Contiguous erosion of the inactive X in human pluripotency concludes with global DNA hypomethylation.

Female human pluripotent stem cells (hPSCs) routinely undergo inactive X (Xi) erosion. This progressive loss of key repressive features follows the loss of XIST expression, the long non-coding RNA driving X inactivation, and causes reactivation of silenced genes across the eroding X (Xe). To date, the sporadic and progressive nature of erosion has obscured its scale, dynamics, and key transition events. To address this problem, we perform an integrated analysis of DNA methylation (DNAme), chromatin accessibility, and gene expression across hundreds of hPSC samples. Differential DNAme orders female hPSCs across a trajectory from initiation to terminal Xi erosion. Our results identify a cis-regulatory element crucial for XIST expression, trace contiguously growing reactivated domains to a few euchromatic origins, and indicate that the late-stage Xe impairs DNAme genome-wide. Surprisingly, from this altered regulatory landscape emerge select features of naive pluripotency, suggesting that its link to X dosage may be partially conserved in human embryonic development.

OFx: A method of 4D image construction from free-breathing non-gated MRI slice acquisitions of the thorax via optical flux.

PURPOSE: Since real-time 4D dynamic magnetic resonance imaging (dMRI) methods with adequate spatial and temporal resolution for imaging the pediatric thorax are currently not available, free-breathing slice acquisitions followed by appropriate 4D construction methods are currently employed. Self-gating methods, which extract breathing signals only from image information without any external gating technology, have much potential for this purpose, such as for use in studying pediatric thoracic insufficiency syndrome (TIS). Patients with TIS frequently suffer from extreme malformations of the chest wall, diaphragm, and spine, leading to breathing that is very complex, including deep or shallow respiratory cycles. Existing 4D construction methods cannot perform satisfactorily in this scenario, and most are not fully automatic, requiring manual interactive operations. In this paper, we propose a novel fully automatic 4D image construction method based on an image-derived concept called flux to address these challenges. METHODS: We utilized 25 dMRI data sets from 25 pediatric subjects with no known thoracic anomalies and 58 dMRI data sets from 29 patients with TIS where each patient had a dMRI scan before and after surgery. A time sequence of 80 slices are acquired at each sagittal location continuously at a rate of ~480 ms per slice under free-breathing conditions, with 30-40 sagittal locations across the chest for each subject depending on the thoracic size. In our approach, we first extract the breathing signal for each sagittal location based on the flux of the optical flow vector field of the body region from the image time series. Here, for each time point of respiratory phase, the net flux of the body region can be regarded as the flux going into or out of the body region, which we term Optical Flux (OFx). OFx provides a very robust representation of the real breathing motion of the thorax. OFx allows us to perform a full analysis of all respiratory cycles, extract only normal cycles in a robust manner, and map all extracted normal cycles on to one cosine respiration model for each sagittal location. Subsequently, we re-sample one normal cycle from the respiration model for each location independently. The normal cycle models associated with the different sagittal locations are finally composited to form the final constructed 4D image. RESULTS: We employ several metrics to evaluate the quality of the 4D construction results: Eie - error in locating time instants corresponding to end inspiration and end expiration; Eto - deviation from correct temporal order in each detected normal cycle; Ess - deviation in spatial smoothness; and Esc - deviation from spatial continuity as scored by a reader. The means and standard deviations of these metrics for normal subjects and TIS patients are found to be, respectively: Eie: 0.25 ± 0.05 and 0.38 ± 0.16 in units of time instance (ideal value = 0); Eto: 2.7% ± 2.3% and 1.8% ± 2% (ideal value = 0%); Ess: 0.5 ± 0.17 and 0.54 ± 0.25 in pixel units (ideal value = 0); Esc: 4.6 ± 0.48 and 4.56 ± 0.98 (score range: best = 5, worst = 1). The results show that the OFx method achieves excellent spatial and temporal continuity and its yield was 100% meaning that it successfully performed 4D construction on every data set tested. Compared to a recently published method, OFx is fully automatic requiring about 5 min of computational time per study starting from acquired dMRI scans. The method achieves high temporal and spatial continuity even on complex TIS data sets that include many abnormal respiratory cycles. CONCLUSIONS: A new 4D dMRI construction method based on the concept of optical flux is presented which is fully automatic and very robust in deriving respiratory signals purely from dynamic image sequences even when presented with complex breathing patterns due to severe disease conditions like TIS. Evaluations show that its accuracy is comparable to the variations found in manual annotations. An important characteristic of the method is that it is independent of the number of sagittal locations used in the construction process, which suggests that it is applicable to imaging techniques where data are acquired at only a few sagittal locations instead of the full width of the thorax. The method is not tied to any specific imaging modality, as demonstrated in this paper on not just dMRI but dynamic computed tomography (CT) as well.

Rib-based Distraction Device Implantation Before Age 3 Associated With Higher Unplanned Rate of Return to the Operating Room.

BACKGROUND: Surgical treatment of early-onset scoliosis (EOS) with rib-based implants such as the vertical expandable prosthetic titanium rib (VEPTR) is associated with a high rate of complications including surgical site infection, skin breakdown, and implant migration. Many of these complications warrant the need for unplanned reoperations, increasing the burden on an already vulnerable patient population, and introducing the further risk of infection. To provide insight into the risks of early intervention, we investigate the relationship between initial device implantation before the age of 3 and the rate of unplanned reoperation. METHODS: A retrospective review was performed of all patients at a single institution who had undergone VEPTR insertion for EOS with at least a 2-year follow-up from 2007 to 2016. Patients were stratified into the case-cohort (0 to 2 y of age) or the comparison cohort (3 to 10 y of age) based on age at the time of device implantation. Multivariate regression accounting for age and scoliosis etiology was performed to identify factors predictive of unplanned reoperation. RESULTS: A total of 137 of 185 patients treated with VEPTR were identified with 76 (56%) undergoing at least 1 unplanned reoperation during the study time period. There were 68 and 69 patients in the age 0- to 2-year and 3- to 10-year cohorts, respectively. Patients aged 0 to 2 years underwent a higher number of total procedures compared with those aged 3 to 10 (13.1±6.5 vs. 10.6±4.8, P=0.032). A significant difference was found in the rate of unplanned reoperation between the 2 cohorts with 44 (65%) patients aged 0 to 2 and 32 (46%) patients aged 3 to 10 undergoing at least 1 unplanned reoperation (P=0.031). Binary logistic multivariate regression accounting for age and scoliosis etiology demonstrated that patients aged 0 to 2 had a significantly greater odds of undergoing an unplanned reoperation (odds ratio=3.050; 95% confidence interval: 1.285-7.241; P=0.011) compared with patients aged 3 to 10 years. CONCLUSION: Overall, EOS patients aged 0 to 2 at initial VEPTR implantation are up to 3 times higher risk of undergoing an unplanned reoperation compared with those aged 3 to 10. LEVEL OF EVIDENCE: Level III.

Concomitant procedures with early-onset scoliosis rib-based surgeries.

STUDY DESIGN: Retrospective cohort study. OBJECTIVE: To determine if additional procedures can be safely performed under the same anesthesia as early-onset scoliosis (EOS) rib-based surgeries. EOS patients with rib-based implants often require repeated invasive surgery to treat their spinal deformity, with associated risks of cognitive development impairment from multiple anesthetic exposures. Many of these patients have comorbidities that require additional procedures, possibly increasing the risk of adverse outcomes. METHODS: We performed a retrospective review of EOS patients treated with rib-based implants with at least 2 year follow up at a single institution. Patients were divided into two cohorts: Group A included patients who never had another procedure in conjunction with a rib-based implant insertion, expansion, and/or revision surgery throughout their treatment and Group B included patients who had at least one other procedure operate in conjunction with rib-based implant surgery. Univariate analysis was performed to compare demographics, surgical parameters, and incidence of complication between the two groups. A subanalysis of patients who experienced an infection requiring surgery in Group A and Group B was also performed to further analyze the impact of concomitant procedures. RESULTS: 147 EOS patients underwent rib-based growing instrumentation at our institution. 98 patients (Group A) did not have another procedure occur under the same anesthetic as a rib-based implant surgery and 49 patients (Group B) had another procedure performed in conjunction with a rib-based implant surgery. Gender, etiology, BMI, length of follow up and number of rib-based implant procedures were similar between both cohorts (p = 0.91, 0.24, 0.28, 0.91, 0.77, respectively). The total surgical and anesthesia time was significantly longer in the patients undergoing concomitant procedures (Group B) (p < 0.0001, p < 0.0001, respectively). Among the patients in Group B, Otolaryngology was the most common department adding surgical care, followed by non-spine orthopaedics and general surgery. Mircrolaryngoscopy and bronchoscopy were performed most often (24 procedures), followed by cerumen removal under anesthesia (22 procedures), ear tube insertion (19 procedures) and bronchoscopy (19 procedures). Infection requiring irrigation and debridement was the most common complication found in both cohorts. Although Group B had a significantly lower number of patients who experienced complications (p = 0.002), the complication rate per procedure was 14% in Group A and 16% in Group B. When performing a subanalysis on patients who experienced an infection requiring irrigation and debridement surgery among Group A (47 patients) and Group B (8 patients), gender, etiology, BMI, length of follow up, and number of rib-based implant procedures were similar (p = 0.71, 0.26, 0.06, 0.69, 0.84 and, respectively). The total surgical and anesthesia time were similar (p = 0.11 and 0.13, respectively). In addition, the number of irrigation and debridement surgeries needed to treat each infection even was similar between the two groups (p = 0.59). CONCLUSION: There was no difference in complication rate per procedure when other services operate in conjunction with rib-based implant surgery. LEVEL OF EVIDENCE: III.

Patient-Reported Outcomes Are Equivalent in Patients Who Receive Vertebral Body Tethering Versus Posterior Spinal Fusion in Adolescent Idiopathic Scoliosis.

Anterior vertebral body tethering (AVBT), or spinal growth tethering, is an emerging technology that recently received Food and Drug Administration (FDA) approval through a humanitarian device exemption designation to treat idiopathic scoliosis patients with remaining growth. This study compared patients who underwent AVBT with those treated with standard-of-care posterior spinal fusion (PSF) to determine inherent differences in patients and families who seek cutting-edge treatments. The authors reviewed 62 PSF patients from a multicenter registry and 20 AVBT patients from an FDA-approved investigational clinical trial. The authors examined demographics, preoperative clinical and radiographic variables, and health-related quality of life (HRQOL). All included patients preoperatively were classified as Lenke type 1 or 2 with a thoracic curve of 35° to 60°, a lumbar curve less than 35°, and a skeletal maturity score of Risser sign 0 or Sanders bone age of 4 or less. Idiopathic scoliosis patients treated with surgical intervention were primarily White females who were 12 years old. No differences in demographics, clinical variables, and radiographic measures were detected between the PSF and AVBT cohorts. The AVBT group showed more thoracic flexibility on bending radiographs, correcting on average 59% compared with 43% for PSF patients (P=.005). Patients had similar HRQOL total scores and scores across each of the 5 domains of the Scoliosis Research Society Questionnaire Version 22. The percentage of patients scoring below 4.0 within each domain was comparable between cohorts. Scoliosis patients who underwent vertebral tethering at a level of deformity magnitude and maturity similar to those who underwent posterior fusion did not differ at baseline regarding demographics, clinical variables, and HRQOL. [Orthopedics. 2021;44(1):24-28.].

Thoracic Quantitative Dynamic MRI to Understand Developmental Changes in Normal Ventilatory Dynamics.

BACKGROUND: A database of normative quantitative measures of regional thoracic ventilatory dynamics, which is essential to understanding better thoracic growth and function in children, does not exist. RESEARCH QUESTION: How to quantify changes in the components of ventilatory pump dynamics during childhood via thoracic quantitative dynamic MRI (QdMRI)? STUDY DESIGN AND METHODS: Volumetric parameters were derived via 51 dynamic MRI scans for left and right lungs, hemidiaphragms, and hemichest walls during tidal breathing. Volume-based symmetry and functional coefficients were defined to compare left and right sides and to compare contributions of the hemidiaphragms and hemichest walls with tidal volumes (TVs). Statistical analyses were performed to compare volume components among four age-based groups. RESULTS: Right thoracic components were significantly larger than left thoracic components, with average ratios of 1.56 (95% CI, 1.41-1.70) for lung TV, 1.81 (95% CI, 1.60-2.03) for hemidiaphragm excursion TV, and 1.34 (95% CI, 1.21-1.47) for hemichest wall excursion TV. Right and left lung volumes at end-expiration showed, respectively, a 44% and 48% increase from group 2 (8 ≤ age < 10) to group 3 (10 ≤ age < 12). These numbers from group 3 to group 4 (12 ≤ age ≤ 14) were 24% and 28%, respectively. Right and left hemichest wall TVs exhibited, respectively, 48% and 45% increases from group 3 to group 4. INTERPRETATION: Normal right and left ventilatory volume components have considerable asymmetry in morphologic features and dynamics and change with age. Chest wall and diaphragm contributions vary in a likewise manner. Thoracic QdMRI can provide quantitative data to characterize the regional function and growth of the thorax as it relates to ventilation.

Automatic labeling of respiratory phases and detection of abnormal respiratory signals in free-breathing thoracic dynamic MR image acquisitions based on deep learning.

4D thoracic images constructed from free-breathing 2D slice acquisitions based on dynamic magnetic resonance imaging (dMRI) provide clinicians the capability of examining the dynamic function of the left and right lungs, left and right hemi-diaphragms, and left and right chest wall separately for thoracic insufficiency syndrome (TIS) treatment [1]. There are two shortcomings of the existing 4D construction methods [2]: a) the respiratory phase corresponding to end expiration (EE) and end inspiration (EI) need to be manually identified in the dMRI sequence; b) abnormal breathing signals due to non-tidal breathing cannot be detected automatically which affects the construction process. Since the typical 2D dynamic MRI acquisition contains ~3000 slices per patient, handling these tasks manually is very labor intensive. In this study, we propose a deep-learning-based framework for addressing both problems via convolutional neural networks (CNNs) [3] and Long Short-Term Memory (LSTM) [4] models. A CNN is used to extract the motion characteristics from the respiratory dMRI sequences to automatically identify contiguous sequences of slices representing exhalation and inhalation processes. EE and EI annotations are subsequently completed by comparing the changes in the direction of motion of the diaphragm. A LSTM network is used for detecting abnormal respiratory signals by exploiting the non-uniform motion feature sequence of abnormal breathing motions. Experimental results show the mean error of labeling EE and EI is ~0.3 dMRI time point unit (much less than one time point). The accuracy of abnormal cycle detection reaches 80.0%. The proposed approach achieves results highly comparable to manual labeling in accuracy but with close to full automation of the whole process. The framework proposed here can be readily adapted to other modalities and dynamic imaging applications.

Lengthening Less Than 7 Months Leads to Greater Spinal Height Gain With Rib-based Distraction.

BACKGROUND: Severe early-onset scoliosis (EOS) has been associated with a multitude of comorbidities, chief among them being deficient thoracic spine growth and pulmonary complications. EOS management with rib-based instrumentation involves repeated lengthening. Despite expansion practice patterns, there is limited literature and no evidence-based guidelines for optimal expansion intervals. Our study evaluates clinical outcomes in relation to lengthening intervals with the aim of optimizing the timing of surgical expansion in EOS patients. METHODS: A single-institution retrospective review of 60 EOS patients treated with rib-based growth instrumentation with a minimum of 3-year follow-up and 3 expansion/revision surgeries. Patients were separated into 2 expansion cohorts: (1) more frequent lengthening [MFL group (≤7 mo)] and (2) less frequent lengthening [LFL group (>7 mo)]. Demographic information and clinical factors were recorded. Univariate and bivariate analyses were performed. RESULTS: Both the MFL group (35 patients) and LFL group (25 patients) were similar in sex distribution, diagnosis, preoperative parameters of interest, and treatment duration. The mean follow-up was 6.0 years. There was an increase in postoperative T1-S1 spine height gained in the MFL group (P=0.006) as well as a higher percent expected spine growth based on normative values (P=0.03) when compared with the LFL group. The MFL group had more expansion/revision surgeries (P=0.003) but no increase in the number of complications (P=0.86). CONCLUSIONS: More frequent lengthenings were associated with statistically significant overall spinal height gain and percent expected growth without a significant increase in complication rates. It was shown that change in major curve and space available for the lungs was not associated with the lengthening intervals. LEVEL OF EVIDENCE: Level III-a comparative retrospective study.

Comparison of T1-S1 Spine Height of Postoperative Rib-based Implant Patients With Age-matched Peers.

BACKGROUND: Severe early-onset scoliosis leads to deficient spine height, thoracic growth inhibition, and ultimately pulmonary compromise. Rib-based growing instrumentation seeks to correct thoracic deformities, in part by correcting the spinal deformity, adding height, increasing thoracic volume, and allowing for continual spinal growth until maturity. However, the amount of growth in these patients relative to their peers is unknown. METHODS: Sixty patients who had undergone surgical intervention for the treatment of early-onset scoliosis were assessed via radiographic measurements of coronal T1-S1 height and major curve angle before implantation and again at most recent follow-up (minimum 2 years). T1-S1 measurements were then compared with age-matched peers to assess growth differences. Clinical information was examined for relevant parameters. RESULTS: The average age of our cohort at initial surgery and most recent follow-up was 4.4±3.8 and 10.0±4.4 years old, respectively. In this patient set, there was an average increase in T1-S1 height of 13.1±11.1 mm per year, with the majority of growth occurring in the first 2 years following implantation, and improvement in a major curve from 68±8 to 53±7 degrees. Overall, 77% of patients saw improvement in the major curve at most recent follow-up. Furthermore, a statistically significant greater percent of expected growth was seen in congenital compared with neuromuscular scoliosis (P<0.001). In addition, a weak negative correlation was observed between a number of surgical lengthenings and T1-S1 growth. CONCLUSIONS: Rib-based implant intervention has been shown to improve the major curve, but only improves growth potential to around 80% of expected growth. Scoliosis diagnosis type also influences growth rate potential, with congenital scoliosis patients being surgically treated earlier in life and having a growth rate approaching that of a healthy individual. LEVEL OF EVIDENCE: Level III-Case control.

Can We Save the Implant: Rib-based Implant Removal Rates and Risk Factors Following Irrigation and Debridement (I&D) Surgery?

BACKGROUND: In the event of a surgical site infection, management includes surgical debridement in an attempt to treat the infection and retain the implant; however they are often unsuccessful in this regard. Although studies have described the incidence of complications, current literature does not have sufficient evidence to provide clear recommendations regarding retention versus removal of implants. This study aims to identify predictive factors associated with the need for implant removal to decrease unnecessary attempts at implant retention. METHODS: A retrospective review of early-onset scoliosis patients at a single institution treated with rib-based vertical expandable prosthetic titanium rib implants who developed infection requiring irrigation and debridement (I&D) due to wound problems including surgical site infection, skin slough, and wound dehiscence. All patients had a minimum of a 2-year follow-up. Univariate and multivariate logistic regression analyses were conducted to determine the odds of implant removal. RESULTS: Fifty-nine of 181 patients (32%) required an I&D due to a wound problem. These patients underwent the initial implant procedure at a mean age of 4.6±3.8 years. In total, 29 patients ultimately underwent implant removal. Significant predictive factors for removal included total number of wound problems, total number of I&Ds, days from identification of wound problem to I&D procedure, days on antibiotics, total number of surgeries, presence of gastrostomy tube, and nonambulatory status (P<0.0001, 0.001, 0.095, 0.093, 0.082, 0.054, and 0.026, respectively). Multiple logistic regression results indicated a total number of wound problems [odds ratio (OR): 6.00, P=0.001], average days from identification of wound problem to I&D (OR: 1.03, P=0.039), and presence of a gastrostomy tube (OR: 5.7, P=0.07) as independent predictors for implant removal. CONCLUSIONS: Data suggests that time from the onset of signs of infection until debridement surgery inversely correlates with the ability to retain the implants. In addition, gastrostomy tube and history of previous wound infections may be predictive clinical factors for implant removal in patients with a rib-based vertical expandable prosthetic titanium rib instrumentation. Such information can be useful for clinicians in deciding on whom to attempt implant retention versus removal when a wound problem presents itself. LEVEL OF EVIDENCE: Level III.

Small heat shock proteins mediate cell-autonomous and -nonautonomous protection in a Drosophila model for environmental-stress-induced degeneration.

Cell and tissue degeneration, and the development of degenerative diseases, are influenced by genetic and environmental factors that affect protein misfolding and proteotoxicity. To better understand the role of the environment in degeneration, we developed a genetic model for heat shock (HS)-stress-induced degeneration in Drosophila This model exhibits a unique combination of features that enhance genetic analysis of degeneration and protection mechanisms involving environmental stress. These include cell-type-specific failure of proteostasis and degeneration in response to global stress, cell-nonautonomous interactions within a simple and accessible network of susceptible cell types, and precise temporal control over the induction of degeneration. In wild-type flies, HS stress causes selective loss of the flight ability and degeneration of three susceptible cell types comprising the flight motor: muscle, motor neurons and associated glia. Other motor behaviors persist and, accordingly, the corresponding cell types controlling leg motor function are resistant to degeneration. Flight motor degeneration was preceded by a failure of muscle proteostasis characterized by diffuse ubiquitinated protein aggregates. Moreover, muscle-specific overexpression of a small heat shock protein (HSP), HSP23, promoted proteostasis and protected muscle from HS stress. Notably, neurons and glia were protected as well, indicating that a small HSP can mediate cell-nonautonomous protection. Cell-autonomous protection of muscle was characterized by a distinct distribution of ubiquitinated proteins, including perinuclear localization and clearance of protein aggregates associated with the perinuclear microtubule network. This network was severely disrupted in wild-type preparations prior to degeneration, suggesting that it serves an important role in muscle proteostasis and protection. Finally, studies of resistant leg muscles revealed that they sustain proteostasis and the microtubule cytoskeleton after HS stress. These findings establish a model for genetic analysis of degeneration and protection mechanisms involving contributions of environmental factors, and advance our understanding of the protective functions and therapeutic potential of small HSPs.