Impact of woodland agri-environment management on woodland structure and target bird species.

Agri-environment incentives form a central mechanism supporting changes to land management to provide public benefits. This study assesses the medium-term effects of woodland management on 13 target, specialist woodland bird species, as well as other woodland birds in a single region of the UK. The abundance of breeding birds (using two methods: point counts and territory mapping) and metrics of woodland structure were recorded on sites with Woodland Improvement Grants (improvement sites) and nearby comparison sites (control sites). Initial measurements were made prior to management and repeated 7-9 years later. A separate comparison of changes in bird abundance was made between the managed woodland sites and woodland from similar landscapes surveyed as part of the national Breeding Bird Survey (BBS). There was an increase in abundance of target species on improvement sites compared to a decrease on both control sites and BBS sites, although this was only evident from point count data. The effects on target species were stronger than for other woodland specialists and there was no apparent effect on woodland generalists, suggesting that the management interventions were appropriate for the target species. Changes in woodland structure were generally consistent with the expected effect of management, with lower tree density and greater Bramble (Rubus sp.) cover. However, contrary to the aim of increasing understorey cover, a reduction was recorded within the 2-10 m height category in improvement sites. This contrast is due to the removal of young trees during thinning affecting this height band and the short time period since management to allow regrowth. Our findings show that bespoke management supported through government agri-environment incentives can have a positive impact on target woodland birds. For managed forests, identifying species requirements and how management can be adapted to improve their habitats can be an effective way of delivering biodiversity gains when financial incentives are provided to achieve policy goals.

Practicing virology: making and knowing a mid-twentieth century experiment with Tobacco mosaic virus.

Tobacco mosaic virus (TMV) has served as a model organism for pathbreaking work in plant pathology, virology, biochemistry and applied genetics for more than a century. We were intrigued by a photograph published in Phytopathology in 1934 showing that Tabasco pepper plants responded to TMV infection with localized necrotic lesions, followed by abscission of the inoculated leaves. This dramatic outcome of a biological response to infection observed by Francis O. Holmes, a virologist at the Rockefeller Institute for Medical Research, was used to score plants for resistance to TMV infection. Our objective was to gain a better understanding of early to mid-twentieth century ideas of genetic resistance to viruses in crop plants. We investigated Holmes' observation as a practical exercise in reworking an experiment, having been inspired by Pamela Smith's innovative Making and Knowing Project. We had a great deal of difficulty replicating Holmes' experiment, finding that biological materials and experimental customs change over time, in ways that ideas do not. Using complementary tools plus careful study and interpretation of the original text and figures, we were able to rework, yet only partially replicate, this experiment. Reading peer-reviewed manuscripts that cited Holmes' 1934 report provided an additional level of insight into the interpretation and replication of this work in the decades that followed. From this, we touch on how experimental reworking can inform our strategies to address the reproducibility "crisis" in twenty-first century science.

Native Processing of Single Guide RNA Transcripts to Create Catalytic Cas9/Single Guide RNA Complexes in Planta.

The present CRISPR/Cas9 gene editing dogma for single guide RNA (sgRNA) delivery is based on the premise that 5'-and 3'-nucleotide overhangs negate Cas9/sgRNA catalytic activity in vivo. This has led to engineering strategies designed to either avoid or remove extraneous nucleotides at the 5' and 3' termini of sgRNAs. Previously, we used a Tobacco mosaic virus viral vector to express both GFP and a sgRNA from a single virus-derived mRNA in Nicotiana benthamiana This vector yielded high levels of GFP and catalytically active sgRNAs. Here, in an effort to understand the biochemical interactions of this result, we used in vitro assays to demonstrate that nucleotide overhangs 5', but not 3', proximal to the sgRNA do in fact inactivate Cas9 catalytic activity at the specified target site. Next we showed that in planta sgRNAs bound to Cas9 are devoid of the expected 5' overhangs transcribed by the virus. Furthermore, when a plant nuclear promoter was used for expression of the GFP-sgRNA fusion transcript, it also produced indels when delivered with Cas9. These results reveal that 5' auto-processing of progenitor sgRNAs occurs natively in plants. Toward a possible mechanism for the perceived auto-processing, we found, using in vitro-generated RNAs and those isolated from plants, that the 5' to 3' exoribonuclease XRN1 can degrade elongated progenitor sgRNAs, whereas the mature sgRNA end products are resistant. Comparisons with other studies suggest that sgRNA auto-processing may be a phenomenon not unique to plants, but present in other eukaryotes as well.

Multiplexed Gene Editing and Protein Overexpression Using a Tobacco mosaic virus Viral Vector.

Development of CRISPR/Cas9 transient gene editing screening tools in plant biology has been hindered by difficulty of delivering high quantities of biologically active single guide RNAs (sgRNAs). Furthermore, it has been largely accepted that in vivo generated sgRNAs need to be devoid of extraneous nucleotides, which has limited sgRNA expression by delivery vectors. Here, we increased cellular concentrations of sgRNA by transiently delivering sgRNAs using a Tobacco mosaic virus-derived vector (TRBO) designed with 5' and 3' sgRNA proximal nucleotide-processing capabilities. To demonstrate proof-of-principle, we used the TRBO-sgRNA delivery platform to target GFP in Nicotiana benthamiana (16c) plants, and gene editing was accompanied by loss of GFP expression. Surprisingly, indel (insertions and deletions) percentages averaged nearly 70% within 7 d postinoculation using the TRBO-sgRNA constructs, which retained 5' nucleotide overhangs. In contrast, and in accordance with current models, in vitro Cas9 cleavage assays only edited DNA when 5' sgRNA nucleotide overhangs were removed, suggesting a novel processing mechanism is occurring in planta. Since the Cas9/TRBO-sgRNA platform demonstrated sgRNA flexibility, we targeted the N. benthamiana NbAGO1 paralogs with one sgRNA and also multiplexed two sgRNAs using a single TRBO construct, resulting in indels in three genes. TRBO-mediated expression of an RNA transcript consisting of an sgRNA adjoining a GFP protein coding region produced indels and viral-based GFP overexpression. In conclusion, multiplexed delivery of sgRNAs using the TRBO system offers flexibility for gene expression and editing and uncovered novel aspects of CRISPR/Cas9 biology.

The management of infected elbow arthroplasty by two-stage revision.

BACKGROUND: Deep prosthetic infection is a potentially devastating complication after total elbow arthroplasty, with an incidence of up to 12%. This study examined the demographics, microbiologic profile, and outcomes of infected total elbow arthroplasty treated with 2-stage revision in a tertiary referral unit. METHODS: We identified 19 consecutive patients (mean age, 65 years) undergoing revision arthroplasty for deep prosthetic infection. All patients underwent a first-stage procedure with removal of implants, débridement, and insertion of an antibiotic-loaded cement spacer, followed by at least 6 weeks of intravenous antibiotics. Fourteen patients required a second-stage revision. RESULTS: Five patients did not undergo a second-stage procedure because of patient choice (n = 2), medical or surgical risk factors (n = 2), and death from an unrelated cause (n = 1). Of the 19 patients undergoing a first-stage procedure, 16 (84%) remained infection free, and 11 of the 14 patients (79%) undergoing reimplantation of an elbow prosthesis remained infection free. Six patients required further surgery (3 for recurrent infection, 3 for noninfective indications). The commonest infecting organism was Staphylococcus aureus (47%). A degree of postoperative ulnar nerve dysfunction occurred in 37% of patients, but all resolved fully without further treatment. CONCLUSIONS: Management of prosthetic joint infection using 2-stage revision can result in high rates of eradication, although rates of reoperation and transient ulnar nerve dysfunction are high.

Shoulder linked arthroplasty in patients with obstetric brachial plexus palsy can improve quality of life and function at short-term follow-up.

BACKGROUND: Patients with obstetric brachial plexus palsy (OBPP) are prone to develop degenerative shoulder disease at a younger age than the general population. To date, no reports have been published on the complexities or outcome of shoulder arthroplasty (SA) in this unique patient group. METHODS: We reviewed of 9 SAs in 9 patients (3 men and 6 women) with OBPP with mean follow-up 5.1 years (range, 2.6-7.6 years). Patients were a mean age of 29 years (range, 16-56 years). Patients had undergone a mean of 3 previous operations (range, 2-6). All patients underwent linked constrained SA. RESULTS: The mean Oxford Shoulder Score increased from 8 (range, 3-10) preoperatively to 21 (range, 12-32) at the final follow-up (P < .001) predominantly due to pain relief. Mean range of active forward elevation and abduction improved from 35° and 39° to 46° and 45°, respectively. Patients improved significantly in 2 of 8 Short-Form 36-Item health-related quality of life domains, bodily pain (P = .013) and mental health (P = .035), and the overall physical component summary score (P = .006). Range of motion had mild improvements. Three required reoperation (33%), comprising 1 excision of heterotopic ossification, 1 trimming of a prominent screw, and 1 deltoid rupture repair. CONCLUSIONS: SA is effective at relieving pain and health-related quality of life for young patients with OBPP; however, compared with the general population, the complication rate is high and functional gains are small.

Clinical and imaging findings in children with non-syndromic lambdoid synostosis.

UNLABELLED: True unilateral lambdoid synostosis is a very rare type of craniosynostosis. Patients present with unilateral posterior plagiocephaly. The differentiation between frequent, simple positional (deformational) posterior plagiocephaly and lambdoid synostosis is not easy and to date subject of controversy. Accurate and early diagnosis is important, because treatment is different. Simple positional plagiocephaly responds to conservative treatment, but craniosynostosis may require neurosurgical intervention. We analyzed clinical presentation of five patients in whom non-syndromic lambdoid synostosis was confirmed by CT imaging, in one additionally by high-resolution ultrasound, and finally neurosurgical intervention. However, clinical inspection alone did not reliably identify infants with lambdoid synostosis. CONCLUSION: Diagnosis of lambdoid synostosis is not always possible based on clinical inspection alone. To confirm the diagnosis imaging is recommended. There is evidence that high-resolution ultrasound can be used first-line in the future.

[Psychosocial integration of inpatients with borderline personality disorder: reflection of a health care system focused on symptom remission]. / Psychosoziale Integration von stationären Patienten mit Borderline-Persönlichkeitsstörung : Abbild eines auf Symptomremission fokussierten Therapie- und Versorgungssystems.

BACKGROUND: Borderline personality disorder (BPD) is characterized by a profound instability of identity, by stress and affect dysregulation and impulsivity, and by cognitive-perceptual disturbances. These deficits seriously impair interpersonal experiences and social interaction. The typical impulsive style and psychosocial malfunctioning of these patients lead to grave disturbances in their everyday areas of life with numerous break-ups in all relevant areas of life and entail inadequate vocational placement. Despite vast demands on the health care system, BPD attracts little attention of care process research and reintegration measures as, for instance, exist for schizophrenia. At the LVR-Klinikum Düsseldorf as representative of a large clinic in North Rhein-Westphalia, we analysed socio-demographic characteristics of BPD inpatients. The aims of the study were to substantiate the high impact of the disease on patient's social and vocational integration and to sensitize for significant aspects of disease-specific therapy and care. METHODS: A retrospective analysis of DGPPN-BADO of all patients (n = 9,425) who were admitted between January 2004 and December 2009 to the LVR-Klinikum Düsseldorf, department of psychiatry and psychotherapy, Heinrich Heine University Hospital, Germany. Data analysis involved the divisions general psychiatry and addictive disorders and covered 999 patients with BPD (ICD-10: F60.3) and 1,937 patients with schizophrenia (ICD-10: F20). The defined characteristics of DGPPN-BADO that depict psychosocial integration were "living situation at admission", "highest academic/professional degree achieved", "working situation" und "livelihood". Besides descriptive statistics of the elected variables, comparative analysis of the diagnoses BPD and schizophrenia involved calculating group differences after adjusting the groups for sex and age. Statistical analysis was performed by the χ2-test. RESULTS: The comparative analysis of psychosocially relevant BADO characteristics reveals a significantly stronger impairment of patients with BPD concerning their academic and professional qualification than patients with schizophrenia. Nevertheless they more often hold a job or cover their living expenses without subsidy, respectively. CONCLUSION: The data suggest a lack of general awareness in the significance of social and vocational disturbances in BPD. The interactional deficits of patients with BPD affect all areas of life but their illness-related nature seems to be disregarded in relevant life areas, such as educational processes and the vocational situation. Furthermore, the impact of BPD on earning capacity is not given adequate consideration in the assessment of pensions or subsidies. Consistent with the recent literature, our socio-demographic data indicate significant deficits of the present health system in handling BPD.

Modular, programmable RNA sensing using ADAR editing in living cells.

With the increasing availability of single-cell transcriptomes, RNA signatures offer a promising basis for targeting living cells. Molecular RNA sensors would enable the study of and therapeutic interventions for specific cell types/states in diverse contexts, particularly in human patients and non-model organisms. Here we describe a modular, programmable system for live RNA sensing using adenosine deaminases acting on RNA (RADAR). We validate, and then expand, our basic design, characterize its performance, and analyze its compatibility with human and mouse transcriptomes. We identify strategies to boost output levels and improve the dynamic range. Additionally, we show that RADAR enables compact AND logic. In addition to responding to transcript levels, RADAR can distinguish disease-relevant sequence alterations of transcript identities, such as point mutations and fusions. Finally, we demonstrate that RADAR is a self-contained system with the potential to function in diverse organisms.

Rationale and study design of the MINERVA study: Multicentre Investigation of Novel Electrocardiogram Risk markers in Ventricular Arrhythmia prediction-UK multicentre collaboration.

INTRODUCTION: The purpose of this study is to assess the ability of two new ECG markers (Regional Repolarisation Instability Index (R2I2) and Peak Electrical Restitution Slope) to predict sudden cardiac death (SCD) or ventricular arrhythmia (VA) events in patients with ischaemic cardiomyopathy undergoing implantation of an implantable cardioverter defibrillator for primary prevention indication. METHODS AND ANALYSIS: Multicentre Investigation of Novel Electrocardiogram Risk markers in Ventricular Arrhythmia prediction is a prospective, open label, single blinded, multicentre observational study to establish the efficacy of two ECG biomarkers in predicting VA risk. 440 participants with ischaemic cardiomyopathy undergoing routine first time implantable cardioverter-defibrillator (ICD) implantation for primary prevention indication are currently being recruited. An electrophysiological (EP) study is performed using a non-invasive programmed electrical stimulation protocol via the implanted device. All participants will undergo the EP study hence no randomisation is required. Participants will be followed up over a minimum of 18 months and up to 3 years. The first patient was recruited in August 2016 and the study will be completed at the final participant follow-up visit. The primary endpoint is ventricular fibrillation or sustained ventricular tachycardia >200 beats/min as recorded by the ICD. The secondary endpoint is SCD. Analysis of the ECG data obtained during the EP study will be performed by the core lab where blinding of patient health status and endpoints will be maintained. ETHICS AND DISSEMINATION: Ethical approval has been granted by Research Ethics Committees Northern Ireland (reference no. 16/NI/0069). The results will inform the design of a definitive Randomised Controlled Trial (RCT). Dissemination will include peer reviewed journal articles reporting the qualitative and quantitative results, as well as presentations at conferences and lay summaries. TRIAL REGISTRATION NUMBER: NCT03022487.

Predictive factors for neurocognitive function and Quality of Life after surgical treatment for Cushing's disease and acromegaly.

BACKGROUND: Cushing's disease (CD) and acromegaly (AC) are associated with impairment in quality of life (QoL) and neurocognition that can persist after successful treatment. AIM: To investigate the influence of current disease status (remission vs no remission) on neurocognitive function and QoL in treated CD and AC patients and to determine predictive factors (e.g. demographic, clinical, neurosurgical, endocrinological) for post-operative neurocognition and QoL. SUBJECTS AND METHODS: Twenty-four CD and 37 AC patients underwent neuropsychological testing 1 to 10 yr following surgical therapy. Additionally, QoL was assessed. An overnight 2-mg dexamethasone suppression test in CD and IGF-I and GH levels in AC patients were assessed to determine current disease status. The results were compared with 28 sex-, education- and age-matched healthy controls (HC). RESULTS: Impaired QoL was more pronounced than neurocognitive decrease in both pathologies compared to HC. This finding was independent of the current status of disease. In AC, persistent comorbidities were associated with impaired QoL (p<0.05). Older age at operation in AC patients was a significant predictor for adverse effects on psychomotor speed and attentional functions (p<0.05). In CD persistent hypocortisolism, not hypercortisolism, had adverse effects on neurocognition (p<0.01). CONCLUSIONS: The current status of disease plays a subordinate role in postoperative outcome concerning QoL and neurocognition in either pathology. A possible explanation might be the considerably improved endocrinopathy after treatment compared to untreated patients, even if no cure is achieved. The lasting impairments might be explained by irreversible changes that have occurred during the active phase of the disease.

[Neonatal intracerebral bleeding as initial symptom of gliomatosis cerebri WHO III--neurological outcome after partial hemispherectomy]. / Eine neonatale intrazerebrale Blutung als Initialsymptom einer Gliomatosis cerebri WHO III--neurologisches Outcome nach partieller Hemisphärektomie.

INTRODUCTION: Anaplastic astrocytomas presenting as gliomatosis cerebri in neonates are extremely rare. Tumours in newborns are mostly of neuroectodermal origin. CASE REPORT: We report about a female newborn at term [birth weight 3 600 g (P 90), head circumference 35 cm (P 95) APGAR 9/10/10] with an intracerebral partially clotted bleeding in the left parieto-occipital region. The bleeding was expansive leading to axial and lateral cerebral herniation. The intracerebral bleeding in the left occipital region was surgically removed. Macroscopically no solid tumour was seen, but small fragments of an anaplastic astrocytic tumour (WHO grade III) were diagnosed histologically. After surgery, no remaining tumour was visible in the MRI. 6 weeks later, a recurrent tumour (4×4 cm) was found in the area of the initial bleeding. Further treatment was initially refused by the parents. The child was readmitted to our hospital at the age of 11 months in good clinical condition and presented with left-sided hemiparesis, right-sided hemianopsia and intermittent strabismus convergens alternans. Because of the good clinical condition further therapeutic treatment was initiated. Due to the final extension of the tumour into the temporal, parietal and occipital lobes, a gliomatosis cerebri WHO III was diagnosed. An extended partial hemispherectomy was done. After neurosurgery, no further neurological failures occurred. In the follow-up examination, MRI showed no relapse of the tumour. Chemotherapy according to the HIT SKK protocol was initiated. A relapse did not occur over a follow-up of 2 years. CONCLUSION: This is a rare case report of a congenital gliomatosis cerebri WHO grade III, treated with partial hemispherectomy, leading to a good clinical and neurological long-term outcome.

RNA silencing suppressor-influenced performance of a virus vector delivering both guide RNA and Cas9 for CRISPR gene editing.

We report on further development of the agroinfiltratable Tobacco mosaic virus (TMV)-based overexpression (TRBO) vector to deliver CRISPR/Cas9 components into plants. First, production of a Cas9 (HcoCas9) protein from a binary plasmid increased when co-expressed in presence of suppressors of gene silencing, such as the TMV 126-kDa replicase or the Tomato bushy stunt virus P19 protein. Such suppressor-generated elevated levels of Cas9 expression translated to efficient gene editing mediated by TRBO-G-3'gGFP expressing GFP and also a single guide RNA targeting the mgfp5 gene in the Nicotiana benthamiana GFP-expressing line 16c. Furthermore, HcoCas9 encoding RNA, a large cargo insert of 4.2 kb, was expressed from TRBO-HcoCas9 to yield Cas9 protein again at higher levels upon co-expression with P19. Likewise, co-delivery of TRBO-HcoCas9 and TRBO-G-3'gGFP in the presence of P19 also resulted in elevated levels percentages of indels (insertions and deletions). These data also revealed an age-related phenomenon in plants whereby the RNA suppressor P19 had more of an effect in older plants. Lastly, we used a single TRBO vector to express both Cas9 and a sgRNA. Taken together, we suggest that viral RNA suppressors could be used for further optimization of single viral vector delivery of CRISPR gene editing parts.

Reported Technical Aspects of Type II SLAP Lesion Repairs in Athletes.

PURPOSE: To systematically review the available literature to further describe and report the available data on SLAP repair techniques and the association with outcomes. METHODS: A systematic review of literature was performed on manuscripts describing type II SLAP repairs in athletes. Selection criteria included studies reporting exclusively type II SLAP tears without concomitant pathology, minimum 2-year postoperative follow-up, use of anchor fixation, and return to previous level of play data available. We extracted patient outcome as well as surgical construct details from each article. Average outcomes and return to play rates were calculated and substratified further by athlete type. Return to play rates were compared by repair constructs with the Student t test. RESULTS: Initial search resulted in 107 articles. After exclusion criteria were applied, 17 articles were included in the final analysis. Overall, 84% of patients had good-to-excellent results. Of all athletes, 66% returned to previous level of play. There was significant variation in reported technique in terms of anchor number, location, material, suture type, and knotless versus knotted constructs. No significant difference was reported in outcomes in comparison of suture type (P-value .96) or knotted versus knotless constructs (P-value .91). Given the significant variability in reporting, no statistical analysis was felt able to be performed on anchor location and number. CONCLUSIONS: Repair of type II SLAP tears in athletes is a difficult problem to treat with overall low return to play despite a high rate of "good" outcomes when assessed by outcome measures. Significant variability exists in surgical technique, as well as reporting of surgical technique, potentially limiting the ability to define the best or most effective technique for SLAP repair. LEVEL OF EVIDENCE: IV, systematic review of level III and level IV studies.

Hydroxychloroquine as an aerosol might markedly reduce and even prevent severe clinical symptoms after SARS-CoV-2 infection.

Covid-19 is a new coronavirus disease first described in December 2019. This respiratory illness is severe and potentially fatal. Severe cases make up to 15%, lethality ranges between 1.5 and more than 10%. What is urgently needed is an efficient pharmacological treatment for the treatment of severe cases. During the infection of alveolar epithelial cells of the lung, the ACE2 receptor has a central function. The antimalarial drugs chloroquine phosphate (CQ) and hydroxychloroquine (HCQ) impair in vitro the terminal glycosylation of ACE2 without significant change of cell-surface ACE2 and, therefore, might be potent inhibitors of SARS-CoV-2 infections. Starting inhibition at 0.1 µM, CQ completely prevented in vitro infections at 10 µM, suggesting a prophylactic effect and preventing the virus spread 5 h after infection. In a first clinical trial, CQ was effective in inhibiting exacerbation of pneumonia, improving lung imaging findings, promotion of virus-negative conversion, and shortening the disease. In addition, HCQ, which is three times more potent than CQ in SARS-CoV-2 infected cells (EC50 0.72 µM), was significantly associated with viral load reduction/disappearance in COVID-19 patients compared to controls. Theoretically, CQ and HCQ could thus be effectively used in the treatment of SARS-CoV pneumonia. From a pharmacological standpoint, however, the major problems of oral treatment with these drugs are possible severe side effects and toxicity. Concretely, this relates to (a) the inconsistent individual bioavailability of these drugs at the alveolar target cells, depending on intestinal resorption, hepatic first-pass metabolism and accumulation in liver, spleen and lung, and (b) the need for a relatively high concentration of 1-5 µM at the alveolar surface. Therefore, we propose in a first dose estimation the use of HCQ as an aerosol in a dosage of 2-4 mg per inhalation in order to reach sufficient therapeutic levels at the alveolar epithelial cells. By using a low-dose non-systemic aerosol, adverse drug reactions will markedly be reduced compared with oral application. This increase in tolerability enables a broader use for prevention and after contact with an infected person, which would be an advantage especially for the high-risk, often multi-morbid and elderly patients. Empirical data on self-medication with a one-week aerosol application by two of the authors is presented. Inhalation was well tolerated without relevant side effects.

Removal of the membrane-anchoring domain of epidermal growth factor leads to intracrine signaling and disruption of mammary epithelial cell organization.

Autocrine EGF-receptor (EGFR) ligands are normally made as membrane-anchored precursors that are proteolytically processed to yield mature, soluble peptides. To explore the function of the membrane-anchoring domain of EGF, we expressed artificial EGF genes either with or without this structure in human mammary epithelial cells (HMEC). These cells require activation of the EGFR for cell proliferation. We found that HMEC expressing high levels of membrane- anchored EGF grew at a maximal rate that was not increased by exogenous EGF, but could be inhibited by anti-EGFR antibodies. In contrast, when cells expressed EGF lacking the membrane-anchoring domain (sEGF), their proliferation rate, growth at clonal densities, and receptor substrate phosphorylation were not affected by anti-EGFR antibodies. The sEGF was found to be colocalized with the EGFR within small cytoplasmic vesicles. It thus appears that removal of the membrane-anchoring domain converts autocrine to intracrine signaling. Significantly, sEGF inhibited the organization of HMEC on Matrigel, suggesting that spatial restriction of EGF access to its receptor is necessary for organization. Our results indicate that an important role of the membrane-anchoring domain of EGFR ligands is to restrict the cellular compartments in which the receptor is activated.

Plant Virus Vectors 3.0: Transitioning into Synthetic Genomics.

Plant viruses were first implemented as heterologous gene expression vectors more than three decades ago. Since then, the methodology for their use has varied, but we propose it was the merging of technologies with virology tools, which occurred in three defined steps discussed here, that has driven viral vector applications to date. The first was the advent of molecular biology and reverse genetics, which enabled the cloning and manipulation of viral genomes to express genes of interest (vectors 1.0). The second stems from the discovery of RNA silencing and the development of high-throughput sequencing technologies that allowed the convenient and widespread use of virus-induced gene silencing (vectors 2.0). Here, we briefly review the events that led to these applications, but this treatise mainly concentrates on the emerging versatility of gene-editing tools, which has enabled the emergence of virus-delivered genetic queries for functional genomics and virology (vectors 3.0).

The fimbria-fornix/cingular bundle pathways: a review of neurochemical and behavioural approaches using lesions and transplantation techniques.

Extensive lesions of the fimbria-fornix pathways and the cingular bundle deprive the hippocampus of a substantial part of its cholinergic, noradrenergic and serotonergic afferents and, among several other behavioural alterations, induce lasting impairment of spatial learning and memory capabilities. After a brief presentation of the neuroanatomical organization of the hippocampus and the connections relevant to the topic of this article, studies which have contributed to characterize the neurochemical and behavioural aspects of the fimbria-fornix lesion "syndrome" with lesion techniques differing by the extent, the location or the specificity of the damage produced, are reviewed. Furthermore, several compensatory changes that may occur as a reaction to hippocampal denervation (sprouting changes in receptor sensitivity and modifications of neurotransmitter turnover in spared fibres) are described and discussed in relation with their capacity (or incapacity) to foster recovery from the lesion-induced deficits. According to this background, experiments using intrahippocampal or "parahippocampal" grafts to substitute for missing cholinergic, noradrenergic or serotonergic afferents are considered according to whether the reported findings concern neurochemical and/or behavioural effects. Taken together, these experiments suggest that appropriately chosen fetal neurons (or other cells such as for instance, genetically-modified fibroblasts) implanted into or close to the denervated hippocampus may substitute, at least partially, for missing hippocampal afferents with a neurochemical specificity that closely depends on the neurochemical identity of the grafted neurons. Thereby, such grafts are able not only to restore some functions as they can be detected locally, namely within the hippocampus, but also to attenuate some of the behavioural (and other types of) disturbances resulting from the lesions. In some respects, also these graft-induced behavioural effects might be considered as occurring with a neurochemically-defined specificity. Nevertheless, if a graft-induced recovery of neurochemical markers in the hippocampus seems to be a prerequisite for also behavioural recovery to be observed, this neurochemical recovery is neither the one and only condition for behavioural effects to be expressed, nor is it the one and only mechanism to account for the latter effects.

Decision framework for chemotherapeutic interventions for metastatic non-small-cell lung cancer.

BACKGROUND: Best supportive care has long been considered to be the standard therapy for metastatic non-small-cell lung cancer (NSCLC). There is now evidence from randomized trials that a number of chemotherapy regimens can palliate cancer-related symptoms and modestly improve survival. We show how cost-effectiveness analyses can be used to make choices between different (ambulatory) chemotherapy regimens. METHODS: Clinical algorithms describing the diagnosis, staging, and treatment of metastatic NSCLC were incorporated into Statistics Canada's Population Health Model. Using consistent methodology, we assessed the cost-effectiveness of several chemotherapeutic interventions: a combination of vindesine (VDS) plus cisplatin, etoposide (VP-16) plus cisplatin, vinblastine (VLB) plus cisplatin, vinorelbine (Navelbine; NVB) plus cisplatin, paclitaxel (Taxol) plus cisplatin, and gemcitabine (GEM) and NVB alone. We calculated the total chemotherapy costs in 1995 Canadian dollars, the cost per case, the average life-years saved, and the cost per life-year saved. Using the Population Health Model, we then constructed an advanced decision framework that rank-ordered the various treatment regimens so as to optimize benefit below various cost-effectiveness thresholds. RESULTS: One regimen (VLB plus cisplatin) appears to result in better survival and lower health care expenditures than best supportive care. By use of cost-effectiveness thresholds of $25,000 and $50,000 per life-year gained, NVB plus cisplatin is the preferred regimen. When quality of life is considered, however, GEM is preferred to NVB plus cisplatin at a threshold value of $50,000. At thresholds of $75 000 and $100,000, paclitaxel plus cisplatin at a dose of 135 mg/m(2) is the preferred regimen. At thresholds of $50,000 and above, best supportive care is the least preferred regimen. CONCLUSIONS: This decision framework allows the comparison of different treatment regimens based on various cost-effectiveness thresholds. Our analysis also supports the use of chemotherapy regimens and the abandonment of best supportive care as the standard of care for patients with advanced NSCLC. [J Natl Cancer Inst 2000;92:1321-9].