Testing a Human Antimicrobial RNase Chimera Against Bacterial Resistance.

The emergence of bacterial resistance to the most commonly used antibiotics encourages the design of novel antimicrobial drugs. Antimicrobial proteins and peptides (AMPs) are the key players in host innate immunity. They exert a rapid and multifaceted action that reduces the development of bacterial adaptation mechanisms. Human antimicrobial RNases belonging to the vertebrate specific RNase A superfamily participate in the maintenance of tissue and body fluid sterility. Among the eight human canonical RNases, RNase 3 stands out as the most cationic and effective bactericidal protein against Gram-negative species. Its enhanced ability to disrupt the bacterial cell wall has evolved in detriment of its catalytic activity. Based on structure-functional studies we have designed an RNase 3/1 hybrid construct that combines the high catalytic activity of RNase 1 with RNase 3 bactericidal properties. Next, we have explored the ability of this hybrid RNase to target the development of bacterial resistance on an Acinetobacter baumannii cell culture. Synergy assays were performed in combination with colistin, a standard antimicrobial peptide used as an antibiotic to treat severe infections. Positive synergism was observed between colistin and the RNase 3/1 hybrid protein. Subsequently, using an in vitro experimental evolution assay, by exposure of a bacterial culture to colistin at incremental doses, we demonstrated the ability of the RNase 3/1 construct to reduce the emergence of bacterial antimicrobial resistance. The results advance the potential applicability of RNase-based drugs as antibiotic adjuvants.

Apheresis causes complement deposition on red blood cells (RBCs) and RBC antigen alterations, possibly inducing enhanced clearance.

BACKGROUND: Apheresis is increasingly being applied to collect cells or plasma, even allowing the collection of multiple blood components during one procedure. Although the quality of the cellular and plasma products that are obtained by apheresis have been extensively studied and shown to be of high quality, the impact of apheresis on the red blood cells (RBCs) that are returned to the donor has not been investigated. STUDY DESIGN AND METHODS: The effect of the plasma- or plateletpheresis procedures by four different devices-MCS+ (Haemonetics), PCS2 (Haemonetics), Trima Accel (Terumo BCT), and Autopheresis-C (Auto-C, Fresenius Kabi)-on the RBCs that are returned to the donor was tested in a blinded, prospective trial in a cohort of 25 donors. RESULTS: A rheologic analysis of donor RBCs before and after plasma- or plateletpheresis showed no differences in outcome. However, a strong increase in hemolysis was found in samples from the Trima Accel devices after plateletpheresis, compared to all other machines tested. Furthermore, an increase in complement deposition on RBCs was seen after all plasmapheresis procedures (MCS+, PCS2, and Auto-C). Finally, a significant decrease in the expression of the complement-regulating protein CD59 was seen in all postapheresis samples as well as a significant decrease of the adhesion molecule CD147. CONCLUSION: The increase in complement deposition and the decrease in the expression of CD59 suggests that RBC clearance might be enhanced after return to the donor. Possible side effects due to an increase in hemolysis after Trima Accel plateletpheresis should be further investigated.

Hemoglobin analyses in the Netherlands reveal more than 80 different variants including six novel ones.

More than 20,000 blood samples of individuals living in The Netherlands and suspected of hemolytic anemia or diabetes were analyzed by high resolution cation exchange high performance liquid chromatography (HPLC). Besides common disease-related hemoglobins (Hbs), rare variants were also detected. The variant Hbs were retrospectively analyzed by capillary zone electrophoresis (CZE) and by isoelectric focusing (IEF). For unambiguous identification, the globin genes were sequenced. Most of the 80 Hb variants detected by initial screening on HPLC were also separated by capillary electrophoresis (CE), but a few variants were only detectable with one of these methods. Some variants were unstable, had thalassemic properties or increased oxygen affinity, and some interfered with Hb A2 measurement, detection of sickle cell Hb or Hb A1c quantification. Two of the six novel variants, Hb Enschede (HBA2: c.308G > A, p.Ser103Asn) and Hb Weesp (HBA1: c.301C > T, p.Leu101Phe), had no clinical consequences. In contrast, two others appeared clinically significant: Hb Ede (HBB: c.53A > T, p.Lys18Met) caused thalassemia and Hb Waterland (HBB: c.428C > T, pAla143Val) was related to mild polycytemia. Hb A2-Venlo (HBD: c.193G > A, p.Gly65Ser) and Hb A2-Rotterdam (HBD: c.38A > C, p.Asn13Thr) interfered with Hb A2 quantification. This survey shows that HPLC analysis followed by globin gene sequencing of rare variants is an effective method to reveal Hb variants.