BACKGROUND

The sentinel lymph node (SLN) concept has been gaining attention for gastrointestinal neoplasms but remains controversial for esophageal cancer. This study evaluated the feasibility of SLN identification using intraoperative indocyanine green (ICG) fluorescence imaging (IGFI) navigated by preoperative computed tomographic lymphography (CTLG) to treat superficial esophageal cancer.

METHODS

Subjects comprised 20 patients clinically diagnosed with superficial esophageal cancer. Five minutes after endoscopic submucosal injection of iopamidol around the primary lesion using a four-quadrant injection pattern with a 23-gauge endoscopic injection sclerotherapy needle, three-dimensional multidetector computed tomography was performed to identify SLNs and lymphatic routes. ICG solution was injected intraoperatively around the tumor. Fluorescence imaging was obtained by infrared ray electronic endoscopy. Thoracoscope-assisted standard radical esophagectomy with lymphadenectomy was performed to confirm fluorescent lymph nodes detected by CTLG.

RESULTS

Lymphatic vessels and SLNs were identified preoperatively using CTLG in all cases. Intraoperative detection rates were 100% using CTLG and 95% using IGFI. Lymph node metastases were found in four cases, including one false-negative case with SLNs occupied by bulky metastatic tumor that were not enhanced with both methods. The other 19 cases, including three cases of metastatic lymph nodes, were accurately identified by both procedures.

CONCLUSIONS

Preoperative CTLG visualized the correct number and site of SLNs in surrounding anatomy during routine computed tomography to evaluate distant metastases. Referring to CTLG, SLNs were identified using IGFI, resulting in successful SLN navigation and saving time and cost. This method appears clinically applicable as a less-invasive method for treating superficial esophageal cancer.

BACKGROUND

We investigated whether peptides involved in cellular proliferation and apoptosis, [insulin-like growth factor I (IGFI) and its major binding protein (insulin-like growth factor binding protein 3)], predicted risk of benign prostate hyperplasia (BPH).

METHODS

We conducted a nested-case-control study in the placebo arm of the prostate cancer prevention trial (PCPT). Cases (n = 727) were men with surgical or medical treatment for BPH; two or more IPSS scores >14; or two scores of at least five points over baseline one of which was>>or=12. Controls (n = 727) were frequency matched by age to cases, reported no BPH treatment, and no IPSS score >8. Cases and controls remained on the PCPT placebo and were followed closely until their 7-year PCPT anniversary. Baseline serum was analyzed for IGFI and IGFBP3. Unconditional logistic regression and polytomous regression estimated the multivariate-adjusted odds ratio (OR) for BPH risk.

RESULTS

IGFBP3 was inversely and the IGFI:IGFBP3 ratio was positively associated with BPH risk, but findings were statistically significant only for men with severe symptoms (OR = 0.60, 95% CI = 0.40-0.90 for the fifth vs. first quintile of IGFBP3, P-trend = 0.01). Associations did not differ by age (<65 or>>or=65 years), and there was a suggestion that the IGFI:IGFBP3 - BPH risk association may be stronger among overweight men.

CONCLUSIONS

A high IGFI:IGFBP3 ratio was associated with increased BPH risk, and high serum IGFBP3 was associated with decreased BPH risk among men with severe symptoms. These results confirm findings from other recent studies.

A short review of the literature first confirms the clinical value of cathepsin D as a prognostic marker in breast cancer, when using well standardized assays. We then summarize results of studies, mostly performed in our laboratory, aimed at understanding the effect of cathepsin D overexpression on metastasis and the molecular mechanisms involved. Cathepsin D-cDNA transfection increases tumor cell proliferation in vitro and the metastatic potential of 3Y1-Ad12 embryonic rat tumorigenic cells when injected in vivo into nude mice. The mechanism by which cathepsin D increases the incidence of clinical metastasis involves increased cell growth and decreased contact inhibition rather than escape of cancer cells through the basement membrane. Different mechanisms are considered to explain this mitogenic activity. Cathepsin D could act as a protease following its activation at an acidic pH, or as a ligand of different membrane receptors at a more neutral pH. In this case cathepsin D can displace IGFII from the mannose-6-phosphate/IGFII receptor to the IGFI receptor or activate another membrane receptor to be identified. The nature of the mechanisms involved in vivo may depend on the micro environment of the tumor cells. These studies should guide in the development of new therapies aimed at inhibiting the deleterious effect of overexpressed cathepsin D.

OBJECTIVE

This phase I, multicenter, open-label, single-arm, dose-escalation, and dose-expansion study evaluated the safety, tolerability, and antitumor activity of MEDI-573 in adults with advanced solid tumors refractory to standard therapy or for which no standard therapy exists.

METHODS

Patients received MEDI-573 in 1 of 5 cohorts (0.5, 1.5, 5, 10, or 15 mg/kg) dosed weekly or 1 of 2 cohorts (30 or 45 mg/kg) dosed every 3 weeks. Primary end points included the MEDI-573 safety profile, maximum tolerated dose (MTD), and optimal biologic dose (OBD). Secondary end points included MEDI-573 pharmacokinetics (PK), pharmacodynamics, immunogenicity, and antitumor activity.

RESULTS

In total, 43 patients (20 with urothelial cancer) received MEDI-573. No dose-limiting toxicities were identified, and only 1 patient experienced hyperglycemia related to treatment. Elevations in levels of insulin and/or growth hormone were not observed. Adverse events observed in >10% of patients included fatigue, anorexia, nausea, diarrhea, and anemia. PK evaluation demonstrated that levels of MEDI-573 increased with dose at all dose levels tested. At doses >5 mg/kg, circulating levels of insulin-like growth factor (IGF)-I and IGFII were fully suppressed. Of 39 patients evaluable for response, none experienced partial or complete response and 13 had stable disease as best response.

CONCLUSIONS

The MTD of MEDI-573 was not reached. The OBD was 5 mg/kg weekly or 30 or 45 mg/kg every 3 weeks. MEDI-573 showed preliminary antitumor activity in a heavily pretreated population and had a favorable tolerability profile, with no notable perturbations in metabolic homeostasis.

Somatomedin-C (SM-C) and insulin-like growth factor I (IGF-I) produced identical curves of competition in a RIA for SM-C, and in placental cell membrane receptor assays for SM-C and insulin. Somatomedin-A (SM-A), insulin-like growth factor II (IGF-II) and two forms of multiplication stimulating activity (MSA) were less than 5% as potent as SM-C/IGF-I in the RIA and less than 50% that of SM-C/IGFi in the SM-C receptor assay. In the insulin receptor assay, IGF-II and MSA III-2 were much more potent than SM-C/IGF-I. The present data suggest that SM-C and IGF-I are functionally identical substances.

We have investigated the role of the C-terminal of the alpha-subunit in the insulin receptor family by characterizing chimeric mini-receptor constructs comprising the first three domains (468 amino acids) of insulin receptor (IR) or insulin-like growth factor I receptor (IGFIR) combined with C-terminal domain from either insulin receptor (IR) (residues 704-719), IGFIR, or insulin receptor-related receptor (IRRR). The constructs were stably expressed in baby hamster kidney cells and purified, and binding affinities were determined for insulin, IGFI, and a single chain insulin/IGFI hybrid. The C-terminal domain of IRRR was found to abolish binding in IR and IGFIR context, whereas other constructs bound ligands. The two constructs with first three domains of the IR demonstrated low specificity for ligands, all affinities ranging from 3.0 to 15 nM. In contrast, the constructs with the first three domains of the IGFIR had high specificity, the affinity of the novel minimized IGFIR for IGFI was 1.5 nM, whereas the affinity for insulin was more than 3000 nM. When swapping the C-terminal domains in either receptor context only minor changes were observed in affinities (<3-fold), demonstrating that the carboxyl-terminal of IR and IGFIR alpha-subunits are interchangeable and suggesting that this domain is part of the common binding site.

Insulin and insulin-like growth factors (IGFs) belong to the most biologically characterized family of peptides involved in metabolism, growth and development. The cellular responses to the IGFs are mediated primarily by the IGF-I receptor. The IGF-I receptor is a member of the family of tyrosine kinase growth factor receptors, and is highly homologous (70%) to the insulin receptor, especially in the tyrosine kinase domain (84%) ADDIN. Upon ligand binding to the extracellular region, the intrinsic tyrosine kinase domain of the receptor is activated. In the past it was believed that insulin activates primarily metabolic processes while IGFs promote cell growth and differentiation. However, in the last two decades many animal models of IGFI deficiency and excess revealed the importance of IGF-I in carbohydrate and lipid metabolism and now it is clear that these peptide hormones together with growth hormone (GH) work in a coordinate and interdependent manner. In the circulation, IGFs are bound in a binary complex with a family of high affinity IGF-binding proteins (IGFBPs) ADDIN. However, most of the circulating IGF-I associates with a high molecular weight complex approximately 150 KDa consisting of IGFBP-3 and the acid labile subunit (ALS) ADDIN. Once the ternary complex dissociates, the binary complexes of IGFBP-IGFs are removed from the circulation and by crossing the endothelium to reach the target tissues and to interact with cell surface receptors. In the present review we will summarize the role of GH and IGF in somatic growth and focus on the metabolic effects of IGF-I deficiency as assessed in various mouse models.

Insulin-like growth factor-I (IGF-I) is emerging as a powerful neuroprotective molecule that is strongly induced in the central nervous system after different insults. We constructed a recombinant adenoviral vector (RAd-IGFI) harboring the gene for rat IGF-I and used it to implement IGF-I gene therapy in the hypothalamus of senile female rats, which display hypothalamic dopaminergic (DA) neurodegeneration and as a consequence, chronic hyperprolactinemia. Restorative IGF-I gene therapy was implemented in young (5 months) and senile (28 months) female rats, which received a single intrahypothalamic injection of 3 x 10(9) plaque-forming units of RAd-betagal (a control adenoviral vector expressing beta-galactosidase) or RAd-IGFI and were killed 17 days post-injection. In the young animals, neither vector modified serum prolactin levels, but in the RAd-IGFI-injected senile rats a nearly full reversion of their hyperprolactinemic status was recorded. Morphometric analysis revealed a significant increase in the total number of tyrosine hydroxylase-positive cells in the hypothalamus of experimental as compared with control senile animals (5874+/-486 and 3390+/-498, respectively). Our results indicate that IGF-I gene therapy in senile female rats is highly effective for restoring their hypothalamic DA dysfunction and thus reversing their chronic hyperprolactinemia.

Apoptosis is an important cellular process by which superfluous or unwanted cells are deleted from an organism during tissue remodeling and differentiation. Recent studies have demonstrated the role of this programmed cell death or "controlled cell suicide" in the physiological function of an organism. Suppression of apoptosis increases the susceptibility of an individual to malignancy whereas uncontrolled cell death is associated with degenerative diseases. Normal development of both female and male gonads is characterized by massive cell death. More than 99% of ovarian follicles endowed at early life are destined to undergo apoptosis and the exhaustion of these follicles serves as a "clock" for female reproductive senescence. In the testis, up to 75% of male germ cells also undergo apoptosis, perhaps as a mechanism to delete superfluous or defective germ cells. Gonadal cell apoptosis provides valuable models to study hormonal regulation of apoptosis. In the ovary, gonadotropins, estrogens, growth hormone, growth factors (IGFI, EGF/TGF-alpha, basic FGF), cytokine (interleukin-1 beta) and nitric oxide act in concert to ensure the survival of preovulatory follicles. In contrast, androgens, interleukin-6 and gonadal GnRH-like peptide are apoptotic factors. Developmental studies further indicate that fractions of endowed follicles are recruited throughout the reproductive life whereas most of the primordial follicles are "arrested" at the initial stage of development for a prolonged time. Because a transcriptional factor WT1 is expressed in high levels in follicles at early stages of development and because WT1 over-expression represses the promoter activity of inhibin-alpha gene, this nuclear protein may be important in the maintenance of follicles at early stages of development. Once a cohort of follicles is recruited to grow, it is destined to undergo apoptosis unless rescued by survival factors. After puberty onset and under gonadotropin stimulation, some of the growing antral follicles are "selected" to continue their final maturation and secrete high levels of estrogens to trigger ovulation. Following repeated cycles of recruitment, atresia or ovulation, the follicle reserve is exhausted, thus signaling the onset of reproductive senescence. Although the somatic granulosa cell is the major cell type undergoing apoptosis in the ovary, the germ cells in the testis also exhibit signs of apoptotic cell demise. In the testis, gonadotropins and androgens act as survival factors whereas exposure to elevated temperature in cryptorchid testes increases apoptosis. In the seasonally breeding hamster model, photoperiod-entrained regression and recrudescence of testis tissue serves as a unique natural model of apoptosis. With recent advances in our understanding of the cellular mechanism of apoptosis, including the elucidation of the Ced9/bc12 and Ced3/ICE family of proteins, further investigation of gonadal apoptosis may lead to a better understanding of gonadal degenerative disorders (such as premature ovarian failure and oligospermia), reproductive senescence and tumorigenesis. The gonadal model should also be valuable in studying the regulation of intracellular apoptosis genes by external hormonal signals.

Caloric restriction (CR), the consumption of fewer calories without malnutrition, and reduced insulin and/or IGFI receptor signaling delay many age-related physiological changes and extend the lifespan of many model organisms. Here, we present and review microarray and biochemical studies indicating that the potent anticancer effects of CR and disrupted insulin/IGFI receptor signaling evolved as a byproduct of the role of many mitotic tissues as reservoirs of metabolic energy. We argue that the longevity effects of CR are derived from repeated cycles of apoptosis and autophagic cell death in mitotically competent tissues and protein turnover and cellular repair in postmitotic tissues. We review studies showing that CR initiated late in life can rapidly induce many of the benefits of lifelong CR, including its anticancer effects. We also discuss evidence from liver and heart indicating that many benefits of lifelong CR are recapitulated in mitotic and postmitotic tissues when CR is initiated late in life.

We investigated the effect of insulin-like growth factors II and I (IGFII and IGFI) on septal primary cultures from mouse embryonic day 15 brains. The addition of IGFII to septal cultures enhanced total choline acetyltransferase (ChAT) activity in a dose-dependent manner. Maximal stimulation of ChAT activity was observed at 10 ng/ml IGFII. The effect of IGFII on ChAT activity was completely blocked by anti-IGFII/M-6-P receptor antibodies, whereas the antisera alone had no effect on the enzyme activity. Double-labeled immunohistochemical studies revealed that most ChAT-positive neurons expressed IGFII/M-6-P receptor immunoreactivity. These results indicate that the trophic effect of IGFII results from the direct action of this molecule through the IGFII/M-6-P receptor in septal cholinergic neurons. IGFI also stimulated ChAT activity, but with less potency than IGFII. Antibodies against the IGFII/M-6-P receptor inhibited approximately 50% of the IGFI response, suggesting that the effect of IGFI is mediated in part by the IGFII/M-6-P receptor. Thus, it appears that IGFII and IGFI are potent trophic factors for central cholinergic neurons and could potentially play a significant role in the differentiation, maintenance and regeneration of these neurons.

Patients diagnosed with HNPCC harbouring a confirmed germline mutation in DNA mismatch repair (MMR) genes have an 80% lifetime risk of developing an epithelial malignancy. There is, however, considerable variation in the age of disease onset in these patients. Insulin-like growth factor-I (IGFI) has been implicated in colorectal cancer (CRC), and elevated plasma IGFI levels are associated with both sporadic and hereditary CRC risk. In this study, we further investigate the cytosine-adenine (CA) dinucleotide repeat polymorphism located near the promoter region of IGF1 and its relation to early onset CRC risk in 443 Australian and Polish MMR gene mutation carriers using DNA sequencing, Kaplan-Meier survival curves and Cox proportional hazard regression analysis. A significantly smaller number of IGF1 CA repeats was observed in the Polish patient population, which was associated with an earlier age of disease onset compared to the Australian patients. The threshold for the observed modifying effect was again shown to be in patients with 17 or less CA repeats compared to those with 18 or more. Furthermore, when MMR mutation group (i.e., MLH1 or MSH2), gender and family clustering were included in the final Cox model we observed a more robust trend for the role of the IGF1 CA repeat in predicting age of disease onset in HNPCC patients. In addition, this effect was shown to be equal in both MLH1 and MSH2 mutation carrier groups and not restricted to a particular MMR subgroup (p = 0.001). We conclude that the IGF1 CA repeat is an important modifier of disease onset in HNPCC and the first polymorphism to yield consistent results across different populations.

BACKGROUND

Increased myocyte apoptosis in diabetic hearts has been previously reported. The purpose of this study was to evaluate the effects of exercise training on cardiac survival pathways in streptozotocin (STZ)-induced diabetic rats.

METHODS

Forty-eight male Wistar rats were randomly divided into control group (Control), STZ-induced (65 mg/kg, i.p.) diabetes (DM), and DM rats with moderate aerobic exercise training (DM-EX) on a treadmill 60 min/day, 5 days/week, for 10 weeks. Histopathological analysis, positive TUNEL assays and Western blotting were performed on the excised cardiac left ventricles from all three groups.

RESULTS

The components of cardiac survival pathway (insulin-like growth factor I (IGFI), IGFI-receptor (IGFI-R), phosphatidylinositol 3'-kinase (PI3K), and Akt) and the pro-survival Bcl-2 family proteins (Bcl-2, Bcl-xL, and p-BAD) were all significantly decreased in the DM group compared with the Control group whereas they were increased in the DM-EX group. In addition, the abnormal myocardial architecture, enlarged interstitial space and increased cardiac TUNEL-positive apoptotic cells were observed in the DM group, but they were reduced in the DM-EX group. The apoptotic key component, caspase-3, was significantly increased in the DM group relative to the Control group whereas it was decreased in the DM-EX group.

CONCLUSIONS

Exercise training enhances cardiac IGFI-R/PI3K/Akt and Bcl-2 family associated pro-survival pathways, which provides one of the new beneficial effects for exercise training in diabetes.

A link between inflammation of the colon in inflammatory bowel disease (IBD) and the increased risk of colon cancer in ulcerative colitis (UC) may be provided by growth factor receptor genes. Their expression may be altered in response to growth factors present in the mucosa, and this, in turn, may induce further genetic changes, linked to carcinogenesis, in the cells of the colonic epithelium. To test this hypothesis, we assayed steady-state levels of eight growth factor receptor mRNAs in colonic epithelial cells of IBD patients and controls. Four of these genes (EGF-R, IGFI-R, CSF1-R, and PDGF-R-beta) were expressed in epithelial cells, whereas four (erbB-2, erbB-3, NGF-R, and met) were not. The level of the former in involved or uninvolved IBD was considerably lower than in normal epithelial cells from either sporadic colon cancer or diverticulitis patients. In contrast, expression was much higher in IBD patients with colon tumors than in active chronic IBD. The level of PDGF-R-beta mRNA was two- to fourfold higher in involved than in uninvolved areas of the colons of two UC patients, but not in one Crohn's disease patient. Message abundance of its ligand, PDGF-beta, however, was the same in paired UC samples. The pattern of expression of PDGF-beta and cripto was identical to that of EGF-R, whereas the level of mRNA of amphiregulin was the same in active chronic IBD and IBD patients with tumors. A fourth growth factor, Kfgf, was not expressed. Increased levels of PDGF-R-beta mRNA in involved UC relative to uninvolved UC may be related to the disease process in UC. Decreased expression of growth factor- and growth factor receptor-encoded mRNA in active chronic IBD may be related to the disease process, or it may be an effect of steroid therapy undergone by these patients. Enhanced expression of these genes in IBD patients with tumors compared to those without tumors suggests that this may be a marker for development of colon cancer in IBD.

Vitamin D3 and metformin are widely used in humans for regulating mineral metabolism and as an antidiabetic drug, respectively; and both of them have been shown to have chemopreventive effects against various tumors. This study was designed to investigate the potential synergistic chemopreventive effects of vitamin D3 and metformin against the development of early colon neoplasia in two models. The first model was a 1,2-dimethylhydrazine dihydrochloride (DMH)-induced colon cancer rat model and the second, a DMH-dextran sodium sulfate (DSS)-induced colitis-associated colon neoplasia mouse model. Compared with either vitamin D3 or metformin alone, combined use of vitamin D3 and metformin showed more pronounced effect in reducing the numbers of aberrant crypt foci (ACF) and tumor in the colon. The most prominent inhibitory effects were observed in the vitamin D3 medium dose (100 IU/kg/d) and metformin medium dose (120 mg/kg/d) combination group. Furthermore, our results showed that enhancement of metformin's chemopreventive effects by vitamin D3 was associated with downregulation of S6P expression, via the AMPK (IGFI)/mTOR pathway. In addition, enhancement of vitamin D3's chemopreventive effects by metformin was associated with inhibition of the protein expressions of c-Myc and Cyclin D1, via the vitamin D receptor/β-catenin pathway. These findings show that the combined use of vitamin D3 and metformin exhibits synergistic effects against the development of early colon neoplasia. They suggest that the combined use of vitamin D3 and metformin may represent a novel strategy for chemoprevention of colorectal cancer.

OBJECTIVE

To investigate the feasibility and efficacy of anatomical liver resection (ALR) guided by fused images comprising a macroscopic view and indocyanine green fluorescence imaging (fusion IGFI).

BACKGROUND

ALR is established in treating hepatocellular carcinoma or other malignancies to achieve curability and functional preservation. However, the conventional demarcation technique (CDT) marks only the organ surface and sometimes fails to execute a completely valid demarcation.

METHODS

Twenty-four consecutive ALRs for focal liver malignancy were studied using fusion IGFI. Indocyanine green was administered systemically after selective inflow clamping in 12 patients or by portal puncture and direct injection in 12 patients, and we compared demarcation findings between fusion IGFI and CDT. The strength of contrast between target and nontarget areas was quantitatively calculated as contrast index and compared between IGFI and CDT according to injection technique or state of the liver surface.

RESULTS

Fusion IGFI achieved valid demarcation in 23 of 24 patients (95.8%), whereas CDT achieved valid demarcation in only 10 patients (41.7%) (P < 0.0001). The contrast index of fusion IGFI was 0.81 (0.18-2.51), which was significantly higher than that of CDT at 0.12 (0.01-0.42) (P < 0.0001), and the same result was obtained regardless of the injection method or liver surface state used. ALR was conducted referring to 3-dimensional staining of target parenchyma, with no related perioperative adverse events.

CONCLUSIONS

Fusion IGFI is a safe imaging technique for ALR that attained valid 3-dimensional parenchymal demarcation with better feasibility and clearer demarcation than CDT.

Both insulin and the related peptides, the insulin-like growth factors/somatomedins, may function as anabolic factors in the regulation of fetal body size. Infants born to women suffering from diabetes mellitus may show increased deposition of subcutaneous fat and enhanced lean body mass, findings reproduced in experimental animal fetuses with induced hyperinsulinaemia. Fetal adiposity may be associated with a life-time tendency to obesity and its associated diseases. Insulin-like growth factors I (IGFI) and II are present in the circulation of the newborn infant and animal fetus and correlate positively with birth size. The fetal tissues are biologically responsive to IGFs in vitro and are rich in specific cell membrane receptors, those predominantly recognizing IGFI being structurally and functionally similar to the insulin receptor. Insulin could theoretically influence fetal tissues by an interaction with either the insulin or IGF receptor. IGF release is a property of multiple fetal tissues in vitro, but, in contrast to postnatal life, is not dependent on growth hormone. Fetal IGF production may be influenced by placental lactogen, especially IGFII which rapidly declines in the circulation following parturition in the rat and sheep. A positive association also exists between circulating levels of insulin and IGFs when the former is experimentally manipulated in the animal fetus. Similarly the infant born with transient diabetes mellitus has low cord blood levels of insulin and IGFI. Insulin has a dual role in prenatal life. In the last trimester insulin functions as a glucoregulatory hormone, but from much earlier in gestation creates an anabolic environment in the fetus supplied with optimal nutrients. This latter mechanism of action is unclear and probably heterogeneous, but in overview is permissive rather than obligatory. In contrast the growth-promoting role of the IGFs is direct but their interaction with fetal tissues, and thus the overall emphasis of fetal growth, may be paracrine.

Positive emotional states have been shown to confer resilience to depression and anxiety in humans, but the molecular mechanisms underlying these effects have not yet been elucidated. In laboratory rats, positive emotional states can be measured by 50-kHz ultrasonic vocalizations (hedonic USVs), which are maximally elicited by juvenile rough-and-tumble play behavior. Using a focused microarray platform, insulin-like growth factor I (IGFI) extracellular signaling genes were found to be upregulated by hedonic rough-and-tumble play but not depressogenic social defeat. Administration of IGFI into the lateral ventricle increased rates of hedonic USVs in an IGFI receptor (IGFIR)-dependent manner. Lateral ventricle infusions of an siRNA specific to the IGFIR decreased rates of hedonic 50-kHz USVs. These results show that IGFI plays a functional role in the generation of positive affective states and that IGFI-dependent signaling is a potential therapeutic target for the treatment of depression and anxiety.

We have compared the DNase I hypersensitivity of the regulatory region of two estrogen-regulated genes, pS2 and cathepsin D in hormone-dependent and -independent breast carcinoma cell lines. This strategy allowed the identification of two important control regions, one in pS2 and the other in cathepsin D genes. In the hormone-dependent MCF7 cell line, within the pS2 gene 5'-flanking region, we detected two major DNase I hypersensitive sites, induced by estrogens and/or IGFI: pS2-HS1, located in the proximal promoter and pS2-HS4, located -10.5 Kb from the CAP site, within a region that has not been cloned. The presence of these two DNase I hypersensitive sites correlates with pS2 expression. Interestingly in MCF7 cells, estrogens and IGFI induced indistinguishable chromatin structural changes over the pS2 regulatory region, suggesting that the two transduction-pathways converge to a unique chromatin target. In two cell lines that do not express pS2, MDA MB 231, a hormone-independent cell line that lacks the estrogen receptor alpha, and HE5, a cell line derived from MDA MB 231 by transfection that expresses estrogen receptor alpha, there was only one hormone-independent DNase I hypersensitive site. This site, pS2-HS2, was located immediately upstream of pS2-HS1. In MCF7 cells, two major DNase I hypersensitive sites were present in the 5'-flanking sequences of the cathepsin D gene, which is regulated by estrogens in these cells. These sites, catD-HS2 and catD-HS3, located at positions -2.3 Kb and -3.45 Kb, respectively, were both hormone-independent. A much weaker site, catD-HS1, covered the proximal promoter. In MDA MB 231 cells, that express cathepsin D constitutively, we detected an additional strong hormone-independent DNase I hypersensitive site, catD-HS4, located at position -4.3 Kb. This region might control the constitutive over-expression of cathepsin D in hormone-independent breast cancer cells. All together, these data demonstrate that a local reorganization of the chromatin structure over pS2 and cathepsin D promoters accompanies the establishment of the hormone-independent phenotype of the cells.

Protein kinase signaling cascades control most aspects of cellular function. The ATP binding domains of signaling protein kinases are the targets of most available inhibitors. These domains are highly conserved from mammals to flies. Herein we describe screening of a library of small molecule inhibitors of protein kinases for their ability to increase Drosophila lifespan. We developed an assay system which allowed screening using the small amounts of materials normally present in commercial chemical libraries. The studies identified 17 inhibitors, the majority of which targeted tyrosine kinases associated with the epidermal growth factor receptor (EGFR), platelet-derived growth factor (PDGF)/vascular endothelial growth factor (VEGF) receptors, G-protein coupled receptor (GPCR), Janus kinase (JAK)/signal transducer and activator of transcription (STAT), the insulin and insulin-like growth factor (IGFI) receptors. Comparison of the protein kinase signaling effects of the inhibitors in vitro defined a consensus intracellular signaling profile which included decreased signaling by p38MAPK (p38), c-Jun N-terminal kinase (JNK) and protein kinase C (PKC). If confirmed, many of these kinases will be novel additions to the signaling cascades known to regulate metazoan longevity.

OBJECTIVE

The aim of this study was to determine the preoperative predictors of the extent of resection and endocrinological remission following endonasal endoscopic removal of growth hormone (GH)-secreting pituitary adenomas.

METHODS

The authors analyzed a prospectively collected database of 24 consecutive acromegalic patients who underwent endoscopic endonasal transsphenoidal surgery. The extent of resection was evaluated on postoperative contrast-enhanced MR imaging. Endocrinological remission was defined as normal insulin-like growth factor I (IGFI) serum levels and either a nadir GH level of < 0.4 ng/ml after an oral glucose load or a basal GH serum level < 1 ng/ml.

RESULTS

The majority of acromegalic patients (83%) had macroadenomas>> 1 cm in maximum diameter. Gross-total resection was achieved in 17 (71%) of 24 patients. Notably, endoscopic transsphenoidal surgery allowed complete resection of all lesions without cavernous sinus invasion, regardless of the suprasellar extent. Biochemical remission was achieved in 11 (46%) of 24 patients. A smaller tumor volume and a postoperative reduction in GH serum levels were associated with a higher rate of biochemical cure (p < 0.05). During a 23-month follow-up period 5 patients (21%) underwent Gamma Knife treatment of any residual disease to further reduce excess GH production. Twenty patients (83%) reported significant relief of their symptoms, while 3 (13%) considered their symptoms stable. Two patients (8%) with large macroadenomas experienced postoperative panhypopituitarism, and 2 patients (8%) suffered from CSF leaks, which were treated with lumbar CSF diversion.

CONCLUSIONS

A purely endoscopic endonasal transsphenoidal adenoma resection leads to a high rate of gross-total tumor resection and endocrinological remission in acromegalic patients, even those harboring macroadenomas with wide suprasellar extension. Extended approaches and angled endoscopes are useful tools for increasing the extent of resection.

Insulin-like growth factor-I (IGF-I) has important growthpromoting and metabolic effects and is expressed in virtually every tissue of the body. The highest expression is found in the liver, but the physiological role of liver-derived IGF-I is unknown. It has been difficult to separate the endocrine effects of liver-derived IGF-I from the autocrine/paracrine effects of locally produced IGF-I in peripheral tissues. Therefore, we have developed a mouse model with a liver-specific inducible deletion of the IGF-I gene (LI-IGF-I-/- mouse). The LI-IGF-I-/- mouse has dramatically reduced (>80%) serum IGF-I levels, demonstrating that the major part of serum IGF-I is liver-derived. Surprisingly, LI-IGFI -/- mice demonstrate a normal appendicular skeletal growth up to at least 12 mo of age despite the dramatic decrease in circulating IGF-I levels, indicating that liver-derived IGF-I is not required for appendicular skeletal growth. However, the adult axial skeletal growth is reduced in the LI-IGF-I-/- mice. Furthermore, the amount of cortical bone is reduced due to decreased radial growth of the cortical bone, while the trabecular bone mineral density is unchanged in the LI-IGFI -/- mice. The decreased levels of circulating IGF-I are associated with increased serum levels of growth hormone (GH), indicating a role for liver-derived IGFI in the negative-feedback regulation of GH secretion. Measurements of factors regulating GH secretion in the pituitary and in the hypothalamus revealed an increased expression of GH-releasing-hormone (GHRH) and GHsecretagogue (GHS) receptors in the pituitary of LI-IGFI -/- mice. This in turn results in an increased sensitivity to systemically administered GHRH and GHS, demonstrating that the regulatory action of liver-derived IGF-I on GH secretion is at the pituitary rather than at the hypothalamic level. The liver is an important metabolic organ and LI-IGF-I-/- mice are markedly hyperinsulinemic and yet normoglycemic, consistent with an adequately compensated insulin resistance. Interestingly, LI-IGF-I-/- mice display a reduced age-dependent fat mass accumulation compared with control mice. Furthermore, LI-IGF-I-/- mice have increased blood pressure attributable to increased peripheral resistance indicating a role for liver-derived IGF-I in the regulation of blood pressure. In conclusion, liver-derived IGF-I is important for carbohydrate and lipid metabolism and for the regulation of GH secretion at the pituitary level. Furthermore, it regulates adult axial skeletal growth and cortical radial growth while it is not required for appendicular skeletal growth.

OBJECTIVE

In laparoscopic colorectal cancer (Lap-CRC) surgery, determination of a suitable mesentery division line and the appropriate degree of lymphadenectomy by tracing the blood supply is critical. We performed visualization of the lymph and blood flow by laparoscopic indocyanine green (ICG) fluorescence imaging (Lap-IGFI).

METHODS

ICG is injected into the submucosa near the tumor via colonoscopy, and the lymph flow is observed. Intestinal blood flow is evaluated by administering ICG intravenously.

RESULTS

For lymph flow, visualization of the main lymph node basin helped to determine the surgical division line for cases in which the blood flow was not completely visualized. Lap-IGFI changed the surgical plan of the lymphadenectomy in 23.5 %. In our experience, the metastatic rate of ICG-positive nodes was 10.0 %, and the metastatic rate of ICG-negative nodes was 5.3 %. Furthermore, there were no metastatic nodes that were ICG negative more than 5 cm from the tumor. For blood flow, the blood flow distribution of the intestinal wall from the last branch of the vasa recta of the anastomotic site was clearly visualized and proved useful in choosing the extent of intestinal resection. Lap-IGFI changed the surgical plan of the extensive intestinal resection in 16.7 %.

CONCLUSIONS

Lap-IGFI can noninvasively provide detailed lymph and blood flow information and is a useful device to aid in the accurate identification of individual patients' lymph drainage. This helps dictate adequate lymphadenectomy and the extent of intestinal resection in Lap-CRC surgery.

BACKGROUND

Glioblastomas (GBMs) are the most common malignant primary brain tumours in adults and are refractory to conventional therapy, including surgical resection, radiotherapy and chemotherapy. The insulin-like growth factor (IGF) system is a complex network that includes ligands (IGFI and IGFII), receptors (IGF-IR and IGF-IIR) and high-affinity binding proteins (IGFBP-1 to IGFBP-6). Many studies have reported a role for the IGF system in the regulation of tumour cell biology. However, the role of this system remains unclear in GBMs.

METHODS

We investigate the prognostic value of both the IGF ligands' and receptors' expression in a cohort of human GBMs. Tissue microarray and image analysis were conducted to quantitatively analyse the immunohistochemical expression of these proteins in 218 human GBMs.

RESULTS

Both IGF-IR and IGF-IIR were overexpressed in GBMs compared with normal brain (P<10(-4) and P=0.002, respectively). Moreover, with regard to standard clinical factors, IGF-IR positivity was identified as an independent prognostic factor associated with shorter survival (P=0.016) and was associated with a less favourable response to temozolomide.

CONCLUSIONS

This study suggests that IGF-IR could be an interesting target for GBM therapy.