Bone Radiation-Induced Sarcomas: Outcomes Based on Histology and Surgical Treatment: A Systematic Review of the Literature.

BACKGROUND: Bone radiation-induced sarcomas (B-RIS) are secondary neoplasms with reportedly worse overall survival than de novo bone sarcoma. Treatment strategy for these neoplasms remains uncertain. Our systematic review sought to assess overall survival based on histology and surgical intervention. METHODS: A systemic review was conducted following Preferred Reporting Items for Systematic reviews and Meta-Analyses guidelines and registered in PROSPERO (438415). Studies describing oncologic outcomes of patients with B-RIS in the appendicular and axial skeleton were included. The Strengthening the Reporting of Observational Studies in Epidemiology checklist was used for quality assessment. Survival analysis by histologic subtype and surgery type was performed in a subset of 234 patients from 11 articles with individualized data. A total of 20 articles with a total of 566 patients were included. The most frequent location was the pelvis (27.7%), and the main histological types were osteosarcoma (69.4%), undifferentiated pleomorphic sarcoma (14.1%), and fibrosarcoma (9.2%). Limb-salvage and amputation were performed in 68.5% and 31.5% of cases, respectively. RESULTS: Local recurrence was 13%, without difference between limb-salvage surgery and amputation (p = 0.51). The metastasis rate was 42.3%. Five-year OS was 43.7% (95% confidence interval [CI], 33.3%-53.5%) for osteosarcoma, 31.5% (95% CI, 11.3%-54.2%) for UPS, and 28.1% (95% CI, 10.6%-48.8%) for fibrosarcoma. Five-year OS was 49.2% (95% CI, 35.3%-61.6%) for limb-salvage and 46.9% (95% CI, 29.1%-62.9%) for amputation. There was no difference in 5-year OS between histologic subtypes (p = 0.18) or treatment type (p = 0.86). CONCLUSION: B-RIS demonstrated poor OS at 5 years after initial management regardless of histology. Limb-salvage surgery was not associated with lower 5-year OS compared with amputation. Future studies should compare both groups while controlling for confounders. LEVEL OF EVIDENCE: Level III. See Instructions for Authors for a complete description of levels of evidence.

ß-Cell Knockout of SENP1 Reduces Responses to Incretins and Worsens Oral Glucose Tolerance in High-Fat Diet-Fed Mice.

SUMOylation reduces oxidative stress and preserves islet mass at the expense of robust insulin secretion. To investigate a role for the deSUMOylating enzyme sentrin-specific protease 1 (SENP1) following metabolic stress, we put pancreas/gut-specific SENP1 knockout (pSENP1-KO) mice on a high-fat diet (HFD). Male pSENP1-KO mice were more glucose intolerant following HFD than littermate controls but only in response to oral glucose. A similar phenotype was observed in females. Plasma glucose-dependent insulinotropic polypeptide (GIP) and glucagon-like peptide 1 (GLP-1) responses were identical in pSENP1-KO and wild-type littermates, including the HFD-induced upregulation of GIP responses. Islet mass was not different, but insulin secretion and ß-cell exocytotic responses to the GLP-1 receptor agonist exendin-4 (Ex4) and GIP were impaired in islets lacking SENP1. Glucagon secretion from pSENP1-KO islets was also reduced, so we generated ß-cell-specific SENP1 KO mice. These phenocopied the pSENP1-KO mice with selective impairment in oral glucose tolerance following HFD, preserved islet mass expansion, and impaired ß-cell exocytosis and insulin secretion to Ex4 and GIP without changes in cAMP or Ca2+ levels. Thus, ß-cell SENP1 limits oral glucose intolerance following HFD by ensuring robust insulin secretion at a point downstream of incretin signaling.

Transient antibiotic-induced changes in the neonatal swine intestinal microbiota impact islet expression profiles reducing subsequent function.

Neonatal antibiotics administered to human infants initiate gut microbiota dysbiosis that may have long-term effects on body weight and metabolism. We examined antibiotic-induced adaptations in pancreatic islets of the piglet, a well-accepted model of human infant microbiota and pancreas development. Neonatal piglets randomized to amoxicillin [30 mg/kg body wt/day; n = 7, antibiotic (ANTI)] or placebo [vehicle control; n = 7, control (CON)] from postnatal day (PND)0-13 were euthanized at PND7, 14, and 49. The metabolic phenotype along with functional, immunohistological, and transcriptional phenotypes of the pancreatic islets were studied. The gut microbiome was characterized by 16S rRNA gene sequencing, and microbial metabolites and microbiome-sensitive host molecules were measured. Compared with CON, ANTI PND7 piglets had elevated transcripts of genes involved in glucagon-like peptide 1 ((GLP-1) synthesis or signaling in islets (P < 0.05) coinciding with higher plasma GLP-1 (P = 0.11), along with increased tumor necrosis factor α (Tnf) (P < 0.05) and protegrin 1 (Npg1) (P < 0.05). Antibiotic-induced relative increases in Escherichia, Coprococcus, Ruminococcus, Dehalobacterium, and Oscillospira of the ileal microbiome at PND7 normalized after antibiotic withdrawal. In ANTI islets at PND14, the expression of key regulators pancreatic and duodenal homeobox 1 (Pdx1), insulin-like growth factor-2 (Igf2), and transcription factor 7-like 2 (Tcf7l2) was downregulated, preceding a 40% reduction of ß-cell area (P < 0.01) and islet insulin content at PND49 (P < 0.05). At PND49, a twofold elevated plasma insulin concentration (P = 0.07) was observed in ANTI compared with CON. We conclude that antibiotic treatment of neonatal piglets elicited gut microbial changes accompanied by phasic alterations in key regulatory genes in pancreatic islets at PND7 and 14. By PND49, reduced ß-cell area and islet insulin content were accompanied by elevated nonfasted insulin despite normoglycemia, indicative of islet stress.

Disruption of Beta-Cell Mitochondrial Networks by the Orphan Nuclear Receptor Nor1/Nr4a3.

Nor1, the third member of the Nr4a subfamily of nuclear receptor, is garnering increased interest in view of its role in the regulation of glucose homeostasis. Our previous study highlighted a proapoptotic role of Nor1 in pancreatic beta cells and showed that Nor1 expression was increased in islets isolated from type 2 diabetic individuals, suggesting that Nor1 could mediate the deterioration of islet function in type 2 diabetes. However, the mechanism remains incompletely understood. We herein investigated the subcellular localization of Nor1 in INS832/13 cells and dispersed human beta cells. We also examined the consequences of Nor1 overexpression on mitochondrial function and morphology. Our results show that, surprisingly, Nor1 is mostly cytoplasmic in beta cells and undergoes mitochondrial translocation upon activation by proinflammatory cytokines. Mitochondrial localization of Nor1 reduced glucose oxidation, lowered ATP production rates, and inhibited glucose-stimulated insulin secretion. Western blot and microscopy images revealed that Nor1 could provoke mitochondrial fragmentation via mitophagy. Our study unveils a new mode of action for Nor1, which affects beta-cell viability and function by disrupting mitochondrial networks.

Evidence of unrestrained beta-cell proliferation and neogenesis in a patient with hyperinsulinemic hypoglycemia after gastric bypass surgery.

Hyperinsulinemic hypoglycemia syndrome (HIHG) is a rare complication of roux-en-Y gastric bypass surgery. The pathology is associated with an excessive function of pancreatic beta-cells, and requires pancreas resection in patients that are recalcitrant to nutritional and pharmacological interventions. The exact prevalence is not clearly understood and the underlying mechanisms not yet fully characterized. We herein sought to perform histological and molecular examination of pancreatic sections obtained from a patient who developed HIHG as a complication of gastric bypass compared to 3 weight-matched controls. We studied markers of cellular replication and beta-cell differentiation by immunohistochemistry and immunofluorescence. HIHG after gastric bypass was characterized by a profound increase in beta-cell mass. Cellular proliferation was increased in islets and ducts compared to controls, suggesting unrestrained proliferation in HIHG. We also detected beta-cell differentiation markers in duct cells and occasional duct cells displaying both insulin and glucagon immunoreactivity. These histological observations suggest that beta-cell differentiation from ductal progenitor cells could also underly beta-cell mass expansion in HIHG. Altogether, our results can be construed to demonstrate that HIHG after gastric bypass is characterized by abnormal beta-cell mass expansion, resulting from both unrestrained beta-cell replication and neogenesis.

Glutamate formation via the leucine-to-glutamate pathway of rat pancreas.

The leucine-to-glutamate (Leu→Glu) pathway, which metabolizes the carbon atoms of l-leucine to form l-glutamate, was studied by incubation of rat tissue segments with l-[U-(14)C]leucine and estimation of the [(14)C]glutamate formed. Metabolism of the leucine carbon chain occurs in most rat tissues, but maximal activity of the Leu→Glu pathway for glutamate formation is limited to the thoracic aorta and pancreas. In rat aorta, the Leu→Glu pathway functions to relax the underlying smooth muscle; its functions in the pancreas are unknown. This report characterizes the Leu→Glu pathway of rat pancreas and develops methods to examine its functions. Pancreatic segments effect net formation of glutamate on incubation with l-leucine, l-glutamine, or a mix of 18 other plasma amino acids at their concentrations in normal rat plasma. Glutamate formed from leucine remains mainly in the tissue, whereas that from glutamine enters the medium. The pancreatic Leu→Glu pathway uses the leucine carbons for net glutamate formation; the α-amino group is not used; the stoichiometry is as follows: 1 mol of leucine yields 2 mol of glutamate (2 leucine carbons per glutamate) plus 2 mol of CO2. Comparison of the Leu→Glu pathway in preparations of whole pancreatic segments, isolated acini, and islets of Langerhans localizes it in the acini; relatively high activity is found in cultures of the AR42J cell line and very little in the INS-1 832/13 cell line. Pancreatic tissue glutamate concentration is homeostatically regulated in the range of ∼1-3 µmol/g wet wt. l-Valine and leucine ethyl, benzyl, and tert-butyl esters inhibit the Leu→Glu pathway without decreasing tissue total glutamate.

A critical role for the neural zinc factor ST18 in pancreatic ß-cell apoptosis.

The tumor suppressor gene ST18 was originally characterized as the third member of the neural zinc finger transcription factor family. However, little is known about its biological functions. Herein, we demonstrate that, in the pancreas, ST18 expression is restricted to endocrine cells. The detection of ST18 expression in pancreatic ß-cells prompted us to investigate its regulation and its role in ß-cell mass and function. We show that ST18 expression and activity are increased by cytotoxic concentrations of fatty acids and cytokines in INS832/13 cells. Furthermore, ST18 is also increased in islets of diet-induced obese animals. Overexpression and RNA interference knockdown studies demonstrate that ST18 induces ß-cell apoptosis and curtails ß-cell replication. Finally, our data suggest that ST18 impairs insulin secretion. Taken together, our findings indicate that ST18 could represent a novel transcriptional mediator of lipotoxicity and cytokine-induced ß-cell death. We suggest that genetic or pharmacologic manipulations of ST18 could help maintain a functional ß-cell mass.

Nov/Ccn3, a novel transcriptional target of FoxO1, impairs pancreatic ß-cell function.

Type 2 diabetes is characterized by both insulin resistance and progressive deterioration of ß-cell function. The forkhead transcription factor FoxO1 is a prominent mediator of insulin signaling in ß-cells. We reasoned that identification of FoxO1 target genes in ß-cells could reveal mechanisms linking ß-cell dysfunction to insulin resistance. In this study, we report the characterization of Nov/Ccn3 as a novel transcriptional target of FoxO1 in pancreatic ß-cells. FoxO1 binds to an evolutionarily conserved response element in the Ccn3 promoter to regulate its expression. Accordingly, CCN3 levels are elevated in pancreatic islets of mice with overexpression of a constitutively active form of FoxO1 or insulin resistance. Our functional studies reveal that CCN3 impairs ß-cell proliferation concomitantly with a reduction in cAMP levels. Moreover, CCN3 decreases glucose oxidation, which translates into inhibition of glucose-stimulated Ca(2+) entry and insulin secretion. Our results identify CCN3, a novel transcriptional target of FoxO1 in pancreatic ß-cells, as a potential target for therapeutic intervention in the treatment of diabetes.

Glucagon-like peptide 1 inhibits the sirtuin deacetylase SirT1 to stimulate pancreatic ß-cell mass expansion.

OBJECTIVE: The glucoincretin hormone glucagon-like peptide 1 (GLP-1) enhances glucose-stimulated insulin secretion and stimulates pancreatic ß-cell mass expansion. We have previously shown that the forkhead transcription factor FoxO1 is a prominent transcriptional effector of GLP-1 signaling in the ß-cell. FoxO1 activity is subject to a complex regulation by Akt-dependent phosphorylation and SirT1-mediated deacetylation. In this study, we aimed at investigating the potential role of SirT1 in GLP-1 action. RESEARCH DESIGN AND METHODS: FoxO1 acetylation levels and binding to SirT1 were studied by Western immunoblot analysis in INS832/13 cells. SirT1 activity was evaluated using an in vitro deacetylation assay and correlated with the NAD(+)-to-NADH ratio. The implication of SirT1 in GLP-1-induced proliferation was investigated by BrdU incorporation assay. Furthermore, we determined ß-cell replication and mass in wild-type and transgenic mice with SirT1 gain of function after daily administration of exendin-4 for 1 week. RESULTS: Our data show that GLP-1 increases FoxO1 acetylation, decreases the binding of SirT1 to FoxO1, and stunts SirT1 activity in ß-INS832/13 cells. GLP-1 decreases both the NAD(+)-to-NADH ratio and SirT1 expression in INS cells and isolated islets, thereby providing possible mechanisms by which GLP-1 could modulate SirT1 activity. Finally, the action of GLP-1 on ß-cell mass expansion is abolished in both transgenic mice and cultured ß-cells with increased dosage of SirT1. CONCLUSIONS: Our study shows for the first time that the glucoincretin hormone GLP-1 modulates SirT1 activity and FoxO1 acetylation in ß-cells. We also identify SirT1 as a negative regulator of ß-cell proliferation.

Glucagon-like peptide-1 (GLP-1) diminishes neuronal degeneration and death caused by NGF deprivation by suppressing Bim induction.

Glucagon-like peptide-1 (GLP-1) is a glucoincretin hormone most intensively studied for its actions on insulin secreting beta-cells. GLP-1 and its receptor are also found in brain and accumulating evidence indicates that GLP-1 has neuroprotective actions. Here, we investigated whether GLP-1 protects neuronal cells from death evoked by nerve growth factor (NGF) withdrawal. Compromised trophic factor signaling may underlie neurodegenerative diseases ranging from Alzheimer disease to diabetic neuropathies. We report that GLP-1 provides sustained protection of cultured neuronal PC12 cells and sympathetic neurons from degeneration and death caused by NGF deprivation. Past work shows that NGF deprivation induces the pro-apoptotic protein Bim which contributes to neuron death. Here, we find that GLP-1 suppresses Bim induction promoted by NGF deprivation. Thus, GLP-1 may protect neurons, at least in part, by suppressing Bim induction. Our findings support the idea that drugs that mimic or elevate GLP-1 represent potential therapeutics for neurodegenerative diseases.

Mutational loss of PTEN induces resistance to NOTCH1 inhibition in T-cell leukemia.

Gain-of-function mutations in NOTCH1 are common in T-cell lymphoblastic leukemias and lymphomas (T-ALL), making this receptor a promising target for drugs such as gamma-secretase inhibitors, which block a proteolytic cleavage required for NOTCH1 activation. However, the enthusiasm for these therapies has been tempered by tumor resistance and the paucity of information on the oncogenic programs regulated by oncogenic NOTCH1. Here we show that NOTCH1 regulates the expression of PTEN (encoding phosphatase and tensin homolog) and the activity of the phosphoinositol-3 kinase (PI3K)-AKT signaling pathway in normal and leukemic T cells. Notch signaling and the PI3K-AKT pathway synergize in vivo in a Drosophila melanogaster model of Notch-induced tumorigenesis, and mutational loss of PTEN is associated with human T-ALL resistance to pharmacological inhibition of NOTCH1. Overall, these findings identify transcriptional control of PTEN and regulation of the PI3K-AKT pathway as key elements of the leukemogenic program activated by NOTCH1 and provide the basis for the design of new therapeutic strategies for T-ALL.

Transcription factor FoxO1 mediates glucagon-like peptide-1 effects on pancreatic beta-cell mass.

The glucoincretin hormone glucagon-like peptide-1 (GLP-1) increases pancreatic beta-cell proliferation and survival through sequential activation of the epidermal growth factor receptor (EGFR), phosphatidylinositol-3 kinase (PI 3-kinase), and Akt. We investigated the role of transcription factor FoxO1 in the proliferative and antiapoptotic actions of GLP-1 in beta-cells. GLP-1 inhibited FoxO1 through phosphorylation-dependent nuclear exclusion in pancreatic beta (INS832/13) cells. The effect of GLP-1 was suppressed by inhibitors of EGFR (AG1478) and PI 3-kinase (LY294002). In contrast, LY294002 but not AG1478 suppressed insulin-induced FoxO1 phosphorylation. Expression of constitutively nuclear FoxO1 in beta-cells prevented the proliferative and antiapoptotic actions of GLP-1 in cultured beta-cells and the increase in pancreatic beta-cell mass in response to Exendin4 in transgenic mice. Gene expression and chromatin immunoprecipitation assays demonstrated that GLP-1 increases pancreatic and duodenal homeobox gene-1 and Foxa2 expression and inhibits FoxO1 binding to both promoters. We propose that FoxO1 mediates the pleiotropic effects of the glucoincretin hormone on cell proliferation and survival.

Glucagon-like peptide 1 induces pancreatic beta-cell proliferation via transactivation of the epidermal growth factor receptor.

We previously provided evidence that glucagon-like peptide 1 (GLP-1) induces pancreatic beta-cell growth nonadditively with glucose in a phosphatidylinositol (PI) 3-kinase- and protein kinase C zeta-dependent manner. However, the exact mechanism by which the GLP-1 receptor (GLP-1R), a member of the G protein-coupled receptor (GPCR) superfamily, activates the PI 3-kinase signaling pathway to promote beta-cell growth remains unknown. We hypothesized that the GLP-1R could activate PI 3-kinase and promote beta-cell proliferation through transactivation of the epidermal growth factor (EGF) receptor (EGFR), an event possibly linked to GPCRs via activation of c-Src and the production of putative endogenous EGF-like ligands. Both the c-Src inhibitor PP1 and the EGFR-specific inhibitor AG1478 blocked GLP-1-induced [(3)H]thymidine incorporation in INS(832/13) cells as well as in isolated rat islets, while only AG1478 inhibited the proliferative action of betacellulin (BTC), an EGFR agonist. Both compounds also suppressed GLP-1-induced PI 3-kinase activation. A time-dependent increase in tyrosine phosphorylation of the EGFR in response to GLP-1 was observed in INS(832/13) cells. This transactivation of the EGFR was sensitive to both the pharmacological agents PP1 and AG1478. The action of GLP-1 and BTC on INS cell proliferation was found to be not additive. Overexpression of a dominant-negative EGFR in INS cells with a retroviral expression vector curtailed GLP-1-induced beta-cell proliferation. GLP-1 treatment of INS cells caused a decrease in cell surface-associated BTC, as shown by FACS analysis. Also, the metalloproteinase inhibitor GM6001 and an anti-BTC neutralizing antibody suppressed the GLP-1 proliferative effect. Finally, coculturing the prostatic cancer cell line LNCaP that lacks GLP-1 responsiveness with INS cells increased LNCaP cell proliferation in the presence of GLP-1, thus revealing that INS cells secrete a growth factor in response to GLP-1. GM6001 and an anti-BTC neutralizing antibody suppressed increased LNCaP cell proliferation in the presence of GLP-1 in the coculture experiments. The results are consistent with a model in which GLP-1 increases PI 3-kinase activity and enhances beta-cell proliferation via transactivation of the EGFR that would require the proteolytic processing of membrane-anchored BTC or other EGF-like ligands.