Transposition of the apophysis of the greater trochanter for reconstruction of the femoral head after septic hip arthritis in children: 4 children followed for more than 15 years

Background and purpose Total necrosis of the femoral head after infection in children during their first months of life gives a dislocated hip with severe leg shortening. A new femoral head can be achieved with subtrochanteric osteotomy and transposition of the apophysis of the greater trochanter into the acetabulum. Previous reports have dealt with short-term results (up to 12 years). Here I present some results of this procedure 15–24 years after operation. Patients and methods 4 children aged 1–6 years with complete necrosis of the femoral head were operated on with transposition of the greater trochanter. Secondary shelf plasty was performed later in 1 child, distal femoral epiphysiodesis in another, and femoral bone lengthening in 1 child. The mean follow-up period was 19 (15–24) years. Results A new femoral head developed in all hips. 2 of them had a spherical head with a good acetabular cover, and without any osteoarthritis except for slight reduction of cartilage height. These hips were painless, with a mobility that allowed good walking function after 16 and 24 years, respectively. In the other 2 patients, in which there was a severe acetabular dysplasia at the primary operation, the new femoral head was somewhat flattened; painful osteoarthritis led to hip replacement 15 and 21 years after trochanter arthroplasty. Even these patients had a relatively good walking function until the last couple of years before hip replacement. Maximum leg length discrepancy was 7 cm. Interpretation Trochanter arthroplasty with subtrochanteric osteotomy in total femoral head necrosis after septic arthritis in children may give satisfactory long-term results provided adequate acetabular cover is obtained. Although the method cannot provide a normal hip, it can contribute to less length discrepancy, less pain, improved gait, and more favorable conditions for later hip replacement

Innate Immune Sensing of DNA

Discusses efforts to understand how DNA triggers immune responses

The Inflammatory Response to Double Stranded DNA in Endothelial Cells Is Mediated by NFκB and TNFα

Endothelial cells represent an important barrier between the intravascular compartment and extravascular tissues, and therefore serve as key sensors, communicators, and amplifiers of danger signals in innate immunity and inflammation. Double stranded DNA (dsDNA) released from damaged host cells during injury or introduced by pathogens during infection, has emerged as a potent danger signal. While the dsDNA-mediated immune response has been extensively studied in immune cells, little is known about the direct and indirect effects of dsDNA on the vascular endothelium. In this study we show that direct dsDNA stimulation of endothelial cells induces a potent proinflammatory response as demonstrated by increased expression of ICAM1, E-selectin and VCAM1, and enhanced leukocyte adhesion. This response was dependent on the stress kinases JNK and p38 MAPK, required the activation of proinflammatory transcription factors NFκB and IRF3, and triggered the robust secretion of TNFα for sustained secondary activation of the endothelium. DNA-induced TNFα secretion proved to be essential in vivo, as mice deficient in the TNF receptor were unable to mount an acute inflammatory response to dsDNA. Our findings suggest that the endothelium plays an active role in mediating dsDNA-induced inflammatory responses, and implicate its importance in establishing an acute inflammatory response to sterile injury or systemic infection, where host or pathogen derived dsDNA may serve as a danger signal.United States. Dept. of Defense (CDMRP Predoctoral Training Award)National Institutes of Health (U.S.) (NIH BioMEMS Resource Center Grant P41 EB-002503)National Institutes of Health (U.S.) (NIH Grant RO1AI063795)Shriners Hospital for Childre

Exposure to delayed visual feedback of the hand changes motor-sensory synchrony perception

We examined whether the brain can adapt to temporal delays between a self-initiated action and the naturalistic visual feedback of that action. During an exposure phase, participants tapped with their index finger while seeing their own hand in real time (~0 ms delay) or delayed at 40, 80, or 120 ms. Following exposure, participants were tested with a simultaneity judgment (SJ) task in which they judged whether the video of their hand was synchronous or asynchronous with respect to their finger taps. The locations of the seen and the real hand were either different (Experiment 1) or aligned (Experiment 2). In both cases, the point of subjective simultaneity (PSS) was uniformly shifted in the direction of the exposure lags while sensitivity to visual-motor asynchrony decreased with longer exposure delays. These findings demonstrate that the brain is quite flexible in adjusting the timing relation between a motor action and the otherwise naturalistic visual feedback that this action engenders

Detection of partial-thickness supraspinatus tendon tears: is a single direct MR arthrography series in ABER position as accurate as conventional MR arthrography?

The purpose of this study was to retrospectively evaluate sensitivity and specificity of a single magnetic resonance (MR) arthrography series in abduction external rotation (ABER) position compared with conventional MR arthrography for detection of supraspinatus tendon tears, with arthroscopy as gold standard, and to assess interobserver variability. Institutional review board approval was obtained; informed consent was waived. MR arthrograms of 250 patients (170 men and 80 women; mean age, 36 years) were retrospectively and independently evaluated by three observers. Oblique coronal T1-weighted fat-suppressed images, proton density, and T2-weighted images and axial T1-weighted images and oblique sagittal T1-weighted fat-suppressed images were analyzed to detect supraspinatus tendon tears. Separately, a single T1-weighted fat-suppressed oblique axial series in ABER position was evaluated. Both protocols were scored randomly without knowledge of patients' clinical history and arthroscopy results. Tears were subclassified, based on articular surface integrity and extension (Lee classification). Interobserver agreement was assessed by kappa statistics for all patients. Ninety-two of 250 patients underwent arthroscopy; sensitivity and specificity of ABER and conventional MR arthrography were calculated and compared using paired McNemar test. Weighted kappa values of ABER and conventional MR arthrography were 0.48-0.65 and 0.60-0.67, respectively. According to arthroscopy, 69 of 92 patients had an intact cuff, and 23 patients had a cuff tear (16 partial thickness and seven full thickness). There were no statistically significant differences between ABER and conventional MR arthrography regarding sensitivity (48-61% and 52-70%, respectively) and specificity (80-94% and 91-95%). Sensitivity and specificity of a single T1-weighted series in ABER position and conventional MR arthrography are comparable for assessment of rotator cuff tear

The Duration of Antigen-Stimulation Significantly Alters the Diversity of Multifunctional CD4 T Cells Measured by Intracellular Cytokine Staining

The assessment of antigen-specific T cell responses by intracellular cytokine staining (ICS) has become a routine technique in studies of vaccination and immunity. Here, we highlight how the duration of in vitro antigen pre-stimulation, combined with the cytokine accumulation period, are critical parameters of these methods. The effect of varying these parameters upon the diversity and frequency of multifunctional CD4 T cell subsets has been investigated using a murine model of TB vaccination and in cattle naturally infected with Mycobacterium bovis. We demonstrate a substantial influence of the duration of the antigen pre-stimulation period on the repertoire of the antigen-specific CD4 T cell responses. Increasing pre-stimulation from 2 to 6 hours amplified the diversity of the seven potential multifunctional CD4 T cell subsets that secreted any combination of IFN-γ, IL-2 and TNF-α. However, increasing pre-stimulation from 6 to 16 hours markedly altered the multifunctional CD4 T cell repertoire to a dominant IFN-γ+ only response. This was observed in both murine and cattle models

The DEAD-box helicase DDX3X is a critical component of the TANK-binding kinase 1-dependent innate immune response

TANK-binding kinase 1 (TBK1) is of central importance for the induction of type-I interferon (IFN) in response to pathogens. We identified the DEAD-box helicase DDX3X as an interaction partner of TBK1. TBK1 and DDX3X acted synergistically in their ability to stimulate the IFN promoter, whereas RNAi-mediated reduction of DDX3X expression led to an impairment of IFN production. Chromatin immunoprecipitation indicated that DDX3X is recruited to the IFN promoter upon infection with Listeria monocytogenes, suggesting a transcriptional mechanism of action. DDX3X was found to be a TBK1 substrate in vitro and in vivo. Phosphorylation-deficient mutants of DDX3X failed to synergize with TBK1 in their ability to stimulate the IFN promoter. Overall, our data imply that DDX3X is a critical effector of TBK1 that is necessary for type I IFN induction

Cross-oncopanel study reveals high sensitivity and accuracy with overall analytical performance depending on genomic regions

BackgroundTargeted sequencing using oncopanels requires comprehensive assessments of accuracy and detection sensitivity to ensure analytical validity. By employing reference materials characterized by the U.S. Food and Drug Administration-led SEquence Quality Control project phase2 (SEQC2) effort, we perform a cross-platform multi-lab evaluation of eight Pan-Cancer panels to assess best practices for oncopanel sequencing.ResultsAll panels demonstrate high sensitivity across targeted high-confidence coding regions and variant types for the variants previously verified to have variant allele frequency (VAF) in the 5-20% range. Sensitivity is reduced by utilizing VAF thresholds due to inherent variability in VAF measurements. Enforcing a VAF threshold for reporting has a positive impact on reducing false positive calls. Importantly, the false positive rate is found to be significantly higher outside the high-confidence coding regions, resulting in lower reproducibility. Thus, region restriction and VAF thresholds lead to low relative technical variability in estimating promising biomarkers and tumor mutational burden.ConclusionThis comprehensive study provides actionable guidelines for oncopanel sequencing and clear evidence that supports a simplified approach to assess the analytical performance of oncopanels. It will facilitate the rapid implementation, validation, and quality control of oncopanels in clinical use.Peer reviewe

Molecular mechanisms and cellular functions of cGAS-STING signalling

The cGAS–STING signalling axis, comprising the synthase for the second messenger cyclic GMP–AMP (cGAS) and the cyclic GMP–AMP receptor stimulator of interferon genes (STING), detects pathogenic DNA to trigger an innate immune reaction involving a strong type I interferon response against microbial infections. Notably however, besides sensing microbial DNA, the DNA sensor cGAS can also be activated by endogenous DNA, including extranuclear chromatin resulting from genotoxic stress and DNA released from mitochondria, placing cGAS–STING as an important axis in autoimmunity, sterile inflammatory responses and cellular senescence. Initial models assumed that co-localization of cGAS and DNA in the cytosol defines the specificity of the pathway for non-self, but recent work revealed that cGAS is also present in the nucleus and at the plasma membrane, and such subcellular compartmentalization was linked to signalling specificity of cGAS. Further confounding the simple view of cGAS–STING signalling as a response mechanism to infectious agents, both cGAS and STING were shown to have additional functions, independent of interferon response. These involve non-catalytic roles of cGAS in regulating DNA repair and signalling via STING to NF-κB and MAPK as well as STING-mediated induction of autophagy and lysosome- dependent cell death. We have also learnt that cGAS dimers can multimerize and undergo liquid–liquid phase separation to form biomolecular condensates that could importantly regulate cGAS activation. Here, we review the molecular mechanisms and cellular functions underlying cGAS–STING activation and signalling, particularly highlighting the newly emerging diversity of this signalling pathway and discussing how the specificity towards normal, damage-induced and infection-associated DNA could be achieved

The impact of transposable element activity on therapeutically relevant human stem cells

Human stem cells harbor significant potential for basic and clinical translational research as well as regenerative medicine. Currently ~ 3000 adult and ~ 30 pluripotent stem cell-based, interventional clinical trials are ongoing worldwide, and numbers are increasing continuously. Although stem cells are promising cell sources to treat a wide range of human diseases, there are also concerns regarding potential risks associated with their clinical use, including genomic instability and tumorigenesis concerns. Thus, a deeper understanding of the factors and molecular mechanisms contributing to stem cell genome stability are a prerequisite to harnessing their therapeutic potential for degenerative diseases. Chemical and physical factors are known to influence the stability of stem cell genomes, together with random mutations and Copy Number Variants (CNVs) that accumulated in cultured human stem cells. Here we review the activity of endogenous transposable elements (TEs) in human multipotent and pluripotent stem cells, and the consequences of their mobility for genomic integrity and host gene expression. We describe transcriptional and post-transcriptional mechanisms antagonizing the spread of TEs in the human genome, and highlight those that are more prevalent in multipotent and pluripotent stem cells. Notably, TEs do not only represent a source of mutations/CNVs in genomes, but are also often harnessed as tools to engineer the stem cell genome; thus, we also describe and discuss the most widely applied transposon-based tools and highlight the most relevant areas of their biomedical applications in stem cells. Taken together, this review will contribute to the assessment of the risk that endogenous TE activity and the application of genetically engineered TEs constitute for the biosafety of stem cells to be used for substitutive and regenerative cell therapiesS.R.H. and P.T.R. are funded by the Government of Spain (MINECO, RYC-2016- 21395 and SAF2015–71589-P [S.R.H.]; PEJ-2014-A-31985 and SAF2015–71589- P [P.T.R.]). GGS is supported by a grant from the Ministry of Health of the Federal Republic of Germany (FKZ2518FSB403)