Conversion of the Liver into a Biofactory for DNaseI Using Adeno-Associated Virus Vector Gene Transfer Reduces Neutrophil Extracellular Traps in a Model of Systemic Lupus Erythematosus.

Adeno-associated virus (AAV) vectors are proving to be clinically transformative tools in the treatment of monogenic genetic disease. Rapid ongoing development of this technology promises to not only increase the number of monogenic disorders amenable to this approach but also to bring diseases with complex multigenic and nongenetic etiologies within therapeutic reach. In this study, we explore the broader paradigm of converting the liver into a biofactory for systemic output of therapeutic molecules using AAV-mediated delivery of the endonuclease DNaseI as an exemplar. DNaseI can clear neutrophil extracellular traps (NETs), which are nuclear-protein structures possessing antimicrobial action, also involved in the pathophysiology of clinically troubling immune-mediated diseases. However, a translational challenge is short half-life of the enzyme in vivo (<5 h). This study demonstrates that AAV-mediated liver-targeted gene transfer stably induces serum DNaseI activity to >190-fold above physiological levels. In lupus-prone mice (NZBWF1), the activity was maintained for longer than 6 months, the latest time point tested, and resulted in a clear functional effect with reduced renal presence of neutrophils, NETs, IgG, and complement C3. However, treatment in this complex disease model did not extend lifespan, improve serological endpoints, or preserve renal function, indicating there are elements of pathophysiology not accessible to DNaseI in the NZBWF1 model. We conclude that a translational solution to the challenge of short half-life of DNaseI is AAV-mediated gene delivery and that this may be efficacious in treating disease where NETs are a dominant pathological mechanism.

Diagnostic Inflammation Biomarkers for Prediction of 30-Day Mortality Rate in Acute Cholangitis.

Background: Acute cholangitis (AC) is an acute inflammation and infection of the biliary tract, a potentially life-threatening infection, which is usually associated with biliary tree obstruction and impairment of bile flow from the liver to the duodenum. AC is classified by severity from mild, moderate to severe infection (grade I to III, respectively). Methods: This study recruited a retrospective cohort from Jan 2015 to July 2018. Overall, 294 patients of age ≥ 18 years with AC were enrolled. The study was conducted according to the World Medical Association Declaration of Helsinki. Demographic and laboratory data were collected for analysis. T-Bilirubin and other laboratory results were collected and analyzed using independent T-test and ANOVA for continuous values and multivariate COX regression for survival analysis for identifying independent factors for early mortality. The cut-off threshold of T-bilirubin was determined by calculating the area under the receiver operating characteristic (ROC) curve. Results: There were 213 male and 81 female patients and mean age ± SD of patients was 49.57 ± 16.1 and 56.12 ± 20.18 respectively. 31.9% patients were found older than 60 years of age and 35% patients were found between 30-45 years of age. T-bilirubin and length of hospital stay (LOS) were found statistically significant (P < 0.05) in relation to mortality in AC patients. The area under ROC curve for T-bilirubin level (P = 0.017, OR = 1.010) was 0.717 (95% CI, 6.25-168.9) and this is consistent with the Cut-off point for more than or equal to 38.6 µmol/L (2.26 mg/dL). Conclusions: In this study, T-bilirubin level is found to be significantly related to short-term mortality in AC. Further studies are still needed with larger cohorts to shed more light on these findings. Highlights: Herein, we report a retrospective observational study aiming to evaluate biomarkers contributing to mortality in AC and to determine the cut-off diagnostic levels that could be easily used in emergency setting. Overall, 294 patients of age ≥ 18 years with AC were enrolled. The study was conducted according to the World Medical Association Declaration of Helsinki and approved by Institutional Review Board (IRB) with approval: MRC-01-20-823 at Hamad Medical Corporation (HMC). Demographic and laboratory data were collected for analysis. Total-Bilirubin and other laboratory results were collected and analyzed using independent T-test and ANOVA for continuous values and multivariate COX regression for survival analysis for identifying independent factors for early mortality. The cut-off threshold of T-bilirubin was determined by calculating the area under the receiver operating characteristic (ROC) curve. There were 213 male and 81 female patients and mean age ± SD of patients was 49.57 ± 16.1 and 56.12 ± 20.18 respectively. 31.9% patients were found older than 60 years of age and 35% patients were found between 30-45 years of age. T-bilirubin and length of hospital stay (LOS) were found statistically significant (P < 0.05) in relation to mortality in AC patients. The area under ROC curve for T-bilirubin level (P = 0.037, OR = 1.010) was 0.717 (95% CI, 6.25-168.9) and this is consistent with the Cut-off point for more than or equal to 38.6 µmol/L (2.26 mg/dL).

Adeno-Associated Virus Vector Gene Delivery Elevates Factor I Levels and Downregulates the Complement Alternative Pathway In Vivo.

The complement system is a key component of innate immunity, but impaired regulation influences disease susceptibility, including age-related macular degeneration and some kidney diseases. While complete complement inhibition has been used successfully to treat acute kidney disease, key unresolved challenges include strategies to modulate rather than completely inhibit the system and to deliver therapy potentially over decades. Elevating concentrations of complement factor I (CFI) restricts complement activation in vitro and this approach was extended in the current study to modulate complement activation in vivo. Sustained increases in CFI levels were achieved using an adeno-associated virus (AAV) vector to target the liver, inducing a 4- to 5-fold increase in circulating CFI levels. This led to decreased activity of the alternative pathway as demonstrated by a reduction in the rate of inactive C3b (iC3b) deposition and more rapid formation of C3 degradation products. In addition, vector application in a mouse model of systemic lupus erythematosus (NZBWF1), where tissue injury is, in part, complement dependent, resulted in reduced complement C3 and IgG renal deposition. Collectively, these data demonstrate that sustained elevation of CFI reduces complement activation in vivo providing proof-of-principle support for the therapeutic application of AAV gene delivery to modulate complement activation.

Gain-of-function factor H-related 5 protein impairs glomerular complement regulation resulting in kidney damage.

Genetic variation within the factor H-related (FHR) genes is associated with the complement-mediated kidney disease, C3 glomerulopathy (C3G). There is no definitive treatment for C3G, and a significant proportion of patients develop end-stage renal disease. The prototypical example is CFHR5 nephropathy, through which an internal duplication within a single CFHR5 gene generates a mutant FHR5 protein (FHR5mut) that leads to accumulation of complement C3 within glomeruli. To elucidate how abnormal FHR proteins cause C3G, we modeled CFHR5 nephropathy in mice. Animals lacking the murine factor H (FH) and FHR proteins, but coexpressing human FH and FHR5mut (hFH-FHR5mut), developed glomerular C3 deposition, whereas mice coexpressing human FH with the normal FHR5 protein (hFH-FHR5) did not. Like in patients, the FHR5mut had a dominant gain-of-function effect, and when administered in hFH-FHR5 mice, it triggered C3 deposition. Importantly, adeno-associated virus vector-delivered homodimeric mini-FH, a molecule with superior surface C3 binding compared to FH, reduced glomerular C3 deposition in the presence of the FHR5mut. Our data demonstrate that FHR5mut causes C3G by disrupting the homeostatic regulation of complement within the kidney and is directly pathogenic in C3G. These results support the use of FH-derived molecules with enhanced C3 binding for treating C3G associated with abnormal FHR proteins. They also suggest that targeting FHR5 represents a way to treat complement-mediated kidney injury.

Global Distribution of Businesses Marketing Stem Cell-Based Interventions.

A structured search reveals that online marketing of stem-cell-based interventions is skewed toward developed economies including the United States, Ireland, Australia, and Germany. Websites made broad, imprecise therapeutic claims and frequently failed to detail procedures. Widespread marketing poses challenges to regulators, bioethicists, and those seeking realistic hope from therapies.

Feedforward interview technique in obstetrics and gynaecology residents: a fact or fallacy.

OBJECTIVE: To determine the role of Feedforward Interview (FFI) technique in motivating residents of Obstetrics and Gynaecology for better learning and performance. STUDY DESIGN: An explorative study with mixed method approach being employed. PLACE AND DURATION OF STUDY: Department of Obstetrics and Gynaecology, Sandeman (Provincial) Hospital, Quetta, from November 2010 till May 2013. METHODOLOGY: Feedforward interview technique was complimented by survey questionnaire employing similar philosophy of FFI to triangulate data through two methods. Survey questionnaire was filled-up by 21 residents and analysed by SPSS version 17. Fourteen of these participants were identified for in-depth Feedforward Interviews (FFI), based on nonprobability purposive sampling after informed consent, and content analysis was done. RESULTS: Feedforward interview technique enabled majority of residents in recalling minimum of 3 positive experiences, mainly related to surgical experiences, which enhanced their motivation to aspire for further improvement in this area. Hard work was the main personal contributing factor both in FFI and survey. In addition to identifying clinical experiences enhancing desire to learn, residents also reported need for more academic support as an important factor which could also boost motivation to attain better performance. CONCLUSION: Feedforward interview technique not only helps residents in recalling positive learning experiences during their training but it also has a significant influence on developing insight about one's performance and motivating residents to achieve higher academic goals.

Developing professionalism: dental students' perspective.

OBJECTIVE: To explore the undergraduate dental students' insight of their professionalism development through Focus Group Discussions (FGD). STUDY DESIGN: Constructivist approach using qualitative phenomenological design. PLACE AND DURATION OF STUDY: Fatima Memorial Hospital, College of Dentistry, Lahore, from April to June 2011. METHODOLOGY: Four FGDs of 1st year (8 students), 2nd year (6 students), 3rd year (6 students) and 4th year (6 students) enrolled in Bachelor of Dental Surgery (BDS) program were conducted to explore how they have developed various elements of professionalism namely altruism, accountability, excellence, duty and service, honor and integrity, and respect for all; and how professionalism can be further developed in them. The FGDs were audio taped, transcribed and analyzed through thematic analysis. Triangulation of themes and trends were done through content analysis by relating to their respective frequency of quotes. Data verification was done through audit by second author. RESULTS: Role models and social responsibility were the main reasons in the students' professionalism development thus far with personal virtues and reasons; religion; and punishment and reward contributing to a lesser degree. Training contributed least but was deemed most in furthering professionalism. Excessive workload (quota) and uncongenial educational environment were considered detrimental to the cause. CONCLUSION: Formal planning and implementation of professionalism curriculum; selection of students with appropriate attributes; control of hidden curriculum, including effective role models, good educational and working environments will foster professionalism among dental students maximally.

Perceptions of doctors on being treated by a doctor just completing the house job.

OBJECTIVE: To determine the percentage of medical teachers and fresh doctors who feel that graduating doctors are competent or incompetent to handle common ailments and to evaluate the reasons for their perceptions. STUDY DESIGN: Sequential mixed method. PLACE AND DURATION OF STUDY: First phase extending from December 2010 to December 2011 and second phase was conducted in March 2014. METHODOLOGY: First phase comprised electronic distribution of questionnaire to 100 medical teachers and fresh doctors working in hospitals attached with 5 private and 5 public sector medical colleges of Lahore and Karachi to rate an average house officer on a frequency scale of 1 - 6 and do self-assessment, in case of a fresh doctor. The second phase included interviews of 20 medical teachers to explore justification for their rating in the survey questionnaire and for triangulation of data. Quantitative data was analyzed through SPSS version 15 to calculate frequencies and percentages and interviews were analyzed through quasi-statistical approach. RESULTS: In survey, 38.36% out of 73 medical teachers and 30% out of 20 medical teachers in interviews confirmed their confidence on consulting fresh doctors for common ailments as opposed to 61.64% and 70% respectively, expressing their dissatisfaction. Self-assessment of fresh doctors indicated that 40% are confident in handling common medical conditions as opposed to 33.3% out of 75 respondents, who are not confident about their clinical skills. CONCLUSION: Faculty and self-assessment of fresh doctors has a fair overlap, indicating room for further improvement in the house job training program.

Task-based learning versus problem-oriented lecture in neurology continuing medical education.

OBJECTIVE: To determine whether general practitioners learned better with task-based learning or problem-oriented lecture in a Continuing Medical Education (CME) set-up. STUDY DESIGN: Quasi-experimental study. PLACE AND DURATION OF STUDY: The Aga Khan University, Karachi campus, from April to June 2012. METHODOLOGY: Fifty-nine physicians were given a choice to opt for either Task-based Learning (TBL) or Problem Oriented Lecture (PBL) in a continuing medical education set-up about headaches. The TBL group had 30 participants divided into 10 small groups, and were assigned case-based tasks. The lecture group had 29 participants. Both groups were given a pre and a post-test. Pre/post assessment was done using one-best MCQs. The reliability coefficient of scores for both the groups was estimated through Cronbach's alpha. An item analysis for difficulty and discriminatory indices was calculated for both the groups. Paired t-test was used to determine the difference between pre- and post-test scores of both groups. Independent t-test was used to compare the impact of the two teaching methods in terms of learning through scores produced by MCQ test. RESULTS: Cronbach's alpha was 0.672 for the lecture group and 0.881 for TBL group. Item analysis for difficulty (p) and discriminatory indexes (d) was obtained for both groups. The results for the lecture group showed pre-test (p) = 42% vs. post-test (p) = 43%; pre- test (d) = 0.60 vs. post-test (d) = 0.40. The TBL group showed pre -test (p) = 48% vs. post-test (p) = 70%; pre-test (d) = 0.69 vs. post-test (d) = 0.73. Lecture group pre-/post-test mean scores were (8.52 ± 2.95 vs. 12.41 ± 2.65; p < 0.001), where TBL group showed (9.70 ± 3.65 vs. 14 ± 3.99; p < 0.001). Independent t-test exhibited an insignificant difference at baseline (lecture 8.52 ± 2.95 vs. TBL 9.70 ± 3.65; p = 0.177). The post-scores were not statistically different lecture 12.41 ± 2.65 vs. TBL 14 ± 3.99; p = 0.07). CONCLUSION: Both delivery methods were found to be equally effective, showing statistically insignificant differences. However, TBL groups' post-test higher mean scores and radical increase in the post-test difficulty index demonstrated improved learning through TBL delivery and calls for further exploration of longitudinal studies in the context of CME.

Evaluating neurology CME in two educational methods using Patton's utilization focused model.

BACKGROUND: Generally in continuing education medical education (CME) the most time is consumed for in the planning and preparation of the event. This planning and preparation, however, needs recognition through an evaluative process. The purpose of this study was to evaluate neurology CME in two educational methods, lecture vs task-based learning, using Patton's utilisation focused model. METHODS: This was an observational, cross-sectional inquiry. The questionnaire evaluated the educational elements such as learning objectives met, content covered, presentations at the level of understanding, level of interaction, knowledge gained, time management, queries responded, organisation, quality of learning material and overall grading of the educational event. General Practitioners were the key participants in this evaluation and consisted of 60 self-selected physicians distributed equally in both the TBL and lecture groups. Patton's utilization focused model was used to produce findings for effective decision making. The data were analysed using Mann-Whitney U test to know the value of the learning method that satisfied the most participants. RESULTS: A total of 58 evaluations were returned, 29 from the TBL group and 29 from the lecture. The analysis of the elements showed higher mean ranks for TBL method ranging between 32.2 and 38.4 versus lecture (20.6-26.8). Most of the elements assessed were statistically significant (p > 0.05), except time management (p = 0.22). However, elements as 'objectives of the activity met' (p = 0.07), 'overall grading of the event' (p = 0.06) and 'presentations at the level of understanding' (p = 0.06) were at border line. Of the 29 respondents in the TBL group, 75% rated all the elements of the program above very good. In the lecture group, 22 (75%) respondents out of 29 rated almost half of the elements above very good. CONCLUSION: Majority of respondents in the TBL group rated all program elements as exceptional compared to the lecture group in which only half of the elements were rated above very good. Task-based learning method made the most impact on participants' satisfaction.

The relationship between asthma and allergic rhinitis in the Iraqi population.

BACKGROUND: Recently, extensive research has established that epidemiologic and therapeutic links exist between allergic rhinitis and asthma. The objective of this study was to clarify this association in Iraq. METHODS: The data included in this study were collected from five surveys for asthma and allergic rhinitis that were performed during the period from September 2000 to July 2008. These surveys were parts of Tikrit University College of Medicine PHC program. RESULTS: The frequency of allergic rhinitis (AR) was 61.6% among individuals with asthma versus 6% among non-asthmatic (control) subjects (Odd Ratio [OR] = 25.5; P < 0.0001). All studies indicated a significant frequency of AR among asthmatic patients in comparison with non-asthmatic subjects, whether the patients were adults or children (OR for adults = 14.9 and 22.5, for children 34.7 and 48.4; P < 0.001 for all). Furthermore, the high frequency of AR in asthmatic patients was seen whether the study was a community based study (CBS)(OR = 14.9 and 48.4; P < 0.0001) or a hospital based study (HBS)(OR = 22.5 & 34.7; P < 0.0001). The frequency of current asthma was 51.8% among individuals with AR versus 5.4% among control subjects (OR = 23.1; P < 0.0001). CONCLUSIONS: This study provided evidence that AR and asthma are strongly associated with each other and the treatment approach should consider the entire airway rather than only a part.