Fetal Cardiovascular Magnetic Resonance Feature Tracking Myocardial Strain Analysis in Congenital Heart Disease.

BACKGROUND: Cardiovascular magnetic resonance (CMR) is an emerging imaging modality for assessing anatomy and function of the fetal heart in congenital heart disease (CHD). This study aimed to evaluate myocardial strain using fetal CMR feature tracking (FT) in different subtypes of CHD. METHODS: Fetal CMR FT analysis was retrospectively performed on four-chamber cine images acquired with Doppler US gating at 3 Tesla. Left ventricular (LV) global longitudinal strain (GLS), LV global radial strain (GRS), LV global longitudinal systolic strain rate (SR), and right ventricular (RV) GLS were quantified using a dedicated software optimized for fetal strain analysis. Analysis was performed in normal fetuses and different CHD subtypes (d-Transposition of the great arteries (dTGA), hypoplastic left heart syndrome (HLHS), coarctation of the aorta (CoA), tetralogy of Fallot (TOF), RV-dominant atrioventricular septal defect (AVSD), and critical pulmonary stenosis or atresia (PS/PA)). Analyses of variance (ANOVA) with Tukey post-hoc test was used for group comparisons. RESULTS: A total of 60 fetuses were analyzed (8/60 (13%) without CHD, 52/60 (87%) with CHD). Myocardial strain was successfully assessed in 113/120 ventricles (94%). Compared to controls, LV GLS was significantly reduced in fetuses with HLHS (-18.6±2.7% vs. -6.2±5.6%; p<0.001) and RV-dominant AVSD (-18.6±2.7% vs. -7.7±5.0%; p=0.003) and higher in fetuses with CoA (-18.6±2.7% vs. -25.0±4.3%; p=0.038). LV GRS was significantly reduced in fetuses with HLHS (25.7±7.5% vs. 11.4±9.7%; p=0.024). Compared to controls, RV GRS was significantly reduced in fetuses with PS/PA (-16.1±2.8% vs. -8.3±4.2%; p=0.007). Across all strain parameters, no significant differences were present between controls and fetuses diagnosed with dTGA and TOF. CONCLUSIONS: Fetal myocardial strain assessment with CMR FT in CHD is feasible. Distinct differences are present between various types of CHD, suggesting potential implications for clinical decision-making and prognostication in fetal CHD.

CRISPR/Cas9-based double-strand oligonucleotide insertion strategy corrects metabolic abnormalities in murine glycogen storage disease type-Ia.

Glycogen storage disease type-Ia (GSD-Ia), characterized by impaired blood glucose homeostasis, is caused by a deficiency in glucose-6-phosphatase-α (G6Pase-α or G6PC). Using the G6pc-R83C mouse model of GSD-Ia, we explored a CRISPR/Cas9-based double-strand DNA oligonucleotide (dsODN) insertional strategy that uses the nonhomologous end-joining repair mechanism to correct the pathogenic p.R83C variant in G6pc exon-2. The strategy is based on the insertion of a short dsODN into G6pc exon-2 to disrupt the native exon and to introduce an additional splice acceptor site and the correcting sequence. When transcribed and spliced, the edited gene would generate a wild-type mRNA encoding the native G6Pase-α protein. The editing reagents formulated in lipid nanoparticles (LNPs) were delivered to the liver. Mice were treated either with one dose of LNP-dsODN at age 4 weeks or with two doses of LNP-dsODN at age 2 and 4 weeks. The G6pc-R83C mice receiving successful editing expressed ~4% of normal hepatic G6Pase-α activity, maintained glucose homeostasis, lacked hypoglycemic seizures, and displayed normalized blood metabolite profile. The outcomes are consistent with preclinical studies supporting previous gene augmentation therapy which is currently in clinical trials. This editing strategy may offer the basis for a therapeutic approach with an earlier clinical intervention than gene augmentation, with the additional benefit of a potentially permanent correction of the GSD-Ia phenotype.

What Is the Long-term Survivorship of Primary and Revision Cemented Distal Femoral Replacements for Limb Salvage of Patients With Sarcoma?

BACKGROUND: Cemented endoprosthetic reconstruction after resection of primary bone sarcomas has been in common use for decades. Although multiple studies have reported the survivorship of primary endoprostheses, implant survivorship after revision surgery is less well established. Given that earlier advances in systemic therapy improved survival of patients with sarcoma, the usage of revision endoprostheses can be expected to increase and, as such, understanding revision implant survivorship will help to inform patient and surgeon expectations. Additionally, as new implants are developed that allow alternative reconstruction options, a normative dataset establishing accurate expectations for revision cemented endoprostheses is a critical benchmark by which to measure progress. QUESTIONS/PURPOSES: (1) What is the implant survivorship free of all-cause revision for primary and revision cemented distal femoral replacements (DFRs) used in the treatment of malignant or benign tumors? (2) What are the most common indications for revision of primary and revision DFRs in an oncology population with mean follow-up of more than 10 years? (3) How does the indication for revision of a primary DFR affect the subsequent risk for and type of revision DFR complication? (4) What patient, tumor, or implant characteristics are associated with improved survivorship free of revision in cemented DFRs used in patients treated initially for primary malignant or benign tumors? METHODS: This was a retrospective, comparative study using our institution's longitudinally-maintained database of 806 cemented endoprostheses starting in 1980 and assessed through December 31, 2018. In all, 365 DFRs were inserted during this time, but 14% (51 of 365) were placed for nonprimary bone tumors and 1% (5 of 365) were cementless reconstructions, leaving 309 cemented DFRs. Seventy-one percent (218 of 309) were primary implants and 29 percent (91 of 309) were revision implants (used to revise a prior DFR in all patients). During this time period, our strong bias was to use cemented stems and, thus, nearly all of our patients had cemented stems. Six percent (13 of 218) of primary DFRs were implanted more than 2 years before the study end; however, they lacked 2 years of follow-up data and, thus, were considered lost to follow-up, leaving 205 implants in the primary DFR analysis group. Only the first revision after primary DFR revision surgery was included in the revision cohort analysis. Thirty-two percent (29 of 91) of revision DFRs were second or more revision patients and were excluded, leaving 62 implants in the revision analysis group. Most patients in both groups were men (57% [117 of 205] for primary and 71% [44 of 62] for revision) who had been diagnosed with osteosarcoma (75% [153 of 205] and 73% [45 of 62] for primary and revision, respectively). The primary cohort had mean age of 26 ± 16 years with a mean follow-up of 136 ± 122 months, and the revision cohort had mean age of 31 ± 13 years (p = 0.02) with 141 ± 101 months of follow-up. Study endpoints included all-cause implant revision and cause-specific revision for soft tissue complications, aseptic loosening, structural complications (defined as periprosthetic or implant fracture), infection, or tumor progression. Planned surgery for implant lengthening procedures was excluded. Implant survivorship free from all-cause revision was calculated using a competing risk (cumulative incidence) estimator with death as a competing risk. A log-rank test using chi-square analysis was used to evaluate the differences in implant survivorship between primary DFRs and first revisions. The cause-specific incidences of implant revision were tabulated for primary and revision DFRs. Cox regression analysis investigated the odds of subsequent all-cause revision surgery for revision cemented DFRs based on the primary implant complication. A binary logistic regression analysis using age, gender, indication for revision, tumor type, infection, perioperative chemotherapy, and radiation was performed to identify factors associated with a second DFR reoperation. Relative effect sizes are reported as ORs. RESULTS: The revision DFR cohort had a shorter mean survival to all-cause revision than the primary cohort (mean 10 years [95% CI 7 to 12] versus 18 years [95% CI 15 to 20]; p < 0.001). The most common complications necessitating revision for revision implants were periprosthetic or implant fracture in 37% (23 of 62) and aseptic loosening in 15% (9 of 62), and the type of primary implant complication was not associated with risk of subsequent all-cause revision surgery for revision implants. Stem diameter less than 15 mm was associated with repeat all-cause revision in cemented revision DFRs after controlling for resection length, stem length, implant fabrication (custom or modular), and presence of a porous collar (OR 4 [95% CI 1 to 17]; p = 0.03). No other parameters that we explored, including patient age, gender, chemoradiation history, or primary tumor diagnosis, were associated with repeat revision surgery. CONCLUSION: Understanding modifiable factors that can improve revision DFR survival is critical to achieving long-term limb salvage for patients with tumors around the knee. Our data suggest that utilizing implants with the largest possible stems-or at a minimum increasing the stem size over the primary implant-is important to revision cemented DFR survivorship and is an important part of our revision practice. Improving revision implants' resistance to aseptic loosening through designs that resist torsion (a common mode of cemented fixation failure)-such as with the use of custom cross-pin fabrication-may be one method to improve survivorship. Another will be improved implant metallurgy that is resistant to fatigue fracture. Next steps may include understanding the optimal ratio of femoral diaphyseal width to implant diameter in patients where anatomic constraints preclude the insertion of cemented stems 15 mm or more in diameter. LEVEL OF EVIDENCE: Level IV, therapeutic study.

Is High-dose Radiation Therapy Associated With Early Revision Due to Aseptic Loosening in Patients With a Sarcoma of the Lower Extremities Reconstructed With a Cemented Endoprosthesis?

BACKGROUND: The durability of endoprostheses after limb salvage surgery is influenced by surgical factors (resection length, implant location, and residual bone quality), implant design (modular versus custom design, rotating versus fixed hinge, coating, collars, and the use of cross pins), and host factors (patient's immune status, activity levels, and age). In general, radiation therapy increases the risk of fractures, infection, delayed wound healing, and impaired osseointegration. Several studies have shown exposure to radiation to be associated with higher endoprosthesis revision rates and higher periprosthetic infection rates, but results are inconsistent. Although radiation therapy is not routinely used in the treatment of many bone sarcomas in current practice, it is still used in high doses after resection and prosthetic reconstruction in patients who have Ewing sarcoma with close or positive margins and in patients with soft tissue sarcoma. It is also used in varying doses after prosthetic reconstruction in patients with myeloma or bone metastasis after resection of periarticular destructive tumors. These patients may be at an increased risk of complications due to their radiation exposure, but this is a difficult question to study given the rarity of these diagnoses and poor overall survival of these patients. We therefore leveraged a large, longitudinally collected, 40-year endoprosthesis database that included patients who received radiation to the extremity for many bone and soft tissue sarcomas to investigate the association between preoperative or postoperative radiation therapy and endoprosthesis survival. QUESTIONS/PURPOSES: (1) Is receiving preoperative or postoperative radiation therapy in low or high doses for the treatment of bone or soft tissue malignancy of the lower extremities associated with decreased implant survivorship free from amputation or revision due to any cause? (2) Is receiving preoperative or postoperative radiation therapy in low or high doses for the treatment of bone or soft tissue malignancy of the lower extremities associated with decreased implant survivorship free from revision specifically due to aseptic loosening? (3) Is receiving preoperative or postoperative radiation therapy for the treatment of Ewing sarcoma of the femur specifically associated with decreased implant survivorship free from revision specifically due to aseptic loosening? METHODS: This was a retrospective, comparative study using our institution's database of 822 endoprostheses. Between 1980 and 2019, we treated 541 patients with primary cemented endoprostheses of the extremities. Of those patients, 8% (45 of 541) were excluded due to unknown radiation status, 3% (17 of 541) because of prior failed allograft, 15% (83 of 541) due to metastatic disease from a carcinoma, 1% (6 of 541) due to a nononcologic diagnosis, 4% (20 of 541) due to benign tumor diagnosis, 16% (87 of 541) due to upper extremity tumor location, 9% (49 of 541) due to not receiving chemotherapy, and 3% (14 of 541) due to expandable prostheses. Of the remaining 220 patients, 6% (13) were considered missing because they did not have 2 years of follow-up and did not reach a study endpoint. No patients had surgery within the last 2 years of the study end date. In all, 207 patients met inclusion criteria and were eligible for analysis. Patients who had received radiation to the lower extremities at any point in their treatment course were included in the radiation group and were compared with patients who did not receive radiation. For patients where radiation dose was available, the radiation group was subdivided into a low-dose (≤ 3000 cGy) and high-dose (> 3000 cGy) group. Revision surgery was defined as any surgery necessitating removal or replacement of the tibial or femoral stem. The complications necessitating revision or amputation were poor wound healing, aseptic loosening, implant breakage, deep infection, and tumor progression. The primary outcome of interest was implant survival free from revision or amputation due to any cause. The secondary outcome of interest was implant survival free from revision or amputation specifically due to aseptic loosening. The Kaplan-Meier survivorship curves were generated with implant survival free from revision or amputation as the endpoint and patient death as a competing risk. A log-rank test was used to identify differences in survivorship between the patients who received radiation and those who did not. Multivariate regression was used to identify factors associated with decreased implant survival. An odds ratio was used to determine relative effect size among the factors associated with decreased implant survival. RESULTS: The mean implant survival time for patients who did not receive radiation was 18.3 years (95% confidence interval [CI] 15.4 to 21.3) whereas the mean implant survival time for patients who received low- and high-dose radiation were 19.1 years (95% CI 14.5 to 23.7; p = 0.59) and 13.8 years (95% CI 8.2 to 19.5; p = 0.65), respectively. The mean implant survival free from revision for aseptic loosening for patients who did not receive radiation was 27.1 years (95% CI 24.1 to 30.1) whereas the mean implant survival for patients who received low- and high-dose radiation were 24.1 years (95% CI 19.1 to 29.1; p = 0.34) and 16.4 years (95% CI 10.6 to 22.2; p = 0.01), respectively. Patients who received high-dose radiation had decreased 5-year implant survivorship free from amputation or revision due to aseptic loosening (73% [95% CI 44% to 89%]) compared with patients who did not receive radiation (95% [95% CI 90% to 99%]; p = 0.01). For patients treated for Ewing sarcoma of the femur, the 5-year implant survival free from amputation or revision due to aseptic loosening for patients who did not receive radiation (100% [95% CI 100% to 100%]) was no different compared with patients who received radiation (71% [95% CI 35% to 90%]; p = 0.56). CONCLUSION: The results of this study may apply to scenarios where radiation is used, such as Ewing sarcoma with positive margins or local recurrence and after prosthetic reconstruction in patients with myeloma or bone metastasis after resection of periarticular destructive tumors. Surgeons may consider closer monitoring for early clinical and radiographic signs of aseptic loosening in patients who received high-dose radiation. These patients may also benefit from constructs that have increased resistance to aseptic loosening such as cross-pin or side plate fixation. The association between radiation and aseptic loosening should be further studied with larger studies with homogeneity in tumor diagnosis and prosthesis. The dose-dependent relationship between radiation and bone-related complications may also benefit from controlled, laboratory-based biomechanical studies. LEVEL OF EVIDENCE: Level III, therapeutic study.

Non-contrast free-breathing 3D cardiovascular magnetic resonance angiography using REACT (relaxation-enhanced angiography without contrast) compared to contrast-enhanced steady-state magnetic resonance angiography in complex pediatric congenital heart disease at 3T.

BACKGROUND: To evaluate the great vessels in young children with complex congenital heart disease (CHD) using non-contrast cardiovascular magnetic resonance angiography (CMRA) based on three-dimensional relaxation-enhanced angiography without contrast (REACT) in comparison to contrast-enhanced steady-state CMRA. METHODS: In this retrospective study from April to July 2021, respiratory- and electrocardiogram-gated native REACT CMRA was compared to contrast-enhanced single-phase steady-state CMRA in children with CHD who underwent CMRA at 3T under deep sedation. Vascular assessment included image quality (1 = non-diagnostic, 5 = excellent), vessel diameter, and diagnostic findings. For statistical analysis, paired t-test, Pearson correlation, Bland-Altman analysis, Wilcoxon test, and intraclass correlation coefficients (ICC) were applied. RESULTS: Thirty-six young children with complex CHD (median 4 years, interquartile range, 2-5; 20 males) were included. Native REACT CMRA was obtained successfully in all patients (mean scan time: 4:22 ± 1:44 min). For all vessels assessed, diameters correlated strongly between both methods (Pearson r = 0.99; bias = 0.04 ± 0.61 mm) with high interobserver reproducibility (ICC: 0.99 for both CMRAs). Native REACT CMRA demonstrated comparable overall image quality to contrast-enhanced CMRA (3.9 ± 1.0 vs. 3.8 ± 0.9, P = 0.018). With REACT CMRA, better image quality was obtained at the ascending aorta (4.8 ± 0.5 vs. 4.3 ± 0.8, P < 0.001), coronary roots (e.g., left: 4.1 ± 1.0 vs. 3.3 ± 1.1, P = 0.001), and inferior vena cava (4.6 ± 0.5 vs. 3.2 ± 0.8, P < 0.001). In all patients, additional vascular findings were assessed equally with native REACT CMRA and the contrast-enhanced reference standard (n = 6). CONCLUSION: In young children with complex CHD, REACT CMRA can provide gadolinium-free high image quality, accurate vascular measurements, and equivalent diagnostic quality compared to standard contrast-enhanced CMRA.

Neuregulin-1 beta 1 is implicated in pathogenesis of multiple sclerosis.

Multiple sclerosis is characterized by immune mediated neurodegeneration that results in progressive, life-long neurological and cognitive impairments. Yet, the endogenous mechanisms underlying multiple sclerosis pathophysiology are not fully understood. Here, we provide compelling evidence that associates dysregulation of neuregulin-1 beta 1 (Nrg-1ß1) with multiple sclerosis pathogenesis and progression. In the experimental autoimmune encephalomyelitis model of multiple sclerosis, we demonstrate that Nrg-1ß1 levels are abated within spinal cord lesions and peripherally in the plasma and spleen during presymptomatic, onset and progressive course of the disease. We demonstrate that plasma levels of Nrg-1ß1 are also significantly reduced in individuals with early multiple sclerosis and is positively associated with progression to relapsing-remitting multiple sclerosis. The functional impact of Nrg-1ß1 downregulation preceded disease onset and progression, and its systemic restoration was sufficient to delay experimental autoimmune encephalomyelitis symptoms and alleviate disease burden. Intriguingly, Nrg-1ß1 therapy exhibited a desirable and extended therapeutic time window of efficacy when administered prophylactically, symptomatically, acutely or chronically. Using in vivo and in vitro assessments, we identified that Nrg-1ß1 treatment mediates its beneficial effects in EAE by providing a more balanced immune response. Mechanistically, Nrg-1ß1 moderated monocyte infiltration at the blood-CNS interface by attenuating chondroitin sulphate proteoglycans and MMP9. Moreover, Nrg-1ß1 fostered a regulatory and reparative phenotype in macrophages, T helper type 1 (Th1) cells and microglia in the spinal cord lesions of EAE mice. Taken together, our new findings in multiple sclerosis and experimental autoimmune encephalomyelitis have uncovered a novel regulatory role for Nrg-1ß1 early in the disease course and suggest its potential as a specific therapeutic target to ameliorate disease progression and severity.

Recent insights on astrocyte mechanisms in CNS homeostasis, pathology, and repair.

Astrocytes play essential roles in development, homeostasis, injury, and repair of the central nervous system (CNS). Their development is tightly regulated by distinct spatial and temporal cues during embryogenesis and into adulthood throughout the CNS. Astrocytes have several important responsibilities such as regulating blood flow and permeability of the blood-CNS barrier, glucose metabolism and storage, synapse formation and function, and axon myelination. In CNS pathologies, astrocytes also play critical parts in both injury and repair mechanisms. Upon injury, they undergo a robust phenotypic shift known as "reactive astrogliosis," which results in both constructive and deleterious outcomes. Astrocyte activation and migration at the site of injury provides an early defense mechanism to minimize the extent of injury by enveloping the lesion area. However, astrogliosis also contributes to the inhibitory microenvironment of CNS injury and potentiate secondary injury mechanisms, such as inflammation, oxidative stress, and glutamate excitotoxicity, which facilitate neurodegeneration in CNS pathologies. Intriguingly, reactive astrocytes are increasingly a focus in current therapeutic strategies as their activation can be modulated toward a neuroprotective and reparative phenotype. This review will discuss recent advancements in knowledge regarding the development and role of astrocytes in the healthy and pathological CNS. We will also review how astrocytes have been genetically modified to optimize their reparative potential after injury, and how they may be transdifferentiated into neurons and oligodendrocytes to promote repair after CNS injury and neurodegeneration.

Free-breathing high resolution modified Dixon steady-state angiography with compressed sensing for the assessment of the thoracic vasculature in pediatric patients with congenital heart disease.

BACKGROUND: Cardiovascular magnetic resonance angiography (CMRA) is a non-invasive imaging modality of choice in pediatric patients with congenital heart disease (CHD). This study was aimed to evaluate the diagnostic utility of a respiratory- and electrocardiogram-gated steady-state CMRA with modified Dixon (mDixon) fat suppression technique and compressed sensing in comparison to standard first-pass CMRA in pediatric patients with CHD at 3 T. METHODS: In this retrospective single center study, pediatric CHD patients who underwent CMR with first-pass CMRA followed by mDixon steady-state CMRA at 3 T were analyzed. Image quality using a Likert scale from 5 (excellent) to 1 (non-diagnostic) and quality of fat suppression were assessed in consensus by two readers. Blood-to-tissue contrast and quantitative measurements of the thoracic vasculature were assessed separately by two readers. CMRA images were reevaluated by two readers for additional findings, which could be identified only on either one of the CMRA types. Paired Student t test, Wilcoxon test, and intraclass correlation coefficients (ICCs) were used for statistical analysis. RESULTS: 32 patients with CHD (3.3 ± 1.7 years, 13 female) were included. Overall image quality of steady-state mDixon CMRA was higher compared to first-pass CMRA (4.5 ± 0.5 vs. 3.3 ± 0.5; P < 0.001). Blood-to-tissue contrast ratio of steady-state mDixon CMRA was comparable to first-pass CMRA (7.85 ± 4.75 vs. 6.35 ± 2.23; P = 0.133). Fat suppression of steady-state mDixon CMRA was perfect in 30/32 (94%) cases. Vessel diameters were greater in first-pass CMRA compared to steady-state mDixon CMRA with the greatest differences at the level of pulmonary arteries and veins (e.g., right pulmonary artery for reader 1: 10.4 ± 2.4 vs. 9.9 ± 2.3 mm, P < 0.001). Interobserver agreement was higher for steady-state mDixon CMRA for all measurements compared to first-pass CMRA (ICCs > 0.92). In 9/32 (28%) patients, 10 additional findings were identified on mDixon steady-state CMRA (e.g., partial anomalous venous return, abnormalities of coronary arteries, subclavian artery stenosis), which were not depicted using first-pass CMRA. CONCLUSIONS: Steady-state mDixon CMRA offers a robust fat suppression, a high image quality, and diagnostic utility for the assessment of the thoracic vasculature in pediatric CHD patients.

Operative Versus Nonoperative Management of Acute Pediatric Monteggia Injuries With Complete Ulna Fractures.

BACKGROUND: The treatment of acute pediatric Monteggia injuries involving a complete fracture of the ulna remains controversial. The purpose of this study is to compare the outcomes of immediate operative fixation to a trial of closed reduction and casting of acute pediatric Monteggia fractures involving complete ulna fractures. METHODS: We performed a retrospective analysis of 73 patients with Monteggia injuries with complete ulna fractures presenting to 2 pediatric trauma centers from 2008 to 2018. Patients were divided in 2 groups based on the treatment received: patients in group 1 (n=37, 51%) received surgical treatment; patients in group 2 (n=36, 49%) received a trial of closed reduction and casting. The mean follow-up of 15.2 weeks (range, 4.1 to 159 wk). The incidence of radiocapitellar joint redislocation, need for further intervention, complications, and recovery of range of motion was compared between the groups. RESULTS: There were no significant differences between groups 1 and 2 with regards to age (6 vs. 5.8 y, P=0.69), sex (54% vs. 47% female, P=0.64), or the mean maximal ulnar angulation (23 vs. 19 degrees, P=0.94). There was a higher proportion of proximal ulna fractures in group 1 versus 2 (62% vs. 33%, respectively, P=0.02). Bado type III and IV fractures were associated with operative management [odds ratio=22 (95% confidence interval: 1.68-288.7) and 14.9 (95% confidence interval: 2.09-106), respectively]. In group 2, 5 patients (13.9%) sustained a loss of radiocapitellar joint reduction following closed reduction and casting and ultimately received operative treatment. At final follow-up, there were no cases of recurrent radiocapitellar dislocation in either group, all patients achieved fracture union and regained full elbow range of motion. CONCLUSIONS: Even in the presence of a complete ulna fracture, a trial of nonoperative management of acute pediatric Monteggia fractures with closed reduction and casting can result in comparable outcomes to those obtained with immediate surgical management. The nonoperative management of Monteggia fractures requires close clinical follow-up to ensure no loss of reduction. LEVEL OF EVIDENCE: Level IV-therapeutic studies, case series.

Antisense Oligonucleotides Targeting Jagged 1 Reduce House Dust Mite-induced Goblet Cell Metaplasia in the Adult Murine Lung.

Goblet cell metaplasia, excessive mucus production, and inadequate mucus clearance accompany and exacerbate multiple chronic respiratory disorders, such as asthma and chronic obstructive pulmonary disease. Notch signaling plays a central role in controlling the fate of multiple cell types in the lung, including goblet cells. In the present study, we explored the therapeutic potential of modulating the Notch pathway in the adult murine lung using chemically modified antisense oligonucleotides (ASOs). To this end, we designed and characterized ASOs targeting the Notch receptors Notch1, Notch2, and Notch3 and the Notch ligands Jag1 (Jagged 1) and Jag2 (Jagged 2). Pulmonary delivery of ASOs in healthy mice or mice exposed to house dust mite, a commonly used mouse model of asthma, resulted in a significant reduction of the respective mRNAs in the lung. Furthermore, ASO-mediated knockdown of Jag1 or Notch2 in the lungs of healthy adult mice led to the downregulation of the club cell marker Scgb1a1 and the concomitant upregulation of the ciliated cell marker FoxJ1 (forkhead box J1). Similarly, ASO-mediated knockdown of Jag1 or Notch2 in the house dust mite disease model led to reduced goblet cell metaplasia and decreased mucus production. Because goblet cell metaplasia and excessive mucus secretion are a common basis for many lung pathologies, we propose that ASO-mediated inhibition of JAG1 could provide a novel therapeutic path for the treatment of multiple chronic respiratory diseases.

The Ecology of Individual Differences Empirically Applied to Space-Use and Movement Tactics.

Movement provides a link between individual behavioral ecology and the spatial and temporal variation in an individual's landscape. Individual variation in movement traits is an important axis of animal personality, particularly in the context of foraging ecology. We tested whether individual caribou (Rangifer tarandus) displayed plasticity in movement and space-use behavior across a gradient of resource aggregation. We quantified first-passage time and range-use ratio as proxies for movement-related foraging behavior and examined how these traits varied at the individual level across a foraging resource gradient. Our results suggest that individuals adjusted first-passage time but not range-use ratio to maximize access to high-quality foraging resources. First-passage time was repeatable, and intercepts for first-passage time and range-use ratio were negatively correlated. Individuals matched first-passage time but not range-use ratio to the expectations of our patch-use model that maximized access to foraging resources, a result that suggests that individuals acclimated their movement patterns to accommodate both intra- and interannual variation in foraging resources on the landscape. Collectively, we highlight repeatable movement and space-use tactics and provide insight into how individual plasticity in movement interacts with landscape processes to affect the distribution of behavioral phenotypes and potentially fitness and population dynamics.

Local and Intra-articular Administration of Nonsteroidal Anti-inflammatory Drugs for Pain Management in Orthopedic Surgery.

BACKGROUND: Although growing evidence demonstrates the benefits of locally administered nonsteroidal anti-inflammatory drugs (NSAIDs) for postoperative pain management, there is ongoing debate regarding NSAID use in orthopedic surgery. AREAS OF UNCERTAINTY: Current data largely support a local site of NSAID action and suggest that effective pain control can be achieved with delivery of NSAIDs intra-articularly (IA) and/or locally at the site of injury, where they can block peripheral production of inflammatory mediators and may desensitize nociceptors. Improvements in postoperative pain control with locally administered NSAIDs have been widely reported in the total joint arthroplasty literature and may offer benefits in patient's undergoing arthroscopic procedures and those with osteoarthritis as well. The purpose of this review is to examine the available evidence in the literature regarding the efficacy and safety profile of the use of local and IA NSAIDs in orthopedic surgery. DATA SOURCES: Narrative literature review using keywords, expert opinion, either during or from live conference. THERAPEUTIC ADVANCES: Local and IA administration of NSAIDs for pain management in orthopedic surgery. CONCLUSION: There is convincing evidence that NSAIDs administered locally in and around the joint reduce postoperative pain scores and opioid consumption in patients undergoing total joint arthroplasty, yet further research is required regarding the risks of potential chondrotoxicity and the inhibition of bone and soft-tissue healing with locally administered NSAIDs.

A modular analysis of microglia gene expression, insights into the aged phenotype.

BACKGROUND: Microglia are multifunctional cells that are key players in brain development and homeostasis. Recent years have seen tremendous growth in our understanding of the role microglia play in neurodegeneration, CNS injury, and developmental disorders. Given that microglia show diverse functional phenotypes, there is a need for more precise tools to characterize microglial states. Here, we experimentally define gene modules as the foundation for describing microglial functional states. RESULTS: In an effort to develop a comprehensive classification scheme, we profiled transcriptomes of mouse microglia in a stimulus panel with 96 different conditions. Using the transcriptomic data, we generated fine-resolution gene modules that are robustly preserved across datasets. These modules served as the basis for a combinatorial code that we then used to characterize microglial activation under various inflammatory stimulus conditions. CONCLUSIONS: The microglial gene modules described here were robustly preserved, and could be applied to in vivo as well as in vitro conditions to dissociate the signaling pathways that distinguish acutely inflamed microglia from aged microglia. The microglial gene modules presented here are a novel resource for classifying and characterizing microglial states in health and disease.

Specific Tumor Characteristics Predict Upstaging in Early-Stage Esophageal Cancer.

BACKGROUND: Early-stage esophageal cancer (stages 0-1) has been shown to have relatively good outcomes after local endoscopic or surgical resection. For this reason, neoadjuvant chemoradiation usually is reserved for higher-stage disease. Some early tumors, however, are found after resection to be more advanced than predicted based on initial clinical staging, termed pathologic upstaging. Such tumors may have benefited from alternate treatment models had their true stage been known preoperatively. This study aimed to identify high-risk features in early esophageal cancers that might predict tumor upstaging and guide more individualized treatment algorithms. METHODS: Through retrospective review of a single-institution foregut disease registry, we evaluated patients who underwent esophagectomy for high-grade dysplasia (Tis) or stage 1 esophageal cancer, searching for factors associated with pathologic upstaging. RESULTS: The review included 110 patients (88% male, median age at diagnosis, 64.5 years) treated between January 2000 and June 2016. Upstaging occurred for 20.9% of the patients, and was more common for patients with angiolymphatic invasion (odds ratio [OR], 11.07; 95% confidence interval [CI], 2.96-41.44; P < 0.001) or signet-ring features (OR, 23.9; 95% CI, 2.6-216.8; P = 0.005). In the absence of other predictors, upstaging was associated with decreased overall survival (P = 0.006). CONCLUSIONS: Approximately 20% of patients with early-stage esophageal cancer may be upstaged at resection. Angiolymphatic invasion and signet-ring features may predict tumors likely to be upstaged, resulting in decreased overall survival.

Viable RNaseH1 knockout mice show RNaseH1 is essential for R loop processing, mitochondrial and liver function.

Viable constitutive and tamoxifen inducible liver-specific RNase H1 knockout mice that expressed no RNase H1 activity in hepatocytes showed increased R-loop levels and reduced mitochondrial encoded DNA and mRNA levels, suggesting impaired mitochondrial R-loop processing, transcription and mitochondrial DNA replication. These changes resulted in mitochondrial dysfunction with marked changes in mitochondrial fusion, fission, morphology and transcriptional changes reflective of mitochondrial damage and stress. Liver degeneration ensued, as indicated by apoptosis, fibrosis and increased transaminase levels. Antisense oligonucleotides (ASOs) designed to serve as substrates for RNase H1 were inactive in the hepatocytes from the RNase H1 knockout mice and in vivo, demonstrating that RNase H1 is necessary for the activity of DNA-like ASOs. During liver regeneration, a clone of hepatocytes that expressed RNase H1 developed and partially restored mitochondrial and liver function.

Hepatotoxicity of high affinity gapmer antisense oligonucleotides is mediated by RNase H1 dependent promiscuous reduction of very long pre-mRNA transcripts.

High affinity antisense oligonucleotides (ASOs) containing bicylic modifications (BNA) such as locked nucleic acid (LNA) designed to induce target RNA cleavage have been shown to have enhanced potency along with a higher propensity to cause hepatotoxicity. In order to understand the mechanism of this hepatotoxicity, transcriptional profiles were collected from the livers of mice treated with a panel of highly efficacious hepatotoxic or non-hepatotoxic LNA ASOs. We observed highly selective transcript knockdown in mice treated with non-hepatotoxic LNA ASOs, while the levels of many unintended transcripts were reduced in mice treated with hepatotoxic LNA ASOs. This transcriptional signature was concurrent with on-target RNA reduction and preceded transaminitis. Remarkably, the mRNA transcripts commonly reduced by toxic LNA ASOs were generally not strongly associated with any particular biological process, cellular component or functional group. However, they tended to have much longer pre-mRNA transcripts. We also demonstrate that the off-target RNA knockdown and hepatotoxicity is attenuated by RNase H1 knockdown, and that this effect can be generalized to high affinity modifications beyond LNA. This suggests that for a certain set of ASOs containing high affinity modifications such as LNA, hepatotoxicity can occur as a result of unintended off-target RNase H1 dependent RNA degradation.

Stoichiometric distribution models: ecological stoichiometry at the landscape extent.

Human activities are altering the fundamental geography of biogeochemicals. Yet we lack an understanding of how the spatial patterns in organismal stoichiometry affect biogeochemical processes and the tools to predict the impacts of global changes on biogeochemical processes. In this contribution we develop stoichiometric distribution models (StDMs), which allow us to map spatial structure in resource elemental composition across a landscape and evaluate spatial responses of consumers. We parameterise StDMs for a consumer-resource (moose-white birch) system and demonstrate that we can develop predictive models of resource stoichiometry across a landscape and that such models could improve our predictions of consumer space use. With results from our study system application, we argue that explicit consideration of the spatial patterns in organismal elemental composition may uncover emergent individual, population, community and ecosystem properties that are not revealed at the local extents routinely used in ecological stoichiometry. We discuss perspectives for further developments and application of StDMs to advance three emerging frameworks for spatial ecosystem ecology in an era of global change; meta-ecosystem theory, macroecological stoichiometry and remotely sensed biogeochemistry. Progress on these emerging frameworks will allow for the integration of ecological stoichiometry and individual space use and fitness.

The Cell Wall Arabinose-Deficient Arabidopsis thaliana Mutant murus5 Encodes a Defective Allele of REVERSIBLY GLYCOSYLATED POLYPEPTIDE2.

Traditional marker-based mapping and next-generation sequencing was used to determine that the Arabidopsis (Arabidopsis thaliana) low cell wall arabinose mutant murus5 (mur5) encodes a defective allele of REVERSIBLY GLYCOSYLATED POLYPEPTIDE2 (RGP2). Marker analysis of 13 F2 confirmed mutant progeny from a recombinant mapping population gave a rough map position on the upper arm of chromosome 5, and deep sequencing of DNA from these 13 lines gave five candidate genes with G→A (C→T) transitions predicted to result in amino acid changes. Of these five, only insertional mutant alleles of RGP2, a gene that encodes a UDP-arabinose mutase that interconverts UDP-arabinopyranose and UDP-arabinofuranose, exhibited the low cell wall arabinose phenotype. The identities of mur5 and two SALK insertional alleles were confirmed by allelism tests and overexpression of wild-type RGP2 complementary DNA placed under the control of the 35S promoter in the three alleles. The mur5 mutation results in the conversion of cysteine-257 to tyrosine-257 within a conserved hydrophobic cluster predicted to be distal to the active site and essential for protein stability and possible heterodimerization with other isoforms of RGP.