Mutations inducing divergent shifts of constitutive activity reveal different modes of binding among catecholamine analogues to the beta(2)-adrenergic receptor

1. We compared the changes in binding energy generated by two mutations that shift in divergent directions the constitutive activity of the human beta(2) adrenergic receptor (beta(2)AR). 2. A constitutively activating mutant (CAM) and the double alanine replacement (AA mutant) of catechol-binding serines (S204A, S207A) in helix 5 were stably expressed in CHO cell lines, and used to measure the binding affinities of more than 40 adrenergic ligands. Moreover, the efficacy of the same group of compounds was determined as intrinsic activity for maximal adenylyl cyclase stimulation in wild-type beta(2)AR. 3. Although the two mutations had opposite effects on ligand affinity, the extents of change were in both cases largely correlated with the degree of ligand efficacy. This was particularly evident if the extra loss of binding energy due to hydrogen bond deletion in the AA mutant was taken into account. Thus the data demonstrate that there is an overall linkage between the configuration of the binding pocket and the intrinsic equilibrium between active and inactive receptor forms. 4. We also found that AA mutation-induced affinity changes for catecholamine congeners gradually lacking ethanolamine substituents were linearly correlated to the loss of affinity that such modifications of the ligand cause for wild-type receptor. This indicates that the strength of bonds between catechol ring and helix 5 is critically dependent on the rest of interactions of the beta-ethanolamine tail with other residues of the beta(2)-AR binding pocket.     

Role of gemcitabine-based combination therapy in the management of advanced pancreatic cancer: A meta-analysis of randomised trials

BackgroundPancreatic cancer is the fourth leading cause of cancer-related death worldwide. Gemcitabine is the mainstay treatment for advanced disease. However, almost all up-to-date trials, that evaluated the benefit of gemcitabine-combination schedules, failed to demonstrate an improvement in overall survival (OS). In this study, we performed a systematic review and a meta-analysis of randomised clinical trials (RCTs) to investigate the efficacy and safety of gemcitabine-based combination regimens as compared to gemcitabine alone in the management of pancreatic cancer.MethodsClinical trials were collected by searching different databases (PubMed, Embase and the Central Registry of Controlled Trials of the Cochrane Library) and abstracts from major cancer meetings. We considered period ranging from January 1997 to January 2012. Primary end-point was OS, secondary end-points were response rate (RR), disease control rate (DCR) and safety. Hazard ratios (HRs) of OS, odds-ratios (ORs) of RR, DCR and risk ratios of grade 3–4 toxicity rates (TRs), were extracted as presented in retrieved studies and used for statistical analysis. Meta-analytic estimates were derived using random-effects model.FindingsThirty-four trials for a total of 10,660 patients were selected and included in the final analysis. The analysis showed that combination chemotherapy confers benefit in terms of OS (HR: 0.93; 95% confidence interval (CI): 0.89–0.97; p = 0.001). ORs for both RR and DCR demonstrated a significant advantage for combination therapy (OR for RR: 0.60, 95%CI: 0.47–0.76, p < 0.001; OR for DCR: 0.79; 95%CI: 0.66–0.93; p = 0.006). Toxicities were more frequent with the combination treatment and significance in terms of risk ratio was reached for diarrhoea (0.53, 95%CI: 0.36–0.79), nausea (0.74, 95%CI: 0.56–0.96), neutropenia (0.71, 95%CI: 0.59–0.85) and thrombocytopenia (0.57, 95%CI: 0.43–0.75).InterpretationThe combination chemotherapy as compared to gemcitabine alone significantly improves OS in advanced pancreatic cancer (APC). However, this advantage is marginal whereas the treatment-related toxicity is increased, suggesting the use of gemcitabine-based combination regimens only in selected patient populations. New prospective trials, based on translational approaches and innovative validated biomarkers, are eagerly awaited on this topic.     

Possible role of milky spots in mesothelial transplantation

Milky spots are very small omental organs, in contact with peritoneal membrane, devoid of capsule and consisting of macrophages, lymphocytes and a few plasma cells supported by blood and lymphatic vessels. The exact role of these particular organs is still not clear, but they are similar to lymphatic structures and it is clear that they play a role in peritoneal infection and abdominal tumors. Peritoneal dialysis seems to activate the milky spots changing their morphology. The authors try to formulate some hypotheses on the role played by these little omental organs during autologous mesothelial transplant.     

DNA damage induced by 7,12-dimethylbenz[a]anthracene in the liver and the mammary gland of rats exposed to polycyclic aromatic hydrocarbon enzyme inducers during perinatal life

The long-lasting modulating effect induced by the prenatal or neonatal exposure to phenobarbital (PB) and aroclor on the genotoxic activity of 7,12-dimethylbenz[a]anthracene (DMBA) in female Sprague-Dawley rats was studied. The effect was measured as DNA damage evaluated in the liver and in the mammary gland of 55-day-old animals, 4 and 24 h after an i.g. injection of 80 mg/kg of DMBA. PB was given per os, i.g. or in drinking water to pregnant females and by i.g. only to neonates or in adult progeny. Aroclor was injected i.g. in prenatal and in neonatal life, and a second dose was given in adult life. Under these experimental conditions it was shown that DNA damage kinetics caused by DMBA are modulated by exposure to PB and, to a minor extent, by aroclor. The amount and persistence of DNA damage were highest when PB was administered to neonates. An average 2-fold increase in the elution constants (K) of DNA in the liver and the mammary gland was observed 4 h after DMBA treatment, as compared to uninduced animals. Repeated enzyme induction by PB seems to reduce DMBA genotoxicity, as shown by a decrease in DNA damage and persistence in the liver and mammary gland. The inducibility of the monooxygenase enzyme system in perinatal life favouring metabolic activation or inactivation of polycyclic aromatic hydrocarbons might be critical in determining individual susceptibility of adult progeny to chemical carcinogenesis by DMBA.     

Anatomy of the Internal Iliac Vein: Implications for Uterine Transplant

Study Objective

Uterine transplantation has proven feasible since the first live birth reported in 2014. To enable attachment of the uterus in the recipient, long vascular pedicles of the uterine and internal iliac vessels were obtained during donor hysterectomy, which required a prolonged laparotomy to the living donors. To assist further attempts at uterine transplantation, our video serves to review literature reports of internal iliac vein anatomy and demonstrate a laparoscopic dissection of cadaver pelvic vascular anatomy.