The efficacy and safety of tadalafil in the management of erectile dysfunction with diabetes and blood circulation issues.

BACKGROUND: Erectile dysfunction (ED) is a common diabetes-related complication. MATERIALS AND METHODS: This study examined the effect of daily low-dose tadalafil (5 mg) on patients' quality of life (including that of sex life) and blood circulation. Erectile dysfunction questionnaires were administered to 20 patients with type 2 diabetes (T2DM) and ED. The safety and efficacy of tadalafil were evaluated using laboratory tests, and the effect on blood circulation was measured through nail fold capillaroscopy. RESULTS: Daily tadalafil use by patients with T2DM and ED showed a statistically significant increase in the erectile reliability score from of 1.15 to 3.20 (p < .00012). Capillary blood circulation improvement tests showed a statistically significant increase in apical limb width from 13.1 to 14.64 µm (p = .04829) and flow from 9035 to 11946 µm3/s (p = .04405). Although not significant, increased capillary width and speed (rate of blood flow) confirmed improved blood circulation. There were no significant changes in the cardiac indicators (troponin, prostate-specific antigen, or electrocardiogram tests) before and after tadalafil administration, supporting the safety of its low-dose daily administration. CONCLUSIONS: A small dose of daily tadalafil was shown to safely improve erectile dysfunction and peripheral blood flow in patients with T2DM, in which peripheral arterial diseases should not be considered separately but rather as complex entities.

Implementing Point-of-Care Ultrasound for Acute Scrotal Pain in the Pediatric Emergency Department: Screening Testicular Torsion and Patient Flow Analysis.

OBJECTIVES: Testicular torsion (TT) is a pediatric surgical emergency that requires prompt treatment. This study investigated the feasibility of point-of-care ultrasound (POCUS) for diagnosing TT in the pediatric emergency department (ED). METHODS: We retrospectively reviewed the medical records of patients, aged 18 years or younger, who visited a university-affiliated hospital pediatric ED with acute scrotal pain without trauma history and underwent diagnostic ultrasounds between January 2010 and October 2022. RESULTS: This study included 731 patients (median age: 9 years), Of these, 315 (43%) were in the POCUS-performed group: 188 in the POCUS-only group, and 127 in the POCUS-and-RADUS group. The other 416 patients (56.9%) were in the RADUS-only group. In total, 45 patients (6.2%) were diagnosed with TT (19 in the POCUS-performed group and 26 in the RADUS-only group). The sensitivity, specificity, and positive and negative predictive values of POCUS for diagnosing TT were 94.7%, 92.9%, 46.2%, and 99.6%, respectively. The median time to perform POCUS was shorter than RADUS (23 versus 61 minutes, P < .001). The POCUS-performed group had a shorter ED length of stay than the RADUS-only group (93 versus 170 minutes, P < .001). Among the patients diagnosed with TT, performing POCUS first did not significantly delay the ED process, including time to operation (250 versus 205 minutes, P = .142). CONCLUSIONS: For patients with acute scrotal pain, evaluation performed by pediatric emergency physicians using POCUS performs well in screening TT, and can decrease length of stay in the ED.

Vertical Nanowire Electrode Array for Enhanced Neurogenesis of Human Neural Stem Cells via Intracellular Electrical Stimulation.

Extracellular electrical stimulation (ES) can provide electrical potential from outside the cell membrane, but it is often ineffective due to interference from external factors such as culture medium resistance and membrane capacitance. To address this, we developed a vertical nanowire electrode array (VNEA) to directly provide intracellular electrical potential and current to cells through nanoelectrodes. Using this approach, the cell membrane resistivity and capacitance could be excluded, allowing effective ES. Human fetal neural stem cells (hfNSCs) were cultured on the VNEA for intracellular ES. Combining the structural properties of VNEA and VNEA-mediated ES, transient nanoscale perforation of the electrode was induced, promoting cell penetration and delivering current to the cell. Intracellular ES using VNEA improved the neuronal differentiation of hfNSCs more effectively than extracellular ES and facilitated electrophysiological functional maturation of hfNSCs because of the enhanced voltage-dependent ion-channel activity. The results demonstrate that VNEA with advanced nanoelectrodes serves as a highly effective culture and stimulation platform for stem-cell neurogenesis.

Clinical utility of procalcitonin in febrile infants younger than 3 months of age visiting a pediatric emergency room: a retrospective single-center study.

BACKGROUND: Fever in infants under 90 days of age is highly likely to be caused by a severe bacterial infection (SBI) and it accounts for a large number of patients visiting the pediatric emergency room. In order to predict the bacterial infection and reduce unnecessary treatment, the classic classification system is based on white blood cell (WBC) count, urinalysis, and x-ray, and it is modified and applied at each center by incorporating recently studied biomarkers such as c-reactive protein (CRP) or procalcitonin (PCT). This study analyzed the usefulness of PCT in predicting SBI when applied along with the existing classification system, including CRP, among infants less than 90 days old who visited with a fever at a single institution pediatric emergency center. METHODS: We retrospectively reviewed the medical records of patients younger than 3 months of age who presented with fever at the Seoul Asan Medical Center pediatric emergency room between July 2017 and October 2018. RESULTS: A total of 317 patients were analyzed, and 61 were diagnosed with SBI, among which urinary tract infection (UTI) accounted for the largest proportion (55/61, 90.2%). There were differences in WBC, neutrophil proportion, CRP, and PCT between the SBI group and the non-SBI group, and the AUC values of WBC, CRP, and PCT were 0.651, 0.804, and 0.746, respectively. When using the cut-off values of CRP and PCTs as 2.0 mg/dL and 0.3 ng/mL, respectively, the sensitivity and specificity for SBI were 49.2/89.5, and 54.1/87.5, respectively. WBC, CRP, and PCT were statistically significant for predicting SBI in multivariate analysis (odds ratios 1.066, 1.377, and 1.291, respectively). When the subjects were classified using the existing classification criteria, WBC and CRP, the positive predictive value (PPV) and negative predictive value (NPV) were 29.3 and 88.7%, respectively, and when PCT was added, the PPV and NPV were 30.7 and 92%, respectively, both increased. CONCLUSION: PCT is useful for predicting SBI in children aged 3 months or less who visit the emergency room with a fever. It is useful as a single biomarker, and when used in conjunction with classic biomarkers, its diagnostic accuracy is further increased.

Pediatric Sedation in the Emergency Department: Trends from a Nationwide Population-based Study in Korea, 2007-2018.

BACKGROUND: Pediatric sedation in the emergency department (ED) is widely performed in Korea; thus exploring the trends of its use is necessary. This study aimed to investigate the characteristics of patients and sedatives use in the ED and verify their changes over recent years. METHODS: A nationwide population-based retrospective study was conducted including pediatric patients aged ≤ 15 years who received sedative medication in the ED and were discharged during 2007-2018, using the Korean Health Insurance Review and Assessment Service database. Patient characteristics (age, sex, level of ED, and diagnosis) and type of sedative used were analyzed. RESULTS: Sedation was performed in total 468,221 visits during 2007-2018 (399,320 visits, at least 3.8% of overall ED visits during 2009-2018). Among these, 71.0% were children aged 1-3 years and 93.5% were sedated to support diagnosis of injury. An increase in total sedation was observed in patients aged 4-6 years during the study period (from 13.8% to 21.8%). A gradual decrease in the use of chloral hydrate (CH) compared with an increase in ketamine use was observed (CH, from 70.6% to 28.6%; ketamine, from 23.8% to 60.7%). Therefore, ketamine was the most used sedative since 2014. The most frequently used sedatives over the study period differed according to age groups (CH in <1 year and 1-3 years; ketamine in 4-6 years and 7-10 years; and midazolam in 11-15 years). CONCLUSIONS: The characteristics of patients related to sedatives use in the ED have changed over time. These changes should be considered in the development of future Korean guidelines regarding pediatric sedation in the ED.

Feasibility of Point-of-Care Ultrasound for Diagnosing Hypertrophic Pyloric Stenosis in the Emergency Department.

OBJECTIVES: This study aimed to investigate the feasibility of point-of-care ultrasound (POCUS) for diagnosing hypertrophic pyloric stenosis (HPS) in the emergency department (ED). METHODS: A retrospective study was conducted in infants aged younger than 90 days who were brought to the ED due to vomiting between January 2015 and December 2019. Of these, infants who were clinically suspected of having HPS and underwent ultrasound were included and categorized into 3 groups: POCUS only, POCUS followed by radiologist-performed ultrasound (RADUS), and RADUS only. All confirmative diagnoses of HPS were made by RADUS. The diagnostic performance of POCUS was analyzed, and the ED patient flow was compared between the POCUS-performed (POCUS only or POCUS followed by RADUS) and RADUS-only groups. RESULTS: Overall, 171 patients with a median age of 34 days were included. Of these, 79 patients (46.2%) underwent POCUS only, and none had HPS; 50 patients (29.2%) underwent POCUS followed by RADUS; and 42 patients (24.5%) underwent RADUS only. Overall, 41 patients (24.0%) were diagnosed with HPS, and POCUS showed a sensitivity of 96.6% and specificity of 94.0%. In the total cohort, length of stay in the ED (EDLOS) was shorter in the POCUS-performed group than in the RADUS-only group (2.6 vs 3.8 hours, P = 0.015). Among non-HPS patients, time to disposition (1.8 vs 2.7 hours, P = 0.005) and EDLOS (2.0 vs 3.0 hours, P = 0.004) were shorter in the POCUS-performed group than in the RADUS-only group. Performing POCUS followed by RADUS did not significantly delay the treatment among HPS patients. CONCLUSIONS: Point-of-care ultrasound is accurate and useful for diagnosing HPS and improved the ED patient flow by reducing EDLOS and door-to-disposition time in non-HPS patients.

Initial Characteristics and Clinical Severity of Hemophagocytic Lymphohistiocytosis in Pediatric Patients Admitted in the Emergency Department.

OBJECTIVES: The diagnosis and management of children with hemophagocytic lymphohistiocytosis (HLH) admitted in the emergency department (ED) are challenging. The present study aimed at describing the initial characteristics of pediatric patients with HLH upon admission in the ED. Moreover, the clinical severity of the condition was assessed. METHODS: We performed a retrospective study of patients who visited the pediatric ED and were newly diagnosed with HLH during hospitalization between February 2012 and January 2017. The patients were classified in the clinically unstable group if at least 1 of the following conditions was observed upon admission in the ED: hypoxia requiring oxygen supplementation, hypotension requiring inotropic support, coagulopathy with prothrombin time (international normalized ratio, ≥1.5), and seizures or altered consciousness. RESULTS: We enrolled 31 pediatric patients with HLH, with a median age of 6.53 years (interquartile range, 1.35-13.24 years). Abdominal discomfort along with fever (74.2%) was the most common presenting symptom in patients admitted in the ED. Based on the HLH-2004 diagnostic criteria, fever (96.8%), hyperferritinemia (96.8%), splenomegaly (74.2%), hypertriglyceridemia and/or hypofibrinogenemia (67.7%), and bicytopenia (41.9%) were observed in the patients. However, only 8 patients (25.8%) met the criteria. Nineteen patients (61.3%) were included in the clinically unstable group. This group had lower albumin (2.3 vs 3.3 g/dL, P = 0.002) and fibrinogen levels and higher ferritin level and neutrophil count than the clinically stable group. Meanwhile, the number of clinical findings that met the diagnostic criteria was not different between the 2 groups. Lower albumin level was a significant risk factor in the clinically unstable group (odds ratio, 0.040; P = 0.004). CONCLUSIONS: Pediatric patients with HLH often have clinically unstable conditions upon admission in the ED. However, only few patients meet the HLH-2004 diagnostic criteria. Lower albumin level may be useful in assessing clinically unstable patients and preparing for possible deterioration.

Point-of-care ultrasound may be useful for detecting pediatric intussusception at an early stage.

BACKGROUND: This study aimed to verify the usefulness of point-of-care ultrasound (POCUS) performed by pediatric emergency physicians for detecting intussusception at an early stage. METHODS: This retrospective study included 1-month- to 6-year-old children with clinically suspected intussusception, who underwent POCUS in the pediatric emergency department between December 2016 and February 2018. The criteria for performing POCUS were set to broader standards: presenting any one of intermittent abdominal pain/irritability or bloody stool, or ≥ 2 symptoms among nonspecific abdominal pain/irritability, abdominal mass/distension, vomiting, or lethargy. POCUS results were interpreted and categorized as "negative" or "suspicious," and a radiologist performed confirmatory ultrasound in "suspicious" cases. RESULTS: We analyzed 575 POCUS scans from 549 patients (mean age, 25.5 months). Among the 92 "suspicious" cases (16.0%), 70 (12.2%) were confirmed to have intussusception. POCUS showed 100% sensitivity, 95.6% specificity, and 97.8% accuracy. Patients with confirmed intussusception were mainly diagnosed in the early stages, with a mean symptom duration of 11.7 h, and most patients (97.1%) were treated successfully via air enema reduction. Compared to the non-intussusception group, the intussusception group had more intermittent abdominal pain (P < 0.001), but less vomiting (P = 0.001); the other clinical features showed no intergroup differences. CONCLUSION: POCUS performed using the criteria set to broader standards by pediatric emergency physicians may be useful for detecting intussusception at an early stage, which may present with obscure clinical symptoms.

Lactic acid level as an outcome predictor in pediatric patients with intussusception in the emergency department.

BACKGROUND: Intussusception decreases blood flow to the bowel, and tissue hypoperfusion results in increased lactic acid levels. We aimed to determine whether lactic acid levels are associated with pediatric intussusception outcomes. METHODS: The electronic medical records of our emergency department pediatric patients diagnosed with intussusception, between January 2015 and October 2018, were reviewed. An outcome was considered poor when intussusception recurred within 48 h of reduction or when surgical reduction was required due to air enema failure. RESULTS: A total of 249 patients were included in the study, including 39 who experienced intussusception recurrence and 11 who required surgical reductions; hence, 50 patients were included in the poor outcome group. The poor and good outcome groups showed significant differences in their respective blood gas analyses for pH (7.39 vs. 7.41, P = .001), lactic acid (1.70 vs. 1.30 mmol/L, P < .001), and bicarbonate (20.70 vs. 21.80 mmol/L, P = .036). Multivariable logistic regression analyses showed that pH and lactic acid levels were the two factors significantly associated with poor outcomes. When the lactic acid level cutoff values were ≥ 1.5, ≥2.0, ≥2.5, and ≥ 3.0 mmol/L, the positive predictive values for poor outcomes were 30.0, 34.6, 50.0, and 88.9%, respectively. CONCLUSION: Lactic acid levels affect outcomes in pediatric patients with intussusception; higher lactic acid levels are associated with higher positive predictive values for poor outcomes.

Point-of-Care Ultrasound Could Streamline the Emergency Department Workflow of Clinically Nonspecific Intussusception.

OBJECTIVES: The aim of this study was to evaluate whether point-of-care ultrasound (POCUS) for intussusception screening streamlines the workflow of clinically nonspecific intussusception (CNI), an intussusception presenting with only 1 manifestation of the classic triad, and/or vomiting. METHODS: We reviewed 274 consecutive children with intussusception, aged 6 years or younger, who visited a tertiary care hospital emergency department between May 2012 and April 2016. This period was dichotomized by May 2014 (the "PRE" and "POST" groups), starting point of implementation of the POCUS protocol for intussusception screening. All children with CNI who had positive results on or forwent POCUS underwent radiologist-performed ultrasound (US). We measured and compared emergency department length of stay (EDLOS), the sum of door-to-reduction and observation times, and the frequency of POCUS and positive US results between the 2 groups. RESULTS: Of 160 children with CNI, 93 visited the emergency department since May 2014. The POST group showed a shorter median EDLOS (856 vs 630 minutes, P < 0.001), door-to-reduction time (137 vs 111 minutes, P = 0.002), and observation time (700 vs 532 minutes, P < 0.001). The POST group had a higher frequency of POCUS (12% vs 60%, P < 0.001) with positive US results (33% vs 59%, P < 0.001). The PRE group had a higher frequency of severe bowel edema (16% vs 1%, P < 0.001). No significant differences were found in the severity, recurrence, admission, and surgery. One child had a false-negative result on POCUS. CONCLUSIONS: Point-of-care ultrasound could streamline the workflow of CNI via decrease in EDLOS and unnecessary referrals for US.

Implications of Increased Weight Status for the Occurrence of Fall-Induced Intracranial Hemorrhage in Children Aged 4 Years or Younger.

OBJECTIVES: The aim of this study was to investigate the association between increased weight status (IWS), a weight for age/sex at greater than the 95th percentile, and fall-induced intracranial hemorrhage (ICH) in children aged 4 years or younger. METHODS: In 7072 children aged 4 years or younger with head injury who visited a tertiary care hospital emergency department in Korea from 2013 through 2015, the presence of fall-induced ICH was reviewed. The association between IWS and ICH was investigated by multivariable logistic regression. We retrospectively validated the Pediatric Emergency Care Applied Research Network rule alone and in combination with IWS for predicting ICH. RESULTS: Of 7072 children, 547 (7.7%) underwent computed tomography, of whom 451 (6.4%) were enrolled. Of these, 41 (9.1%; estimated event rate, 0.6%) had ICHs, and 26 (5.8%) had IWS. Increased weight status was more common in the children with ICH (P = 0.023). The association between IWS and ICH remained significant after adjustment (odds ratio, 5.24; 95% confidence interval [CI], 1.49-18.46; P = 0.010). The validation of the rule in combination with IWS showed no significant increases in a sensitivity (92.7% [95% CI, 80.1%-98.5%] to 95.7% [95% CI, 83.5%-99.4%]) and negative predictive value (98.2% [95% CI, 94.7%-99.4%] to 98.8% [95% CI, 95.4%-99.8%]). CONCLUSIONS: Increased weight status is associated with fall-induced ICH in children aged 4 years or younger. Information on weight status could be potentially helpful in predicting ICH in young children with fall-induced head injury.

Magnetic Control of Axon Navigation in Reprogrammed Neurons.

While neural cell transplantation represents a promising therapy for neurodegenerative diseases, the formation of functional networks of transplanted cells with host neurons constitutes one of the challenging steps. Here, we introduce a magnetic guidance methodology that controls neurite growth signaling via magnetic nanoparticles (MNPs) conjugated with antibodies targeting the deleted in colorectal cancer (DCC) receptor (DCC-MNPs). Activation of the DCC receptors by clusterization and subsequent axonal growth of the induced neuronal (iN) cells was performed in a spatially controlled manner. In addition to the directionality of the magnetically controlled axon projection, axonal growth of the iN cells assisted the formation of functional connections with pre-existing primary neurons. Our results suggest magnetic guidance as a strategy for improving neuronal connectivity by spatially guiding the axonal projections of transplanted neural cells for synaptic interactions with the host tissue.

High Concentration of C-Reactive Protein Is Associated With Serious Bacterial Infection in Previously Healthy Children Aged 3 to 36 Months With Fever and Extreme Leukocytosis.

OBJECTIVES: The aim of the study was to investigate the diagnostic efficacy of C-reactive protein (CRP) in predicting serious bacterial infection (SBI) in febrile children aged 3 to 36 months with extreme leukocytosis (EL), defined as a peripheral white blood cell count of 25,000 to 49,999/mm. METHODS: The presence of SBI was reviewed in previously healthy children aged 3 to 36 months with a fever of 39°C or higher and EL, who visited a tertiary care hospital emergency department between September 2010 and August 2015. We measured cutoff values of CRP with corresponding likelihood ratios (LRs) and posttest probabilities (PPs). RESULTS: Of 9989 febrile children, 4252 (42.6%) underwent assays for white blood cell and CRP. Of 233 (5.5%) children with EL, 133 (3.1%) were enrolled, of whom 43 (32.3%; 95% confidence interval [CI], 25.0%-40.7%) had SBI, including 33 pyelonephritis, 5 deep abscesses, 3 lobar pneumonia, and 2 soft tissue infections. With the cutoff of 7.8 mg/dL, the sensitivity of CRP in predicting SBI was 81.4% (95% CI, 67.4%-90.3%); the specificity, 80.0% (95% CI, 70.6%-87.0%); the positive predictive value, 66.0% (95% CI, 52.6%-77.3%); the negative predictive value, 90.0% (95% CI, 81.5%-94.9%); the positive LR, 4.1; and the PP, 66.0% (95% CI, 55.6%-75.0%). A CRP concentration of 16.1 mg/dL or higher yielded an LR of 11.2 and a PP of 84.2% (95% CI, 62.1%-94.5%). A CRP concentration of lower than 3.4 mg/dL yielded an LR of 0.05 and a PP of 2.4% (95% CI, 0.3%-14.6%). CONCLUSIONS: High concentration of CRP is strongly associated with the presence of SBI in febrile children with EL.

Requirement for Head Magnetic Resonance Imaging in Children Who Present to the Emergency Department With Acute Nontraumatic Visual Disturbance.

OBJECTIVES: This study aimed to investigate the clinical features and head magnetic resonance imaging (MRI) findings in children who presented to the emergency department with acute nontraumatic visual disturbance and to study related clinical factors for discovering positive lesions on head MRI. METHODS: We performed a retrospective study of 1-month to 15-year-old children who underwent head MRI as an evaluation for acute nontraumatic visual disturbance as a chief complaint in our pediatric emergency department between March 2010 and March 2015. The symptoms of visual disturbance were blurred vision, diplopia, loss of vision, and visual hallucination. Head MRI findings were considered positive when lesions could explain the symptoms. RESULTS: We identified 39 patients (25 with blurred vision, 9 with diplopia, 3 with loss of vision, and 2 with visual hallucination) with a mean age of 8.35 ± 4.06 years. Positive head MRI findings were identified in 13 patients (33.3%). Brain tumors were most common (53.8%), followed by optic nerve inflammations (23.1%), congenital brain lesions (15.4%), and hypertensive encephalopathy (7.7%). Compared with the negative head MRI group, the positive head MRI group showed significantly less transient visual disturbance (duration <1 hour to complete recovery) (P = 0.001), more limited eye movement (P = 0.003), and more pupillary abnormalities (P = 0.030). CONCLUSIONS: We suggest performing urgent head MRI in children with acute nontraumatic visual disturbance if the symptoms last longer than 1 hour without complete recovery and are accompanied by limited eye movement or pupillary abnormality.

Risk Factors for Recurrent Intussusception After Fluoroscopy-Guided Air Enema.

OBJECTIVES: The aim of this study was to identify risk factors for recurrent intussusception after a successful reduction by fluoroscopy-guided air enema, the time required for recurrence, and the association between delayed reduction and the recurrence. METHODS: Medical records of 479 consecutive children with intussusception who underwent fluoroscopy-guided air enema between January 2004 and September 2014 were reviewed. Recurrent intussusception was defined as a recurrence within 48 hours of a reduction. Symptom-to-door time was defined as the time from symptom onset to emergency department arrival. Door-to-reduction time was defined as the time from emergency department arrival to reduction. Time-to-recurrence was defined as the time required for recurrence from the first ultrasound diagnosis. RESULTS: Of the 360 eligible children, 32 had recurrent intussusceptions (8.9%). Multivariable logistic regression showed that age 2 years or older is an independent predictor of recurrent intussusception (odds ratio, 2.39; 95% confidence interval, 1.13-5.02; P = 0.02). Median time to recurrence was 25 hours (18.0-36.0 hours). Although symptom-to-door and door-to-reduction times tended to be longer in the recurrence group, these differences were not significant (12.5 vs 7.0 hours, P = 0.18; 154.0 vs 143.0 minutes, P = 0.67, respectively). CONCLUSIONS: Our data suggest that provision for early recurrence and extended observation may be beneficial for children 2 years or older. Delayed reduction was not associated with recurrent intussusception, but further studies with larger sample sizes are needed to explain this issue.

Mussel Adhesion-Inspired Reverse Transfection Platform Enhances Osteogenic Differentiation and Bone Formation of Human Adipose-Derived Stem Cells.

Using small interfering RNA (siRNA) to regulate gene expression is an emerging strategy for stem cell manipulation to improve stem cell therapy. However, conventional methods of siRNA delivery into stem cells based on solution-mediated transfection are limited due to low transfection efficiency and insufficient duration of cell-siRNA contact during lengthy culturing protocols. To overcome these limitations, a bio-inspired polymer-mediated reverse transfection system is developed consisting of implantable poly(lactic-co-glycolic acid) (PLGA) scaffolds functionalized with siRNA-lipidoid nanoparticle (sLNP) complexes via polydopamine (pDA) coating. Immobilized sLNP complexes are stably maintained without any loss of siRNA on the pDA-coated scaffolds for 2 weeks, likely due to the formation of strong covalent bonds between amine groups of sLNP and catechol group of pDA. siRNA reverse transfection with the pDA-sLNP-PLGA system does not exhibit cytotoxicity and induces efficient silencing of an osteogenesis inhibitor gene in human adipose-derived stem cells (hADSCs), resulting in enhanced osteogenic differentiation of hADSCs. Finally, hADSCs osteogenically committed on the pDA-sLNP-PLGA scaffolds enhanced bone formation in a mouse model of critical-sized bone defect. Therefore, the bio-inspired reverse transfection system can provide an all-in-one platform for genetic modification, differentiation, and transplantation of stem cells, simultaneously enabling both stem cell manipulation and tissue engineering.

Casein kinase 2 complex: a central regulator of multiple pathobiological signaling pathways in Cryptococcus neoformans.

The casein kinase 2 (CK2) complex has garnered extensive attention over the past decades as a potential therapeutic target for diverse human diseases, including cancer, diabetes, and obesity, due to its pivotal roles in eukaryotic growth, differentiation, and metabolic homeostasis. While CK2 is also considered a promising antifungal target, its role in fungal pathogens remains unexplored. In this study, we investigated the functions and regulatory mechanisms of the CK2 complex in Cryptococcus neoformans, a major cause of fungal meningitis. The cryptococcal CK2 complex consists of a single catalytic subunit, Cka1, and two regulatory subunits, Ckb1 and Ckb2. Our findings show that Cka1 plays a primary role as a protein kinase, while Ckb1 and Ckb2 have major and minor regulatory functions, respectively, in growth, cell cycle control, morphogenesis, stress response, antifungal drug resistance, and virulence factor production. Interestingly, triple mutants lacking all three subunits (cka1Δ ckb1Δ ckb2Δ) exhibited more severe phenotypic defects than the cka1Δ mutant alone, suggesting that Ckb1/2 may have Cka1-independent functions. In a murine model of systemic cryptococcosis, cka1Δ and cka1Δ ckb1Δ ckb2Δ mutants showed severely reduced virulence. Transcriptomic, proteomic, and phosphoproteomic analyses further revealed that the CK2 complex controls a wide array of effector proteins involved in transcriptional regulation, cell cycle control, nutrient metabolisms, and stress responses. Most notably, CK2 disruption led to dysregulation of key signaling cascades central to C. neoformans pathogenicity, including the Hog1, Mpk1 MAPKs, cAMP/PKA, and calcium/calcineurin signaling pathways. In summary, our study provides novel insights into the multifaceted roles of the fungal CK2 complex and presents a compelling case for targeting it in the development of new antifungal drugs.IMPORTANCEThe casein kinase 2 (CK2) complex, crucial for eukaryotic growth, differentiation, and metabolic regulation, presents a promising therapeutic target for various human diseases, including cancer, diabetes, and obesity. Its potential as an antifungal target is further highlighted in this study, which explores CK2's functions in C. neoformans, a key fungal meningitis pathogen. The CK2 complex in C. neoformans, comprising the Cka1 catalytic subunit and Ckb1/2 regulatory subunits, is integral to processes like growth, cell cycle, morphogenesis, stress response, drug resistance, and virulence. Our findings of CK2's role in regulating critical signaling pathways, including Hog1, Mpk1 MAPKs, cAMP/PKA, and calcium/calcineurin, underscore its importance in C. neoformans pathogenicity. This study provides valuable insights into the fungal CK2 complex, reinforcing its potential as a target for novel antifungal drug development and pointing out a promising direction for creating new antifungal agents.

Toxicity of a novel antifungal agent (ATB1651 gel) in Yucatan minipigs (Sus scrofa) following 4 weeks of daily dermal administration.

ATB1651 gel is an antifungal drug candidate that enhances antifungal activity through substitution of several aryl rings, alkyl chains, and methyl groups. To ensure safety of use of ATB1651 gel, assessment of its potentially toxic side effects is necessary. In this study, we examined the repeated-dose toxicity of ATB1651 gel to Yucatan minipigs (Sus scrofa) in accordance with the Good Laboratory Practice guidelines. Five doses of ATB1651 gel (0%, 0.2%, 0.5%, 1.0%, 3.0%) were administered dermally to the left and right flanks of 38 minipigs daily for 4 weeks. Mortality, clinical symptoms, dermal scores, body weights, and physiological, biochemical, pathological, and toxicokinetic analyses were performed after the treatment period. No systemic toxicological damage was observed in either male or female minipigs regardless of dose; however, dermal application of ATB1651 gel caused some skin alterations at the application sites. Specifically, erythema and eschar formation, edema, and scabs or raise spots were observed at the application site(s) in males in the 3.0% ATB1651 gel treatment group and in females at ATB1651 gel concentrations ≥ 1.0%, with dermal scores ranging from grade 1 to 2. Additionally, histopathological assay indicated infiltration of different types of inflammatory cells and the presence of pustule/crust at the application site(s) in both males and females at ATB1651 gel concentrations ≥ 0.5%. However, these changes were reversible after a 2-week recovery period and were considered a local irritation effect of ATB1651 gel. The no-observed-adverse-effect level of ATB1651 gel was 3.0% with regard to topical and systemic toxicity in both male and female minipigs. Collectively, our results imply that ATB1651 gel is a safe candidate for clinical development as an antifungal drug with a wide therapeutic window.

Effect of Non-invasive Spinal Stimulation on Self-sustained Firing Motoneuron Model: In-Silico Study Using Human Body Model.

Transcutaneous spinal electrical stimulation (tSCS) is a non-invasive neuromodulation approach using a low intensity direct current. Recent developments in the technique have opened the possibility that tSCS can help restore motor function after spinal cord injury (SCI). However, the exact mechanism of action tSCS has on the spinal circuits is still unknown. Due to the complexity of experimental synthesis in a human model to delineate the mechanisms, models that link the stimulation paradigm and circuit behaviors are advantageous. Thus, this study aims to simulate the underlying changes in motor circuit firing rates in response to external stimuli induced by tSCS. Serial stimulations combining a high-fidelity finite element model with the human torso and spinal cord with a lumped motor neuron model is constructed. The parameters for both components of the model were derived from previous studies. We focused our analysis on a lumped motor neuron model that describes sustained firing behavior of the motor neuron driven primarily by persistent inward current (PIC), a signature behavior of the motor neuron after SCI. Modulation of the PIC behaviors was achieved by stimulating voltage-dependent calcium and sodium channels in the dendrite using a tSCS-induced electric field (E-field) expressed at different a spatial locations of the motor neuron in the gray matter. The PIC behaviors of spinal motor neurons in the left ventral horn were suppressed, while for the most part invariant in the right ventral horn. These initial simulations will provide a steppingstone for future examinations that incorporate additional neuronal models of inhibitory and excitatory interneurons to access the circuit-level effect of spinal stimulation.

Adenylyl-Sulfate Kinase (Met14)-Dependent Cysteine and Methionine Biosynthesis Pathways Contribute Distinctively to Pathobiological Processes in Cryptococcus neoformans.

Blocking of nutrient uptake and amino acid biosynthesis are considered potential targets for next-generation antifungal drugs against pathogenic fungi, including Cryptococcus neoformans. In this regard, the sulfate assimilation pathway is particularly attractive, as it is only present in eukaryotes such as plants and fungi, yet not in mammals. Here, we demonstrated that the adenylyl sulfate kinase (Met14) in the sulfate assimilation pathway is not essential yet is required for the viability of C. neoformans due to its involvement in biosynthesis of two sulfur-containing amino acids, cysteine and methionine. Met14-dependent cysteine and methionine biosynthesis was found to significantly contribute to a diverse range of pathobiological processes in C. neoformans. Met14-dependent cysteine rather than methionine biosynthesis was also found to play pivotal roles in cell growth and tolerance to environmental stresses and antifungal drugs. In contrast, the Met14-dependent methionine biosynthesis was found to be more important than cysteine biosynthesis for the production of major cryptococcal virulence factors of melanin pigments and polysaccharide capsules. Finally, we also found that despite its attenuated virulence in an insect model, Galleria mellonella, the met14Δ mutant yielded no difference in virulence in a murine model of systemic cryptococcosis. Hence, clinical inhibition of Met14-dependent amino acid biosynthetic pathways may not be advantageous for the treatment of systemic cryptococcosis. IMPORTANCE Current antifungal drugs have several limitations, such as drug resistance, severe side effects, and a narrow spectrum. Therefore, novel antifungal targets are urgently needed. To this end, fungal sulfur amino acid biosynthetic pathways are considered potential targets for development of new antifungal agents. Here, we demonstrated that Met14 in the sulfate assimilation pathway promotes growth, stress response, and virulence factor production in C. neoformans via synthesis of sulfur-containing amino acids methionine and cysteine. Met14-dependent cysteine rather than methionine synthesis was found to be critical for growth and stress responses, whereas Met14-dependent methionine synthesis was more important for the production of antiphagocytic capsules and antioxidant melanin in C. neoformans. Surprisingly, deletion of the MET14 gene was found to attenuate cryptococcal virulence in an insect model, yet not in a murine model. Collectively, our results showed that Met14-dependent cysteine and methionine biosynthesis play roles that are distinct from each other in C. neoformans. Moreover, Met14 is unlikely to be a suitable anticryptococcal drug target.