial correlation coefficients between log-transformed IGF and IGFBP concentrations among control participants, adjusted for age at blood collection (<50, 50 to 59, 60 to 69, or 70 years) and study. For each IGF and IGFBP, we categorized men into quintiles of IGF and IGFBP serum concentrations, with cut-points defined by the study-specific quintiles of the distribution within control participants. For studies with more than 1 publication or in which the serum assays were done at different times, resulting in different absolute levels of IGFs (5, 6, 8, 10, 16), we calculated cut-points separately for each substudy. We used a conditional logistic regression stratified by study, age at recruitment (2-year age bands), and date of recruitment (single year) as our main method of analysis. To provide a summary measure of risk, we calculated a linear trend by scoring the quintiles of the serum IGF or IGFBP concentrations as 0, 0.25, 0.5, 0.75, and 1. Under the assumption of linearity, a unit change in this trend variable is equivalent to the odds ratio (OR) comparing the highest with the lowest quintile. All results are unadjusted for participant characteristics, except for those controlled by the stratification variables. We examined the possible influence of 5 participant characteristics by adjusting the relevant conditional logistic regression models for body mass index (BMI) (<22.5, 22.5 to 24.9, 25.0 to 27.4, 27.5 to 29.9, or >30 kg/m2), marital status (married or cohabiting, or not married or cohabiting), educational status (did not attend college or university, or attended college or university), smoking (never, previous, or current), and alcohol consumption (<10 or 10 g/d). We excluded participants from the analysis if they had a missing value for the characteristic under examination. We assessed heterogeneity in linear trends among studies by using a chi-square statistic to test whether the study-specific ORs were statistically different from the overall OR (21). Heterogeneity among studies was also quantified by calculating the H and I 2 statistics (22). To test whether the linear trend OR estimates for each IGF and IGFBP varied according to case patient characteristics, we estimated a series of subsets for each characteristic: stage at diagnosis (localized or advanced), grade at diagnosis (low or high), year of diagnosis (before 1990, 1990 to 1994, or 1995 onward; these year cutoffs were chosen to attempt to reflect differences in the use of the PSA test for cancer detection), age at diagnosis (<60, 60 to 69, or 70 years), and time between blood collection and diagnosis (<3, 3 to 6, or 7 years). We excluded case patients from the analyses of stage and grade at diagnosis if the relevant information was not available. For each of these case patient characteristics, we calculated a heterogeneity chi-square statistic to assess whether the estimated ORs statistically differed from each other (21). To assess whether the OR estimate of the linear trend for each IGF or IGFBP varied according to PSA level at recruitment (<2 g/L or 2 g/L), we entered an interaction term into the conditional logistic regression model for each IGF or IGFBP, and we tested the statistical significance of the interaction term with a likelihood ratio test. Statistical significance was set at the 5% level. All statistical tests were 2-sided. All statistical analyses were done with Stata, version 9.0 (StataCorp, College Station, Texas). Results Table 1 shows the characteristics of the studies. The 12 prospective studies included approximately 3700 case patients with prostate cancer and 5200 control participants. Insulin-like growth factor I and IGFBP-III measurements were available for all and 3600 case patients, respectively. However, IGF-II and IGFBP-II measurements were available for only 379 and 419 case patients, respectively (Table 2). Mean age at blood collection

Association between genetic variation in the gene for insulin-like growth factor-l and low birthweight

Low birthweight is associated with later risk of type 2 diabetes and related disorders. We aimed to show that a polymorphism in the gene for insulin-like growth factor-I, which has proved to raise risk of type 2 diabetes and myocardial infarction, is associated with low birthweight. We recorded birthweight and obtained DNA for 463 adults. Individuals who did not have the wild-type allele of the polymorphism had a 215 g lower birthweight than those homozygous for this allele (95% CI -411 to -10). Our data lend support to the hypothesis that genetic variation affecting fetal growth could account for the association between low birthweight and susceptibility to diabetes and cardiovascular disease in later life.

Serum free IGF-I, total IGF-I, IGFBP-1 and IGFBP-3 levels in an elderly population: relation to age and sex steroid levels.

Most previous studies concerning the relationship between IGF‐I and age used assays measuring total IGF‐I. Although free IGF‐I is considered of greater biological relevance, little is known about its relationship with sex steroids levels in elderly healthy subjects.

Unacylated ghrelin is active on the INS-1E rat insulinoma cell line independently of the growth hormone secretagogue receptor type 1a and the corticotropin releasing factor 2 receptor

Both unacylated ghrelin (UAG) and acylated ghrelin (AG) exert metabolic effects. To investigate the interactions between AG and UAG on ghrelin receptors we evaluated the effects of AG and UAG on INS-1E rat insulinoma cells, using insulin secretion after 30min static incubation as a read-out. A possible involvement of the growth hormone secretagogue receptor type 1a (GHS-R1a) or the corticotropin-releasing factor 2 (CRF2) receptor (CRF2R), as a putative receptor for UAG, was also studied determining their mRNA expression and the functional effects of receptor antagonists on insulin release. Both UAG and AG stimulated insulin release dose-dependently in the nanomolar range. The AG-induced insulin output was antagonized by two GHS-R1a antagonists ([d-Lys(3)]GHRP-6 and BIM28163), which did not block UAG actions. These effects occurred in the presence of low levels of GHS-R1a mRNA. Neither CRF2R expression nor effects of the CRF2R antagonist (astressin(2)B) on insulin output were observed. In conclusion, we provide a sensitive and reproducible assay for specific effects of UAG, which in this study is responsible for insulin release by INS-1E cells. Our data support the existence of a specific receptor for UAG, other than the CRF2R and GHS-R1a. The stimulatory effect on insulin secretion by AG in this cell line is mediated by the GHS-R1a.

The influence of an insulin-like growth factor I gene promoter polymorphism on hip bone geometry and the risk of nonvertebral fracture in the elderly: the Rotterdam Study.

The absence of the wildtype allele of a promoter polymorphism of the IGF‐I gene is associated with increased risk (1.5; 95% CI, 1.1‐2.0) of fragility fracture in women (n = 4212) but not in men (n = 2799). An approximation of hip bone geometry (from DXA) suggested the polymorphism is associated with bone strength and stability in gender‐specific ways.

Endocrine responses to ghrelin in adult patients with isolated childhood-onset growth hormone deficiency

objective Ghrelin, a 28 amino acid acylated peptide, is a natural ligand of the GH secretagogues (GHS) receptor (GHS‐R), which is specific for synthetic GHS. Similar to synthetic GHS, ghrelin strongly stimulates GH secretion but also displays significant stimulatory effects on lactotroph and corticotroph secretion. It has been hypothesized that isolated GH deficiency (GHD) could reflect hypothalamic impairment that would theoretically involve defect in ghrelin activity.

The Asp727Glu polymorphism in the TSH receptor is associated with insulin resistance in healthy elderly men

Background  Variations in thyroid function within the normal range are associated with differences in metabolism and body composition. For instance, TSH is positively associated with body mass index (BMI). This could be due to alterations in thyroid hormone activity, or to direct effects of TSH, as the TSH receptor (TSHR) is also expressed in adipose tissue. The TSHR‐Asp727Glu polymorphism is associated with lower serum TSH levels in vivo. In this study, we analysed whether serum thyroid parameters and the TSHR‐Asp727Glu polymorphism were associated with glucose metabolism and insulin resistance. In addition, we analysed the Thr92Ala polymorphism in the type 2 deiodinase (D2), which was recently associated with insulin resistance.

Effects of type I interferons on IGF-mediated autocrine/paracrine growth of human neuroendocrine tumor cells

We recently demonstrated that interferon (IFN)-beta has a more potent antitumor activity than IFN-alpha in BON cells, a neuroendocrine tumor (NET) cell line. The present study showed the role of type I IFNs in the modulation of the insulin-like growth factor (IGF) system in NETs. BON cells expressed IGF-I, IGF-II, IGF-I receptor, and insulin receptor mRNA. In addition, IGF-I and IGF-II stimulated the proliferation of BON cells and induced an inhibition of DNA fragmentation (apoptosis). As evaluated by quantitative RT-PCR, treatment with IFN-alpha (100 IU/ml) or IFN-beta (100 IU/ml) inhibited the expression of IGF-II mRNA (-42% and -65%, respectively, both P < 0.001), whereas IGF-I receptor mRNA was significantly upregulated by IFN-alpha (+28%, P < 0.001) and downregulated by IFN-beta (-47%, P < 0.001). Immunoreactive IGF-II concentration decreased in the conditioned medium during IFN-alpha (-16%, P < 0.05) and IFN-beta (-69%, P < 0.001) treatment. Additionally, IGF-I receptor bioactivity was reduced (-54%) after IFN-beta treatment. Scatchard analysis of (125)I-labeled IGF-I binding to cell membrane of BON cells revealed a dramatic suppression of maximum binding capacity only in the presence of IFN-beta. Finally, the proapoptotic activity of IFN-beta was partially counteracted by the coadministration of IGF-I and IGF-II (both at 50 nM). In conclusion, these data demonstrate that the IGF system has an important role in autocrine/paracrine growth of BON cells. The more potent antitumor activity of IFN-beta compared with IFN-alpha could be explained by several effects on this system: 1) both IFNs inhibit the transcription of IGF-II, but the suppression is significantly higher after IFN-beta than IFN-alpha and 2) only IFN-beta inhibits the expression of IGF-I receptor.

Genomewide meta-analysis identifies loci associated with IGF-I and IGFBP-3 levels with impact on age-related traits

The growth hormone/insulin‐like growth factor (IGF) axis can be manipulated in animal models to promote longevity, and IGF‐related proteins including IGF‐I and IGF‐binding protein‐3 (IGFBP‐3) have also been implicated in risk of human diseases including cardiovascular diseases, diabetes, and cancer. Through genomewide association study of up to 30 884 adults of European ancestry from 21 studies, we confirmed and extended the list of previously identified loci associated with circulating IGF‐I and IGFBP‐3 concentrations (IGF1, IGFBP3, GCKR, TNS3, GHSR, FOXO3, ASXL2, NUBP2/IGFALS, SORCS2, and CELSR2). Significant sex interactions, which were characterized by different genotype–phenotype associations between men and women, were found only for associations of IGFBP‐3 concentrations with SNPs at the loci IGFBP3 and SORCS2. Analyses of SNPs, gene expression, and protein levels suggested that interplay between IGFBP3 and genes within the NUBP2 locus (IGFALS and HAGH) may affect circulating IGF‐I and IGFBP‐3 concentrations. The IGF‐I‐decreasing allele of SNP rs934073, which is an eQTL of ASXL2, was associated with lower adiposity and higher likelihood of survival beyond 90 years. The known longevity‐associated variant rs2153960 (FOXO3) was observed to be a genomewide significant SNP for IGF‐I concentrations. Bioinformatics analysis suggested enrichment of putative regulatory elements among these IGF‐I‐ and IGFBP‐3‐associated loci, particularly of rs646776 at CELSR2. In conclusion, this study identified several loci associated with circulating IGF‐I and IGFBP‐3 concentrations and provides clues to the potential role of the IGF axis in mediating effects of known (FOXO3) and novel (ASXL2) longevity‐associated loci.