Beta-cell Tumors Research Articles

Mast cells are required for angiogenesis and macroscopic expansion of Myc-induced pancreatic islet tumors

An association between inflammation and cancer has long been recognized, but the cause and effect relationship linking the two remains unclear. Myc is a pleiotropic transcription factor that is overexpressed in many human cancers and instructs many extracellular aspects of the tumor tissue phenotype, including remodeling of tumor stroma and angiogenesis. Here we show in a beta-cell tumor model that activation of Myc in vivo triggers rapid recruitment of mast cells to the tumor site-a recruitment that is absolutely required for macroscopic tumor expansion. In addition, treatment of established beta-cell tumors with a mast cell inhibitor rapidly triggers hypoxia and cell death of tumor and endothelial cells. Inhibitors of mast cell function may therefore prove therapeutically useful in restraining expansion and survival of pancreatic and other cancers.

Temozolomide/capecitabine therapy for metastatic neuroendocrine tumors of the pancreas. A retrospective review

14023 Background: Our laboratory studies suggest additive to synergistic cytotoxicity when neuroendocrine (NET) cells were given continuous exposure to 5 fluorouracil (5-FU) which preceded exposure to temozolomide (T). Based on this in vitro efficacy, salvage treatment with capecitabine (C), an orally-available fluoropyrimidine that is metabolized to 5-FU, and T was initiated in 17 patients (pts) with advanced NET of the pancreas. Methods: A retrospective review was conducted of the 17 pts who received C 1000mg bid on days # 1–14 and T 150–200mg/m2 on days #10–14 of a 28-day cycle, beginning in October, 2002. Pts had imaging performed after every 2 cycles, and serum tumor markers (TM), when present, were measured at every cycle. Median Age 54 (range 33–70). There were 9 men and 8 women. All of the pts had failed first-line treatment with escalating doses of sandostatin LAR. 11 pts had failed multiagent chemotherapy (range 1–5 regimens). Pathology: 10 pts had NET, 3 had non-secretory carcinoid, 1 had secretory carcinoid, one had a glucagonoma; 2 had non-secretory beta cell tumors and one pt had Multiple Endocrine Neoplasia Type I. Results: 13 of the pts are still alive and 12 remain on therapy with SD or PR. One pt did not complete cycle #1 and is not evaluable for response, but is included in an intent-to-treat (ITT) analysis. Grade 3 toxicity was thrombocytopenia in 2 pts, 1 of whom had splenectomy with resolution, and hand-foot syndrome in 1 pt who resumed therapy at reduced dose of C. Of 17 pts with unidimensionally measurable disease, 1 (6%) achieved a complete pathological response. 9 pts (53%) achieved a partial response by RECIST criteria, with a median duration of PR of 284 days (range 85–364 days). Of the 7 pts with evaluable TM, 6 (86%) had a greater than 50% decline. Median overall survival by ITT analysis in the 4 pts now deceased was 268 days (range 111–613 days). Median time to treatment failure was 340 days (range 111–470 days) in 5 pts. Conclusion: The results of this retrospective review suggest that a combination of T + C is active, well tolerated and may prolong survival and palliate symptoms in pts with NET. [Table: see text]

Pericytes limit tumor cell metastasis

Previously we observed that neural cell adhesion molecule (NCAM) deficiency in beta tumor cells facilitates metastasis into distant organs and local lymph nodes. Here, we show that NCAM-deficient beta cell tumors grew leaky blood vessels with perturbed pericyte-endothelial cell-cell interactions and deficient perivascular deposition of ECM components. Conversely, tumor cell expression of NCAM in a fibrosarcoma model (T241) improved pericyte recruitment and increased perivascular deposition of ECM molecules. Together, these findings suggest that NCAM may limit tumor cell metastasis by stabilizing the microvessel wall. To directly address whether pericyte dysfunction increases the metastatic potential of solid tumors, we studied beta cell tumorigenesis in primary pericyte-deficient Pdgfb(ret/ret) mice. This resulted in beta tumor cell metastases in distant organs and local lymph nodes, demonstrating a role for pericytes in limiting tumor cell metastasis. These data support a new model for how tumor cells trigger metastasis by perturbing pericyte-endothelial cell-cell interactions.

Changes in the expression and cytological localization of betacellulin and its receptors (ErbB-1 and ErbB-4) in the trophoblasts in human placenta over the course of pregnancy.

Betacellulin (BTC), purified and cloned from mouse beta cell tumor (BTC-JC10), is regarded as a new member of the epidermal growth factor family. The present study was conducted to clarify the expression of BTC and its receptors, ErbB-1 and ErbB-4, in the trophoblasts in the human placenta over the course of pregnancy. Human placental tissues were obtained from 4 pregnant women at the 4th to 5th week of pregnancy (very early placentas), 10 women at the 6th to 12th week (early placentas), 5 women at the 18th to 21st week (mid placentas) and 8 women at the 38th and 40th week (term placentas). The mRNA expressions of BTC, erbB-1 and erbB-4 were evaluated by quantitative RT-PCR with Southern blotting and the expression of the soluble form of BTC was determined by western immunoblot with a specific antibody to BTC protein. Immunohistochemical staining of BTC, ErbB-1 and ErbB-4 was also performed. The levels of BTC mRNA expression in early and mid placentas were significantly higher than those in term placentas. The soluble form of BTC protein with an estimated molecular mass of 9.5 kDa was expressed in early and mid placentas, whereas the soluble form was not detected in term placentas. BTC from very early placentas until mid placentas was immunolocalized in syncytiotrophoblasts (S-cells), and was most abundant in early placentas. In contrast, BTC was immunolocalized in extravillous trophoblasts (EVTs), but not in villous trophoblasts in term placentas. The levels of erbB-1 mRNA in the early and mid placentas were significantly higher than those in term placentas, whereas the levels of erbB-4 mRNA in early placentas were significantly lower than those in mid and term placentas. ErbB-1 was immunolocalized in cytotrophoblasts in very early placentas, whereas it was immunolocalized in S-cells from early until term placentas. ErbB-4 from very early placentas until mid placentas was immunolocalized in S-cells, whereas ErbB-4 in the term placentas was detected in EVTs, but not in villous trophoblasts. These findings provide evidence for changes in expression and cytological localization of BTC and its receptors in the trophoblasts in human placenta over the course of pregnancy. BTC may play a pivotal role as a local growth factor in promoting the differentiated villous trophoblastic function via ErbB-1 in early placentas and in contributing to placental growth through the maintenance of EVT cell function via ErbB-4 in term placentas.

Brief inactivation of c-Myc is not sufficient for sustained regression of c-Myc-induced tumours of pancreatic islets and skin epidermis

BackgroundTumour regression observed in many conditional mouse models following oncogene inactivation provides the impetus to develop, and a platform to preclinically evaluate, novel therapeutics to inactivate specific oncogenes. Inactivating single oncogenes, such as c-Myc, can reverse even advanced tumours. Intriguingly, transient c-Myc inactivation proved sufficient for sustained osteosarcoma regression; the resulting osteocyte differentiation potentially explaining loss of c-Myc's oncogenic properties. But would this apply to other tumours?ResultsWe show that brief inactivation of c-Myc does not sustain tumour regression in two distinct tissue types; tumour cells in pancreatic islets and skin epidermis continue to avoid apoptosis after c-Myc reactivation, by virtue of Bcl-xL over-expression or a favourable microenvironment, respectively. Moreover, tumours progress despite reacquiring a differentiated phenotype and partial loss of vasculature during c-Myc inactivation. Interestingly, reactivating c-Myc in β-cell tumours appears to result not only in further growth of the tumour, but also re-expansion of the accompanying angiogenesis and more pronounced β-cell invasion (adenocarcinoma).ConclusionsGiven that transient c-Myc inactivation could under some circumstances produce sustained tumour regression, the possible application of this potentially less toxic strategy in treating other tumours has been suggested. We show that brief inactivation of c-Myc fails to sustain tumour regression in two distinct models of tumourigenesis: pancreatic islets and skin epidermis. These findings challenge the potential for cancer therapies aimed at transient oncogene inactivation, at least under those circumstances where tumour cell differentiation and alteration of epigenetic context fail to reinstate apoptosis. Together, these results suggest that treatment schedules will need to be informed by knowledge of the molecular basis and environmental context of any given cancer.

Insulinomas and expression of an insulin splice variant

Insulinomas are beta-cell tumours characterised by uncontrolled insulin secretion even in the presence of hypoglycaemia. However, the mechanisms allowing such excessive insulin secretion are not known. Insulin secretion can occur only when the beta-cell insulin stores have been replenished by insulin biosynthesis, which is mainly controlled by translation. Such specific translational regulation often involves the 5' untranslated region. We have identified an insulin splice variant in isolated human pancreatic islets of non-diabetic donors that retains 26 bp of intron 1 and thereby changes the 5' untranslated region, but leaves the coding region unchanged. This splice variant has increased translation efficiency in vitro and in vivo compared with native insulin mRNA. However, splice variant expression is less than 1% of native insulin mRNA in normal islets. To test whether this splice variant is involved in insulin production by human insulinomas, we extracted RNA from nine laser-captured surgical insulinoma samples and from isolated islets of nine donors who did not have diabetes. We then determined the ratio of splice variant to native insulin mRNA by quantitative real-time RT-PCR. The mean ratio of the splice variant to native insulin mRNA was increased more than 50-fold in insulinomas compared with normal islets, and this difference was present in all nine human insulinomas. Overexpression of the splice variant therefore seems to be a general characteristic of insulinomas and is estimated to contribute about 90% to insulin synthesis by these tumours. Overexpression of the insulin splice variant with increased translation efficiency in insulinomas might explain how these tumours maintain high levels of insulin synthesis and secretion leading to hyperinsulinaemia-the hallmark of this disease.

Of mice and MEN1: Insulinomas in a conditional mouse knockout.

Patients with multiple endocrine neoplasia type 1 (MEN1) develop multiple endocrine tumors, primarily affecting the parathyroid, pituitary, and endocrine pancreas, due to the inactivation of the MEN1 gene. A conditional mouse model was developed to evaluate the loss of the mouse homolog, Men1, in the pancreatic beta cell. Men1 in these mice contains exons 3 to 8 flanked by loxP sites, such that, when the mice are crossed to transgenic mice expressing cre from the rat insulin promoter (RIP-cre), exons 3 to 8 are deleted in beta cells. By 60 weeks of age, >80% of mice homozygous for the floxed Men1 gene and expressing RIP-cre develop multiple pancreatic islet adenomas. The formation of adenomas results in elevated serum insulin levels and decreased blood glucose levels. The delay in tumor appearance, even with early loss of both copies of Men1, implies that additional somatic events are required for adenoma formation in beta cells. Comparative genomic hybridization of beta cell tumor DNA from these mice reveals duplication of chromosome 11, potentially revealing regions of interest with respect to tumorigenesis.

Immunohistochemical localization of amylin in human pancreas, thyroid, pituitary and their tumors

The aim of the present study was to investigate immunohistochemically expression of amylin, a 37 amino acid peptide, cosecreted with insulin by beta cells in pancreatic islets in 12 non-tumorous pancreatic tissues, 22 pancreatic islet tumors, 14 non-tumorous thyroids, 14 medullary carcinomas of the thyroid, 10 non-tumorous pituitaries and 50 pituitary adenomas including 10 amyloid-forming prolactin-cell adenomas using the streptavidin-biotin-peroxidase complex method. Amylin was expressed in non-tumorous pancreatic islets but not in non-tumorous thyroids and pituitaries. Since amylin plays an important role in amyloid formation in pancreatic islets, those tumor types were selected to study which may produce amyloid. Amylin was widely expressed in one insulin producing beta cell tumor. Few tumor cells were immunopositive in 8 islet-cell tumors and in 5 medullary thyroid carcinomas. Immunostaining was not found in pituitary adenomas, including those which produced amyloid. It can be concluded that amylin is not a satisfactory immunohistochemical marker to identify pancreatic islet tumors, medullary thyroid carcinomas and pituitary adenomas.

Antitumorigenic and antiinsulinogenic effects of calcitriol on insulinoma cells and solid beta-cell tumors.

Malignant insulinoma is a rare form of cancer with a poor prognosis because of metastatic dissemination and untreatable hypoglycemia. Effective chemotherapy of patients who are not cured by surgery is needed. Calcitriol has known anticancer properties on different neoplastic cell lines, but no data are available regarding its activity on tumorigenic pancreatic beta-cells. We analyzed the in vitro effects of calcitriol on the murine insulinoma cell line betaTC(3) and primary cultures of human isolated islets and benign insulinoma. The effect of in vivo calcitriol administration on insulinoma of recombinant insulin/Simian virus 40 oncogene-expressing transgenic mice was also investigated. In betaTC(3), calcitriol induced growth inhibition; apoptosis; down-regulation of insulin gene expression; and nongenomic activation of the MAPK pathway. MAPK kinase inhibitor (UO126) and staurosporine reduced calcitriol-mediated betaTC(3) death, and down-regulation of insulin gene transcription was prevented by staurosporine but not UO126. Calcitriol significantly decreased insulin release and mRNA levels of human islets and insulinoma cells. Finally, recombinant insulin/Simian virus 40 oncogene-expressing transgenic mice treated with calcitriol showed reduced insulinoma volumes because of increased apoptosis of adenomatous cells. Together, these findings provide the rationale for testing the efficacy of calcitriol in the treatment of patients with solid beta-cell tumors.

Ferret soft tissue surgery

Private veterinary practitioners are seeing pet ferrets with increasing frequency, and many common disease syndromes of the pet ferret require surgical intervention. This article discusses adrenal gland disease and adrenalectomy, prostatic and paraurethral cysts and their surgical management, perineal urethrostomy, pancreatic beta-cell tumor surgery, and splenectomy in the ferret. The surgical procedures are described with anatomic and physiologic information included when specific to the ferret. Indications for surgery, recomended presurgical diagnostics, and perioperative medical therapy are reviewed. Successful ferret soft tissue surgery depends on the surgeon's skill in combination with a working knowledge of ferret anatomy and the unique pathophysiology associated with the common ferret disease syndromes.

Vascular endothelial growth factor-C-mediated lymphangiogenesis promotes tumour metastasis.

Metastasis is a frequent and lethal complication of cancer. Vascular endothelial growth factor-C (VEGF-C) is a recently described lymphangiogenic factor. Increased expression of VEGF-C in primary tumours correlates with dissemination of tumour cells to regional lymph nodes. However, a direct role for VEGF-C in tumour lymphangiogenesis and subsequent metastasis has yet to be demonstrated. Here we report the establishment of transgenic mice in which VEGF-C expression, driven by the rat insulin promoter (Rip), is targeted to beta-cells of the endocrine pancreas. In contrast to wild-type mice, which lack peri-insular lymphatics, RipVEGF-C transgenics develop an extensive network of lymphatics around the islets of Langerhans. These mice were crossed with Rip1Tag2 mice, which develop pancreatic beta-cell tumours that are neither lymphangiogenic nor metastatic. Double-transgenic mice formed tumours surrounded by well developed lymphatics, which frequently contained tumour cell masses of beta-cell origin. These mice frequently developed pancreatic lymph node metastases. Our findings demonstrate that VEGF-C-induced lymphangiogenesis mediates tumour cell dissemination and the formation of lymph node metastases.

Roles of Insulin Receptor Substrate-1, Phosphatidylinositol 3-Kinase, and Release of Intracellular Ca2+ Stores in Insulin-stimulated Insulin Secretion in β-Cells

The signaling pathway by which insulin stimulates insulin secretion and increases in intracellular free Ca(2+) concentration ([Ca(2+)](i)) in isolated mouse pancreatic beta-cells and clonal beta-cells was investigated. Application of insulin to single beta-cells resulted in increases in [Ca(2+)](i) that were of lower magnitude, slower onset, and longer lifetime than that observed with stimulation with tolbutamide. Furthermore, the increases in [Ca(2+)](i) originated from interior regions of the cell rather than from the plasma membrane as with depolarizing stimuli. The insulin-induced [Ca(2+)](i) changes and insulin secretion at single beta-cells were abolished by treatment with 100 nm wortmannin or 1 micrometer thapsigargin; however, they were unaffected by 10 micrometer U73122, 20 micrometer nifedipine, or removal of Ca(2+) from the medium. Insulin-stimulated insulin secretion was also abolished by treatment with 2 micrometer bisindolylmaleimide I, but [Ca(2+)](i) changes were unaffected. In an insulin receptor substrate-1 gene disrupted beta-cell tumor line, insulin did not evoke either [Ca(2+)](i) changes or insulin secretion. The data suggest that autocrine-activated increases in [Ca(2+)](i) are due to release of intracellular Ca(2+) stores, especially the endoplasmic reticulum, mediated by insulin receptor substrate-1 and phosphatidylinositol 3-kinase. Autocrine activation of insulin secretion is mediated by the increase in [Ca(2+)](i) and activation of protein kinase C.

Auto-induction and growth stimulatory effect of betacellulin in human pancreatic cancer cells.

Betacellulin (BTC) was identified in mouse pancreatic beta cell tumors as a member of the epidermal growth factor (EGF) family, and was found to bind and activate the EGF receptor. BTC is also expressed in some human malignancies and may have an important role in tumor growth progression. We examined whether BTC and EGF have a growth stimulatory effect on human pancreatic cancer cell lines both in vitro and in vivo. We also investigated the BTC expression and autonomous induction of BTC in pancreatic cancer cells. in vitro, both BTC and EGF had almost the same proliferative effect on Panc-1, MIA PaCa-2 and AsPC-1. in vivo, in a Panc-1 inoculated athymic mice model, BTC-treated tumors grew approximately five times larger than in control. Immunocytochemistry showed that BTC expression occurred in three pancreatic cancer cell lines, with MIA PaCa-2 showing the strongest intensity. Semi-quantitative RT-PCR of MIA Paca-2 showed that mRNA levels of BTC gradually increased after treatment with 1 nM BTC. Immunocytochemistry also demonstrated that the intensity of BTC-like immunoreactivity was increased when treated with 1 nM BTC but was reduced after treatment with 100 nM of AG1478, an EGF receptor tyrosine kinase inhibitor. BTC has thus a significant growth stimulatory effect on pancreatic cancer cells and might function as an autocrine and paracrine growth factor. BTC expression in pancreatic cancer cells is, at least in part, controlled by an auto-induction mechanism.

Frequent loss of heterozygosity at chromosome 3p14.2-3p21 in human pancreatic islet cell tumours.

Pancreatic islet betacell tumours occur either sporadically or as part of inherited neoplastic syndromes, most commonly multiple endocrine neoplasia (MEN) type 1. Recently, a transgenic mouse model has been established in which the expression of the SV40 large T antigen was targeted to betacells by the rat insulin promoter, leading to the development of multiple pancreatic betacell tumours. In the advanced stages of tumour evolution, these tumours exhibited a high prevalence of loss of heterozygosity (LOH) on mouse chromosomes 9 and 16, at regions syntenic with regions 3q, 3p21, 6q12, 15q24 and 22q of the human genome. Loss of heterozygosity in human islet cell tumours was analysed in a PCR based approach at regions of the human genome syntenic with the mouse loci linked to pancreatic betacell tumours as well as the MEN1 gene on chromosome 11q13. These included 35 microsatellite markers in the human chromosomal regions 3q, 3p21, 6q12, 11q13, 15q24 and 22q. 21 patients diagnosed with insulinoma were analysed. Histologically, 16 tumours were benign, while 5 were malignant insulinomas. Thirteen of 21 (62%) tumours were found to have loss of genetic material on chromosome 3. The shortest region of overlap implicated a deletion at 3p14.2-3p21 region, corresponding to the marker D3S1295. We did not detect a substantial frequency of LOH in the other syntenic regions, except for the region of MEN 1 gene on 11q13 found to be deleted in 6 (29%) cases, including 3 of 4 tumours from MEN 1 families. Deletions of 3p14. 2-3p21 were observed in 8 of 15 (53%) benign tumours, and in 5 of 6 (83%) malignant neoplasms. These results indicate the high frequency of 3p14.2-3p21 deletions in human pancreatic betacell neoplasms. These finding suggest the presence of a tumour suppressor gene in this region, that may be important in the microevolution of these tumours towards malignancy.

Islet amyloid polypeptide/amylin messenger RNA and protein expression in human insulinomas in relation to amyloid formation.

Islet amyloid polypeptide (IAPP), also named amylin, is the predominant protein component of amyloid deposits in human islet beta cell tumours of the pancreas (insulinomas). IAPP is co-produced with insulin by islet beta cells. We investigated IAPP expression in relation to insulin expression and to amyloid formation in eleven insulinomas. RNA and protein extracts were prepared from the same pieces of tumour tissue, and from specimens of two normal human pancreata. IAPP and insulin mRNA and peptide content were quantified using Northern blot analysis and radioimmunoassay (RIA) respectively. Molecular forms of IAPP immunoreactivity were analysed by reversed-phase high-performance liquid chromatography (HPLC). The presence of islet hormones and of amyloid was assessed by (immuno)histochemical staining of paraffin sections. Plasma levels of IAPP and insulin prior to tumour resection were determined by RIA. IAPP and insulin mRNA and peptide content varied widely between the tumour specimens, and there was considerable intratumour heterogeneity of peptide content. HPLC analysis indicated correct proteolytic processing of the IAPP precursor protein. Amyloid deposits were detected only in the three tumours with the highest IAPP content. In contrast to insulin, plasma levels of IAPP were not elevated in the insulinoma patients. The spectrum of hormone production by insulinomas cannot be inferred from only a few tissue sections due to intratumour heterogeneity. Expression of the IAPP and insulin genes is not coupled in insulinomas, which produce properly processed mature IAPP. In addition to IAPP overproduction, additional factors such as intracellular accumulation of IAPP are involved in amyloidogenesis in insulinomas.

Immunohistochemical localization of betacellulin, a new member of the EGF family, in normal human pancreas and islet tumor cells.

Betacellulin (BTC) purified from mouse beta cell tumor (betaTC-3) is a new member of the epidermal growth factor (EGF) family which can bind receptor tyrosine kinase, EGF receptor (erbB1) and erbB4. It has been demonstrated that proBTC mRNA was abundantly expressed in human pancreas tissue, and that BTC converted amylase-secreting rat acinar cell line (AR42J) into insulin-secreting cells, suggesting that BTC might be important for the growth and/or differentiation of islet cells. However, the cell type producing BTC in the pancreas has not been clarified. In this study, we examined the localization of BTC in human pancreas and islet cell tumors. Immunohistochemistry using specific antibodies to human BTC revealed that this protein was produced in alpha cells and duct cells, and probably in beta cells in normal adult pancreas. Furthermore, strong immunoreactivity to BTC was detected in primitive duct cells of the fetal pancreas, and both insulinoma and glucagonoma cells also showed positive immunoreactivity to BTC. EGF receptor (erbB1) and erbB4 were expressed mainly in islet and duct cells, and duct cells, respectively. These results demonstrate the localization of BTC and its receptors, and suggest that BTC may be one of the factors that have physiologically important roles such as growth and differentiation of islet cells in the human pancreas.

Expression of IGF system genes during T-antigen driven pituitary tumorigenesis.

Expression of IGF-I, IGF-II, the Type-I IGF receptor and six IGF binding proteins were examined in three different T-ag-driven mouse tumors. Unlike the widespread expression of IGF-II in pancreatic beta-cell tumors, IGF-II was not widely expressed in the two different pituitary tumors examined indicating that a mechanism independent of focal IGF-II expression can also drive T-antigen tumorigenesis. In addition, multiple IGF binding proteins were expressed in all three tumor types. This expression, however, was generally heterogeneous with no specific changes to indicate a required role for any IGF binding protein in T-antigen tumorigenesis.

A novel glucose-responsive element in the human insulin gene functions uniquely in primary cultured islets

Insulin gene transcription is limited to the beta cells within the mammalian pancreas and, like insulin secretion, is regulated by glucose. Our previous studies in primary cultured beta cells suggested the presence of a strong glucose-responsive enhancer element between base pairs -341 and -260 of the human insulin promoter, the same region in which a transcriptional repressor had been identified in beta-cell tumor lines. In an attempt to map these promoter activities and resolve these conflicting data, we designed minienhancer constructs spanning this region, and tested them in primary cultured and immortalized cells. One sequence, the Z element (base pairs -292 to -243), functions as both a potent glucose-responsive transcriptional enhancer in primary cultured islet cells and as a transcriptional repressor in immortalized beta and nonbeta cells and in primary fibroblasts. In addition, the Z element binds a novel glucose-responsive protein complex that is found in the nuclei of primary cultured islet cells, but not in the nuclei of tumor cells or primary cultured fibroblasts. These data demonstrate a critical role for the Z element in human insulin gene transcription and its regulation by glucose.

Cyclic ADP-ribose and the pancreatic beta cell: where do we stand?

[3]. Oka-moto et al’s interest in the NAD metabolite cADPRappears to stem from their earlier works where theydemonstrate a crucial role for the cellular level ofNAD in mediating diverse processes such as betacell damage and beta cell tumour formation [4].BackgroundLeeetal.[5]identifiedthestructureofanactivemeta-bolite, i.e. cADPR responsible for the Ca

Novel forms of acidic fibroblast growth factor-1 are constitutively exported by beta tumor cell lines independent from conventional secretion and apoptosis.

Acidic and basic fibroblast growth factors (FGF-1 and FGF-2) are strong mitogens for many tumor cell types and potent inducers of angiogenesis in vitro and in vivo. It is notable that these proteins lack classical signal sequences for secretion; the mechanism by which they are released from cells remains obscure. We demonstrate here that FGF-1 is constitutively exported by tumor cell lines derived from highly angiogenic beta cell tumors of transgenic mice. Remarkably, FGF-1 is sequestered as a latent form in the conditioned medium, as assessed by a lack of mitogenic activity and heparin affinity. High salt treatment of conditioned medium unveils the sequestered FGF-1 as novel high molecular weight forms with reduced heparin affinity and a molecular mass of approximately 30-40kDa; we refer to these as exported forms of FGF-1 (XP-FGF-1). Reducing and denaturing agents convert XP-FGF-1 into the 18kDa monomeric form of FGF-1, indicating it represents tight aggregates of FGF-1. XP-FGF-1 is found in cell lysate as well as conditioned medium, suggesting that they represent export intermediates. Brefeldin A, an inhibitor of conventional secretion, does not interfere with FGF-1 export. Moreover, cell lysis or apoptosis are not involved in this export pathway. Therefore, these data demonstrate that FGF-1 is constitutively exported by beta tumor cell lines via a novel secretory pathway so as to facilitate tumor growth and angiogenesis.