Predictors of clinical outcomes in patients with sepsis: A retrospective study from a tertiary care hospital in Pakistan.

OBJECTIVE: To assess associations between various clinic-demographic factors and clinical outcomes among patients treated for sepsis. METHODS: The retrospective study was conducted at the Aga Khan University Hospital, Karachi, and comprised data of all patients aged >18 years diagnosed with sepsis from January to December 2019. Multivariable logistic regression was used to evaluate independent associations between predictors and outcomes. Data was analysed using R packages. RESULTS: Of the 1,136 patients, 621(54.6%) were male and 515(45.3%) were female. The overall mean age was 59.05±16.91 years. Female gender (odds ratio: 1.029; 95% confidence interval: 1.03-1.64) was found to be an independent predictor of septic shock, while hypertension (odds ratio0.75; 95% confidence interval: 0.59-0.95) emerged as a protective factor. Chronic kidney disease (odds ratio: 1.539; 95% confidence interval: 1.14-2.07) was an independent predictor of prolonged length of stay, while older age appeared to be protective (odds ratio: 0.98; 95% confidence interval: 0.98-0.99). Mortality was associated with a significantly lower odds of Escherichia coli on culture (odds ratio: 0.26; 95% confidence interval: 0.12-0.54). CONCLUSIONS: Independent associations were found between specific patient characteristics and adverse clinical outcomes.

Discriminating spatialised speech in complex environments in multiple sclerosis.

People with multiple sclerosis (pwMS) frequently present with deficits in binaural processing used for sound localization. This study examined spatial release from speech-on-speech masking in pwMS, which involves binaural processing and additional higher level mechanisms underlying streaming, such as spatial attention. 26 pwMS with mild severity (Expanded Disability Status Scale score <3) and 20 age-matched controls listened via headphones to pre-recorded sentences from a standard list presented simultaneously with eight-talker babble. Virtual acoustic techniques were used to simulate sentences originating from 0°, 20°, or 50° on the interaural horizontal plane around the listener whilst babble was presented continuously at 0° azimuth, and participants verbally repeated the target sentence. In a separate task, two simultaneous sentences both containing a colour and number were presented, and participants were required to report the target colour and number. Both competing sentences could originate from 0°, 20°, or 50° on the azimuthal plane. Participants also completed a series of neuropsychological assessments, an auditory questionnaire, and a three-alternative forced-choice task that involved the detection of interaural time differences (ITDs) in noise bursts. Spatial release from masking was observed in both pwMS and controls, as response accuracy in the two speech discrimination tasks improved in the spatially separated conditions (20° and 50°) compared with the co-localised condition. However, pwMS demonstrated significantly less spatial release (18%) than controls (28%) when discriminating colour/number coordinates. At 50° separation, pwMS discriminated significantly fewer coordinates (77%) than controls (89%). In contrast, pwMS had similar performances to controls when sentences were presented in babble, and for the basic ITD discrimination task. Significant correlations between speech discrimination performance and standardized neuropsychological scores were observed across all spatial conditions. Our findings suggest that spatial hearing is likely to be implicated in pwMS, thereby affecting the perception of competing speech originating from various locations.

Enrichment of Nanofiber Hydrogel Composite with Fractionated Fat Promotes Regenerative Macrophage Polarization and Vascularization for Soft-Tissue Engineering.

BACKGROUND: Fractionated fat has been shown to promote dermal regeneration; however, the use of fat grafting for reconstruction of soft-tissue defects is limited because of volume loss over time. The authors have developed a novel approach for engineering of vascularized soft tissue using an injectable nanofiber hydrogel composite enriched with fractionated fat. METHODS: Fractionated fat was generated by emulsification of groin fat pads from rats and mixed in a 3:1 ratio with nanofiber hydrogel composite (nanofiber hydrogel composite with fractionated fat). Nanofiber hydrogel composite with fractionated fat or nanofiber hydrogel composite alone was placed into isolation chambers together with arteriovenous loops, which were subcutaneously implanted into the groin of rats (n = 8 per group). After 21 days, animals were euthanized and systemically perfused with ink, and tissue was explanted for histologic analysis. Immunofluorescent staining and confocal laser scanning microscopy were used to quantify CD34+ progenitor cell and macrophage subpopulations. RESULTS: Nanofiber hydrogel composite with fractionated fat tissue maintained its shape without shrinking and showed a significantly stronger functional vascularization compared to composite alone after 21 days of implantation (mean vessel count, 833.5 ± 206.1 versus 296.5 ± 114.1; p = 0.04). Tissue heterogeneity and cell count were greater in composite with fractionated fat (mean cell count, 49,707 ± 18,491 versus 9263 ± 3790; p = 0.005), with a significantly higher number of progenitor cells and regenerative CD163+ macrophages compared to composite alone. CONCLUSIONS: Fractionated fat-enriched nanofiber hydrogel composite transforms into highly vascularized soft tissue over 21 days without signs of shrinking and promotes macrophage polarization toward regenerative phenotypes. Enrichment of injectable nanofiber hydrogel composite with fractionated fat represents a promising approach for durable reconstruction of soft-tissue defects. CLINICAL RELEVANCE STATEMENT: The authors' approach for tissue engineering may ultimately lay the groundwork for clinically relevant applications with the goal of generating large volumes of vascularized soft tissue for defect reconstruction without donor site morbidity.

Challenges in updating habitat suitability models: An example with the lesser prairie-chicken.

Habitat loss from land-use change is one of the top causes of declines in wildlife species of concern. As such, it is critical to assess and reassess habitat suitability as land cover and anthropogenic features change for both monitoring and developing current information to inform management decisions. However, there are obstacles that must be overcome to develop consistent assessments through time. A range-wide lek habitat suitability model for the lesser prairie-chicken (Tympanuchus pallidicinctus), currently under review by the U. S. Fish and Wildlife Service for potential listing under the Endangered Species Act, was published in 2016. This model was based on lek data from 2002 to 2012, land cover data ranging from 2001 to 2013, and anthropogenic features from circa 2011, and has been used to help guide lesser prairie-chicken management and anthropogenic development actions. We created a second iteration model based on new lek surveys (2015 to 2019) and updated predictors (2016 land cover and cleaned/updated anthropogenic data) to evaluate changes in lek suitability and to quantify current range-wide habitat suitability. Only three of 11 predictor variables were directly comparable between the iterations, making it difficult to directly assess what predicted changes resulted from changes in model inputs versus actual landscape change. The second iteration model showed a similar positive relationship with land cover and negative relationship with anthropogenic features to the first iteration, but exhibited more variation among candidate models. Range-wide, more suitable habitat was predicted in the second iteration. The Shinnery Oak Ecoregion, however, exhibited a loss in predicted suitable habitat that could be due to predictor source changes. Iterated models such as this are important to ensure current information is being used in conservation and development decisions.

Speech discrimination impairments as a marker of disease severity in multiple sclerosis.

BACKGROUND: Multiple Sclerosis (MS) pathology is likely to disrupt central auditory pathways, thereby affecting an individual's ability to discriminate speech from noise. Despite the importance of speech discrimination in daily communication, it's characterization in the context of MS remains limited. This cross-sectional study evaluated speech discrimination in MS under "real world" conditions where sentences were presented in ecologically valid multi-talker speech or broadband noise at several signal-to-noise ratios (SNRs). METHODS: Pre-recorded Bamford-Kowal-Bench sentences were presented at five signal-to-noise ratios (SNR) in one of two background noises: speech-weighted noise and eight-talker babble. All auditory stimuli were presented via headphones to control (n = 38) and MS listeners with mild (n = 20), moderate (n = 16) and advanced (n = 10) disability. Disability was quantified by the Kurtzke Expanded Disability Status Scale (EDSS) and scored by a neurologist. All participants passed a routine audiometric examination. RESULTS: Despite normal hearing, MS psychometric discrimination curves which model the relationship between signal-to-noise ratio (SNR) and sentence discrimination accuracy in speech-weighted noise and babble did not change in slope (sentences/dB) but shifted to higher SNRs (dB) compared to controls. The magnitude of the shift in the curve systematically increased with greater disability. Furthermore, mixed-effects models identified EDSS score as the most significant predictor of speech discrimination in noise (odds ratio = 0.81; p < 0.001). Neither age, sex, disease phenotype or disease duration were significantly associated with speech discrimination performance in noise. Only MS listeners with advanced disability self-reported audio-attentional difficulty in a questionnaire designed to reflect auditory processing behaviours in daily life. CONCLUSION: Speech discrimination performance worsened systematically with greater disability, independent of age, sex, education, disease duration or disease phenotype. These results identify novel auditory processing deficits in MS and highlight that speech discrimination tasks may provide a viable non-invasive and sensitive means for disease monitoring in MS.

Comparing optimized exoskeleton assistance of the hip, knee, and ankle in single and multi-joint configurations.

Exoskeletons that assist the hip, knee, and ankle joints have begun to improve human mobility, particularly by reducing the metabolic cost of walking. However, direct comparisons of optimal assistance of these joints, or their combinations, have not yet been possible. Assisting multiple joints may be more beneficial than the sum of individual effects, because muscles often span multiple joints, or less effective, because single-joint assistance can indirectly aid other joints. In this study, we used a hip-knee-ankle exoskeleton emulator paired with human-in-the-loop optimization to find single-joint, two-joint, and whole-leg assistance that maximally reduced the metabolic cost of walking. Hip-only and ankle-only assistance reduced the metabolic cost of walking by 26 and 30% relative to walking in the device unassisted, confirming that both joints are good targets for assistance (N = 3). Knee-only assistance reduced the metabolic cost of walking by 13%, demonstrating that effective knee assistance is possible (N = 3). Two-joint assistance reduced the metabolic cost of walking by between 33 and 42%, with the largest improvements coming from hip-ankle assistance (N = 3). Assisting all three joints reduced the metabolic cost of walking by 50%, showing that at least half of the metabolic energy expended during walking can be saved through exoskeleton assistance (N = 4). Changes in kinematics and muscle activity indicate that single-joint assistance indirectly assisted muscles at other joints, such that the improvement from whole-leg assistance was smaller than the sum of its single-joint parts. Exoskeletons can assist the entire limb for maximum effect, but a single well-chosen joint can be more efficient when considering additional factors such as weight and cost.

Speech discrimination performance in multiple sclerosis dataset.

The most complex interactions between human beings occur through speech, and often in the presence of background noise. Understanding speech in noisy environments requires the integrity of highly integrated and widespread auditory networks likely to be impacted by multiple sclerosis (MS) related neurogenic injury. Despite the impact auditory communication has on a person's ability to navigate the world, build relationships, and maintain employability; studies of speech-in-noise (SiN) perception in people with MS (pwMS) have been minimal to date. Thus, this paper presents a dataset related to the acquisition of pure-tone thresholds, SiN performance and questionnaire responses in age-matched controls and pwMS. Bilateral pure-tone hearing thresholds were obtained at frequencies of 250 hertz (Hz), 500 Hz, 750 Hz, 1000 Hz, 1500 Hz, 2000 Hz, 4000 Hz, 6000 Hz and 8000 Hz, and hearing thresholds were defined as the lowest level at which the tone was perceived 50% of the time. Thresholds at 500 Hz, 1000 Hz, 2000 Hz and 4000 Hz were used to calculate the four-tone average for each participant, and only those with a bilateral four tone average of ≤ 25 dB HL were included in the analysis. To investigate SiN performance in pwMS, pre-recorded Bamford-Kowal-Bench (BKB) sentences were presented binaurally through headphones at five signal-to-noise ratios (SNR) in two noise conditions: speech-weighted noise and multi-talker babble. Participants were required to verbally repeat each sentence they had just heard; or indicate their inability to do so. A 33-item questionnaire, based on validated inventories for specific adult clinical populations with abnormal auditory processing, was used to evaluate auditory processing in daily life for pwMS. For analysis, pwMS were grouped according to their Expanded Disability Status Scale (EDSS) score as rated by a neurologist. PwMS with EDSS scores ≤ 1.5 were classified as 'mild' (n = 20); between 2 and 4.5 as 'moderate' (n = 16) and between 5 and 7 as 'advanced' (n = 10) and were compared to neurologically healthy controls (n = 38). The outcomes of the SiN task conducted in pwMS can be found in Iva et al., (2021). The present data has important implications for the timing and delivery of preparatory education to patients, family, and caregivers about communication abilities in pwMS. This dataset will also be valuable for the reuse/reanalysis required for future investigations into the clinical utility of SiN tasks to monitor disease progression.

Tissue Engineering of Axially Vascularized Soft-Tissue Flaps with a Poly-(ɛ-Caprolactone) Nanofiber-Hydrogel Composite.

Objective: To develop a novel approach for tissue engineering of soft-tissue flaps suitable for free microsurgical transfer, using an injectable nanofiber hydrogel composite (NHC) vascularized by an arteriovenous (AV) loop. Approach: A rat AV loop model was used for tissue engineering of vascularized soft-tissue flaps. NHC or collagen-elastin (CE) scaffolds were implanted into isolation chambers together with an AV loop and explanted after 15 days. Saphenous veins were implanted into the scaffolds as controls. Neoangiogenesis, ultrastructure, and protein expression of SYNJ2BP, EPHA2, and FOXC1 were analyzed by immunohistochemistry and compared between the groups. Rheological properties were compared between the two scaffolds and native human adipose tissue. Results: A functional neovascularization was evident in NHC flaps with its amount being comparable with CE flaps. Scanning electron microscopy revealed a strong mononuclear cell infiltration along the nanofibers in NHC flaps and a trend toward higher fiber alignment compared with CE flaps. SYNJ2BP and EPHA2 expression in endothelial cells (ECs) was lower in NHC flaps compared with CE flaps, whereas FOXC1 expression was increased in NHC flaps. Compared with the stiffer CE flaps, the NHC flaps showed similar rheological properties to native human adipose tissue. Innovation: This is the first study to demonstrate the feasibility of tissue engineering of soft-tissue flaps with similar rheological properties as human fat, suitable for microsurgical transfer using an injectable nanofiber hydrogel composite. Conclusions: The injectable NHC scaffold is suitable for tissue engineering of axially vascularized soft-tissue flaps with a solid neovascularization, strong cellular infiltration, and biomechanical properties similar to human fat. Our data indicate that SYNJ2BP, EPHA2, and FOXC1 are involved in AV loop-associated angiogenesis and that the scaffold material has an impact on protein expression in ECs.

The effect of a nanofiber-hydrogel composite on neural tissue repair and regeneration in the contused spinal cord.

An injury to the spinal cord causes long-lasting loss of nervous tissue because endogenous nervous tissue repair and regeneration at the site of injury is limited. We engineered an injectable nanofiber-hydrogel composite (NHC) with interfacial bonding to provide mechanical strength and porosity and examined its effect on repair and neural tissue regeneration in an adult rat model of spinal cord contusion. At 28 days after treatment with NHC, the width of the contused spinal cord segment was 2-fold larger than in controls. With NHC treatment, tissue in the injury had a 2-fold higher M2/M1 macrophage ratio, 5-fold higher blood vessel density, 2.6-fold higher immature neuron presence, 2.4-fold higher axon density, and a similar glial scar presence compared with controls. Spared nervous tissue volume in the contused segment and hind limb function was similar between groups. Our findings indicated that NHC provided mechanical support to the contused spinal cord and supported pro-regenerative macrophage polarization, angiogenesis, axon growth, and neurogenesis in the injured tissue without any exogenous factors or cells. These results motivate further optimization of the NHC and delivery protocol to fully translate the potential of the unique properties of the NHC for treating spinal cord injury.

Collaborations on blood transfusion research in sub-Saharan Africa: who, what and where.

BACKGROUND AND OBJECTIVES: Children and pregnant women use 75% of the blood supply in sub-Saharan Africa (SSA) but face widespread blood shortages. To increase safe blood supply, Africa-specific evidence and strengthened capacity for transfusion research are needed. Our study analysed seven years of SSA transfusion publications, compared researched topics against priorities and enumerated SSA transfusion research collaborations. MATERIALS AND METHODS: Data on research topic, journal type, authors' institutions and country were extracted from transfusion-related SSA articles published between 2008 and 14 and used to construct a quantitative, graphic visualization of collaborations. Research topics were compared to those identified as priorities for SSA blood services in 2008 and 2015. RESULTS: Of the 2176, 267 articles (average 38/year) met criteria for analysis. They involved 1245 authors, 673 institutions, 59 countries (35 SSA) and 1375 collaborations. About 41% were on transfusion-transmitted infections. About 34% were published in specialist transfusion journals. Only 7% involved exclusively collaborations within SSA. Two of the top fifteen institutions by publication quantity were from outside SSA. CONCLUSION: Despite a general paucity of SSA-relevant transfusion research, Francophone SSA was well-represented. Published research topics are not well matched to SSA research priorities; research on supply, distribution, financing and systems is particularly neglected. The study provides a baseline against which to track any refocusing of research activity to better meet SSA's needs. Transfusion research hubs within and beyond SSA have been identified as a springboard network for expanding SSA transfusion research capacity.

Hyperaldosteronism from a large adrenal adenoma in a patient with bilateral adrenal nodules.

Primary aldosteronism (PA) is a potentially reversible cause of uncontrolled hypertension. Early diagnosis and timely management of PA can prevent end-organ damage. Aldosteronoma Resolution Score (ARS) is a useful tool to predict cure rates and resolution of hypertension after adrenalectomy.

Speech Discrimination Tasks: A Sensitive Sensory and Cognitive Measure in Early and Mild Multiple Sclerosis.

There is a need for reliable and objective measures of early and mild symptomology in multiple sclerosis (MS), as deficits can be subtle and difficult to quantify objectively in patients without overt physical deficits. We hypothesized that a speech-in-noise (SiN) task would be sensitive to demyelinating effects on precise neural timing and diffuse higher-level networks required for speech intelligibility, and therefore be a useful tool for monitoring sensory and cognitive changes in early MS. The objective of this study was to develop a SiN task for clinical use that sensitively monitors disease activity in early (<5 years) and late (>10 years) stages of MS subjects with mild severity [Expanded Disability Status Scale (EDSS) score < 3]. Pre-recorded Bamford-Kowal-Bench sentences and isolated keywords were presented at five signal-to-noise ratios (SNR) in one of two background noises: speech-weighted noise and eight-talker babble. All speech and noise were presented via headphones to controls (n = 38), early MS (n = 23), and late MS (n = 12) who were required to verbally repeat the target speech. MS subjects also completed extensive neuropsychological testing which included: Paced Auditory Serial Addition Test, Digit Span Test, and California Verbal Learning Test. Despite normal hearing thresholds, subjects with early and late mild MS displayed speech discrimination deficits when sentences and words were presented in babble - but not speech-weighted noise. Significant correlations between SiN performance and standardized neuropsychological assessments indicated that MS subjects with lower functional scores also had poorer speech discrimination. Furthermore, a quick 5-min task with words and keywords presented in multi-talker babble at an SNR of -1 dB was 82% accurate in discriminating mildly impaired MS individuals (median EDSS = 0) from healthy controls. Quantifying functional deficits in mild MS will help clinicians to maximize the opportunities to preserve neurological reserve in patients with appropriate therapeutic management, particularly in the earliest stages. Given that physical assessments are not informative in this fully ambulatory cohort, a quick 5-min task with words and keywords presented in multi-talker babble at a single SNR could serve as a complementary test for clinical use due to its ease of use and speed.

Oligodendrocyte precursors gain Schwann cell-like phenotype and remyelinate axons upon engraftment into peripheral nerves.

The ability to treat large peripheral nerve injuries may be greatly advanced if an accessible source of human myelinating cells is identified, as it overcomes one of the major limitations of acellular or synthetic nerve guides compared with autografts, the gold standard for large defect repair. Methods to derive oligodendrocyte precursor cells (OPCs) from human pluripotent stem cells have advanced to the point where they have been shown capable of myelination and are being evaluated in clinical trials. Here, we test the hypothesis that OPCs can survive and remyelinate axons in the peripheral nervous system during a repair process. Using freshly isolated OPCs from mouse post-natal brains, we engrafted these OPCs into the tibial nerve immediately after it being subjected to cryolesioning. At 1-month postengraftment, we found numerous graft-derived cells that survived in this environment, and many transplanted cells expressed Schwann cell markers such as periaxin and S100ß coexpressed with myelin basic protein, whereas oligodendrocyte markers O4 and Olig2 were virtually absent. Our results demonstrate that OPCs can survive in a peripheral nervous system micro-environment and undergo niche-dependent transdifferentiation into Schwann cell-like cells as has previously been observed in central nervous system focal demyelination models, suggesting that OPCs constitute an accessible source of cells for peripheral nerve cell therapies.

Poly(ε-Caprolactone) Nanofiber Wrap Improves Nerve Regeneration and Functional Outcomes after Delayed Nerve Repair.

BACKGROUND: The purpose of this study was to assess the efficacy of biodegradable, electrospun poly(ε-caprolactone) nanofiber nerve conduits in improving nerve regeneration. METHODS: The authors used a rat forelimb chronic denervation model to assess the effects of poly(ε-caprolactone) conduits on improving nerve regeneration and upper extremity function. Three groups of rats were examined: (1) negative-control animals (n = 5), which underwent 8 weeks of median nerve chronic denervation injury followed by repair with no conduit; (2) experimental animals (n = 5), which underwent 8 weeks of median nerve chronic denervation followed by repair and poly(ε-caprolactone) nerve conduit wrapping of the nerve coaptation site; and (3) positive-control animals (n = 5), which were naive controls. All animals underwent compound muscle action potential and functional testing. At 14 weeks after repair, the median nerve and flexor muscles were harvested for histologic analysis. RESULTS: Histomorphometric analysis of regenerating median nerves demonstrated augmented axonal regeneration in experimental versus negative control animals (total axon count, 1769 ± 672 versus 1072 ± 123.80; p = 0.0468). With regard to functional recovery, experimental and negative-control animals (1.67 ± 0.04 versus 0.97 ± 0.39; p = 0.036) had regained 34.9 percent and 25.4 percent, respectively, of baseline hand grip strength at 14 weeks after repair. Lastly, less collagen deposition at the nerve coaptation site of experimental animals was found when compared to control animals (p < 0.05). CONCLUSION: Biodegradable, poly(ε-caprolactone) nanofiber nerve conduits can improve nerve regeneration and subsequent physiologic extremity function in the setting of delayed nerve repair by decreasing the scar burden at nerve coaptation sites.

Glial Cell Line-Derived Neurotrophic Factor and Chondroitinase Promote Axonal Regeneration in a Chronic Denervation Animal Model.

Functional recovery following nerve injury declines when target re-innervation is delayed. Currently, no intervention exists to improve outcomes after prolonged denervation. We explored the neuroregenerative effects of glial cell line-derived neurotrophic factor (GDNF) and chondroitinase (CDN) in a chronic denervation animal model. A fibrin-based sustained delivery method for growth factors was optimized in vitro and in vivo, and then tested in our animal model. GDNF, CDN, and GDNF+CDN were injected into the denervated stump at the time of nerve repair. Histomorphometry and retrograde labeling were used to assess axonal regeneration. The mechanisms promoting such regeneration were explored with immunofluorescence. Five weeks after repair, the GDNF+CDN group had the highest number and maturity of axons. GDNF was noted to preferentially promote axonal maturity, whereas CDN predominantly increased the number of axons. GDNF favored motor neuron regeneration, and upregulated Ki67 in Schwann cells. CDN did not favor motor versus sensory regeneration and was noted to cleave inhibitory endoneurial proteoglycans. Early measures of nerve regeneration after delayed repair are improved by activating Schwann cells and breaking down the inhibitory proteoglycans in the distal nerve segment, suggesting a role for GDNF+CDN to be translated for human nerve repairs.

Nanofiber-hydrogel composite-mediated angiogenesis for soft tissue reconstruction.

Soft tissue losses from tumor removal, trauma, aging, and congenital malformation affect millions of people each year. Existing options for soft tissue restoration have several drawbacks: Surgical options such as the use of autologous tissue flaps lead to donor site defects, prosthetic implants are prone to foreign body response leading to fibrosis, and fat grafting and dermal fillers are limited to small-volume defects and only provide transient volume restoration. In addition, large-volume fat grafting and other tissue-engineering attempts are hampered by poor vascular ingrowth. Currently, there are no off-the-shelf materials that can fill the volume lost in soft tissue defects while promoting early angiogenesis. Here, we report a nanofiber-hydrogel composite that addresses these issues. By incorporating interfacial bonding between electrospun poly(ε-caprolactone) fibers and a hyaluronic acid hydrogel network, we generated a composite that mimics the microarchitecture and mechanical properties of soft tissue extracellular matrix. Upon subcutaneous injection in a rat model, this composite permitted infiltration of host macrophages and conditioned them into the pro-regenerative phenotype. By secreting pro-angiogenic cytokines and growth factors, these polarized macrophages enabled gradual remodeling and replacement of the composite with vascularized soft tissue. Such host cell infiltration and angiogenesis were also observed in a rabbit model for repairing a soft tissue defect filled with the composite. This injectable nanofiber-hydrogel composite augments native tissue regenerative responses, thus enabling durable soft tissue restoration outcomes.

Drug repositioning in epilepsy reveals novel antiseizure candidates.

Objective: Epilepsy treatment falls short in ~30% of cases. A better understanding of epilepsy pathophysiology can guide rational drug development in this difficult to treat condition. We tested a low-cost, drug-repositioning strategy to identify candidate epilepsy drugs that are already FDA-approved and might be immediately tested in epilepsy patients who require new therapies. Methods: Biopsies of spiking and nonspiking hippocampal brain tissue from six patients with unilateral mesial temporal lobe epilepsy were analyzed by RNA-Seq. These profiles were correlated with transcriptomes from cell lines treated with FDA-approved drugs, identifying compounds which were tested for therapeutic efficacy in a zebrafish seizure assay. Results: In spiking versus nonspiking biopsies, RNA-Seq identified 689 differentially expressed genes, 148 of which were previously cited in articles mentioning seizures or epilepsy. Differentially expressed genes were highly enriched for protein-protein interactions and formed three clusters with associated GO-terms including myelination, protein ubiquitination, and neuronal migration. Among the 184 compounds, a zebrafish seizure model tested the therapeutic efficacy of doxycycline, metformin, nifedipine, and pyrantel tartrate, with metformin, nifedipine, and pyrantel tartrate all showing efficacy. Interpretation: This proof-of-principle analysis suggests our powerful, rapid, cost-effective approach can likely be applied to other hard-to-treat diseases.

Macroporous nanofiber wraps promote axonal regeneration and functional recovery in nerve repair by limiting fibrosis.

Functional outcomes following nerve repair remain suboptimal. Scarring at the repair site is a major impediment to regeneration. A biomaterial scaffold applied around the coaptation site that decreases inflammation holds great potential in reducing scarring, enhancing axonal growth, and improving functional recovery. In this study, we evaluated the effect of a macroporous nanofiber wrap, comprised of nonwoven electrospun poly-ε-caprolactone (PCL), in improving axonal regeneration in a rat sciatic nerve cut and direct repair model. Controls consisted of conventional epineurial repair. We also evaluated our wrap against the commercially available AxoGuard wrap. At five weeks following repair, the nanofiber wrap group showed a significantly decreased intraneural macrophage invasion and collagen deposition at the repair site. This was associated with increased expression of the anti-inflammatory cytokine (IL-10), decreased expression of the pro-inflammatory cytokine (TNF-α), and a decrease in the M1:M2 macrophage phenotype ratio. These findings suggest that this nanofiber wrap, with its unique macroporosity, is modulating the inflammatory response at the repair site by polarizing macrophages towards a pro-regenerative M2 phenotype. Concomitantly, a higher number of regenerated axons was noted. At sixteen weeks, the nanofiber wrap resulted in enhanced functional recovery as demonstrated by electrophysiology, neuromuscular re-innervation, and muscle histology. When compared to the AxoGuard wrap, the nanofiber wrap showed similar inflammation at the repair site and similar nerve morphometric findings, but there was a trend towards a lower overall number of macrophages invading the wrap wall. These results demonstrate favorable outcomes of the macroporous nanofiber wrap in promoting neuroregeneration and functional recovery following nerve repair. STATEMENT OF SIGNIFICANCE: Electrospun nanofiber scaffolds, with specific fiber and pore sizes, were shown to modulate the immune response and create a regenerative environment. In this paper, we present a macroporous nanofiber wrap, made of poly-ε-caprolactone, to be applied at the coaptation site in primary nerve repair. We show that it regulates the inflammatory response at the repair site and decreases scarring/fibrosis. This results in enhanced axonal regeneration, allowing a higher number of axons to cross the suture line and reach the target muscle in a timely fashion. Functional outcomes are thus improved.

Quality control project of NGS HLA genotyping for the 17th International HLA and Immunogenetics Workshop.

The 17th International HLA and Immunogenetics Workshop (IHIW) organizers conducted a Pilot Study (PS) in which 13 laboratories (15 groups) participated to assess the performance of the various sequencing library preparation protocols, NGS platforms and software in use prior to the workshop. The organizers sent 50 cell lines to each of the 15 groups, scored the 15 independently generated sets of NGS HLA genotyping data, and generated "consensus" HLA genotypes for each of the 50 cell lines. Proficiency Testing (PT) was subsequently organized using four sets of 24 cell lines, selected from 48 of 50 PS cell lines, to validate the quality of NGS HLA typing data from the 34 participating IHIW laboratories. Completion of the PT program with a minimum score of 95% concordance at the HLA-A, HLA-B, HLA-C, HLA-DRB1 and HLA-DQB1 loci satisfied the requirements to submit NGS HLA typing data for the 17th IHIW projects. Together, these PS and PT efforts constituted the 17th IHIW Quality Control project. Overall PT concordance rates for HLA-A, HLA-B, HLA-C, HLA-DPA1, HLA-DPB1, HLA-DQA1, HLA-DQB1, HLA-DRB1, HLA-DRB3, HLA-DRB4 and HLA-DRB5 were 98.1%, 97.0% and 98.1%, 99.0%, 98.6%, 98.8%, 97.6%, 96.0%, 99.1%, 90.0% and 91.7%, respectively. Across all loci, the majority of the discordance was due to allele dropout. The high cost of NGS HLA genotyping per experiment likely prevented the retyping of initially failed HLA loci. Despite the high HLA genotype concordance rates of the software, there remains room for improvement in the assembly of more accurate consensus DNA sequences by NGS HLA genotyping software.

In Vivo Bioluminescence Imaging in a Rabbit Model of Orthopaedic Implant-Associated Infection to Monitor Efficacy of an Antibiotic-Releasing Coating.

BACKGROUND: In vivo bioluminescence imaging (BLI) provides noninvasive monitoring of bacterial burden in animal models of orthopaedic implant-associated infection (OIAI). However, technical limitations have limited its use to mouse and rat models of OIAI. The goal of this study was to develop a larger, rabbit model of OIAI using in vivo BLI to evaluate the efficacy of an antibiotic-releasing implant coating. METHODS: A nanofiber coating loaded with or without linezolid-rifampin was electrospun onto a surgical-grade locking peg. To model OIAI in rabbits, a medial parapatellar arthrotomy was performed to ream the femoral canal, and a bright bioluminescent methicillin-resistant Staphylococcus aureus (MRSA) strain was inoculated into the canal, followed by retrograde insertion of the coated implant flush with the articular surface. In vivo BLI signals were confirmed by ex vivo colony-forming units (CFUs) from tissue, bone, and implant specimens. RESULTS: In this rabbit model of OIAI (n = 6 rabbits per group), implants coated without antibiotics were associated with significantly increased knee width and in vivo BLI signals compared with implants coated with linezolid-rifampin (p < 0.001 and p < 0.05, respectively). On day 7, the implants without antibiotics were associated with significantly increased CFUs from tissue (mean [and standard error of the mean], 1.4 × 10 ± 2.1 × 10 CFUs; p < 0.001), bone (6.9 × 10 ± 3.1 × 10 CFUs; p < 0.05), and implant (5.1 × 10 ± 2.2 × 10 CFUs; p < 0.05) specimens compared with implants with linezolid-rifampin, which demonstrated no detectable CFUs from any source. CONCLUSIONS: By combining a bright bioluminescent MRSA strain with modified techniques, in vivo BLI in a rabbit model of OIAI demonstrated the efficacy of an antibiotic-releasing coating. CLINICAL RELEVANCE: The new capability of in vivo BLI for noninvasive monitoring of bacterial burden in larger-animal models of OIAI may have important preclinical relevance.