AAV5 delivery of CRISPR/Cas9 mediates genome editing in the lungs of young rhesus monkeys.

Genome editing has the potential to treat genetic diseases in a variety of tissues including the lung. We have previously developed and validated a dual adeno-associated virus (AAV) CRISPR platform that supports effective editing in the airways of mice. To validate this delivery vehicle in a large animal model, we have shown that intratracheal instillation of CRISPR/Cas9 in AAV5 can edit a housekeeping gene or a disease-related gene in the lungs of young rhesus monkeys. We observed up to 8% editing of ACE2 in lung lobes after single-dose administration. Single-nuclear RNA-sequencing revealed that AAV5 transduces multiple cell types in the caudal lung lobes, including alveolar cells, macrophages, fibroblasts, endothelial cells, and B cells. These results demonstrate that AAV5 is efficient in the delivery of CRISPR/Cas9 in the lung lobes of young rhesus monkeys.

Prenatal and postnatal methamphetamine exposure alters prefrontal cortical gene expression and behavior in mice.

Methamphetamine is a highly abused psychostimulant that substantially impacts public health. Prenatal and postnatal methamphetamine exposure alters gene expression, brain development, and behavior in the offspring, although the underlying mechanisms are not fully defined. To assess these adverse outcomes in the offspring, we employed a mouse model of prenatal and postnatal methamphetamine exposure. Juvenile offspring were behaviorally assessed on the open field, novel object recognition, Y-maze, and forced swim tests. In addition, RNA sequencing was used to explore potential alterations in prefrontal cortical gene expression. We found that methamphetamine-exposed mice exhibited decreased locomotor activity and impaired cognitive performance. In addition, differential expression of genes involved in neurotransmission, synaptic plasticity, and neuroinflammation were found with notable changes in dopaminergic signaling pathways. These data suggest potential neural and molecular mechanisms underlying methamphetamine-exposed behavioral changes. The altered expression of genes involved in dopaminergic signaling and synaptic plasticity highlights potential targets for therapeutic interventions for substance abuse disorders and related psychiatric complications.

Use of a Novel Hypercrosslinked Carbohydrate Scaffold for Vocal Fold Medialization in an Ovine Model.

Objectives: Vocal fold medialization is commonly performed for glottic insufficiency and vocal fold immobility. Currently available materials are temporary injectables or synthetic implants. Acellular scaffolds may allow vocal fold augmentation with autologous tissue via host cell migration. The purpose of this investigation was to evaluate the use of a novel carbohydrate scaffold as a medialization implant. Study Design: Animal model. Setting: Academic medical center. Methods: Unilateral type I medialization thyroplasty was performed in 3 Dorper cross ewes using a hypercrosslinked carbohydrate polymer (HCCP) scaffold. Animals were monitored for 4 weeks for general well-being, dyspnea, and weight loss. The animals were euthanized at 4 weeks and the larynges harvested. Histologic evaluation was performed to assess for adverse tissue reaction, migration, degradation, and biocompatibility. Results: No adverse events were reported. No animals lost weight or displayed evidence of dyspnea. Histology demonstrated ingrowth of host cells and neovascularization with minimal peri-implant inflammatory reaction. Cellular ingrowth into the scaffold was predominately made up of fibroblasts and early inflammatory cells. Scaffold shape was grossly maintained as it underwent degradation and replacement with host tissue. Migration of the implant material was not observed. Conclusion: Vocal fold medialization in an ovine model with an HCCP scaffold resulted in the ingrowth of host cells with minimal peri-implant inflammation. Scaffold shape was maintained without evidence of migration as it underwent replacement with host tissue. Further research is required to assess long-term efficacy in comparison to currently available implants.

Use of Thromboprophylaxis after Autologous Breast Reconstruction: A Cost-Effective Break-Even Analysis.

PURPOSE: Post-operative venous thromboembolism (VTE) is a major source of morbidity and mortality. The use of thromboprophylaxis amongst surgeons is not well studied in autologous breast reconstruction. The purpose of this study was to determine the rate of VTE in breast cancer patients undergoing autologous breast reconstruction and to compare the cost-effectiveness of postoperative chemoprophylactic agents. METHODS: The TriNetX LLC. National Health Research Network database was used to identify patients with breast cancer who underwent autologous breast reconstruction surgery between 2002-2022. The incidence of occurrence of VTE within the first 30 days of surgery was calculated. Then a break-even analysis was performed to determine the break-even rate of VTE at which the chemoprophylactic agent would be cost effective. RESULTS: A cohort of 8,221 patients was identified in this study. The rate of VTE was significantly higher in those without anticoagulation (4.0%) compared to those who received anticoagulation (2.6%) (*p=0.0008). The break-even analysis for heparin and enoxaparin's cost-effectiveness yielded ARRs of 0.73% and 1.63% for high risk patients requiring 30 days of therapy and 0.20% and 0.43% for moderate risk patients requiring 7 days of therapy, respectively. CONCLUSION: The use of thromboprophylaxis significantly lowered the risk of VTE within 30 days after autologous breast reconstruction. Heparin appeared to be more cost-effective at preventing VTE compared to enoxaparin for both high and moderate risk patients. The presented model holds potential for other institution-specific variables that can be easily applied by plastic surgeons to determine the cost-effectiveness of any therapy of their choice.

Polyhydroxyalkanoate production in Pseudomonas putida from alkanoic acids of varying lengths.

Many studies have been conducted to produce microbial polyhydroxyalkanoates (PHA), a biopolymer, from Pseudomonas sp. fed with various alkanoic acids. Because this previous data was collected using methodologies that varied in critical aspects, such as culture media and size range of alkanoic acids, it has been difficult to compare the results for a thorough understanding of the relationship between feedstock and PHA production. Therefore, this study utilized consistent culture media with a wide range of alkanoic acids (C7-C14) to produce medium chain length PHAs. Three strains of Pseudomonas putida (NRRL B-14875, KT2440, and GN112) were used, and growth, cell dry weight, PHA titer, monomer distribution, and molecular weights were all examined. It was determined that although all the strains produced similar PHA titers using C7-C9 alkanoic acids, significant differences were observed with the use of longer chain feedstocks. Specifically, KT2440 and its derivative GN112 produced higher PHA titers compared to B-14875 when fed longer chain alkanoates. We also compared several analytical techniques for determining amounts of PHA and found they produced different results. In addition, the use of an internal standard had a higher risk of calculating inaccurate concentrations compared to an external standard. These observations highlight the importance of considering this aspect of analysis when evaluating different studies.

Neuroanatomical Alterations in the CNTNAP2 Mouse Model of Autism Spectrum Disorder.

Autism spectrum disorder (ASD) is associated with neurodevelopmental alterations, including atypical forebrain cellular organization. Mutations in several ASD-related genes often result in cerebral cortical anomalies, such as the abnormal developmental migration of excitatory pyramidal cells and the malformation of inhibitory neuronal circuitry. Notably here, mutations in the CNTNAP2 gene result in ectopic superficial cortical neurons stalled in lower cortical layers and alterations to the balance of cortical excitation and inhibition. However, the broader circuit-level implications of these findings have not been previously investigated. Therefore, we assessed whether ectopic cortical neurons in CNTNAP2 mutant mice form aberrant connections with higher-order thalamic nuclei, potentially accounting for some autistic behaviors, such as repetitive and hyperactive behaviors. Furthermore, we assessed whether the development of parvalbumin-positive (PV) cortical interneurons and their specialized matrix support structures, called perineuronal nets (PNNs), were altered in these mutant mice. We found alterations in both ectopic neuronal connectivity and in the development of PNNs, PV neurons and PNNs enwrapping PV neurons in various sensory cortical regions and at different postnatal ages in the CNTNAP2 mutant mice, which likely lead to some of the cortical excitation/inhibition (E/I) imbalance associated with ASD. These findings suggest neuroanatomical alterations in cortical regions that underlie the emergence of ASD-related behaviors in this mouse model of the disorder.

Behavioral regulation by perineuronal nets in the prefrontal cortex of the CNTNAP2 mouse model of autism spectrum disorder.

Autism spectrum disorders (ASDs) arise from altered development of the central nervous system, and manifest behaviorally as social interaction deficits and restricted and repetitive behaviors. Alterations to parvalbumin (PV) expressing interneurons have been implicated in the neuropathological and behavioral deficits in autism. In addition, perineuronal nets (PNNs), specialized extracellular matrix structures that enwrap the PV-expressing neurons, also may be altered, which compromises neuronal function and susceptibility to oxidative stress. In particular, the prefrontal cortex (PFC), which regulates several core autistic traits, relies on the normal organization of PNNs and PV-expressing cells, as well as other neural circuit elements. Consequently, we investigated whether PNNs and PV-expressing cells were altered in the PFC of the CNTNAP2 knockout mouse model of ASD and whether these contributed to core autistic-like behaviors in this model system. We observed an overexpression of PNNs, PV-expressing cells, and PNNs enwrapping PV-expressing cells in adult CNTNAP2 mice. Transient digestion of PNNs from the prefrontal cortex (PFC) by injection of chondroitinase ABC in CNTNAP2 mutant mice rescued some of the social interaction deficits, but not the restricted and repetitive behaviors. These findings suggest that the neurobiological regulation of PNNs and PVs in the PFC contribute to social interaction behaviors in neurological disorders including autism.

Comparison of Tumescent Anesthesia Versus Pectoral Nerve Block in Bilateral Reduction Mammaplasty.

INTRODUCTION: With an increasing focus on multimodal pain control to reduce opioid requirements, regional and local anesthesia techniques have been investigated in bilateral reduction mammaplasty with variable results. The purpose of this study is to compare tumescent anesthesia with pectoral nerve block II (PECS II) in patients undergoing bilateral reduction mammaplasty with respect to postoperative pain and nausea, opioid consumption, length of stay, and cost. METHODS: A retrospective review of patients undergoing bilateral reduction mammaplasty for macromastia between November 2020 and December 2021 was performed. Demographic information, operative and anesthesia times, antiemetic and morphine equivalent requirements, postoperative numeric pain rating scales, and time until hospital discharge were compared between groups. χ2 and Fisher exact tests examined subgroup differences in categorical variables. Two-sample t test and Wilcoxon rank-sum test evaluated differences in continuous parametric and nonparametric variables, respectively. RESULTS: Fifty-three patients underwent bilateral reduction mammaplasty by 3 surgeons, 71.7% (n = 38) with tumescent anesthesia infiltrated by the operating surgeon before the start of the procedure and 28.3% (n = 15) with bilateral PECS II blocks performed by anesthesia before the start of the procedure. There was no difference in age, body mass index, weight resected, intraoperative medication, or immediate postoperative complications. Postoperative pain scores and opioid requirements were similar between the 2 groups. Twenty-one percent (n = 8) of tumescent patients compared with 66.7% (n = 10) of block patients required 1 or more doses of postoperative antiemetics ( P = 0.002). Patients who received blocks spent longer in the postoperative recovery area (5.3 vs 7.1 hours, P < 0.01). However, this did not translate to a significant increase in overnight stays. The block group had significantly higher hospitalization cost by an average of $4000, driven by pharmacy and procedural cost ( P < 0.01). CONCLUSION: In this cohort of multimodal perioperative pain-controlled reduction mammaplasty patients, tumescent anesthesia was associated with decreased antiemetic requirements, less time in recovery before discharge, and lower cost compared with PECS II blocks. Therefore, tumescent anesthesia may be favored over PECS II blocks when considering multimodal pain control strategies in reduction mammaplasty patients.

Potential of Industrial Hemp for Phytoremediation of Heavy Metals.

The accumulation of anthropogenic heavy metals in soil is a major form of pollution. Such potentially toxic elements are nonbiodegradable and persist for many years as threats to human and environmental health. Traditional forms of remediation are costly and potentially damaging to the land. An alternative strategy is phytoremediation, where plants are used to capture metals from the environment. Industrial hemp (Cannabis sativa) is a promising candidate for phytoremediation. Hemp has deep roots and is tolerant to the accumulation of different metals. In addition, the crop biomass has many potential commercial uses after harvesting is completed. Furthermore, the recent availability of an annotated genome sequence provides a powerful tool for the bioengineering of C. sativa for better phytoremediation.

Vitamin D attenuated 6-OHDA-induced behavioural deficits, dopamine dysmetabolism, oxidative stress, and neuro-inflammation in mice.

Background: L-DOPA, the predominant therapy for Parkinson's disease (PD) is associated with motor deficits after prolonged use. The nigrostriatal tract, a primary target of neurodegeneration in PD, contains abundant Vitamin-D receptors, suggesting a potential role for VD in the disease. Therefore, we tested the impact of Vitamin D3 (VD3) in a mouse model of PD.Methods: PD was induced in adult male C57BL6 mice by a single intrastriatal injection of 6-hydroxydopamine. Two weeks post lesion, these mice received injections of a vehicle, VD3, L-DOPA, or a combination of VD3/L-DOPA and compared with sham controls. Treatment lasted three weeks, during which motor-cognitive neurobehaviour was assessed. Five weeks post lesion, brains were collected and striatal levels of the following proteins assessed: tyrosine hydroxylase (TH), dopamine decarboxylase (DDC), monoamine oxidase (MAO-B), Catechol-O-methyl transferase (COMT), dopamine transporter (DAT), brain-derived neurotrophic factor (BDNF), microglia marker (CD11b), inflammation (IL-1ß), apoptotic signaling (BAX) and oxidative stress (p47phox).Results: Treatment with VD3 attenuated behavioural deficits induced by 6-OHDA, protein associated with dopamine metabolism and biomarkers of oxidative stress. VD3 significantly increased contralateral wall touches, exploratory motor and cognitive activities. VD3 significantly enhanced the expression of TH, DAT, BDNF, while significantly reducing expression of MAO-B, CD11b, IL-I ß and p47phox.Conclusion: VD3 reversed some of the 6-OHDA induced changes in proteins involved in modulating the dopamine system, behavioural deficits and oxidative stress biomarkers. The data suggests that VD3 might be beneficial in reducing L-DOPA dosage, thereby reducing problems associated with dosage and prolonged use of L-DOPA in PD management.

Evaluation of manganese application after soil stabilization to effectively reduce cadmium in rice.

In order to produce safe rice from cadmium (Cd) contaminated soils, a special pot experiment in the field was proposed to facilitate the study of multiple remediation measures. In the field experiments, four treatments were selected for the first half of the year: rice without soil treatment (R); rice with zeolite stabilization (RZ); oilseed rape phytoremediation (OR); and inter-cropping Sedum alfredii and maize phytoremediation (IC). As the early rice with zeolite stabilization still contained elevated Cd, manganese (Mn) fertilizer was added in the late rice with a special pot experiment in the field. Results showed that, in the first crops, the grains of maize and oilseed rape contained Cd below the food standard limit, while Cd in rice grain exceeded the limit of 0.2 mg/kg. The RZ treatment did not reduce Cd in rice but decreased significantly Mn in rice straw. In the late rice, Mn fertilizer additionally reduced Cd in rice grain to 0.12 mg/kg in combination with the RZ treatment. Mn accumulation in rice straw was enhanced by Mn fertilizer. These results indicate that the pot experiment in the field provides a useful tool to further evaluate effective treatment combinations to reduce Cd in rice.

CA1 Spike Timing is Impaired in the 129S Inbred Strain During Cognitive Tasks.

A spontaneous mutation of the disrupted in schizophrenia 1 (Disc1) gene is carried by the 129S inbred mouse strain. Truncated DISC1 protein in 129S mouse synapses impairs the scaffolding of excitatory postsynaptic receptors and leads to progressive spine dysgenesis. In contrast, C57BL/6 inbred mice carry the wild-type Disc1 gene and exhibit more typical cognitive performance in spatial exploration and executive behavioral tests. Because of the innate Disc1 mutation, adult 129S inbred mice exhibit the behavioral phenotypes of outbred B6 Disc1 knockdown (Disc1-/-) or Disc1-L-100P mutant strains. Recent studies in Disc1-/- and L-100P mice have shown that impaired excitation-driven interneuron activity and low hippocampal theta power underlie the behavioral phenotypes that resemble human depression and schizophrenia. The current study compared the firing rate and connectivity profile of putative neurons in the CA1 of freely behaving inbred 129S and B6 mice, which have mutant and wild-type Disc1 genes, respectively. In cognitive behavioral tests, 129S mice had lower exploration scores than B6 mice. Furthermore, the mean firing rate for 129S putative pyramidal (pyr) cells and interneurons (int) was significantly lower than that for B6 CA1 neurons sampled during similar tasks. Analysis of pyr/int connectivity revealed a significant delay in synaptic transmission for 129S putative pairs. Sampled 129S pyr/int pairs also had lower detectability index scores than B6 putative pairs. Therefore, the spontaneous Disc1 mutation in the 129S strain attenuates the firing of putative pyr CA1 neurons and impairs spike timing fidelity during cognitive tasks.

Cholecalciferol (VD3) Attenuates L-DOPA-Induced Dyskinesia in Parkinsonian Mice Via Modulation of Microglia and Oxido-Inflammatory Mechanisms.

L-DOPA, the gold standard for managing Parkinson's disease (PD) is fraught by motor fluctuations termed L-Dopa-Induced Dyskinesia (LID). LID has very few therapeutic options. Hence, the need for preclinical screening of new interventions. Cholecalciferol (VD3) treatment reportedly improves motor deficit in experimental Parkinsonism. Therefore, the novel anti-dyskinetic effect of VD3 and its underlying mechanisms in LID was investigated. Dyskinesia was induced by chronic L-DOPA administration in parkinsonian (6-OHDA- lesioned) mice. The experimental groups: Control, Dyskinesia, Dyskinesia/VD3, and Dyskinesia/Amantadine were challenged with L-DOPA to determine the abnormal involuntary movements (AIMs) score during 14 days of VD3 (30 mg/kg) or Amantadine (40 mg/kg) treatment. Behavioral Axial, Limb & Orolingual (ALO) AIMs were scored for 1 min at every 20 mins interval, over a duration of 100 mins on days 1,3,7,11 and 14. Using western blot, striatum was assessed for expression of dopamine metabolic enzymes: Tyrosine Hydroxylase (TH) and Monoamine Oxidase-B (MAO-B); CD11b, BAX, P47phox, and IL-1ß. Cholecalciferol significantly attenuated AIMs only on days 11 & 14 with maximal reduction of 32.7%. Expression of TH and MAO-B was not altered in VD3 compared with dyskinetic mice. VD3 significantly inhibited oxidative stress (P47phox), apoptosis (BAX), inflammation (IL-1ß) and microglial activation (CD11b). VD3 showed anti-dyskinetic effects behaviorally by attenuating abnormal involuntary movements, modulation of striatal oxidative stress, microglial responses, inflammation, and apoptotic signaling; without affecting dopamine metabolic enzymes. Its use in the management of dyskinesia is promising. More studies are required to further evaluate these findings. Keywords: Cholecalciferol; L-DOPA-Induced Dyskinesia; Parkinson's Disease; Microglial; Oxidative stress; Inflammation.

Crossed Connections From Insular Cortex to the Contralateral Thalamus.

Sensory information in all modalities, except olfaction, is processed at the level of the thalamus before subsequent transmission to the cerebral cortex. This incoming sensory stream is refined and modulated in the thalamus by numerous descending corticothalamic projections originating in layer 6 that ultimately alter the sensitivity and selectivity for sensory features. In general, these sensory thalamo-cortico-thalamic loops are considered strictly unilateral, i.e., no contralateral crosstalk between cortex and thalamus. However, in contrast to this canonical view, we characterize here a prominent contralateral corticothalamic projection originating in the insular cortex, utilizing both retrograde tracing and cre-lox mediated viral anterograde tracing strategies with the Ntsr1-Cre transgenic mouse line. From our studies, we find that the insular contralateral corticothalamic projection originates from a separate population of layer 6 neurons than the ipsilateral corticothalamic projection. Furthermore, the contralateral projection targets a topographically distinct subregion of the thalamus than the ipsilateral projection. These findings suggest a unique bilateral mechanism for the top-down refinement of ascending sensory information.

An in vivo Cell-Based Delivery Platform for Zinc Finger Artificial Transcription Factors in Pre-clinical Animal Models.

Zinc finger (ZF), transcription activator-like effectors (TALE), and CRISPR/Cas9 therapies to regulate gene expression are becoming viable strategies to treat genetic disorders, although effective in vivo delivery systems for these proteins remain a major translational hurdle. We describe the use of a mesenchymal stem/stromal cell (MSC)-based delivery system for the secretion of a ZF protein (ZF-MSC) in transgenic mouse models and young rhesus monkeys. Secreted ZF protein from mouse ZF-MSC was detectable within the hippocampus 1 week following intracranial or cisterna magna (CM) injection. Secreted ZF activated the imprinted paternal Ube3a in a transgenic reporter mouse and ameliorated motor deficits in a Ube3a deletion Angelman Syndrome (AS) mouse. Intrathecally administered autologous rhesus MSCs were well-tolerated for 3 weeks following administration and secreted ZF protein was detectable within the cerebrospinal fluid (CSF), midbrain, and spinal cord. This approach is less invasive when compared to direct intracranial injection which requires a surgical procedure.

Production of polyhydroxyalkanoate copolymers containing 4-hydroxybutyrate in engineered Bacillus megaterium.

Despite being used as a common platform for the commercial production of many biochemicals, Bacilli are often overlooked as a source of industrial polyhydroxyalkanoates (PHAs), biodegradable plastic replacements. In addition to having a robust expression system, the lack of lipopolysaccharides and ease of lysis make Bacilli an attractive host for the production of PHAs. In this work, a Bacillus megaterium strain was engineered to generate poly(3-hydroxybutyrate-co-4-hydroxybutryate) (P[3HB-co-4HB]) copolymers, which are among the most useful and industrially-relevant copolymers. These copolymers had lower modulus and increased toughness, thus making the copolymer suitable for a broader range of applications. Due to high metabolic flux through succinate, the engineered B. megaterium strain produced P(3HB-co-4HB) with >10% mol fraction 4HB from glucose, without the use of highly regulated and expensive precursors or potentially damaging truncation of central biochemical pathways.

Distinct expression trend of signature antigens of Staphylococcus aureus osteomyelitis correlated with clinical outcomes.

The major limitations of clinical outcome predictions of osteomyelitis mediated by Staphylococcus aureus (S. aureus) are not specific and definitive. To this end, current studies aim to investigate host immune responses of trend changes of the iron-regulated surface determinant (Isd) of IsdA, IsdB, IsdH, cell wall-modifying proteins of amidase (Amd) and glucosaminidase (Gmd), and secreted virulence factor of chemotaxis inhibitory protein S. aureus (CHIPS) and staphylococcal complement inhibitor (SCIN) longitudinally to discover their correlationship with clinical outcomes. A total of 55 patients with confirmed S. aureus infection of the long bone by clinical and laboratory methods were recruited for the study. Whole blood was collected at 0, 6, 12 months for the serum that was used to test IsdA, IsdB, IsdH, Gmd, Amd, CHIPS, and SCIN using a customized Luminex assay after clinical standard care parameters were collected. The patients were then divided into two groups: (1) infection controlled versus (2) adverse outcome based on clinical criteria for statistical analysis. We found that standard clinical parameters were unable to distinguish therapeutic outcomes. Significant overexpression of all antigens was confirmed in infection patients at 0-, 6-, and 12-month time points. A distinct expression trend and dynamic changes of IsdB, Amd, Gmd, and CHIPS were observed between infection controlled and adverse outcome patients, while the IsdA, IsdH, SCIN remained demonstrated no statistical significance. We conclude that dynamic changes of specific antigens could predict clinical outcomes of S. aureus osteomyelitis. Clinical Relevance: The trend changes of host immune responses to S. aureus specific antigens of IsdB, Gmd, Amd, and CHIPS could predict clinical outcomes of S. aureus osteomyelitis.

Positive Modulation of SK Channel Impedes Neuron-Specific Cytoskeletal Organization and Maturation.

N-methyl-D-aspartate receptor (NMDAR) modulates the structural plasticity of dendritic spines by impacting cytoskeletal organization and kinase signaling. In the developing nervous system, activation of NMDAR is pertinent for neuronal migration, neurite differentiation, and cellular organization. Given that small conductance potassium channels (SK2/3) repress NMDAR ionotropic signaling, this study highlights the impact of neonatal SK channel potentiation on adult cortical and hippocampal organization. Neonatal SK channel potentiation was performed by one injection of SK2/3 agonist (CyPPA) into the pallium of mice on postnatal day 2 (P2). When the animals reached adulthood (P55), the hippocampus and cortex were examined to assess neuronal maturation, lamination, and the distribution of synaptic cytoskeletal proteins. Immunodetection of neuronal markers in the brain of P2-treated P55 mice revealed the presence of immature neurons in the upper cortical layers (layers II-IV) and CA1 (hippocampus). Also, layer-dependent cortical-cell density was attenuated due to the ectopic localization of mature (NeuN+) and immature (Doublecortin+ [DCX+]) neurons in cortical layers II-IV. Similarly, the decreased count of NeuN+ neurons in the CA1 is accompanied by an increase in the number of immature DCX+ neurons. Ectopic localization of neurons in the upper cortex and CA1 caused the dramatic expression of neuron-specific cytoskeletal proteins. In line with this, structural deformity of neuronal projections and the loss of postsynaptic densities suggests that postsynaptic integrity is compromised in the SK2/3+ brain. From these results, we deduced that SK channel activity in the developing brain likely impacts neuronal maturation through its effects on cytoskeletal formation.

A Bioinformatic Approach to Utilize a Patient's Antibody-Secreting Cells against Staphylococcus aureus to Detect Challenging Musculoskeletal Infections.

Noninvasive diagnostics for Staphylococcus aureus musculoskeletal infections (MSKI) remain challenging. Abs from newly activated, pathogen-specific plasmablasts in human blood, which emerge during an ongoing infection, can be used for diagnosing and tracking treatment response in diabetic foot infections. Using multianalyte immunoassays on medium enriched for newly synthesized Abs (MENSA) from Ab-secreting cells, we assessed anti-S. aureus IgG responses in 101 MSKI patients (63 culture-confirmed S. aureus, 38 S. aureus-negative) and 52 healthy controls. MENSA IgG levels were assessed for their ability to identify the presence and type of S. aureus MSKI using machine learning and multivariate receiver operating characteristic curves. Eleven S. aureus-infected patients were presented with prosthetic joint infections, 15 with fracture-related infections, 5 with native joint septic arthritis, 15 with diabetic foot infections, and 17 with suspected orthopedic infections in the soft tissue. Anti-S. aureus MENSA IgG levels in patients with non-S. aureus infections and healthy controls were 4-fold (***p = 0.0002) and 8-fold (****p < 0.0001) lower, respectively, compared with those with culture-confirmed S. aureus infections. Comparison of MENSA IgG responses among S. aureus culture-positive patients revealed Ags predictive of active MSKI (IsdB, SCIN, Gmd) and Ags predictive of MSKI type (IsdB, IsdH, Amd, Hla). When combined, IsdB, IsdH, Gmd, Amd, SCIN, and Hla were highly discriminatory of S. aureus MSKI (area under the ROC curve = 0.89 [95% confidence interval 0.82-0.93, p < 0.01]). Collectively, these results demonstrate the feasibility of a bioinformatic approach to use a patient's active immune proteome against S. aureus to diagnose challenging MSKI.

Disc1 Carrier Mice Exhibit Alterations in Neural pIGF-1Rß and Related Kinase Expression.

Mutation of the disc1 gene underlies a broad range of developmental neuropsychiatric defects, including schizophrenia, depression, and bipolar disorder. The pathophysiological phenotypes linked with disc1 mutation are due to the truncation of the DISC1 primary protein structure. This leads to a defective post-synaptic scaffolding and kinase-GSK3ß and Erk1/2-signaling. As a result, synaptic function and maintenance are significantly impaired in the disc1 mutant brain. Among several other pathways, GSK3ß and Erk1/2 are involved in insulin-like growth factor 1 receptor (IGF-1Rß) kinase signaling. Although disc1 mutation alters these kinases, it is unclear if the mutation impacts IGF-1R expression and activity in the brain. Here, we demonstrate that the expression of active IGF-1Rß (pIGF-1Rß) is altered in the hippocampus and prefrontal cortex (PFC) of disc1 mutant mice and vary with the dose of the mutation (homozygous and heterozygous). The expression of pIGF-1Rß decreased significantly in 129S (hom, disc1 -/-) brains. In contrast, 129S:B6 (het, disc1 +/-) brains were characterized by an increase in pIGF-1Rß when compared with the C57BL/6 (disc1 +/+) level. The decrease in pIGF-1Rß level for the 129S brains was accompanied by the loss of Akt activity (S473 pAkt) and decreased Ser9 phosphorylation of GSK3ß (increased basal GSK3ß). Additionally, hippocampal and cortical pErk1/2 activity increased in the 129S hippocampus and cortex. Although 129S:B6 recorded alterations in pIGF-1Rß-pAkt-GSK3ß (like 129S), there was no observable change in pErk1/2 activity for the heterozygote (disc1 +/-) mutant. In addition to GSK3ß inhibition, we conclude that pIGF-1R, pAkt, and pErk1/2 are potential targets in disc1 -/- mutant brain. On the other hand, pIGF-1R and pAkt can be further explored in disc1 +/- brain.