Structure and Processes of Existing Practice in Radiotherapy Peer Review: A Systematic Review of the Literature.

Peer review in radiotherapy is an essential step in clinical quality assurance to avoid planning-related errors that can impact on patient safety and treatment outcomes. Despite recommendations that radiotherapy centres should include peer review in their regular quality assurance pathway, adoption of the practice has not been universal, and to date there have been no formal guidelines set out to standardise the process. We undertook a systematic review of the literature to determine existing practice in radiotherapy peer review internationally, with respect to meeting structure and processes, in order to define a standardised framework. A PubMed and Web of Science search identified 17 articles detailing peer review practice. The results revealed significant variation in peer review processes between institutions, and a lack of consensus on documentation and reporting. Variations in the grading of outcomes of peer review were also noted. Taking into account the results of this review, a framework for standardising the process and outcome documentation for peer review has been developed. This can be utilised by radiotherapy centres introducing or updating peer review practice, and can facilitate meaningful evaluation of the clinical impact of peer review in the future.

Web-based Conferencing: What Radiology Educators Need to Know.

Advances in technology have resulted in the significant growth of web-based conferencing and teaching. While these remote sessions have many advantages, they may result in challenges and frustration for both host and attendees when there are technological issues, poor or distracting audio, or ineffective presentation styles. Knowing a few basic concepts behind web conferencing and preparing in advance can markedly improve the experience and facilitate effective distance learning and collaboration.

A systematic review of the prevalence and incidence of prescribing errors with high-risk medicines in hospitals.

WHAT IS KNOWN: Prescribing errors are the most common type of error in the medication use process. However, there is a paucity of literature regarding the prevalence or incidence of prescribing errors in high-risk medicines (HRMs). HRMs bear a heightened risk of causing significant patient harm when they are used in error. OBJECTIVE: The aim of this research was to systematically investigate the literature regarding the prevalence and incidence of prescribing errors in HRMs in inpatient settings. METHODS: A search strategy was developed based on four categories of keywords: prescribing errors, HRMs, hospital inpatients, and prevalence or incidence. All keywords were searched for in Medline, Embase, Cochrane and the International Pharmaceutical Abstracts. The search was limited to English quantitative studies that reported the incidence or prevalence of prescribing errors by medical prescribers, whether they were seniors or juniors, since 1985. RESULTS: Of the 3507 records identified, nine studies met the review criteria. The most frequent denominator in the included studies was medication orders, in eight studies, ranged from 0·24 to 89·6 errors per 100 orders of HRMs. Two studies reported 107 and 218 errors per 100 admissions prescribed HRMs, and one study reported 27·2 errors per 100 prescriptions with a HRM. The incidence of prescribing errors could not be calculated. WHAT IS NEW AND CONCLUSION: The prevalence of prescribing errors in HRMs in the inpatient setting has a very wide range that reflects the different data collection methods used within the included studies. Future studies in prescribing errors should use standardized approaches to enable comparison.

Serum microRNA biomarkers for drug-induced liver injury.

New biomarkers of drug-induced liver injury (DILI) are required in the clinic and in preclinical pharmaceutical evaluation. Liver-enriched microRNAs are promising serum biomarkers of acetaminophen-induced acute liver injury in mice. The utility of circulating microRNAs as biomarkers of human acute DILI is discussed in the context of correlation with existing biomarkers of liver injury and patient outcomes in acetaminophen toxicity, mechanisms of cellular microRNA release, and their potential advantages over current clinical biomarkers of DILI.

Transcription factor dynamics.

Gene expression is a fundamental process that is highly conserved from humans to bacteria. The first step in gene expression, transcription, is performed by structurally conserved DNA-dependent RNA polymerases (RNAPs), which results in the synthesis of an RNA molecule from a DNA template. In bacteria, a single species of RNAP is responsible for transcribing both stable RNA (i.e. t- and rRNA) and protein-encoding genes (i.e. mRNA), unlike eukaryotic systems, which use three distinct RNAP species to transcribe the different gene classes (RNAP I transcribes most rRNA, RNAP II transcribes mRNA, and RNAP III transcribes tRNA and 5S rRNA). The versatility of bacterial RNAP is dependent on both dynamic interactions with co-factors and the coding sequence of the template DNA, which allows RNAP to respond appropriately to the transcriptional needs of the cell. Although the majority of the research on gene expression has focused on the initiation stage, regulation of the elongation phase is essential for cell viability and represents an important topic for study. The elongation factors that associate with RNAP are unique and highly conserved among prokaryotes, making disruption of their interactions a potentially important target for antibiotic development. One of the most significant advances in molecular biology over the last decade has been the use of green fluorescent protein (GFP) and its spectral variants to observe the subcellular localization of proteins in live intact cells. This review discusses transcription dynamics with respect to RNAP and its associated transcription elongation factors in the two best-studied prokaryotes, Escherichia coli and Bacillus subtilis.

Different patterns of integral membrane protein localization during cell division in Bacillus subtilis.

Cell division in rod-shaped bacteria nearly always occurs exactly at mid-cell and is dependent on the formation of the cytokinetic FtsZ ring and its associated division proteins. Many thousands of copies of division, or septum-specific proteins assemble at this site and may lead to the exclusion of other integral membrane proteins that are normally able to diffuse freely throughout the cytoplasmic membrane. In this study we have investigated the localization of a series of integral membrane proteins in Bacillus subtilis and we show that the recruitment of division and septum-specific proteins does not necessarily preclude the diffusion of other integral membrane proteins. However, some proteins, namely ATP synthase and succinate dehydrogenase, are reduced/absent from the mid-cell region at the onset of cell division, which may reflect an association with lipid domains rich in phosphatidylglycerol that are thought to be present at diminished levels at sites of cell division.

Dynamic localization of membrane proteins in Bacillus subtilis.

The subcellular localization of membrane proteins in Bacillus subtilis was examined by using fluorescent protein fusions. ATP synthase and succinate dehydrogenase were found to localize within discrete domains on the membrane rather than being homogeneously distributed around the cell periphery as expected. Dual labelling of cells indicated partial colocalization of ATP synthase and succinate dehydrogenase. Further analysis using an ectopically expressed phage protein gave the same localization patterns as ATP synthase and succinate dehydrogenase, implying that membrane proteins are restricted to domains within the membrane. 3D reconstruction of images of the localization of ATP synthase showed that domains were not regular and there was no bias for localization to cell poles or any other positions. Further analysis revealed that this localization was highly dynamic, but random, implying that integral membrane proteins are free to diffuse two-dimensionally around the cytoplasmic membrane.

Fusion of C3d molecule with bovine rotavirus VP7 or bovine herpesvirus type 1 glycoprotein D inhibits immune responses following DNA immunization.

The binding of the complement C3d molecule with receptors on B cells and/or follicular dendritic cells (FDCs) influences the induction of humoral immune responses. For example, C3d fused to an antigen has been shown to have a strong adjuvant effect on antibody production. We investigated the possibility that co-expression of antigen and C3d as a fusion protein could enhance antigen-specific immune responses, following plasmid immunization. One or two copies of murine C3d-cDNA, C3d or (C3d)(2), respectively, were cloned together with bovine rotavirus (BRV) VP7 or bovine herpesvirus type 1 (BHV-1) glycoprotein D (gD) genes. All constructs contained a signal peptide that resulted in the secretion of the expressed proteins. In vitro, the characterization of the chimeric proteins indicated that both VP7 and gD retained their antigenicity and the C3d remained biologically active. However, immunization with plasmids encoding VP7-C3d chimeras did not enhance rotavirus-specific antibody responses and the frequency of BRV-specific IFN-gamma secreting cells in the spleens were significantly lower in mice immunized with pVP7-(C3d)(2) when compared with mice immunized with plasmid encoding VP7. The same pattern of immune responses was observed for plasmids encoding gD-C3d. Both gD-specific antibody responses and the frequency of gD-specific IFN-gamma secreting cells were significantly lower in mice immunized with plasmid expressing gD-C3d chimeras when compared with mice immunized with plasmid encoding gD alone. These results indicate that co-expression of C3d with an antigen actually inhibit both humoral and cell-mediated antigen-specific immune responses.

Medically intractable, localization-related epilepsy with normal MRI: presurgical evaluation and surgical outcome in 43 patients.

PURPOSE: High-resolution magnetic resonance imaging (MRI) plays a crucial role in the presurgical evaluation of patients with medically refractory partial epilepsy. Although MRI detects a morphologic abnormality as the cause of the epilepsy in the majority of patients, some patients have a normal MRI. This study was undertaken to explore the hypothesis that in patients with normal MRI, invasive monitoring can lead to localization of the seizure-onset zone and successful epilepsy surgery. METHODS: A series of 115 patients with partial epilepsy who had undergone intracranial electrode evaluation (subdural strip, subdural grid, and/or depth electrodes) between February 1992 and February 1999 was analyzed retrospectively. Of these, 43 patients (37%) had a normal MRI. RESULTS: Invasive monitoring detected a focal seizure onset in 25 (58%) patients, multifocal seizure origin in 12 (28%) patients, and in six patients, no focal seizure origin was found. Of the 25 patients with a focal seizure origin, cortical resection was performed in 24, of whom 20 (83%) had a good surgical outcome with respect to seizure control. Six of the 12 patients with multifocal seizure origin underwent other forms of epilepsy surgery (palliative cortical resection in two, anterior callosotomy in two, and vagal nerve stimulator placement in two). CONCLUSIONS: Successful epilepsy surgery is possible in patients with normal MRIs, but appropriate presurgical evaluations are necessary. In patients with evidence of multifocal seizure origin during noninvasive evaluation, invasive monitoring should generally be avoided.

Improved plasmid vectors for the production of multiple fluorescent protein fusions in Bacillus subtilis.

The intrinsically fluorescent green fluorescent protein has been used in many laboratories as a cytological marker to monitor protein localisation in live cells. Multiple spectrally modified mutant versions and novel fluorescent proteins from other species have subsequently been reported and used for labelling cells with multiple fluorescent protein fusions. In this work we report the design and use of vectors containing some of these spectral variants of GFP for use in the Gram positive bacterium Bacillus subtilis. These vectors complement those previously described (Lewis and Marston, 1999. Gene 227, 101-109) to provide a large suite of plasmid vectors for use in this and other related Gram positive organisms. Using these vectors we have been able to directly demonstrate the sequential assembly/disassembly of proteins involved in the generation of cellular asymmetry during development.

Investigation of extra-temporal epilepsy.

Medically intractable epilepsy of extra-temporal origin can represent a difficult therapeutic challenge. Our Epilepsy Service has managed these patients using standard investigative methods as well as ictal SPECT and intracranial electrode recording. In the present series of patients, image-guided surgery was used for all electrode implantation and resective surgery. Seizure localization and successful resection were achieved in 70-80% of 42 patients with follow-up of at least one year. Normal MRI and previous failed intracranial investigation were not associated with poorer outcome.

Does performing image registration and subtraction in ictal brain SPECT help localize neocortical seizures?

UNLABELLED: Ictal brain SPECT (IS) findings in neocortical epilepsy (patients without mesiotemporal sclerosis) can be subtle. This study is aimed at assessing how the seizure focus identification was improved by the inclusion of individual IS and interictal brain SPECT (ITS)-MRI image registration as well as performing IS - ITS image subtraction. METHODS: The study involved the posthoc analysis of 64 IS scans using 99mTc-ethyl cysteinate dimer that were obtained in 38 patients without mesiotemporal sclerosis but with or without other abnormalities on MRI. Radiotracer injection occurred during video-electroencephalographic (EEG) monitoring. Patients were injected 2-80 s (median time, 13 s) after clinical or EEG seizure onset. All patients had sufficient follow-up to correlate findings with the SPECT results. All patients had ITS and MRI, including a coronal volume sequence used for registration. Image registration (IS and ITS to MRI) was performed using automated software. After normalization, IS - ITS subtraction was performed. The IS, ITS, and subtraction studies were read by 2 experienced observers who were unaware of the clinical data and who assessed the presence and localization of an identifiable seizure focus before and after image registration and subtraction. Correlation was made with video-EEG (surface and invasive) and clinical and surgical follow-up. RESULTS: Probable or definite foci were identified in 38 (59%) studies in 33 (87%) patients. In 52% of the studies, the image registration aided localization, and in 58% the subtraction images contributed additional information. In 9%, the subtraction images confused the interpretation. In follow-up after surgery, intracranial EEG or video-EEG monitoring (or both) has confirmed close or reasonable localization in 28 (74%) patients. In 6 (16%) patients, SPECT indicated false seizure localization. CONCLUSION: Image registration and image subtraction improve the localization of neocortical seizure foci using IS, but close correlation with the original images is required. False localizations occur in a minority of patients.

Dynamic relocalization of phage phi 29 DNA during replication and the role of the viral protein p16.7.

We have examined the localization of DNA replication of the Bacillus subtilis phage phi 29 by immunofluorescence. To determine where phage replication was localized within infected cells, we examined the distribution of phage replication proteins and the sites of incorporation of nucleotide analogues into phage DNA. On initiation of replication, the phage DNA localized to a single focus within the cell, nearly always towards one end of the host cell nucleoid. At later stages of the infection cycle, phage replication was found to have redistributed to multiple sites around the periphery of the nucleoid, just under the cell membrane. Towards the end of the cycle, phage DNA was once again redistributed to become located within the bulk of the nucleoid. Efficient redistribution of replicating phage DNA from the initial replication site to various sites surrounding the nucleoid was found to be dependent on the phage protein p16.7.

What progress has been made in meeting the needs of seriously maltreated children? The course of 200 cases through the Boston Juvenile Court.

OBJECTIVES: The study examined child, parent, and case characteristics in a sample of 200 cases of serious child maltreatment brought before the Boston Juvenile Court (BJC) on Care and Protection petitions in 1994. Whether recent changes in Massachusetts law have been effective in reducing delays in adjudication and helping children achieve permanent placements more quickly was also examined. METHOD: Data were abstracted from court records by the research team. The 200 cases were followed prospectively for 4 years. Retrospective data on the families' previous involvement with the protective service system were also abstracted from the records. Data from the 1994 cases were compared to that obtained from a sample of cases brought before the BJC in 1985-1986. RESULTS: Children permanently removed from parental custody in the 1994 sample required less time post-disposition to achieve permanent placements. However, overall, time frames for the 1994 cases remained remarkably similar to those in 1985-1986: children were in the protective service system an average of 5 years; cases required an average of 1.6 years in court; and half of the children permanently removed from parental custody were still in "temporary" foster care at 4-year follow-up. CONCLUSIONS: Although some improvements have occurred since 1985-1986, the system still fails to meet the needs of seriously maltreated children to achieve permanent placements promptly. The implications of the findings for system reform are discussed.

Compartmentalization of transcription and translation in Bacillus subtilis.

Using fusions of green fluorescent protein to subunits of RNA polymerase (RNAP) and ribosomes, we have investigated the subcellular localization of the transcriptional and translational machinery in the bacterium Bacillus subtilis. Unexpectedly, we found that RNAP resides principally within the nucleoid. Conversely, ribosomes localized almost exclusively outside the nucleoid, concentrating particularly towards sites of cell division. This zonal localization was not dependent on cell division and is probably due, at least in part, to exclusion from the nucleoid. Dual labelling of RNAP and ribosomes was used to confirm the spatial separation of the two processes. We conclude that, even in the absence of a nuclear membrane, transcription and translation occur predominantly in separate functional domains. At higher growth rates, concentrations of RNAP developed, probably representing the sites of rRNA synthesis. These may represent a further spatial specialization, possibly equivalent to the eukaryotic nucleolus.

Induction of immune responses to bovine herpesvirus type 1 gD in passively immune mice after immunization with a DNA-based vaccine.

The potential for plasmids encoding a secreted form of bovine herpesvirus type 1 (BHV-1) glycoprotein D (gD) to elicit immune responses in passively immune mice following intramuscular immunization was investigated. In these experiments, 6- to 8-week-old female C3H/HeN or C57BL/6 mice were passively immunized with hyperimmune antisera raised against BHV-1 recombinant, truncated (secreted) gD immediately prior to immunization with plasmids. A single immunization of passively immune mice with plasmid encoding the secreted form of BHV-1 gD resulted in rapid development of both cell-mediated immunity and antibody responses. Furthermore, 50% of mice immunized with a suboptimal dose of recombinant gD formulated into an adjuvant developed significant levels of serum antibodies if mice were pre-treated with hyperimmune antisera. The apparent failure of passive polyclonal antisera to suppress the induction of immune responses to pSLRSV may be related to the immunoglobulin subtypes present in the hyperimmune sera.

Altering the cellular location of an antigen expressed by a DNA-based vaccine modulates the immune response.

The potential for DNA vaccines encoding mutated versions of the same antigen to modulate immune responses in C3H/HeN mice was investigated. We created expression plasmids that encoded several versions of glycoprotein D (gD) from bovine herpesvirus 1, including authentic membrane-anchored glycoprotein (pSLRSV.AgD), a secreted glycoprotein (pSLRSV.SgD), and an intracellular protein (pSLRSV.CgD). Immunization of an inbred strain of mice with these plasmids resulted in highly efficacious and long-lasting humoral and cell-mediated immunity. We also demonstrated that the cell compartment in which plasmid-encoded gD was expressed caused a deviation in the serum immunoglobulin (Ig) isotype profile as well as the predominant cytokines secreted from the draining lymph node. Immunization of C3H/HeN mice with DNA vaccines encoding cell-associated forms of gD resulted in a predominance of serum IgG2a and gamma interferon-secreting cells within the spleens and draining lymph nodes. In contrast, mice immunized with a secreted form of this same antigen displayed immune responses characterized by greater levels of interleukin 4 in the draining lymph node and IgG1 as the predominant serum isotype. We also showed evidence of compartmentalization of distinct immune responses within different lymphoid organs.

DNA vaccines: a review.

Therapeutic and prophylactic DNA vaccine clinical trials for a variety of pathogens and cancers are underway (Chattergoon et al., 1997; Taubes, 1997). The speed with which initiation of these trials occurred is no less than astounding; clinical trials for a human immunodeficiency virus (HIV) gp160 DNA-based vaccine were underway within 36 months of the first description of "genetic immunization" (Tang et al., 1992) and within 24 months of publication of the first article describing intramuscular delivery of a DNA vaccine (Ulmer et al., 1993). Despite the relative fervor with which clinical trials have progressed, it can be safely stated that DNA-based vaccines will not be an immunological "silver bullet." In this regard, it was satisfying to see a publication entitled "DNA Vaccines--A Modern Gimmick or a Boon to Vaccinology?" (Manickan et al., 1997b). There is no doubt that this technology is well beyond the phenomenology phase of study. Research niches and models have been established and will allow the truly difficult questions of mechanism and application to target species to be studied. These two aspects of future studies are intricately interwoven and will ultimately determine the necessity for mechanistic understanding and the evolution of target species studies. The basic science of DNA vaccines has yet to be clearly defined and will ultimately determine the success or failure of this technology to find a place in the immunological arsenal against disease. In a commentary on a published study describing DNA vaccine-mediated protection against heterologous challenge with HIV-1 in chimpanzees, Ronald Kennedy (1997) states, "As someone who has been in the trenches of AIDS vaccine research for over a decade and who, together with collaborators, has attempted a number of different vaccine approaches that have not panned out, I have a relatively pessimistic view of new AIDS vaccine approaches." Kennedy then goes on to summarize a DNA-based multigene vaccine approach and the subsequent development of neutralizing titers and potent CTL activity in immunized chimpanzees (Boyer et al., 1997). Dr. Kennedy closes his commentary by stating. "The most exciting aspect of this report is the experimental challenge studies.... Viraemia was extremely transient and present at low levels during a single time point. These animals remained seronegative ... for one year after challenge" and "Overall, these observations engender some excitement". (Kennedy, 1997). Although this may seem a less than rousing cheer for DNA vaccine technology, it is a refreshingly hopeful outlook for a pathogen to which experience has taught humility. It has also been suggested that DNA vaccine technology may find its true worth as a novel alternative option for the development of vaccines against diseases that conventional vaccines have been unsuccessful in controlling (Manickan et al., 1997b). This is a difficult task for any vaccine, let alone a novel technology. DNA-based vaccine technology represents a powerful and novel entry into the field of immunological control of disease. The spinoff research has also been dramatic, and includes the rediscovery of potent bacterially derived immunomodulatory DNA sequences (Gilkeson et al., 1989), as well as availability of a methodology that allows extremely rapid assessment and dissection of both antigens and immunity. The benefits of potent Th1-type immune responses to DNA vaccines must not be overlooked, particularly in the light of suggestions that Western culture immunization practices may be responsible for the rapid increases in adult allergic and possibly autoimmune disorders (Rook and Stanford, 1998). The full utility of this technology has not yet been realized, and yet its broad potential is clearly evident. Future investigations of this technology must not be hindered by impatience, misunderstanding, and lack of funding or failure of an informed collective and collaborative effort.