Perceived barriers to computerised quality documentation during anaesthesia: a survey of anaesthesia staff

BACKGROUND: Underreporting of intraoperative events in anaesthesia is well-known and compromises quality documentation. The reasons for such omissions remain unclear. We conducted a questionnaire-based survey of anaesthesia staff to explore perceived barriers to reliable documentation during anaesthesia. METHODS: Participants anonymously completed a paper-based questionnaire. Predefined answers referred to potential barriers. Additional written comments were encouraged. Differences between physician and nurse anaesthetists were tested with t-tests and chi-square tests. RESULTS: Twenty-five physician and 30 nurse anaesthetists (81% of total staff) completed the survey. The reported problems referred to three main categories: (I) potential influences related to working conditions and practices of data collection, such as premature entry of the data (indicated by 85% of the respondents), competing duties (87%), and interfering interruptions or noise (67%); (II) problems referring to institutional management of the data, for example lacking feedback on the results (95%) and lacking knowledge about what the data are used for (75%); (III) problems related to specific attitudes, e.g., considering these data not useful for quality improvement (47%). Physicians were more sceptical than nurses regarding the relevance of these data for quality and patient safety. CONCLUSIONS: The common perceived difficulties reported by physician and nurse anaesthetists resemble established barriers to incident reporting and may similarly act as barriers to quality documentation during anaesthesia. Further studies should investigate if these perceived obstacles have a causal impact on quality reporting in anaesthesia. TRIAL REGISTRATION: ClinicalTrials.gov identifier is NCT01524484. Registration date: January 21, 2012

Cultivation of Chinese hamster ovary (CHO) cells in protein-free media supplemented with plant peptones is sufficient to prevent proteolysis of secreted recombinant glycoproteins

Chinese hamster ovary (CHO) cells are widely utilized to produce complex recombinant (glyco)proteins due to their ability to perform efficiently post-translational modifications and to grow in suspension at very high densities. For therapeutic purposes, biosafety has to be ensured at each step of the protein purification, but also during the cultivation of the cells. Consequently, serum and animal components are progressively banned from new culture medium formulations and now protein-free (PF) media are available for the cultivation of CHO cells. Unfortunately, a decrease in cell density and productivity is often associated with the shift from serum-containing media to PF media as well as an increase in the risk of proteolysis due to the lack of serum protease inhibitors. We have used CHO-320 cells that are secreting recombinant human interferon-ã (IFN-ã), as a cellular model to study cell growth, glycoprotein secretion, glycosylation heterogeneities, as well as the extent of proteolysis in such media. After a direct adaptation from adhesion in serum to suspension in a homemade PF medium supplemented with different plant protein hydrolysates (peptones), CHO-320 cells grew faster than in serum-containing medium. IFN-ã production was similar from a quantitative viewpoint, but most the IFN-ã was bi-glycosylated whereas in serumcontaining medium, the non-, mono-, and bi-glycosylated glycoforms were found in equal proportions. Further supplementation of this PF medium with some plant peptones increased the IFN-ã secretion by up to 60% and cell density by up to 30% upon cultivation in suspension. The cultivation of CHO-320 cells inside microcarriers in PF media with plant peptones was also investigated. It appears that in such culture conditions, plant peptones were only able to increase the IFN-ã secretion while the cell density remains apparently unaffected. Depending on the type of plant peptones utilized, IFN-ã underwent proteolysis in the culture medium. Some extracellular proteases expressed by CHO cells were identified and their relationships to this proteolytic event was investigated. CHO cells express members of the matrix metalloproteinases (MMP) and of serine proteases, either at the cell surface i.e. (pro)MMP-14, dipeptidyl peptidase IV (DPPIV) and tripeptidylpeptidase II (TPPII) or released them in the extracellular medium i.e. proMMP-9, urinary-type (uPA) and tissue-type plasminogen activators (tPA). In addition and probably more importantly, the presence of their natural inhibitors was also detected i.e. TIMP-1 and -2 as well as plasminogen activator inhibitor type 1 (PAI-1). It has also been shown that peptones could act as a source of exogenous inhibitors of tripeptidylpeptidase II (TPPII). Our results indicate that the proteolytic activity of these proteases was tightly regulated at a posttranslational level by (i) their expression as inactive precursors (proMMP-9 and proMMP-14), (ii) by the presence of some endogenous inhibitors and (iii) by the presence of some peptides in plant peptones acting as exogenous TPPII inhibitors. Nevertheless, when some plant peptones were added to the nutritive medium, IFN-ã proteolysis was observed. We showed that this proteolytic event was related to a cysteine protease already present as residual enzymatic contamination of the peptones and not to extracellular proteases naturally expressed by CHO cells. Finally, at a molecular level, we have some evidence that plant peptones could act as a nutritional source. Indeed, plant peptones could behave as competitive substrates of peptide transporters and/or TPPII. Consequently, di- and or tripeptides present in the peptones can directly be absorbed by peptide transporters, whereas some larger peptides in the peptones could be cleaved into tripeptides by TPPII and afterwards be internalised. Nevertheless, the effects of the peptones cannot be reproduced by amino acids supplementation and some of their properties have to be interpreted as an indication of the presence of bioactive substances. To conclude, plant peptones are highly beneficial supplements for the cultivation of CHO cells in protein-free media. Unfortunately, their undefined nature represents an important limitation, which can have adverse effects and, therefore, the biologically active components present in the peptones should be identified to substitute peptones from the formulation of culture media as well as the transduction cascades should be dissected at the molecular level.Doctorat en sciences (sciences chimiques) (CHIM 3)--UCL, 200

Plant peptones: Nutritional properties sustaining recombinant protein secretion by CHO cells

The network metabolic model of High-FiveTM insect cells based on the consumption / production of metabolites and the protein, DNA/RNA, lipid and carbohydrates content of the cells has made it possible to study the internal fluxes of these cells’ metabolism

Origin of rice protein hydrolysates added to protein-free media alters secretion and extracellular proteolysis of recombinant interferon-gamma as well as CHO-320 cell growth

CHO-320 cells, cultivated in suspension in a protein-free medium supplemented with rice protein hydrolysates (peptones), secrete recombinant interferon-gamma (IFN-gamma) that undergo will or will not proteolysis, depending on the origin of the peptones. This proteolytic event, as well as the appearance of an unidentified 70 kDa gelatinase-like protease, are attributed to a cysteine protease. Casein zymographies revealed that one rice protein hydrolysate, but not another, contains a papain-like cysteine protease whose activity is undetectable in solution. This work underlines the significance of the origin of peptones when considered as supplements in serum- and protein-free media for overproduction of recombinant proteins

Toxicology, biodistribution and shedding profile of a recombinant measles vaccine vector expressing HIV-1 antigens, in cynomolgus macaques

As a new human immunodeficiency virus type 1 (HIV-1) vaccine approach, the live-attenuated measles virus (MV) Schwarz vaccine strain was genetically engineered to express the F4 antigen (MV1-F4). F4 is a fusion protein comprising HIV-1 antigens p17 and p24, reverse transcriptase and Nef. This study assessed the toxicity, biodistribution and shedding profiles of MV1-F4. Cynomolgus macaques were intramuscularly immunized one or three times with the highest dose of MV1-F4 intended for clinical use, the reference (Schwarz) measles vaccine or saline, and monitored clinically for 11 or 85 days. Toxicological parameters included local and systemic clinical signs, organ weights, haematology, clinical and gross pathology and histopathology. Both vaccines were well tolerated, with no morbidity, clinical signs or gross pathological findings observed. Mean spleen weights were increased after three doses of either vaccine, which corresponded with increased numbers and/or sizes of germinal centers. This was likely a result of the immune response to the vaccines. Either vaccine virus replicated preferentially in secondary lymphoid organs and to a lesser extent in epithelium-rich tissues (e.g., intestine, urinary bladder and trachea) and the liver. At the expected peak of viremia, viral RNA was detected in some biological fluid samples from few animals immunized with either vaccine, but none of these samples contained infectious virus. In conclusion, no shedding of infectious viral particles was identified in cynomolgus monkeys after injection of MV1-F4 or Schwarz measles vaccines. Furthermore, no toxic effect in relation to the MV vaccination was found with these vaccines in this study