A novel mechanism of CD40-induced apoptosis of carcinoma cells involving TRAF3 and JNK/AP-1 activation

Membrane-presented CD40 agonists can induce apoptosis in carcinoma, but not normal homologous epithelial cells, whereas soluble agonists are growth inhibitory but not proapoptotic unless protein synthesis is blocked. Here we demonstrate that membrane-presented CD40 ligand (CD154) (mCD40L), but not soluble agonists, triggers cell death in malignant human urothelial cells via a direct mechanism involving rapid upregulation of TNFR-associated factor (TRAF)3 protein, without concomitant upregulation of TRAF3 mRNA, followed by activation of the c-Jun N-terminal kinase (JNK)/activator protein-1 (AP-1) pathway and induction of the caspase-9/caspase-3-associated intrinsic apoptotic machinery. TRAF3 knockdown abrogated JNK/AP-1 activation and prevented CD40-mediated apoptosis, whereas restoration of CD40 expression in CD40-negative carcinoma cells restored apoptotic susceptibility via the TRAF3/AP-1-dependent mechanism. In normal human urothelial cells, mCD40L did not trigger apoptosis, but induced rapid downregulation of TRAF2 and 3, thereby paralleling the situation in B-lymphocytes. Thus, TRAF3 stabilization, JNK activation and caspase-9 induction define a novel pathway of CD40-mediated apoptosis in carcinoma cells.

Induced alkalosis and caffeine supplementation : effects on 2,000-m rowing performance

Introduction: The purpose of this investigation was to determine the effect of ingested caffeine, sodium bicarbonate, and their combination on 2,000-m rowing performance, as well as on induced alkalosis (blood and urine pH and blood bicarbonate concentration [HCO3 -]), blood lactate concentration ([La-]), gastrointestinal symptoms, and rating of perceived exertion (RPE). Methods: In a double-blind, crossover study, 8 well-trained rowers performed 2 baseline tests and 4 × 2,000-m rowing-ergometer tests after ingesting 6 mg/kg caffeine, 0.3 g/kg body mass (BM) sodium bicarbonate, both supplements combined, or a placebo. Capillary blood samples were collected at preingestion, pretest, and posttest time points. Pairwise comparisons were made between protocols, and differences were interpreted in relation to the likelihood of exceeding the smallest-worthwhile- change thresholds for each variable. A likelihood of >75% was considered a substantial change. Results: Caffeine supplementation elicited a substantial improvement in 2,000-m mean power, with mean (± SD) values of 354 ± 67 W vs. placebo with 346 ± 61 W. Pretest [HCO3 -] reached 29.2 ± 2.9 mmol/L with caffeine + bicarbonate and 29.1 ± 1.9 mmol/L with bicarbonate. There were substantial increases in pretest [HCO3 -] and pH and posttest urine pH after bicarbonate and caffeine + bicarbonate supplementation compared with placebo, but unclear performance effects. Conclusions: Rowers' performance in 2,000-m efforts can improve by ~2% with 6 mg/kg BM caffeine supplementation. When caffeine is combined with sodium bicarbonate, gastrointestinal symptoms may prevent performance enhancement, so further investigation of ingestion protocols that minimize side effects is required. ABSTRACT FROM AUTHOR

G-CSF treatment can attenuate dexamethasone-induced reduction in C2C12 myotube protein synthesis

Granulocyte-colony stimulating factor (G-CSF) has been demonstrated to enhance skeletal muscle recovery following injury and increases muscle function in the context of neuromuscular disease in rodent models. However, understanding of the underlying mechanisms used by G-CSF to mediate these functions remains poor. G-CSF acts on responsive cells through binding to a specific membrane spanning receptor, G-CSFR. Recently identified, the G-CSFR is expressed in myoblasts, myotubes and mature skeletal muscle tissue. Therefore, elucidating the actions of G-CSF in skeletal muscle represents an important prerequisite to consider G-CSF as a therapeutic agent to treat skeletal muscle. Here we show for the first time that treatment with moderate doses (4 and 40ng/ml) of G-CSF attenuates the effects of dexamethasone in reducing protein synthesis in C2C12 myotubes. However, a higher dose (100ng/ml) of G-CSF exacerbates the dexamethasone-induced reduction in protein synthesis. In contrast, G-CSF had no effect on basal or dexamethasone-induced protein degradation, nor did G-CSF influence the phosphorylation of Akt, STAT3, Erk1/2, Src, Lyn and Erk5 in C2C12 myotubes. In conclusion, physiologically relevant doses of G-CSF may attenuate reduced skeletal muscle protein synthesis during catabolic conditions, thereby improving recovery.

Efficacy of nefazodone in the treatment of neuroleptic induced extrapyramidal side effects: a double-blind randomised parallel group placebo-controlled trial.

Many atypical antipsychotics show antagonism at both serotonergic and dopaminergic neurones and show fewer extrapyramidal side effects (EPS). Nefazodone blocks postsynaptic 5HT2A receptors and weakly inhibits serotonin reuptake. This study aimed to elucidate the role of nefazodone in the treatment of antipsychotic-induced EPS. The trial was a double-blind, randomised, placebo-controlled trial of patients requiring antipsychotic treatment with haloperidol 10mg daily; from which a subgroup of patients who developed EPS were selected for the study. Patients were randomised to add-on therapy with either placebo (n=24) or nefazodone (n=25) 100mg bd. EPS were measured on days 0, 3 and 7 using the Simpson Angus, Barnes akathisia, abnormal involuntary movement and Chouinard scales. Nefazodone significantly reduced EPS as measured by both the Simpson Angus scale and CGI (p=0.007 and 0.0247, respectively). Akathisia and tardive dyskinesia did not differ between the two groups (p=0.601; p=0.507, respectively). These results suggest the role of 5HT2 antagonism in the mechanism of action of atypical antipsychotics with respect to lowering rates of drug-induced EPS. In addition, a therapeutic role for nefazodone is suggested in the treatment of antipsychotic-induced EPS.

WISE-2005: bed-rest induced changes in bone mineral density in women during 60 days simulated microgravity

To better understand the effects of prolonged bed-rest in women, 24 healthy women aged 25 to 40 years participated in 60-days of strict 6° head-down tilt bed-rest (WISE-2005). Subjects were assigned to either a control group (CON, n=8) which performed no countermeasure, an exercise group (EXE, n=8) undertaking a combination of resistive and endurance training or a nutrition group (NUT, n=8), which received a high protein diet. Using peripheral quantitative computed tomography (pQCT) and dual X-ray absorptiometry (DXA), bone mineral density (BMD) changes at various sites, body-composition and lower-leg and forearm muscle cross-sectional area were measured up to 1-year after bed-rest. Bone loss was greatest at the distal tibia and proximal femur, though losses in trabecular density at the distal radius were also seen. Some of these bone losses remained statistically significant one-year after bed-rest. There was no statistically significant impediment of bone loss by either countermeasure in comparison to the control-group. The exercise countermeasure did, however, reduce muscle cross-sectional area and lean mass loss in the lower-limb and also resulted in a greater loss of fat mass whereas the nutrition countermeasure had no impact on these parameters. The findings suggest that regional differences in bone loss occur in women during prolonged bed-rest with incomplete recovery of this loss one-year after bed-rest. The countermeasures as implemented were not optimal in preventing bone loss during bed-rest and further development is required.

The effects of bed-rest and countermeasure exercise on the endocrine system in male adults: evidence for immobilization-induced reduction in sex hormone-binding globulin levels

BACKGROUND AND AIM: There is limited data on the effects of inactivity (prolonged bed-rest) on parameters of endocrine and metabolic function; we therefore aimed to examine changes in these systems during and after prolonged (56- day) bed-rest in male adults. SUBJECTS AND METHODS: Twenty healthy male subjects underwent 8 weeks of strict bed-rest and 12 months of follow-up as part of the Berlin Bed Rest Study. Subjects were randomized to an inactive group or a group that performed resistive vibration exercise (RVE) during bed-rest. All outcome parameters were measured before, during and after bed-rest. These included body composition (by whole body dual X-ray absorptiometry), SHBG, testosterone (T), estradiol (E2), PRL, cortisol (C), TSH and free T3 (FT3). RESULTS: Serum SHBG levels decreased in inactive subjects but remained unchanged in the RVE group (p<0.001). Serum T concentrations increased during the first 3 weeks of bed-rest in both groups (p<0.0001), while E2 levels sharply rose with re-mobilization (p<0.0001). Serum PRL decreased in the control group but increased in the RVE group (p=0.021). C levels did not change over time (p≥0.10). TSH increased whilst FT3 decreased during bed-rest (p all ≤0.0013). CONCLUSIONS: Prolonged bed-rest has significant effects on parameters of endocrine and metabolic function, some of which are related to, or counteracted by physical activity.

Cisplatin-induced formation of biocompatible and biodegradable polypeptide-based vesicles for targeted anticancer drug delivery

Novel cisplatin (CDDP)-loaded, polypeptide-based vesicles for the targeted delivery of cisplatin to cancer cells have been prepared. These vesicles were formed from biocompatible and biodegradable maleimide-poly(ethylene oxide)114-b-poly(L-glutamic acid)12 (Mal-PEG114-b-PLG12) block copolymers upon conjugation with the drug itself. CDDP conjugation forms a short, rigid, cross-linked, drug-loaded, hydrophobic block in the copolymer, and subsequently induces self-assembly into hollow vesicle structures with average hydrodynamic diameters (Dh) of ∼ 270 nm. CDDP conjugation is critical to the formation of the vesicles. The reactive maleimide-PEG moieties that form the corona and inner layer of the vesicles were protected via formation of a reversible Diels-Alder (DA) adduct throughout the block copolymer synthesis so as to maintain their integrity. Drug release studies demonstrated a low and sustained drug release profile in systemic conditions (pH = 7.4, [Cl(-)] = 140 mM) with a higher "burst-like" release rate being observed under late endosomal/lysosomal conditions (pH = 5.2, [Cl(-)] = 35 mM). Further, the peripheral maleimide functionalities on the vesicle corona were conjugated to thiol-functionalized folic acid (FA) (via in situ reduction of a novel bis-FA disulfide, FA-SS-FA) to form an active targeting drug delivery system. These targeting vesicles exhibited significantly higher cellular binding/uptake into and dose-dependent cytotoxicity toward cancer cells (HeLa) compared to noncancerous cells (NIH-3T3), which show high and low folic acid receptor (FR) expression, respectively. This work thus demonstrates a novel approach to polypeptide-based vesicle assembly and a promising strategy for targeted, effective CDDP anticancer drug delivery.

PKR is not obligatory for high-fat diet-induced obesity and its associated metabolic and inflammatory complications

Protein kinase R (PKR) has previously been suggested to mediate many of the deleterious consequences of a high-fat diet (HFD). However, previous studies have observed substantial phenotypic variability when examining the metabolic consequences of PKR deletion. Accordingly, herein, we have re-examined the role of PKR in the development of obesity and its associated metabolic complications in vivo as well as its putative lipid-sensing role in vitro. Here we show that the deletion of PKR does not affect HFD-induced obesity, hepatic steatosis or glucose metabolism, and only modestly affects adipose tissue inflammation. Treatment with the saturated fatty acid palmitate in vitro induced comparable levels of inflammation in WT and PKR KO macrophages, demonstrating that PKR is not necessary for the sensing of pro-inflammatory lipids. These results challenge the proposed role for PKR in obesity, its associated metabolic complications and its role in lipid-induced inflammation.