Tumour Suppressive Function and Modulation of Programmed Cell Death 4 (PDCD4) in Ovarian Cancer

Background: Programmed cell death 4 (PDCD4), originally identified as the neoplastic transformation inhibitor, was attenuated in various cancer types. Our previous study demonstrated a continuous down-regulation of PDCD4 expression in the sequence of normal-borderline-malignant ovarian tissue samples and a significant correlation of PDCD4 expression with disease-free survival. The objective of the current study was to further investigate the function and modulation of PDCD4 in ovarian cancer cells. Principal Findings: We demonstrated that ectopic PDCD4 expression significantly inhibited cell proliferation by inducing cell cycle arrest at G1 stage and up-regulation of cell cycle inhibitors of p27 and p21. Cell migration and invasion were also inhibited by PDCD4. PDCD4 over-expressing cells exhibited elevated phosphatase and tensin homolog (PTEN) and inhibited protein kinase B (p-Akt). In addition, the expression of PDCD4 was up-regulated and it was exported to the cytoplasm upon serum withdrawal treatment, but it was rapidly depleted via proteasomal degradation upon serum re-administration. Treatment of a phosphoinositide 3-kinase (PI3K) inhibitor prevented the degradation of PDCD4, indicating the involvement of PI3K-Akt pathway in the modulation of PDCD4. Conclusion: PDCD4 may play a critical function in arresting cell cycle progression at key checkpoint, thus inhibiting cell proliferation, as well as suppressing tumour metastasis. The PI3K-Akt pathway was implied to be involved in the regulatio

Drugs developed to treat diabetes, liraglutide and lixisenatide, cross the blood brain barrier and enhance neurogenesis

<p>Abstract</p> <p>Background</p> <p>Type 2 diabetes is a risk factor for Alzheimer's disease (AD), most likely linked to an impairment of insulin signalling in the brain. Therefore, drugs that enhance insulin signalling may have therapeutic potential for AD. Liraglutide (Victoza) and exenatide (Byetta) are novel long-lasting analogues of the GLP-1 incretin hormone and are currently available to treat diabetes. They facilitate insulin signalling via the GLP-1 receptor (GLP-1R). Numerous <it>in vitro </it>and <it>in vivo </it>studies have shown that GLP-1 analogues have a range of neuroprotective properties. GLP-1Rs are expressed in the hippocampal area of the brain an important site of adult neurogenesis and maintenance of cognition and memory formation. Therefore, if GLP-1 analogues can cross the blood brain barrier, diffuse through the brain to reach the receptors and most importantly activate them, their neuroprotective effects may be realized.</p> <p>Results</p> <p>In the present study we profiled the GLP-1 receptor agonists liraglutide (Victoza) and lixisenatide (Lyxumia). We measured the kinetics of crossing the blood brain barrier (BBB), activation of the GLP-1R by measuring cAMP levels, and physiological effects in the brain on neuronal stem cell proliferation and neurogenesis. Both drugs were able to cross the BBB. Lixisenatide crossed the BBB at all doses tested (2.5, 25, or 250 nmol/kg bw ip.) when measured 30 min post-injection and at 2.5-25 nmol/kg bw ip. 3 h post-injection. Lixisenatide also enhanced neurogenesis in the brain. Liraglutide crossed the BBB at 25 and 250 nmol/kg ip. but no increase was detectable at 2.5 nmol/kg ip. 30 min post-injection, and at 250 nmol/kg ip. at 3 h post-injection. Liraglutide and lixisenatide enhanced cAMP levels in the brain, with lixisenatide being more effective.</p> <p>Conclusions</p> <p>Our results suggest that these novel incretin analogues cross the BBB and show physiological activity and neurogenesis in the brain, which may be of use as a treatment of neurodegenerative diseases.</p

Impaired Hyperglycemia-Induced Delay in Gastric Emptying in Patients With Type 1 Diabetes Deficient for Islet Amyloid Polypeptide

OBJECTIVE—Slowing of gastric emptying by hyperglycemia, a physiological response to minimize postprandial hyperglycemia, may be impaired in patients with type 1 diabetes. The causes and consequences on glucose homeostasis are unknown

PANIC-ATTAC: A Mouse Model for Inducible and Reversible β-Cell Ablation

OBJECTIVE—Islet transplantations have been performed clinically, but their practical applications are limited. An extensive effort has been made toward the identification of pancreatic β-cell stem cells that has yielded many insights to date, yet targeted reconstitution of β-cell mass remains elusive. Here, we present a mouse model for inducible and reversible ablation of pancreatic β-cells named the PANIC-ATTAC (pancreatic islet β-cell apoptosis through targeted activation of caspase 8) mouse

Regulation of GIP and GLP1 Receptor Cell Surface Expression by N-Glycosylation and Receptor Heteromerization

In response to a meal, Glucose-dependent Insulinotropic Polypeptide (GIP) and Glucagon-like Peptide-1 (GLP-1) are released from gut endocrine cells into the circulation and interact with their cognate G-protein coupled receptors (GPCRs). Receptor activation results in tissue-selective pleiotropic responses that include augmentation of glucose-induced insulin secretion from pancreatic beta cells. N-glycosylation and receptor oligomerization are co-translational processes that are thought to regulate the exit of functional GPCRs from the ER and their maintenance at the plasma membrane. Despite the importance of these regulatory processes, their impact on functional expression of GIP and GLP-1 receptors has not been well studied. Like many family B GPCRs, both the GIP and GLP-1 receptors possess a large extracellular N-terminus with multiple consensus sites for Asn-linked (N)-glycosylation. Here, we show that each of these Asn residues is glycosylated when either human receptor is expressed in Chinese hamster ovary cells. N-glycosylation enhances cell surface expression and function in parallel but exerts stronger control over the GIP receptor than the GLP-1 receptor. N-glycosylation mainly lengthens receptor half-life by reducing degradation in the endoplasmic reticulum. N-glycosylation is also required for expression of the GIP receptor at the plasma membrane and efficient GIP potentiation of glucose-induced insulin secretion from the INS-1 pancreatic beta cell line. Functional expression of a GIP receptor mutant lacking N-glycosylation is rescued by co-expressed wild type GLP1 receptor, which, together with data obtained using Bioluminescence Resonance Energy Transfer, suggests formation of a GIP-GLP1 receptor heteromer

A human ribonuclease induces apoptosis associated with p21WAF1/CIP1 induction and JNK inactivation

<p>Abstract</p> <p>Background</p> <p>Ribonucleases are promising agents for use in anticancer therapy. Among the different ribonucleases described to be cytotoxic, a paradigmatic example is onconase which manifests cytotoxic and cytostatic effects, presents synergism with several kinds of anticancer drugs and is currently in phase II/III of its clinical trial as an anticancer drug against different types of cancer. The mechanism of cytotoxicity of PE5, a variant of human pancreatic ribonuclease carrying a nuclear localization signal, has been investigated and compared to that of onconase.</p> <p>Methods</p> <p>Cytotoxicity was measured by the MTT method and by the tripan blue exclusion assay. Apoptosis was assessed by flow cytometry, caspase enzymatic detection and confocal microscopy. Cell cycle phase analysis was performed by flow cytometry. The expression of different proteins was analyzed by western blot.</p> <p>Results</p> <p>We show that the cytotoxicity of PE5 is produced through apoptosis, that it does not require the proapoptotic activity of p53 and is not prevented by the multiple drug resistance phenotype. We also show that PE5 and onconase induce cell death at the same extent although the latter is also able to arrest the cell growth. We have compared the cytotoxic effects of both ribonucleases in the NCI/ADR-RES cell line by measuring their effects on the cell cycle, on the activation of different caspases and on the expression of different apoptosis- and cell cycle-related proteins. PE5 increases the number of cells in S and G₂/M cell cycle phases, which is accompanied by the increased expression of cyclin E and p21^WAF1/CIP1together with the underphosphorylation of p46 forms of JNK. Citotoxicity of onconase in this cell line does not alter the cell cycle phase distribution and it is accompanied by a decreased expression of XIAP</p> <p>Conclusions</p> <p>We conclude that PE5 kills the cells through apoptosis associated with the p21^WAF1/CIP1induction and the inactivation of JNK. This mechanism is significantly different from that found for onconase.</p

Differential hypoglycaemic, anorectic, autonomic and emetic effects of the glucagon-like peptide receptor agonist, exendin-4, in the conscious telemetered ferret.

Background: Rodents are incapable of emesis and consequently the emetic potential of glucagon-like peptide-1 receptor (GLP-1R) agonists in studies designed to assess a potential blood glucose lowering action of the compound was missed. Therefore, we investigated if the ferret, a carnivore with demonstrated translation capability in emesis research, would identify the emetic potential of the GLP-1R agonist, exendin-4, and any associated effects on gastric motor function, appetite and cardiovascular homeostasis. Methods: The biological activity of the GLP-1R ligands was investigated in vivo using a glucose tolerance test in pentobarbitone-anesthetised ferrets and in vitro using organ bath studies. Radiotelemetry was used to investigate the effect of exendin-4 on gastric myoelectric activity (GMA) and cardiovascular function in conscious ferrets; behaviour was also simultaneously assessed. Western blot was used to characterize GLP-1R distribution in the gastrointestinal and brain tissues. Results: In anesthetised ferrets, exendin-4 (30 nmol/kg, s.c.) reduced experimentally elevated blood glucose levels by 36.3%, whereas the GLP-1R antagonist, exendin (9–39) (300 nmol/kg, s.c.) antagonised the effect and increased AUC0–120 by 31.0% when injected alone (P < 0.05). In animals with radiotelemetry devices, exendin-4 (100 nmol/kg, s.c.) induced emesis in 1/9 ferrets, but inhibited food intake and decreased heart rate variability (HRV) in all animals (P < 0.05). In the animals not exhibiting emesis, there was no effect on GMA, mean arterial blood pressure, heart rate, or core body temperature. In the ferret exhibiting emesis, there was a shift in the GMA towards bradygastria with a decrease in power, and a concomitant decrease in HRV. Western blot revealed GLP-1R throughout the gastrointestinal tract but exendin-4 (up to 300 nM) and exendin (9–39), failed to contract or relax isolated ferret gut tissues. GLP-1R were found in all major brain regions and the levels were comparable those in the vagus nerve. Conclusions: Peripherally administered exendin-4 reduced blood glucose and inhibited feeding with a low emetic potential similar to that in humans (11% vs 12.8%). A disrupted GMA only occurred in the animal exhibiting emesis raising the possibility that disruption of the GMA may influence the probability of emesis occurring in response to treatment with GLP-1R agonists