DNA and Gene Therapy: Uncoating of Polyoma Pseudovirus in Mouse Embryo Cells

Pseudovirions (host DNA fragments encapsidated by polyoma virus-coat protein) and polyoma virions are produced after infection of primary mouse embryo cells with polyoma virus. Purified pseudovirions are adsorbed to, and uncoated by, mouse embryo cells. The evidence for this uncoating is based on the conversion by pancreatic DNase of the uncoated pseudovirus DNA from an acid-insoluble to an acid-soluble form. The implications of these experiments to the eventual use of DNA for gene therapy are discussed.     

1 Versus 2-cm Excision Margins for pT2-pT4 Primary Cutaneous Melanoma (MelMarT): A Feasibility Study

Background

There is a lack of consensus regarding optimal surgical excision margins for primary cutaneous melanoma?>?1 mm in Breslow thickness (BT). A narrower surgical margin is expected to be associated with lower morbidity, improved quality of life (QoL), and reduced cost. We report the results of a pilot international study (MelMarT) comparing a 1 versus 2-cm surgical margin for patients with primary melanoma?>?1 mm in BT.

Methods

This phase III, multicentre trial [NCT02385214] administered by the Australia & New Zealand Medical Trials Group (ANZMTG 03.12) randomised patients with a primary cutaneous melanoma?>?1 mm in BT to a 1 versus 2-cm wide excision margin to be performed with sentinel lymph node biopsy. Surgical closure technique was at the discretion of the treating surgeon. Patients’ QoL was measured (FACT-M questionnaire) at baseline, 3, 6, and 12 months after randomisation.

Results

Between January 2015 and June 2016, 400 patients were randomised from 17 centres in 5 countries. A total of 377 patients were available for analysis. Primary melanomas were located on the trunk (56.9%), extremities (35.6%), and head and neck (7.4%). More patients in the 2-cm margin group required reconstruction (34.9 vs. 13.6%; p?<?0.0001). There was an increased wound necrosis rate in the 2-cm arm (0.5 vs. 3.6%; p?=?0.036). After 12 months’ follow-up, no differences were noted in QoL between groups.

Discussion

This pilot study demonstrates the feasibility of a large international RCT to provide a definitive answer to the optimal excision margin for patients with intermediate- to high-risk primary cutaneous melanoma.

     

Pre-Implantation Multiple Cytokine mRNA Expression Analysis of Donor Lung Grafts Predicts Survival After Lung Transplantation in Humans

While current donor selection with clinical findings is generally effective, the imprecise nature of the assessment forces clinicians to remain on the conservative side. A reliable biological marker would assist donor selection and would improve donor organ utilization. We collected biopsies from 169 donor lungs before implantation. Expression levels of IL-6, IL-8, IL-10, TNF-alpha, IFN-gamma and IL-1beta were measured by quantitative real-time RT-PCR (qRT-PCR). Seventeen cases died within 30 days after transplantation. No donor factor was significantly associated with 30-day mortality. Univariate analysis of the 84 cases for development of the prediction model showed that IL-6, IL-8, TNF-alpha and IL-1beta were risk factors for mortality and IL-10 and IFN-gamma were protective factors. We analyzed the cytokine expression ratios of risk to protective cytokines. A stepwise logistic regression for 30-day mortality demonstrated that a model containing the ratio of IL-6/IL-10 was the most predictive (p = 0.0013). When applied to the remaining 85 cases for validation, the test of model fit was significant (p = 0.039). Using the cytokine ratio, we were able to define three risk groups with striking differences in survival (p = 0.0003). Multi-cytokine analysis of the donor lung graft with qRT-PCR shows significant promise as a strategy to biologically evaluate the donor lung prior to implantation.     

γ-Aminobutyric acid (GABA) concentration inversely correlates with basal perfusion in human occipital lobe

Commonly used neuroimaging approaches in humans exploit hemodynamic or metabolic indicators of brain function. However, fundamental gaps remain in our ability to relate such hemo-metabolic reactivity to neurotransmission, with recent reports providing paradoxical information regarding the relationship among basal perfusion, functional imaging contrast, and neurotransmission in awake humans. Here, sequential magnetic resonance spectroscopy (MRS) measurements of the primary inhibitory neurotransmitter, γ-aminobutyric acid (GABA+macromolecules normalized by the complex N-acetyl aspartate-N-acetyl aspartyl glutamic acid: [GABA⁺]/[NAA–NAAG]), and magnetic resonance imaging (MRI) measurements of perfusion, fractional gray-matter volume, and arterial arrival time (AAT) are recorded in human visual cortex from a controlled cohort of young adult male volunteers with neurocognitive battery-confirmed comparable cognitive capacity (3 T; n=16; age=23±3 years). Regression analyses reveal an inverse correlation between [GABA⁺]/[NAA–NAAG] and perfusion (R=−0.46; P=0.037), yet no relationship between AAT and [GABA⁺]/[NAA–NAAG] (R=−0.12; P=0.33). Perfusion measurements that do not control for AAT variations reveal reduced correlations between [GABA⁺]/[NAA–NAAG] and perfusion (R=−0.13; P=0.32). These findings largely reconcile contradictory reports between perfusion and inhibitory tone, and underscore the physiologic origins of the growing literature relating functional imaging signals, hemodynamics, and neurotransmission.     

Differences in the perceptions of HIV risk amongst dental care providers in Western Australian clinics

This paper examines the perceived risk of occupationally contracting HIV among dental care providers. This examination is based upon responses to a mailed questionnaire from all registered dentists (76 per cent response rate, n = 550) and dental therapists (63 per cent response rate, n = 208) in Western Australia during 1992. Results indicate that: 1) dental care providers who perceive a high risk of occupationally contracting HIV report a more conservative, cautious approach to HIV infection than do providers who perceive less risk of contracting the virus; 2) dental therapists are more likely than dentists to report a higher degree of perceived risk of occupationally contracting HIV; and 3) dentists are more likely than dental therapists to perceive compliance with universal precaution guidelines in dental practices in which they work. Reasons are presented for a suspicion that both sets of perceptions may be exaggerated. The need to educate dental therapists on the realistic risks of occupationally contracting HIV and to educate dentists on the value of compliance (and of appearing to comply) with universal precaution guidelines in order to help reduce undue stress and hindrances in the provision of safe and effective dental care in this era of AIDS concludes this paper.     

PET for evaluation of differential myocardial perfusion dynamics after VEGF gene therapy and laser therapy in end-stage coronary artery disease.

The purpose of this study was to appraise the value of PET in the assessment of the effect of supposedly proangiogenic new therapies such as gene therapy with vascular endothelial growth factor (VEGF) gene and endomyocardial laser therapy. METHODS: Thirty-five patients with end-stage coronary artery disease and class III (Canadian Cardiovascular Society) angina were included. Myocardial ischemia was evaluated with dipyridamole PET scanning and exercise tolerance with bicycle ergometry. Ten patients were treated with naked plasmid DNA encoding for human VEGF165 (VEGF) and 12 patients were treated with laser therapy (direct myocardial revascularization [DMR]) using an electromechanical mapping system. Thirteen patients were treated with standard medical therapy (control). RESULTS: In both active treatment groups, angina was reduced in most subjects, except in 2 VEGF and 5 DMR patients. In the control group, no improvement in anginal classification was found, except in 3 subjects. On the PET scan, solely in the VEGF group, the stress perfusion was significantly improved (from 57 +/- 33 to 81 +/- 55 mL/min/100 g; P = 0.031). Furthermore, in the VEGF group, the number of ischemic segments was reduced from 274 +/- 41 to 234 +/- 48 segments (P = 0.004) but not in the DMR group (from 209 +/- 43 to 215 +/- 52 segments) or in the control group (from 218 +/- 18 to 213 +/- 28 segments). Bicycle exercise duration showed slight nonsignificant changes in the VEGF group (from 3.6 +/- 2.0 to 4.6 +/- 2.1 min), in the DMR group (from 5.1 +/- 1.5 to 4.7 +/- 1.3 min), and in the control group (from 3.3 +/- 1.8 to 3.5 +/- 2.3 min). CONCLUSION: PET showed that intramyocardial gene therapy with the human VEGF165 gene in contrast to laser DMR treatment effectively reduces myocardial ischemia.     

Effect of Prior Salter or Chiari Osteotomy on THA with Developmental Hip Dysplasia

Background  

Controversy exists regarding the outcome of THA after prior pelvic osteotomy.     

Is Oxidized Zirconium Femoral Head Superior to Other Bearing Types in Total Hip Arthroplasty? A Systematic Review and Meta-Analysis

BackgroundOxidized zirconium (OxZi) is a relatively new type of material that combines the strength of a metal with the surface/wears properties of a ceramic. Our aim was to investigate whether OxZi femoral heads lead to lower polyethylene wear, higher survival rate, and better clinical outcomes than the other bearing types in patients treated with total hip arthroplasty (THA).MethodsTwo reviewers independently conducted a systematic search according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses using the MEDLINE/PubMed database and the Cochrane Database of Systematic Reviews. These databases were queried with the terms: a. “oxidized” AND “zirconium” AND “total” AND “hip” AND “arthroplasty”; b. “oxinium” AND “total” AND “hip” AND “arthroplasty”. The primary outcome measures were the survival rate of the bearing surfaces as well as the polyethylene wear.ResultsThe vast majority (85.7%) of the studies, which reported the mean polyethylene wear rate, showed that there was not any significant difference between OxZi and cobalt-chrome (CoCr) femoral heads (rate ratio: 0.836; 95% confidence interval: 0.362-1.928; P = .674). All studies comparing the survival rate of OxZi and CoCr femoral heads illustrated almost excellent survivorship with both implants.ConclusionOxZi femoral heads did not lead to lower polyethylene wear rate or higher survival rate, when compared with CoCr femoral heads in patients treated with THA. On the basis of these results and taking into account the higher cost of these implants, we would not recommend the routine use of OxZi femoral heads in primary THAs.Level of EvidenceSystematic review and meta-analysis of therapeutic studies I-III.