Outcomes of cell infusion for the treatment of neurological sequelae induced by spinal anesthesia-associated subdural hematoma: A case report.

Background: Subdural hematoma following spinal anesthesia for cesarean delivery is a rare complication. Surgical removal of the hematoma is the standard treatment. However, there are still many patients who suffer permanent nerve damage of varying degrees after surgery. Cell therapy has recently shown great potential for treating nerve damage. Case presentation: This report described a case of paraplegia due to an epidural hematoma occurring after spinal anesthesia for cesarean section. The patient underwent surgery to remove the hematoma and rehabilitation afterward. However, no improvement was noted. Paralysis of the lower extremities associated with urinary retention and constipation persisted. The patient received three administrations of cell infusion: the first time with autologous bone marrow-derived mononuclear cells and the following two with autologous adipose mesenchymal/stromal cells. After three cell infusions, the patient was able to walk and could urinate and defecate voluntarily. Sensory and motor function were improved and MRI showed a decrease in adherence of the nerve roots and spinal cord. Conclusions: Our results demonstrated that cell therapy may ameliorate paralysis of the lower extremities as well as fecal and urinary function following spinal hematoma associated with spinal anesthesia.

Inflammatory mediators drive neuroinflammation in autism spectrum disorder and cerebral palsy.

Inflammation conditions are associated with autism spectrum disorder (ASD) and cerebral palsy (CP), primarily observed in the peripheral immune system. However, the extent of neuro-inflammation and neuro-immune dysregulation remains poorly studied. In this study, we analyzed the composition of cerebrospinal fluid (CSF) to uncover the inflammatory mediators driving the neuro-immune system in ASD and CP patients. Our findings revealed that ASD patients had elevated levels of four inflammatory cytokines (TNF-α, IL-4, IL-21, and BAFF) compared to controls, while CP patients exhibited increased levels of eight inflammatory cytokines (IFN-γ, GM-CSF, TNF-α, IL-2, IL-4, IL-6, IL-17A and IL-12), one anti-inflammatory cytokine (IL-10), and five growth factors (GFs) (NGF-ß, EGF, GDF-15, G-CSF and BMP-9) compared to both controls and ASD patients. Additionally, intrathecal infusion of autologous bone marrow mononuclear cells (BMMNCs) led to a slight decrease in TGF-ß and GDF-15 levels in the CSF of ASD and CP patients, respectively. Our study provides new insights into the molecular composition of CSF in ASD and CP patients, with the potential to develop more effective diagnosis methods and improved treatment for these diseases.Clinical trial registration CSF samples used in this study are from clinical trials NCT03225651, NCT05307536, NCT02569775, NCT03123562, NCT02574923, NCT05472428 and previous reports [7, 9, 17-19].

Implementation of the Canadian syncope pathway: a pilot non-randomized stepped wedge trial.

BACKGROUND: We developed the Canadian Syncope Pathway (CSP) based on the Canadian Syncope Risk Score (CSRS) to aid emergency department (ED) syncope management. This pilot implementation study assessed patient inclusion, length of transition period, as well as process measures (engagement, reach, adoption, and fidelity) to prepare for multicenter implementation. METHODS: A non-randomized stepped wedge trial at two hospitals was conducted over a 7-month period. After 2-3 months in the control condition, the hospitals crossed over in a stepwise fashion to the intervention condition. Study participants were ED and non-ED physicians, or their delegates, and patients (aged ≥ 18 years) with syncope. We aimed to analyze patient characteristics, ED management including disposition decision, and CSRS recommendations application for all eligible patients during the intervention period. Our targets were 95% inclusion rate, 70% adoption (proportion of physicians who applied the pathway), 60% reach (intervention applied to eligible patients) and 70% fidelity (appropriate recommendations application) for all eligible patients. Clinical Trials registration NCT04790058. RESULTS: 1002 eligible patients (mean age 56.6 years; 51.0% males) were included: 349 patients during the control and 653 patients during the intervention period. Physician engagement varied from 39.7% to 97.1% for presentation at meetings. Process measures for the first month and the end of the intervention were: adoption 70.7% (58/82) and 84.4% (103/122), reach 67.5% (108/160) and 55.0% (359/653), fidelity among patients with physician data form completion 86.3% (88/102) and 88.3% (294/333), versus fidelity among all eligible patients 83.8% (134/160) and 83.3% (544/653) respectively with no significant differences in fidelity at one month and the end of the intervention period. CONCLUSION: In this pilot study, we achieved all prespecified benchmarks for proceeding to the multicenter CSP implementation except reach. Our results indicate a 1-month transition period will be adequate though regular reminders will be needed during full-scale implementation.

RéSUMé: CONTEXTE: Nous avons mis au point la Canadian Syncope Pathway (CSP) basée sur le Canadian Syncope Risk Score (CSRS) pour aider les services d'urgence à gérer la syncope. Cette étude pilote de mise en œuvre a évalué l'inclusion des patients, la durée de la période de transition, ainsi que les mesures de processus (engagement, portée, adoption et fidélité) pour se préparer à la mise en œuvre multicentrique MéTHODES: Un essai par étapes non randomisé dans deux hôpitaux a été mené sur une période de 7 mois. Après 2 à 3 mois dans l'état de contrôle, les hôpitaux sont passés progressivement à l'état d'intervention. Les participants à l'étude étaient des médecins du service de l'urgence et non du service de l'urgence, ou leurs délégués, et des patients (âgés de 18 ans) atteints de syncope. Nous avons cherché à analyser les caractéristiques des patients, la prise en charge des urgences, y compris la décision de disposition, et l'application des recommandations du CSRS pour tous les patients admissibles pendant la période d'intervention. Nos cibles étaient le taux d'inclusion de 95 %, l'adoption de 70 % (proportion de médecins qui ont appliqué la voie), la portée de 60 % (intervention appliquée aux patients admissibles) et la fidélité de 70 % (application des recommandations appropriées) pour tous les patients admissibles. Enregistrement des essais cliniques NCT04790058. RéSULTATS: 1002 patients éligibles (âge moyen 56,6 ans; 51,0% d'hommes) ont été inclus : 349 patients pendant le contrôle et 653 patients pendant la période d'intervention. La participation des médecins variait de 39,7 % à 97,1 % pour la présentation aux réunions. Les mesures du processus pour le premier mois et la fin de l'intervention étaient les suivantes : adoption 70,7 % (58/82) et 84,4 % (103/122), atteinte de 67,5 % (108/160) et 55,0 % (359/653), fidélité chez les patients ayant rempli le formulaire de données médicales 86,3 % (88/102) et 88,3 % (294/333), versus fidélité chez tous les patients admissibles 83,8 % (134/160) et 83,3 % (544/653) respectivement, sans différence significative de fidélité à un mois et à la fin de la période d'intervention. CONCLUSION: Dans cette étude pilote, nous avons atteint tous les points de repère prédéterminés pour procéder à la mise en œuvre du PSC multicentrique, sauf la portée. Nos résultats indiquent qu'une période de transition d'un mois sera adéquate, bien que des rappels réguliers seront nécessaires pendant la mise en œuvre à grande échelle.

Developing Implementation Strategies to Support the Uptake of a Risk Tool to Aid Physicians in the Clinical Management of Patients With Syncope: Systematic Theoretical and User-Centered Design Approach.

BACKGROUND: The Canadian Syncope Risk Score (CSRS) was developed to improve syncope management in emergency department settings. Evidence-based tools often fail to have the intended impact because of suboptimal uptake or poor implementation. OBJECTIVE: In this paper, we aimed to describe the process of developing evidence-based implementation strategies to support the deployment and use of the CSRS in real-world emergency department settings to improve syncope management among physicians. METHODS: We followed a systematic approach for intervention development, including identifying who needs to do what differently, identifying the barriers and enablers to be addressed, and identifying the intervention components and modes of delivery to overcome the identified barriers. We used the Behaviour Change Wheel to guide the selection of implementation strategies. We engaged CSRS end users (ie, emergency medicine physicians) in a user-centered design approach to generate and refine strategies. This was achieved over a series of 3 qualitative user-centered design workshops lasting 90 minutes each with 3 groups of emergency medicine physicians. RESULTS: A total of 14 physicians participated in the workshops. The themes were organized according to the following intervention development steps: theme 1-identifying and refining barriers and theme 2-identifying the intervention components and modes of delivery. Theme 2 was subdivided into two subthemes: (1) generating high-level strategies and developing strategies prototypes and (2) refining and testing strategies. The main strategies identified to overcome barriers included education in the format of meetings, videos, journal clubs, and posters (to address uncertainty around when and how to apply the CSRS); the development of a web-based calculator and integration into the electronic medical record (to address uncertainty in how to apply the CSRS); a local champion (to address the lack of team buy-in); and the dissemination of evidence summaries and feedback through email communications (to address a lack of evidence about impact). CONCLUSIONS: The ability of the CSRS to effectively improve patient safety and syncope management relies on broad buy-in and uptake across physicians. To ensure that the CSRS is well positioned for impact, a comprehensive suite of strategies was identified to address known barriers.

Development of practice recommendations based on the Canadian Syncope Risk Score and identification of barriers and facilitators for implementation.

BACKGROUND: Wide variations in emergency department (ED) syncope management exist. The Canadian Syncope Risk Score (CSRS) was developed to predict the probability of 30-day serious outcomes after ED disposition. Study objectives were to evaluate the acceptability of proposed CSRS practice recommendations among providers and patients, and identify barriers and facilitators for CSRS use to guide disposition decisions. METHODS: We conducted semi-structured interviews with 41 physicians involved in ED syncope and 35 ED patients with syncope. We used purposive sampling to ensure a variety of physician specialties and CSRS patient risk levels. Thematic analysis was completed by two independent coders with consensus meetings to resolve conflicts. Analysis proceeded in parallel with interviews until data saturation. RESULTS: The majority (97.6%; 40/41) of physicians agreed with discharge of low risk (CSRS ≤ 0) but opined that 'no follow up' changed to 'follow-up as needed'. Physicians indicated current practices do not align with the medium-risk recommendation to discharge patients with 15-day monitoring (CSRS = 1-3; due to lack of access to monitors and timely follow-up) and the high-risk recommendation (CSRS ≥ 4) to potentially discharge patients with 15-day monitoring. Physicians recommended brief hospitalization of high-risk patients due to patient safety concerns. Facilitators included the CSRS-based patient education and scores supporting their clinical gestalt. Patients reported receiving varying levels of information regarding syncope and post-ED care, were satisfied with care received and preferred less resource intensive options. CONCLUSION: Our recommendations based on the study results were: discharge of low-risk patients with physician follow-up as needed; discharge of medium-risk patients with 15-day cardiac monitoring and brief hospitalization of high-risk patients with 15-day cardiac monitoring if discharged. Patients preferred less resource intensive options, in line with CSRS recommended care. Implementation should leverage identified facilitators (e.g., patient education) and address the barriers (e.g., monitor access) to improve ED syncope care.

RéSUMé: CONTEXTE: La prise en charge des syncopes par les services d'urgence varie considérablement. Le Canadian Syncope Risk Score (CSRS) a été mis au point pour prédire la probabilité d'une issue grave à 30 jours après la prise en charge par le service des urgences. Les objectifs de l'étude étaient d'évaluer l'acceptabilité des recommandations pratiques proposées par le CSRS parmi les prestataires et les patients, et d'identifier les barrières et les facilitateurs de l'utilisation du CSRS pour guider les décisions de disposition. MéTHODES: Nous avons mené des entretiens semi-structurés avec 41 médecins impliqués dans la syncope aux urgences et 35 patients souffrant de syncope aux urgences. Nous avons utilisé un échantillonnage raisonné pour assurer une variété de spécialités médicales et de niveaux de risque pour les patients du CSRS. L'analyse thématique a été réalisée par deux codeurs indépendants, avec des réunions de consensus pour résoudre les conflits. L'analyse s'est déroulée parallèlement aux entretiens jusqu'à saturation des données. RéSULTATS: La majorité (97,6 % ; 40/41) des médecins étaient d'accord avec la sortie des patients à faible risque (CSRS ≤ 0), mais ont estimé que " pas de suivi " devait être remplacée par " suivi en fonction des besoins ". Les médecins ont indiqué que leurs pratiques actuelles ne sont pas conformes à la recommandation à risque moyen de faire sortir les patients avec une surveillance de 15 jours (CSRS = 1-3 ; en raison du manque d'accès aux moniteurs et au suivi en temps opportun) et à la recommandation à risque élevé (CSRS ≥ 4) de potentiellement faire sortir les patients avec une surveillance de 15 jours. Les médecins ont recommandé une brève hospitalisation des patients à haut risque pour des raisons de sécurité. Les facilitateurs comprenaient l'éducation des patients basée sur le CSRS et les scores soutenant leur gestalt clinique. Les patients ont déclaré avoir reçu différents niveaux d'information concernant la syncope et les soins post-urgence, étaient satisfaits des soins reçus et préféraient des options moins gourmandes en ressources. CONCLUSIONS: Nos recommandations basées sur les résultats de l'étude sont les suivantes : sortie des patients à faible risque avec suivi par un médecin si nécessaire ; la sortie des patients à risque moyen avec une surveillance cardiaque de 15 jours et une brève hospitalisation des patients à risque élevé avec une surveillance cardiaque de 15 jours en cas de sortie. Les patients ont préféré des options moins gourmandes en ressources, conformément aux soins recommandés par le CSRS. La mise en œuvre devrait s'appuyer sur les facilitateurs identifiés (par exemple, l'éducation des patients) et s'attaquer aux obstacles (par exemple, le contrôle de l'accès) pour améliorer les soins aux urgences en cas de syncope.

The Growth of Extended Melem Units on g-C3N4 by Hydrothermal Treatment and Its Effect on Photocatalytic Activity of g-C3N4 for Photodegradation of Tetracycline Hydrochloride under Visible Light Irradiation.

In this work, the growth of extended tri-s-triazine units (melem units) on g-C3N4 (CN) by hydrothermal treatment and its effect on the photodegradation efficiency of tetracycline hydrochloride (TC) is investigated. The CN-180-x and CN-200-6 samples were prepared using different hydrolysis times and temperatures, and they were characterized by multiple physicochemical techniques. In addition, their photodegradation performance was evaluated under visible light irradiation. Compared to the CN, CN-180-6 possesses remarkable photocatalytic degradation efficiency at 97.17% towards TC removal in an aqueous solution. The high visible-light-induced photo-reactivity of CN-180-6 directly correlates to charge transfer efficiency, numerous structural defects with a high specific surface area (75.0 m2 g-1), and sufficient O-functional groups over g-C3N4. However, hydrothermal treatment at a higher temperature or during a longer time additionally induces the growth of extended melem units on the surface of g-C3N4, resulting in the inhibition of the charge transfer. In addition, the superoxide radical is proven to be generated from photoexcited reaction and plays a key role in the TC degradation.

Human Umbilical Cord Mesenchymal Stem Cells for Severe Neurological Sequelae due to Anti-N-Methyl-d-Aspartate Receptor Encephalitis: First Case Report.

Anti-N-methyl-d-aspartate (NMDA) receptor encephalitis is caused by altered patient immune reactions. This study reports the first patient with severe neurologic sequelae after NMDA receptor encephalitis treated with allogeneic umbilical cord-derived mesenchymal stem/stromal cells (UC-MSCs). A 5-year-old girl was diagnosed with NMDA receptor encephalitis and treated with immunosuppressive medicaments and intravenous immunoglobulin (IVIG). Despite intensive therapy, the patient's condition worsened so that allogenic UC-MSC therapy was contemplated. The patient received three intrathecal infusions of xeno- and serum-free cultured UC-MSCs at a dose of 106 cells/kg. At baseline and after each UC-MSC administration, the patient was examined by the German Coma Recovery Scale (CRS), the Gross Motor Function Classification System (GMFCS), the Gross Motor Function Measure-88 (GMFM-88), the Manual Ability Classification System (MACS), the Modified Ashworth Scale, and the Denver II test. Before cell therapy, she was in a permanent vegetative state with diffuse cerebral atrophy. Her cognition and motor functions improved progressively after three UC-MSC infusions. At the last visit, she was capable of walking, writing, and counting numbers. Control of urinary and bowel functions was completely recovered. Cerebral atrophy was reduced on brain magnetic resonance imaging (MRI). Overall, the outcomes of this patient suggest a potential cell therapy for autoimmune encephalitis and its neurological consequences.

Solvent Etching Process for Graphitic Carbon Nitride Photocatalysts Containing Platinum Cocatalyst: Effects of Water Hydrolysis on Photocatalytic Properties and Hydrogen Evolution Behaviors.

In this study, we synthesized Pt/g-C3N4 photocatalysts modified by a solvent etching process where ethanol (Pt/CN0), water (Pt/CN100), and a 50:50 mixture (Pt/CN50) were used as a solvent, and investigated the optimal properties of g-C3N4 to prepare the best Pt/g-C3N4 for photocatalytic hydrogen evolution. From diverse characterizations, water was proven to be a stronger solvent agent, resulting in not only the introduction of more O-functional groups onto the g-C3N4 surface, but also the degradation of a regular array of tri-s-triazine units in the g-C3N4 structure. While the addition of O-functional groups positively influenced the oxidation state of the Pt cocatalyst and the hydrogen production rate, the changes to g-C3N4 structure retarded charge transfer on its surface, inducing negative effects such as fast recombination and less oxidized Pt species. Pt/CN50 that was synthesized with the 50:50 solvent mixture exhibited the highest hydrogen production rate of 590.9 µmol g-1h-1, while the hydrogen production rates of Pt/CN0 (with pure ethanol solvent) and Pt/CN100 (with pure water solvent) were 462.7, and 367.3 µmol g-1h-1, respectively.

Ethanol Solvothermal Treatment on Graphitic Carbon Nitride Materials for Enhancing Photocatalytic Hydrogen Evolution Performance.

Recently, Pt-loaded graphic carbon nitride (g-C3N4) materials have attracted great attention as a photocatalyst for hydrogen evolution from water. The simple surface modification of g-C3N4 by hydrothermal methods improves photocatalytic performance. In this study, ethanol is used as a solvothermal solvent to modify the surface properties of g-C3N4 for the first time. The g-C3N4 is thermally treated in ethanol at different temperatures (T = 140 °C, 160 °C, 180 °C, and 220 °C), and the Pt co-catalyst is subsequently deposited on the g-C3N4 via a photodeposition method. Elemental analysis and XPS O 1s data confirm that the ethanol solvothermal treatment increased the contents of the oxygen-containing functional groups on the g-C3N4 and were proportional to the treatment temperatures. However, the XPS Pt 4f data show that the Pt2+/Pt0 value for the Pt/g-C3N4 treated at ethanol solvothermal temperature of 160 °C (Pt/CN-160) is the highest at 7.03, implying the highest hydrogen production rate of Pt/CN-160 is at 492.3 µmol g-1 h-1 because the PtO phase is favorable for the water adsorption and hydrogen desorption in the hydrogen evolution process. In addition, the electrochemical impedance spectroscopy data and the photoluminescence spectra emission peak intensify reflect that the Pt/CN-160 had a more efficient charge separation process that also enhanced the photocatalytic activity.

Myosin-II proteins are involved in the growth, morphogenesis, and virulence of the human pathogenic fungus Mucor circinelloides.

Mucormycosis is an emerging lethal invasive fungal infection. The infection caused by fungi belonging to the order Mucorales has been reported recently as one of the most common fungal infections among COVID-19 patients. The lack of understanding of pathogens, particularly at the molecular level, is one of the reasons for the difficulties in the management of the infection. Myosin is a diverse superfamily of actin-based motor proteins that have various cellular roles. Four families of myosin motors have been found in filamentous fungi, including myosin I, II, V, and fungus-specific chitin synthase with myosin motor domains. Our previous study on Mucor circinelloides, a common pathogen of mucormycosis, showed that the Myo5 protein (ID 51513) belonging to the myosin type V family had a critical impact on the growth and virulence of this fungus. In this study, to investigate the roles of myosin II proteins in M. circinelloides, silencing phenotypes and null mutants corresponding to myosin II encoding genes, designated mcmyo2A (ID 149958) and mcmyo2B (ID 136314), respectively, were generated. Those mutant strains featured a significantly reduced growth rate and impaired sporulation in comparison with the wild-type strain. Notably, the disruption of mcmyo2A led to an almost complete lack of sporulation. Both mutant strains displayed abnormally short, septate, and inflated hyphae with the presence of yeast-like cells and an unusual accumulation of pigment-filled vesicles. In vivo virulence assays of myosin-II mutant strains performed in the invertebrate model Galleria mellonella indicated that the mcmyo2A-knockout strain was avirulent, while the pathogenesis of the mcmyo2B null mutant was unaltered despite the low growth rate and impaired sporulation. The findings provide suggestions for critical contributions of the myosin II proteins to the polarity growth, septation, morphology, pigment transportation, and pathogenesis of M. circinelloides. The findings also implicate the myosin family as a potential target for future therapy to treat mucormycosis.

Personalised risk prediction following emergency department assessment for syncope.

BACKGROUND: Published risk tools do not provide possible management options for syncope in the emergency department (ED). Using the 30-day observed risk estimates based on the Canadian Syncope Risk Score (CSRS), we developed personalised risk prediction to guide management decisions. METHODS: We pooled previously reported data from two large cohort studies, the CSRS derivation and validation cohorts, that prospectively enrolled adults (≥16 years) with syncope at 11 Canadian EDs between 2010 and 2018. Using this larger cohort, we calculated the CSRS calibration and discrimination, and determined with greater precision than in previous studies the 30-day risk of adjudicated serious outcomes not identified during the index ED evaluation depending on the CSRS and the risk category. Based on these findings, we developed an on-line calculator and pictorial decision aids. RESULTS: 8233 patients were included of whom 295 (3.6%, 95% CI 3.2% to 4.0%) experienced 30-day serious outcomes. The calibration slope was 1.0, and the area under the curve was 0.88 (95% CI 0.87 to 0.91). The observed risk increased from 0.3% (95% CI 0.2% to 0.5%) in the very-low-risk group (CSRS -3 to -2) to 42.7% (95% CI 35.0% to 50.7%), in the very-high-risk (CSRS≥+6) group (Cochrane-Armitage trend test p<0.001). Among the very-low and low-risk patients (score -3 to 0), ≤1.0% had any serious outcome, there was one death due to sepsis and none suffered a ventricular arrhythmia. Among the medium-risk patients (score +1 to+3), 7.8% had serious outcomes, with <1% death, and a serious outcome was present in >20% of high/very-high-risk patients (score +4 to+11) including 4%-6% deaths. The online calculator and the pictorial aids can be found at: https://teamvenk.com/csrs CONCLUSIONS: 30-day observed risk estimates from a large cohort of patients can be obtained for management decision-making. Our work suggests very-low-risk and low-risk patients may be discharged, discussion with patients regarding investigations and disposition are needed for medium-risk patients, and high-risk patients should be hospitalised. The online calculator, accompanied by pictorial decision aids for the CSRS, may assist in discussion with patients.

Gene therapy for hemophilia B using CB 2679d-GT: a novel factor IX variant with higher potency than factor IX Padua.

Sustained expression of therapeutic factor IX (FIX) levels has been achieved after adeno-associated viral (AAV) vector-based gene therapy in patients with hemophilia B. Nevertheless, patients are still at risk of vector dose-limiting toxicity, particularly liver inflammation, justifying the need for more efficient vectors and a lower dosing regimen. A novel increased potency FIX (designated as CB 2679d-GT), containing 3 amino acid substitutions (R318Y, R338E, T343R), significantly outperformed the R338L-Padua variant after gene therapy. CB 2679d-GT demonstrated a statistically significant approximately threefold improvement in clotting activity when compared with R338L-Padua after AAV-based gene therapy in hemophilic mice. Moreover, CB 2679d-GT gene therapy showed significantly reduced bleeding time (approximately fivefold to eightfold) and total blood loss volume (approximately fourfold) compared with mice treated with the R338L-Padua, thus achieving more rapid and robust hemostatic correction. FIX expression was sustained for at least 20 weeks with both CB 2679d-GT and R338L-Padua whereas immunogenicity was not significantly increased. This is a novel gene therapy study demonstrating the superiority of CB 2679d-GT, highlighting its potential to obtain higher FIX activity levels and superior hemostatic efficacy following AAV-directed gene therapy in hemophilia B patients than what is currently achievable with the R338L-Padua variant.

Outcomes of bone marrow mononuclear cell transplantation combined with interventional education for autism spectrum disorder.

The aim of this study was to evaluate the safety and efficacy of autologous bone marrow mononuclear cell transplantation combined with educational intervention for children with autism spectrum disorder. An open-label clinical trial was performed from July 2017 to August 2019 at Vinmec International Hospital, Hanoi, Vietnam. Thirty children who fulfilled the autism criteria of the Diagnostic and Statistical Manual of Mental Disorders, Fifth Edition, and had Childhood Autism Rating Scale (CARS) scores >37 were selected. Bone marrow was harvested by anterior iliac crest puncture under general anesthesia. The volume collected was as follows: 8 mL/kg for patients under 10 kg (80 mL + [body weight in kg - 10] × 7 mL) for patients above 10 kg. Mononuclear cells were isolated with a Ficoll gradient and then infused intrathecally. The same procedure was repeated 6 months later. After the first transplantation, all patients underwent 8 weeks of educational intervention based on the Early Start Denver Model. There were no severe adverse events associated with transplantation. The severity of autism spectrum disorder (ASD) was significantly reduced, with the median CARS score decreasing from 50 (range 40-55.5) to 46.5 (range 33.5-53.5) (P < .05). Adaptive capacity increased, with the median Vineland Adaptive Behavior Scales score rising from 53.5 to 60.5. Social communication, language, and daily skills improved markedly within 18 months after transplantation. Conversely, repetitive behaviors and hyperactivity decreased remarkably. Autologous bone marrow mononuclear cell transplantation in combination with behavioral intervention was safe and well tolerated in children with ASD (Trial registration: ClinicalTrials.gov identifier: NCT03225651).

How to select the appropriate method(s) of cytotoxicity analysis of mammalian cells at biointerfaces: A tutorial.

Many individuals perform cell viability assays as a measure of biocompatibility whether the focus of their research is on novel drug discovery, development of novel biomedical devices, or the study of biointerfacial phenomena. In this tutorial paper, the most commonly used methods available to users to perform biocompatibility testing are discussed. This includes a brief introduction into the benefits and drawbacks of the techniques, including which are best used as screening assays, which are better suited to experienced users, the relative cost of the assays per unit, and what detection techniques are most appropriate for use in conjunction with the assays. In addition to helping users ensure the rigor and reproducibility of their research design, this tutorial is meant to assist reviewers of interdisciplinary journals (such as Biointerphases itself), whose expertise is in other areas of this research but do not have the experience with cell-based assays themselves.

Study of in vitro interaction between Fusarium verticillioides and Streptomyces sp. using metabolomics.

The Streptomyces sp. strain AV05 isolated from an organic amendment was found to impact both growth and fumonisin production of Fusarium verticillioides during in vitro direct confrontation. In order to investigate the interactions between the Streptomyces sp. strain AV05 and F. verticillioides, a metabolomic approach was used. The study of the endometabolomes of the microorganisms was carried out in two different conditions: the microorganisms were cultivated alone or in confrontation. The aim of this study was to examine the modifications of the endometabolome of F. verticillioides in confrontation with the Streptomyces strain. The metabolites involved in these modifications were identified using 2D NMR. Many metabolites were found to be overproduced in confrontation assays with the Streptomyces strain, notably 16 proteinogenic amino acids, inosine, and uridine. This suggested that fungal metabolic pathways such as protein synthesis have been affected due to interaction. Thus, metabolomic studies, as well as proteomics or transcriptomics, are useful for deciphering the mechanisms of interactions between biological control agents and mycotoxigenic fungi. This comprehension is one of the key elements of the improvement of the selection and use of antagonistic agents.

Missed detections of influenza A(H1)pdm09 by real-time RT-PCR assay due to haemagglutinin sequence mutation, December 2017 to March 2018, northern Viet Nam.

INTRODUCTION: There are two methods of reverse transcription polymerase chain reaction (RT-PCR) that have been the common methods to detect influenza infections: conventional and real-time RT-PCR. From December 2017 to March 2018, several missed diagnoses of influenza A(H1)pdm09 using real-time RT-PCR were reported in northern Viet Nam. This study investigated how these missed detections occurred to determine their effect on the surveillance of influenza. METHODS: The haemagglutinin (HA) segments of A(H1N1)pdm09 from both real-time RT-PCR positive and negative samples were isolated and sequenced. The primer and probe sets in the HA gene were checked for mismatches, and phylogenetic analyses were performed to determine the molecular epidemiology of these viruses. RESULTS: There were 86 positive influenza A samples; 32 were A(H1)pdm09 positive by conventional RT-PCR but were negative by real-time RT-PCR. Sequencing was conducted on 23 influenza (H1N1)pdm09 isolates that were recovered from positive samples. Eight of these were negative for A(H1)pdm09 by real-time RT-PCR. There were two different mismatches in the probe target sites of the HA gene sequences of all isolates (n = 23) with additional mismatches only at position 7 (template binding site) identified for all eight negative real-time RT-PCR isolates. The prime target sites had no mismatches. Phylogenetic analysis of the HA gene showed that both the positive and negative real-time RT-PCR isolates were grouped in clade 6B.1; however, the real-time RT-PCR negative viruses were located in a subgroup that referred to substitution I295V. CONCLUSION: Constant monitoring of genetic changes in the circulating influenza A(H1N1)pdm09 viruses is important for maintaining the sensitivity of molecular detection assays.

Optimization of electrospun poly(N-isopropyl acrylamide) mats for the rapid reversible adhesion of mammalian cells.

Poly(N-isopropyl acrylamide) (pNIPAM) is a "smart" polymer that responds to changes in altering temperature near physiologically relevant temperatures, changing its relative hydrophobicity. Mammalian cells attach to pNIPAM at 37 °C and detach spontaneously as a confluent sheet when the temperature is shifted below the lower critical solution temperature (∼32 °C). A variety of methods have been used to create pNIPAM films, including plasma polymerization, self-assembled monolayers, and electron beam ionization. However, detachment of confluent cell sheets from these pNIPAM films can take well over an hour to achieve potentially impacting cellular behavior. In this work, pNIPAM mats were prepared via electrospinning (i.e., espNIPAM) by a previously described technique that the authors optimized for cell attachment and rapid cell detachment. Several electrospinning parameters were varied (needle gauge, collection time, and molecular weight of the polymer) to determine the optimum parameters. The espNIPAM mats were then characterized using Fourier-transform infrared, x-ray photoelectron spectroscopy, and scanning electron microscopy. The espNIPAM mats showing the most promise were seeded with mammalian cells from standard cell lines (MC3T3-E1) as well as cancerous tumor (EMT6) cells. Once confluent, the temperature of the cells and mats was changed to ∼25 °C, resulting in the extremely rapid swelling of the mats. The authors find that espNIPAM mats fabricated using small, dense fibers made of high molecular weight pNIPAM are extremely well-suited as a rapid release method for cell sheet harvesting.

Ciliopathy proteins establish a bipartite signaling compartment in a C. elegans thermosensory neuron.

How signaling domains form is an important, yet largely unexplored question. Here, we show that ciliary proteins help establish two contiguous, yet distinct cyclic GMP (cGMP) signaling compartments in Caenorhabditis elegans thermosensory AFD neurons. One compartment, a bona fide cilium, is delineated by proteins associated with Bardet-Biedl syndrome (BBS), Meckel syndrome and nephronophthisis at its base, and requires NPHP-2 (known as inversin in mammals) to anchor a cGMP-gated ion channel within the proximal ciliary region. The other, a subcompartment with profuse microvilli and a different lipid environment, is separated from the dendrite by a cellular junction and requires BBS-8 and DAF-25 (known as Ankmy2 in mammals) for correct localization of guanylyl cyclases needed for thermosensation. Consistent with a requirement for a membrane diffusion barrier at the subcompartment base, we reveal the unexpected presence of ciliary transition zone proteins where no canonical transition zone ultrastructure exists. We propose that differential compartmentalization of signal transduction components by ciliary proteins is important for the functions of ciliated sensory neurons.

A secondary structure in the 5' untranslated region of adhE mRNA required for RNase G-dependent regulation.

Escherichia coli RNase G is involved in the degradation of several mRNAs, including adhE and eno, which encode alcohol dehydrogenase and enolase respectively. Previous research indicates that the 5' untranslated region (5'-UTR) of adhE mRNA gives RNase G-dependency to lacZ mRNA when tagged at the 5'-end, but it has not been elucidated yet how RNase G recognizes adhE mRNA. Primer extension analysis revealed that RNase G cleaved a phosphodiester bond between -19A and -18C in the 5'-UTR (the A of the start codon was defined as +1). Site-directed mutagenesis indicated that RNase G did not recognize the nucleotides at -19 and -18. Random deletion analysis indicated that the sequence from -145 to -125 was required for RNase G-dependent degradation. Secondary structure prediction and further site-directed deletion suggested that the stem-loop structure, with a bubble in the stem, is required for RNaseG-dependent degradation of adhE mRNA.

Gene cluster and regulation system for 1,1-dichloro-2,2-bis(4-chlorophenyl)ethylene (DDE) degradation in Janibacter sp. TYM3221.

A DDE-degrading bacterium, Janibacter sp. TYM3221, is able to grow on biphenyl and degrades 1,1-dichloro-2,2-bis(4-chlorophenyl)ethylene (DDE) via a meta-ring cleavage pathway. The bphAa gene, encoding a biphenyl dioxygenase large subunit, was previously demonstrated to be involved in the degradation of DDE in TYM3221. The bph gene cluster, containing orf2 and bphDAaAbAcAdBCST was cloned and characterized. Reverse transcription-PCR (RT-PCR) analysis indicated that these genes were transcribed as an operon. The real-time RT-PCR on orf2, bphAa, bphC, and bphS suggest the presence of the inducible orf2 promoter (orf2p) and constitutive bphAa promoter (bphAap). The TYM3221 bphST conducted biphenyl-dependent inducible activation plus constitutive basal activation of orf2p and constitutive activation of bphAap in a rhodococcal host strain, Rhodococcus erythropolis IAM1399, suggesting that expression of the TYM3221 bph operon depends on the bphST-coded two-component regulatory system. Both of these promoters were also induced by the bphS1T1 of a biphenyl degrader, Rhodococcus jostii RHA1, and contained the 24-bp consensus sequences of RHA1 bphS1T1-dependent promoters. The replacement of RHA1 bphS1 with TYM3221 bphS in combination with RHA1 bphT1 suggests that TYM3221 bphS is responsible for low inducible and high constitutive activation of orf2p in IAM1399 by the TYM3221 bphST-system. Expression of bphAaAbAcAdBC in IAM1399 resulted in the transformation of DDE to the meta-ring cleavage product via 2,3-hydroxylation, suggesting that these genes are involved in DDE degradation.