A new vector for heterologous gene expression in Escherichia coli with increased stability in the absence of antibiotic.

Expression vectors for industrial production should be stable and allow tight control of protein synthesis. This is necessary to ensure plasmid transmission to daughter cells in order to achieve a stable population capable of synthesizing high amounts of the target protein. A high-copy-number plasmid, pAE, was previously used for laboratory-scale production of recombinant human granulocyte colony-stimulating factor (rhG-CSF) and the Schistosoma mansoni fatty acid binding protein (rSm14), but it was unstable for large-scale production. Therefore, here we evaluated a new expression vector derived from pAE, pAR-KanI, which combines two plasmid replication strategies: a high-copy plasmid pUC origin of replication as pAE, and a par locus sequence derived from pSC101, which is typical of low copy plasmids, for rhG-CSF and rSm14 production in Escherichia coli. Clones bearing these constructs were cultivated in two complex media (2YT and auto-induction) and both yielded higher-than-95% resistant colonies, before and after induction, either with or without antibiotics. In 2YT medium, we obtained 244 µg/mL of rSm14, 181 µg/mL and 392 µg/mL for rhG-CSF, with and without glucose, respectively. In auto-induction medium without antibiotics, 147 µg/mL of rSm14 and 162 µg/mL of rhG-CSF were obtained. The new vector presented high stability for the production of both recombinant proteins in complex media in Escherichia coli, even in the absence of antibiotics, making the pAR-KanI a promising vector for industrial production of recombinant proteins.

Nondipping Blood Pressure Patterns Predict Obstructive Sleep Apnea in Patients Undergoing Ambulatory Blood Pressure Monitoring.

A nondipping blood pressure (BP) pattern is common in patients with obstructive sleep apnea (OSA). However, it is unclear how useful a nondipping BP pattern is in screening for OSA. In this cross-sectional study, we recruited consecutive patients with clinical indications for performing ambulatory BP monitoring evaluating the following dipping patterns: (1) normal: ≥10% but <20%; (2) extreme: ≥20%; (3) reduced: ≥0% but <10%; and (4) reverse (riser): <0%. Sleep questionnaires and sleep studies were performed within 7 days after ambulatory BP monitoring. OSA was defined as an apnea-hypopnea index ≥15 events/h. We evaluated 153 patients (OSA frequency, 50.3%). Patients with OSA had higher BPs during sleep, were taking more antihypertensive drugs, and more frequently used hypertensive drugs during the night than patients without OSA. Considering systolic BP, the frequency of OSA in patients with reverse dippers (73.5%) was higher than normal (37.3%), extreme (46.2%), and reduced dippers (49.1%; P=0.012). For diastolic BP, OSA was more common in reduced (66.7%) and reverse dippers (69.6%) as compared to normal (41.4%) or extreme dippers (33.3%; P=0.007). In the regression analysis, reverse systolic dipper was independently associated with OSA (odds ratio, 3.92; 95% CI, 1.31-11.78). Both reduced and reverse diastolic dippers increased the likelihood of OSA for 2.7-fold and 3.5-fold, respectively. Snoring and positive sleep questionnaire findings were associated with a modest increase in the accuracy of reverse dipping pattern for predicting OSA. In conclusion, reverse systolic, as well as reduced and reverse diastolic dippers are independently associated with OSA.