Findings Associated With Nonaccidental Trauma in Children With Isolated Femoral Diaphyseal Fractures.

BACKGROUND: Pediatric patients with isolated femoral diaphyseal fractures are difficult to assess for nonaccidental trauma (NAT). The purpose of this study was to determine (1) if there are any demographic features of isolated femoral diaphyseal fractures associated with suspected NAT and (2) if there are clinical signs associated with isolated femoral diaphyseal fractures associated with suspected NAT. METHODS: All patients with femoral diaphyseal fractures from January 2010 to June 2018 were reviewed. We included patients younger than 4 years old with isolated femoral diaphyseal fractures. We excluded patients 4 years old and older, polytraumas, motor vehicle collisions, and patients with altered bone biology. Diagnosis of suspected NAT was determined by review of a documented social work assessment. We recorded fracture characteristics including location along femur as well as fracture pattern and presence of associated findings on NAT workup including the presence of retinal hemorrhage, subdural hematoma, evidence of prior fracture, or cutaneous lesions. Sensitivity, specificity, positive predictive value (PPV), and negative predictive value (NPV) of these associated findings were calculated. RESULTS: Totally, 144 patients met the inclusion criteria. Social work was consulted on 50 patients (35%). Suspected NAT was diagnosed in 27 patients (19%). The average age of patients with suspected NAT was 0.82 and 2.25 years in patients without NAT (P<0.01). The rate and type of skin lesions present on exam were not different between the 2 groups. Patients with suspected NAT had no findings of retinal hemorrhage or subdural hematoma, but 5 of 27 patients (19%) had evidence of prior fracture on skeletal survey. The sensitivities of retinal hemorrhage, subdural, and skeletal survey were 0%, 0%, and 19% and the specificities of all were 100%. The NPVs were 39%, 27%, and 63%, respectively. The PPV of skeletal survey was 100%. Since there were no patients in this study with positive findings of retinal hemorrhage or subdural hematoma, the PPV for these could not be assessed. CONCLUSIONS: In the current study, signs of NAT such as skin lesions, retinal hemorrhage, subdural hematoma, and evidence of prior fracture on skeletal survey may not be helpful to diagnosis suspected NAT in patients with an isolated femoral diaphyseal fracture. LEVEL OF EVIDENCE: Level III-diagnostic study.

Prediction of CYP Down Regulation after Tusamitamab Ravtansine Administration (a DM4-Conjugate), Based on an In Vitro-In Vivo Extrapolation Approach.

Tusamitamab ravtansine is an antibody-drug conjugate (ADC) composed of a humanized monoclonal antibody (IgG1) and DM4 payload. Even if DM4 and its main metabolite methyl-DM4 (Me-DM4) circulate at low concentrations after ADC administration, their potential as perpetrators of cytochrome P450 mediated drug-drug interaction was assessed. In vitro studies in human hepatocytes indicated that Me-DM4 elicited a clear concentration-dependent down regulation of cytochrome P450 enzymes (CYP3A4, 1A2, and 2B6). Because DM4 was unstable under the incubation conditions studied, the in vitro constants could not be determined for this entity. Thus, to predict the clinical relevance of this observed downregulation, an in vitro-in vivo extrapolation (IVIVE) pharmacokinetic (PK) based approach was developed. To mitigate model prediction errors and because of their similar inhibitory effect on tubulin polymerization, the same downregulation constants were used for DM4 and Me-DM4. This approach describes the time course of decreasing CYP3A4, 1A2, and 2B6 enzyme amounts as a function of circulating concentrations of DM4 and Me-DM4 predicted from a population PK model. The developed IVIVE-PK model showed that the highest CYP abundance decrease was observed for CYP3A4, with a transient reduction of < 10% from baseline. The impact on midazolam exposure, as probe substrate of CYP3A, was then simulated based on a physiologically-based PK static method. The maximal CYP3A4 abundance reduction was associated with a predicted midazolam area under the curve (AUC) ratio of 1.14. To conclude, the observed in vitro downregulation of CYPs by Me-DM4 is not expected to have relevant clinical impact.

BCG vaccination impacts the epigenetic landscape of progenitor cells in human bone marrow.

While the Bacille-Calmette-Guérin (BCG) vaccine is used to prevent tuberculosis, it also offers protection against a diverse range of non-mycobacterial infections. However, the underlying protective mechanisms in humans are not yet fully understood. Here, we surveyed at single-cell resolution the gene expression and chromatin landscape of human bone marrow, aspirated before and 90 days after BCG vaccination or placebo administration. We show that BCG vaccination significantly alters both the gene expression and epigenetic profiles of human hematopoietic stem and progenitor cells (HSPCs). Changes in gene expression occur primarily on the most uncommitted stem cells and are reflective of a persistent myeloid bias. In contrast, BCG-induced changes in chromatin accessibility are most prevalent within differentiated progenitor cells at sites influenced by Kruppel-like factor (KLF)/SP and EGR transcription factors (TFs). These TFs are also activated in the most uncommitted stem cells, indicating that activated TFs, which drive persistent changes in HSC gene expression, likely also drive chromatin dynamics appearing within downstream progenitor cells. This perspective contests the prevailing notion that epigenetic modifications linked to innate immune memory transfer directly from stem cells to their differentiated derivatives. Finally, we show that alterations in gene expression and chromatin accessibility in HSPCs due to BCG vaccination were highly correlated (r>0.8) with the IL-1ß secretion capacity of paired PBMCs upon secondary immune challenge. Overall, our findings shed light on BCG vaccination's profound and lasting effects on HSPCs and its influence on innate immune responses.

Interdisciplinary Orthopedic Management of Pediatric Patella Dislocation and Instability: An Educational Case Series.

Pediatric patellar instability and/or dislocation is a challenging diagnosis category that requires an interdisciplinary team consisting of orthopedic surgeons and physical therapists for optimized patient outcomes. This educational case series outlines core concepts for three patients with unique patellar dislocation presentations. Case 1 is a 16-year-old male who presented with a history of five knee traumatic patellar dislocations with self-reduction and underwent knee arthroscopic surgery with debridement and microfracture of the patella chondral defect, arthroscopic lateral release to improve the patellar tilt, and medial patellofemoral ligament (MPFL) reconstruction. Case 2 is a 15-year-old female who presented with chronic knee pain and patella instability who underwent knee arthroscopic surgery with abrasion arthroplasty, microfracture of the patella, lateral release, tibial tubercle osteotomy medializing osteotomy, and MPFL reconstruction. Case 3 is a 14-year-old male who presented after a single episode of lateral patella dislocation and underwent open reduction and fixation of the lateral femoral condyle osteochondral fracture, a Grammont patellar medialization procedure, and MPFL reconstruction. All three patients received postoperative physical therapy (PT) to improve function and outcomes. These cases represent important concepts of patellar containment, risk factors for recurrent instability, associated pathology, and appropriate surgical care and postoperative rehabilitation. Furthermore, this case series highlights management decisions and pathways for three patients with different symptoms related to patellar instability, subsequent surgical correction, and postoperative physical therapy. Overall, interdisciplinary care of common pediatric orthopedic conditions can help improve patient outcomes and satisfaction. By understanding the biomechanics and decision-making surgical parameters regarding patellofemoral instability, clinicians can provide patients with better care.

Assessment and Validation of Anterior Humeral Line for Diagnosing Supracondylar Humerus Fractures in the Pediatric Population.

INTRODUCTION: Pediatric supracondylar humerus fractures are commonly evaluated using the anterior humeral line (AHL) on a lateral radiograph. Rotational variations in radiographic projection are common due to child discomfort and could lead to changes in management based on where the AHL intersects the capitellum. The purpose of this study was to establish whether rotational variations in elbow rotation leads to significant changes in AHL position and whether drawing the AHL based on the distal humerus versus shaft is more tolerant to rotation. METHODS: Fifty children with nonoperative supracondylar humerus fractures were identified with sub optimally positioned injury and well positioned follow-up lateral radiographs. The proportion of the bone anterior to the intersection of the AHL and the capitellum was measured using the humeral shaft versus distal humerus to guide position of the AHL. This process was repeated on ten pediatric humerus dry cadaveric specimens which were imaged in 5-degree rotational increments along the axis of the humeral shaft from -20 to +20 degrees. RESULTS: AHL position correlated poorly when measured on rotated lateral radiographs of clinical patients versus non-rotated lateral radiographs when using the distal humerus as a guide (intraclass correlation coefficient 0.14), compared with when using the humeral shaft as a guide (intraclass correlation coefficient 0.81). When assessing the pediatric humerus dry cadavers between the 2 techniques, there was greater statistically significant variation in rotated positions compared with the neutral position in the distal humerus AHL measurement approach compared with the humeral shaft AHL measurement approach, with the mean AHL within the central third of the capitellum for more rotational positions when using the shaft compared with the distal humerus. CONCLUSIONS: With rotated lateral elbow radiographs in supracondylar humerus fractures, utilizing the humeral shaft provides more consistent AHL measurements than utilizing the distal humerus, and thus drawing the line starting at the shaft of the humerus is recommended for surgical decision making.

Antigen-driven colonic inflammation is associated with development of dysplasia in primary sclerosing cholangitis.

Primary sclerosing cholangitis (PSC) is an immune-mediated disease of the bile ducts that co-occurs with inflammatory bowel disease (IBD) in almost 90% of cases. Colorectal cancer is a major complication of patients with PSC and IBD, and these patients are at a much greater risk compared to patients with IBD without concomitant PSC. Combining flow cytometry, bulk and single-cell transcriptomics, and T and B cell receptor repertoire analysis of right colon tissue from 65 patients with PSC, 108 patients with IBD and 48 healthy individuals we identified a unique adaptive inflammatory transcriptional signature associated with greater risk and shorter time to dysplasia in patients with PSC. This inflammatory signature is characterized by antigen-driven interleukin-17A (IL-17A)+ forkhead box P3 (FOXP3)+ CD4 T cells that express a pathogenic IL-17 signature, as well as an expansion of IgG-secreting plasma cells. These results suggest that the mechanisms that drive the emergence of dysplasia in PSC and IBD are distinct and provide molecular insights that could guide prevention of colorectal cancer in individuals with PSC.

Reply to Barton et al: signatures of natural selection during the Black Death.

Barton et al.1 raise several statistical concerns regarding our original analyses2 that highlight the challenge of inferring natural selection using ancient genomic data. We show here that these concerns have limited impact on our original conclusions. Specifically, we recover the same signature of enrichment for high FST values at the immune loci relative to putatively neutral sites after switching the allele frequency estimation method to a maximum likelihood approach, filtering to only consider known human variants, and down-sampling our data to the same mean coverage across sites. Furthermore, using permutations, we show that the rs2549794 variant near ERAP2 continues to emerge as the strongest candidate for selection (p = 1.2×10-5), falling below the Bonferroni-corrected significance threshold recommended by Barton et al. Importantly, the evidence for selection on ERAP2 is further supported by functional data demonstrating the impact of the ERAP2 genotype on the immune response to Y. pestis and by epidemiological data from an independent group showing that the putatively selected allele during the Black Death protects against severe respiratory infection in contemporary populations.

Evolution of immune genes is associated with the Black Death.

Infectious diseases are among the strongest selective pressures driving human evolution1,2. This includes the single greatest mortality event in recorded history, the first outbreak of the second pandemic of plague, commonly called the Black Death, which was caused by the bacterium Yersinia pestis3. This pandemic devastated Afro-Eurasia, killing up to 30-50% of the population4. To identify loci that may have been under selection during the Black Death, we characterized genetic variation around immune-related genes from 206 ancient DNA extracts, stemming from two different European populations before, during and after the Black Death. Immune loci are strongly enriched for highly differentiated sites relative to a set of non-immune loci, suggesting positive selection. We identify 245 variants that are highly differentiated within the London dataset, four of which were replicated in an independent cohort from Denmark, and represent the strongest candidates for positive selection. The selected allele for one of these variants, rs2549794, is associated with the production of a full-length (versus truncated) ERAP2 transcript, variation in cytokine response to Y. pestis and increased ability to control intracellular Y. pestis in macrophages. Finally, we show that protective variants overlap with alleles that are today associated with increased susceptibility to autoimmune diseases, providing empirical evidence for the role played by past pandemics in shaping present-day susceptibility to disease.

When Will You Succeed Casting Patients With Early-onset Scoliosis? Prospective Evaluation of Predictive Radiographic Parameters.

BACKGROUND: In a recent retrospective study, in cast correction of the major curve correlated with final curve size in patients with early-onset scoliosis treated with casting. We therefore sought to perform a prospective study with controlled methodology to determine if there are parameters associated with reduction of coronal deformity. METHODS: A prospective, observational study was conducted between 2014 and 2019 at selected sites willing to comply with a standard radiographic and follow-up protocol. Radiographic data was collected at time points of precast, in traction, initial in-cast, and at minimum 1 year follow-up. Multivariate linear regression models were utilized to control for potential confounders using a stepwise procedure. Twenty-nine patients met inclusion criteria. RESULTS: On multivariate analysis, traction major curve (P=0.043) and initial in-cast (P=0.011) major curve Cobb angles were independently associated with final out of cast major curve Cobb angle. The only factor that was independently associated with failure to cure (<15-degree major curve) was traction major curve Cobb angle (P=0.046). A threshold traction major curve Cobb angle of 20 degrees was found to have good accuracy with 81% sensitivity and 73% specificity (receiver operator curve area: 0.869, P<0.001). A traction major curve Cobb angle over 20 degrees would accurately predict failure of casting treatment to cure scoliosis in 79% of cases. A threshold in-cast major curve Cobb angle of 21 degrees was found to have slightly less accuracy than traction with 69% sensitivity, 82% specificity, and 74% accuracy (receiver operator curve area: 0.830, P=0.004). CONCLUSIONS: Radiographic measurements in traction and initially in the cast are predictive of curve size at follow-up for children with early-onset scoliosis treated with casting. The standardization and utility of traction films should be further explored. LEVEL OF EVIDENCE: Level II.

Use of Vancomycin Powder in Spinal Deformity Surgery in Cerebral Palsy Patients is Associated With Proteus Surgical Site Infections.

PURPOSE: Surgical site infection (SSI) rates in pediatric spinal deformity surgery for cerebral palsy (CP) patients are higher than that in idiopathic scoliosis. The use of vancomycin powder is associated with decreased risk of SSI in neuromuscular patients. Prior studies in adult and pediatric early-onset scoliosis patients have shown that vancomycin powder alters microbacterial profile in patients that develop SSI. However, the effects of topical vancomycin powder on microbiology in spinal deformity surgery for CP patients has not been studied. METHODS: An international multicenter database of CP neuromuscular scoliosis patients was used in this retrospective cohort study. All patients that underwent posterior spinal instrumented fusion for CP neuromuscular scoliosis from 2008 to 2019 were queried, and 50 cases complicated by postoperative SSI were identified. Intraoperative antibiotic details were documented in 49 cases (98.0%). Microbiology details were documented in 45 cases (91.8%). Microbiology for patients that received topical vancomycin powder were compared with patients that did not. A multivariate regression model was used to control for potential confounders. RESULTS: There were 45 patients included in this study. There were 27 males (60.0%) and 18 females (40.0%). Mean age at surgery was 14.8±2.4 years. There were 24 patients that received topical vancomycin powder (53.3%). The mean time from index surgery to SSI was 4.3±11.3 months.On univariate analysis of microbiology cultures by vancomycin powder cohort, there were no significant differences in culture types. Proteus spp. trended on significance with association with vancomycin powder use (P=0.078). When controlling for potential confounders on multivariate analysis, intraoperative topical vancomycin powder was associated with increased risk for proteus infection (adjusted odds ratio: 262.900, 95% confidence interval: 1.806-38,267.121, P=0.028). DISCUSSION: In CP patients undergoing pediatric spinal deformity surgery, the use of vancomycin powder was independently associated with increased risk for proteus infections. Further study into antibiotic regimens for spinal deformity surgery in the CP population should be performed. LEVEL OF EVIDENCE: Level III-retrospective cohort study.

Surgical site infections in early onset scoliosis: what are long-term outcomes in patients with traditional growing rods?

PURPOSE: Deep surgical site infections (SSIs) are a common and potentially severe complication in early onset scoliosis (EOS) patients. We sought to identify the long-term outcomes following SSI, specific risk factors associated with recurrent infections, and if instrument retention is a prudent SSI management strategy in EOS. METHODS: We performed a retrospective review of all EOS patients who underwent traditional growing rod spine procedures from 2003 to 2017. Infections were categorized as single or multiple SSIs. All infections were treated with operative irrigation and debridement (I&D) as well as antibiotics. Univariate analysis was performed using chi-square and ANOVA tests to assess differing factors between patients with single versus multiple infections. RESULTS: Eighty-one patients underwent 638 growth-friendly traditional growing rod procedures. There were 21 patients (26%) who developed a total of 27 SSIs (4.2% SSI per procedure). Fifteen patients had a single infection and six patients had multiple infections. Demographics were not significantly different between these two groups. Patients with multiple infections had a significant difference in the number of procedures after initial infection (p value = 0.025) and positive preoperative nasal Staphylococcus aureus screen (p value = 0.0021) when compared to those with a single SSI. Of note, these results were not available at the time of pre-operative antibiotic selection. All 21 patients had resolution of their SSIs. Twenty patients reached final instrumented fusion. Two patients, both of whom had multiple infections, underwent complete removal of instrumentation. Reasons included one each, parental request resulting in termination of treatment and infection > 7 years after final fusion. CONCLUSION: Most patients who develop SSIs during growing spine treatment are able to remain instrumented. Risk factors associated with developing multiple SSIs include infection earlier in the course of growing spine surgery, a resultant higher number of procedures following the initial infection and having a positive preoperative nasal Staphylococcus aureus screen. LEVEL OF EVIDENCE: IV.

Alveolar macrophages from persons living with HIV show impaired epigenetic response to Mycobacterium tuberculosis.

Persons living with HIV (PLWH) are at increased risk of tuberculosis (TB). HIV-associated TB is often the result of recent infection with Mycobacterium tuberculosis (M. tuberculosis) followed by rapid progression to disease. Alveolar macrophages (AMs) are the first cells of the innate immune system that engage M. tuberculosis, but how HIV and antiretroviral therapy (ART) affect the anti-mycobacterial response of AMs is not known. To investigate the impact of HIV and ART on the transcriptomic and epigenetic response of AMs to M. tuberculosis, we obtained AMs by bronchoalveolar lavage from 20 PLWH receiving ART, 16 control subjects who were HIV-free (HC), and 14 subjects who received ART as preexposure prophylaxis (PrEP) to prevent HIV infection. Following in vitro challenge with M. tuberculosis, AMs from each group displayed overlapping but distinct profiles of significantly up- and downregulated genes in response to M. tuberculosis. Comparatively, AMs isolated from both PLWH and PrEP subjects presented a substantially weaker transcriptional response. In addition, AMs from HC subjects challenged with M. tuberculosis responded with pronounced chromatin accessibility changes while AMs obtained from PLWH and PrEP subjects displayed no significant changes in their chromatin state. Collectively, these results revealed a stronger adverse effect of ART than HIV on the epigenetic landscape and transcriptional responsiveness of AMs.

Postoperative Correction in Idiopathic Scoliosis: Which Preoperative Imaging Technique Is Most Predictive?

INTRODUCTION: Preoperative radiographic assessment of curve flexibility in patients with idiopathic scoliosis is important to determine Lenke classification, operative levels, and potential postoperative correction. However, no consensus exists regarding the optimal technique. We compared measurements from supine side bending (SB) and intraoperative traction radiographs under general anesthesia (TUGA) with actual postoperative correction followed for 1 year. METHODS: We identified 235 patients with idiopathic scoliosis who underwent posterior spinal fusion with pedicle screw instrumentation between 2010 and 2018 who had preoperative and postoperative radiographs including standing posterior-anterior (PA) and lateral radiographs, preoperative SB radiographs, and TUGA radiographs. Curves were categorized into proximal thoracic, main thoracic/thoracolumbar (MT), and distal thoracolumbar/lumbar (TL/L) curves. Flexibility was calculated from SB and TUGA radiographs. Correction rates were calculated from 1 month and 1 year radiographs postoperatively. Bending radiographs that correlated significantly with postoperative correction with P<0.10 were eligible for inclusion. Preoperative demographics, etiology, deformity details, and surgical details were included in the multivariate models. RESULTS: On univariate analysis, TUGA radiographs correlated with postoperative correction at 1 month and 1 year on MT curves (r=0.214, P=0.001; r=0.209, P=0.001) and TL/L curves (r=0.280, P<0.001; r=0.181, P=0.006). Supine SB radiographs did not correlate with postoperative correction on either MT or T/TL curves. On multivariate analysis, major curve TUGA radiographs were independently associated with postoperative MT curve correction at 1 month (beta: 0.158, 95% confidence interval: 0.035-0.280, P=0.012) and 1 year (beta: 0.195, 95% confidence interval: 0.049-0.340, P=0.009). MT curve SB radiographs were not associated with postoperative major curve correction at 1 month (P=0.088). CONCLUSIONS: TUGA radiographs independently correlated with postoperative main thoracic and distal thoracolumbar/lumbar curve correction at 1 month and 1 year postoperatively. SB radiographs independently correlated only with TL/L curve correction at 1 year postoperatively. However, this correlation was not as strong as TUGA correction (beta of 0.280 vs. beta of 0.092). TUGA radiographs appear superior to SB radiographs at predicting curve correction after surgery. LEVEL OF EVIDENCE: Level III.

Use of Vancomycin Powder in the Surgical Treatment of Early Onset Scoliosis Is Associated With Different Microbiology Cultures After Surgical Site Infection.

BACKGROUND: The use of vancomycin powder has been shown to decrease risk of surgical site infection (SSI) in early onset scoliosis (EOS). While there is potential benefit in SSI reduction, there is also theoretical risk in creating increased bacterial resistance to standard treatment regimens. However, the effects of topical vancomycin powder on microbiology in these patients has not been studied. METHODS: A multicenter database for EOS patients was retrospectively analyzed. All patients that underwent surgical treatment with traditional growing rods, magnetically controlled growing rods, vertical expandable prosthetic titanium rib, and Shilla for EOS performed after 2010 were identified (n=1115). Patients that sustained at least 1 SSI after guided growth surgery were assessed (n=104, 9.3%). Patients with culture and antibiotic details were included (n=55). Patients that received vancomycin powder at index surgery were compared with patients that did not. A multivariate regression model was used to control for potential confounders. RESULTS: There were 55 patients included in this study, including 26 males (47%) and 29 females (53%). Mean age at index surgery was 7.2±6.9 years. Vancomycin powder was utilized in 18 cases (33%). Mean time from index surgery to SSI was 2.0±1.3 years. There were 2 cases of wound dehiscence (4%), 7 cases of superficial infection (13%), and 46 cases of deep infection (84%).There were significant differences in overall microbiology results between vancomycin and no vancomycin cohorts (P=0.047). On univariate analysis, the vancomycin powder cohort had a significantly high incidence of cultures without growth (n=7, 39% vs. n=4, 11%, relative risk: 2.063, 95% confidence interval: 0.927-4.591, P=0.028). This association remained significant on multivariate analysis (adjusted odds ratio: 9.656, 95% confidence interval: 1.743-53.494, P=0.009). CONCLUSIONS: In EOS patients undergoing procedures complicated by SSI, the use of vancomycin powder was independently associated with increased risk of no culture growth. Surgeons and infectious disease physicians should be aware and adjust diagnostic and treatment strategies appropriately. LEVEL OF EVIDENCE: Level III-retrospective cohort study.

Preclinical in vitro and in vivo pharmacokinetic properties of danicamtiv, a new targeted myosin activator for the treatment of dilated cardiomyopathy.

Dilated cardiomyopathy (DCM) is a disease of the myocardium defined by left ventricular enlargement and systolic dysfunction leading to heart failure. Danicamtiv, a new targeted myosin activator designed for the treatment of DCM, was characterised in in vitro and in vivo preclinical studies. Danicamtiv human hepatic clearance was predicted to be 0.5 mL/min/kg from in vitro metabolic stability studies in human hepatocytes. For human, plasma protein binding was moderate with a fraction unbound of 0.16, whole blood-to-plasma partitioning ratio was 0.8, and danicamtiv showed high permeability and no efflux in a Caco-2 cell line. Danicamtiv metabolism pathways in vitro included CYP-mediated amide-cleavage, N-demethylation, as well as isoxazole- and piperidine-ring-opening. Danicamtiv clearance in vivo was low across species with 15.5, 15.3, 1.6, and 5.7 mL/min/kg in mouse, rat, dog, and monkey, respectively. Volume of distribution ranged from 0.24 L/kg in mouse to 1.7 L/kg in rat. Oral bioavailability ranged from 26% in mouse to 108% in dog. Simple allometric scaling prediction of human plasma clearance, volume of distribution, and half-life was 0.64 mL/min/kg, 0.98 L/kg, and 17.7 h, respectively. Danicamtiv preclinical attributes and predicted human pharmacokinetics supported advancement toward clinical development.

IL-15, gluten and HLA-DQ8 drive tissue destruction in coeliac disease.

Coeliac disease is a complex, polygenic inflammatory enteropathy caused by exposure to dietary gluten that occurs in a subset of genetically susceptible individuals who express either the HLA-DQ8 or HLA-DQ2 haplotypes1,2. The need to develop non-dietary treatments is now widely recognized3, but no pathophysiologically relevant gluten- and HLA-dependent preclinical model exists. Furthermore, although studies in humans have led to major advances in our understanding of the pathogenesis of coeliac disease4, the respective roles of disease-predisposing HLA molecules, and of adaptive and innate immunity in the development of tissue damage, have not been directly demonstrated. Here we describe a mouse model that reproduces the overexpression of interleukin-15 (IL-15) in the gut epithelium and lamina propria that is characteristic of active coeliac disease, expresses the predisposing HLA-DQ8 molecule, and develops villous atrophy after ingestion of gluten. Overexpression of IL-15 in both the epithelium and the lamina propria is required for the development of villous atrophy, which demonstrates the location-dependent central role of IL-15 in the pathogenesis of coeliac disease. In addition, CD4+ T cells and HLA-DQ8 have a crucial role in the licensing of cytotoxic T cells to mediate intestinal epithelial cell lysis. We also demonstrate a role for the cytokine interferon-γ (IFNγ) and the enzyme transglutaminase 2 (TG2) in tissue destruction. By reflecting the complex interaction between gluten, genetics and IL-15-driven tissue inflammation, this mouse model provides the opportunity to both increase our understanding of coeliac disease, and develop new therapeutic strategies.

Seromas Following Pediatric Spinal Deformity Surgery: Is Operative Management Necessary?

BACKGROUND: Seromas are known complications after pediatric spinal deformity surgery. Although many surgeons perform an early debridement to prevent deep surgical site infections (SSIs), a less invasive approach to seroma management has not been studied. We hypothesized that a conservative approach to seroma management would be safe and yield equivalent outcomes. METHODS: We performed a retrospective review of patients who developed a postoperative seroma with or without nonpurulent drainage. Inclusion criteria were patients below 21 years who underwent primary posterior spinal fusion from 1996 to 2016 and developed a postoperative wound seroma. Seromas were clinically defined as an afebrile patient with a fluid collection that was soft and nontender to palpation and without induration or erythema. Growing spine surgeries and revision procedures were excluded from this study. RESULTS: Twenty-five of 790 total patients with a mean follow-up of 57.8 months (±48.5 mo) developed a seroma. Seromas were identified at a mean of 13.6 days postoperatively and resolved after a mean of 12.2 days following the presentation. Seromas occurred in 12 patients with idiopathic scoliosis, 12 with neuromuscular scoliosis, and 1 patient with Scheuermann kyphosis. All cases were managed conservatively with monitoring of the incision without an operative procedure. In cases of spontaneous drainage, a sterile dressing was applied to the wound and changed as needed until drainage ceased. Two patients underwent bedside needle aspiration and 5 patients received prophylactic antibiotics at the treating surgeon's discretion. All cases resolved spontaneously without development of an acute SSI. Three cases subsequently developed a late SSI (range, 18 to 38 mo postoperatively). Two had idiopathic scoliosis and 1 had neuromuscular scoliosis. None of these seromas drained spontaneously. CONCLUSIONS: Conservative management of postoperative seromas after pediatric spinal deformity surgery is appropriate. It is unclear if seromas contributed to the development of the 3 late infections. Further studies are needed regarding the relationship of late infections in seroma patients. LEVEL OF EVIDENCE: Level IV-case series.

Natural selection contributed to immunological differences between hunter-gatherers and agriculturalists.

The shift from a hunter-gatherer to an agricultural mode of subsistence is believed to have been associated with profound changes in the burden and diversity of pathogens across human populations. Yet, the extent to which the advent of agriculture affected the evolution of the human immune system remains unknown. Here we present a comparative study of variation in the transcriptional responses of peripheral blood mononuclear cells to bacterial and viral stimuli between Batwa rainforest hunter-gatherers and Bakiga agriculturalists from Uganda. We observed increased divergence between hunter-gatherers and agriculturalists in the early transcriptional response to viruses compared with that for bacterial stimuli. We demonstrate that a significant fraction of these transcriptional differences are under genetic control and we show that positive natural selection has helped to shape population differences in immune regulation. Across the set of genetic variants underlying inter-population immune-response differences, however, the signatures of positive selection were disproportionately observed in the rainforest hunter-gatherers. This result is counter to expectations on the basis of the popularized notion that shifts in pathogen exposure due to the advent of agriculture imposed radically heightened selective pressures in agriculturalist populations.

Gene activation precedes DNA demethylation in response to infection in human dendritic cells.

DNA methylation is considered to be a relatively stable epigenetic mark. However, a growing body of evidence indicates that DNA methylation levels can change rapidly; for example, in innate immune cells facing an infectious agent. Nevertheless, the causal relationship between changes in DNA methylation and gene expression during infection remains to be elucidated. Here, we generated time-course data on DNA methylation, gene expression, and chromatin accessibility patterns during infection of human dendritic cells with Mycobacterium tuberculosis We found that the immune response to infection is accompanied by active demethylation of thousands of CpG sites overlapping distal enhancer elements. However, virtually all changes in gene expression in response to infection occur before detectable changes in DNA methylation, indicating that the observed losses in methylation are a downstream consequence of transcriptional activation. Footprinting analysis revealed that immune-related transcription factors (TFs), such as NF-κB/Rel, are recruited to enhancer elements before the observed losses in methylation, suggesting that DNA demethylation is mediated by TF binding to cis-acting elements. Collectively, our results show that DNA demethylation plays a limited role to the establishment of the core regulatory program engaged upon infection.

Rel-Dependent Immune and Central Nervous System Mechanisms Control Viral Replication and Inflammation during Mouse Herpes Simplex Encephalitis.

Herpes simplex encephalitis (HSE), caused by HSV type 1 (HSV-1) infection, is an acute neuroinflammatory condition of the CNS and remains the most common type of sporadic viral encephalitis worldwide. Studies in humans have shown that susceptibility to HSE depends in part on the genetic make-up of the host, with deleterious mutations in the TLR3/type I IFN axis underlying some cases of childhood HSE. Using an in vivo chemical mutagenesis screen for HSV-1 susceptibility in mice, we identified a susceptible pedigree carrying a causal truncating mutation in the Rel gene (RelC307X ), encoding for the NF-κB transcription factor subunit c-Rel. Like Myd88-/- and Irf3-/- mice, RelC307X mice were susceptible to intranasal HSV-1 infection. Reciprocal bone marrow transfers into lethally irradiated hosts suggested that defects in both hematopoietic and CNS-resident cellular compartments contributed together to HSE susceptibility in RelC307X mice. Although the RelC307X mutation maintained cell-intrinsic antiviral control, it drove increased apoptotic cell death in infected fibroblasts. Moreover, reduced numbers of CD4+CD25+Foxp3+ T regulatory cells, and dysregulated NK cell and CD4+ effector T cell responses in infected RelC307X animals, indicated that protective immunity was also compromised in these mice. In the CNS, moribund RelC307X mice failed to control HSV-1 viral replication in the brainstem and cerebellum, triggering cell death and elevated expression of Ccl2, Il6, and Mmp8 characteristic of HSE neuroinflammation and pathology. In summary, our work implicates c-Rel in both CNS-resident cell survival and lymphocyte responses to HSV-1 infection and as a novel cause of HSE disease susceptibility in mice.