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During hibernation in the 13-lined ground squirrel, Ictidomys tridecemlineatus, the cerebral cortex is electrically silent, yet the brainstem continues to regulate cardiorespiratory function. Previous work showed that neurons in slices through the medullary ventral respiratory column (VRC) but not the cortex are insensitive to high doses of pentobarbital during hibernation, leading to the hypothesis that GABAA receptors (GABAAR) in the VRC undergo a seasonal modification in subunit composition. To test whether alteration of GABAAR subunits are responsible for hibernation-associated pentobarbital insensitivity, we examined an array of subunits using RT-PCR and Western blots and identified changes in ϵ- and δ-subunits in the medulla but not the cortex. Using immunohistochemistry, we confirmed that during hibernation, the expression of ϵ-subunit-containing GABAARs nearly doubles in the VRC. We also identified a population of δ-subunit-containing GABAARs adjacent to the VRC that were differentially expressed during hibernation. As δ-subunit-containing GABAARs are particularly sensitive to ethanol (EtOH), multichannel electrodes were inserted in slices of medulla and cortex from hibernating squirrels and EtOH was applied. EtOH, which normally inhibits neuronal activity, excited VRC but not cortical neurons during hibernation. This excitation was prevented by bicuculline pretreatment, indicating the involvement of GABAARs. We propose that neuronal activity in the VRC during hibernation is unaffected by pentobarbital due to upregulation of ϵ-subunit-containing GABAARs on VRC neurons. Synaptic input from adjacent inhibitory interneurons that express δ-subunit-containing GABAARs is responsible for the excitatory effects of EtOH on VRC neurons during hibernation. [ABSTRACT FROM AUTHOR]

ANP was determined in 13 patients with DDD pacemakers due to total AV block, at rest and during bicycle ergomeny under both pacing modes, DDD and VVI (70/min]. Under DDD pacing, the mean ANP level (normal range 5-95 pg/ml) at rest was 36 ± 18 pg/mL (mean ± SD) and increased significantly by the/actor of 3.5 fo I 27 pg/mL during exercise (p < 0.01). By just changing the pacing mode to WI, the ANP levels rose to 73 ± 28 pg/mL (p <0.01) at rest (= 203% of DDD resting level] and to 216 ± 184 pg/mL (= 170% of DDD peak level) during exercise P < 0.01). These results show that under AV synchronous pacing, the ANP levels we generally lower. A possible explanation for this increased release of ANP during asynchronous WI pacing is the acute increase of the atrial wall tension which occurs when the atria contacts during the systole against closed AV valves. [ABSTRACT FROM AUTHOR]

Objectives: Radiation pneumonitis (RP) is a local inflammatory response, and we hypothesize that RP serves as an immune stimulator and is a protective factor against disease progression.  Methods: We analyzed patients with early-stage non-small cell lung cancer (NSCLC) treated with stereotactic body radiation therapy (SBRT) at two institutions. Radiographic RP (RRP) was evaluated and maximal axial dimensions were measured at three-, six-, and twelve-month timepoints with surveillance CT. RRP was measured using radiographic markers such as ground-glass opacities and airspace consolidation. Disease recurrence was evaluated and categorized as local, regional, and distant.  Results: Seventy-seven unique patient records were randomly selected from the database, 72 patients (93.5%) had RRP and five patients (6.5%) did not. The median follow-up was 24.3 months (IQR: 12.0 - 41.9). Disease failure occurred in 28.6% of patients with 6.5% local only, 2.6% regional only, 7.8% distant only, and 11.7% with multiple recurrences. Patients with RRP demonstrated a lower rate of disease failure with 25.0% of those with RRP experiencing disease failure and 80% of those without RRP experiencing disease failure (p=0.02). Patients with RRP had a 71% reduced risk of disease recurrence, compared to patients with no RRP, after adjusting for maximum tumor dimension (HR 0.29, p = 0.05). Among patients with RRP, there was no significant difference in recurrence based on extent of RRP (maximal area of RRP on CT). RRP did not correlate with overall survival.  Discussion: Most patients who received SBRT had RRP, and this study suggests that it may be protective of cancer recurrence. These results are hypothesis-generating and will need to be validated in larger and independent datasets., Competing Interests: The authors have declared that no competing interests exist., (Copyright © 2022, Wolf et al.)

Introduction Recent data suggest synergy of chemoradiotherapy and metformin in locally-advanced non-small cell lung cancer (NSCLC). It remains unclear if similar synergy exists with stereotactic lung body radiation therapy (SBRT) and metformin. We analyzed the role of metformin on progression-free survival (PFS) and toxicity in the setting of lung SBRT. Methods We identified 31 patients on metformin-treated with SBRT for early-stage NSCLC. Eighty-nine similarly treated patients were chosen as controls. Kaplan-Meier method was used to estimate cumulative PFS probabilities. Results Median follow-up was 30.7 months. Forty-two patients had diabetes, 31 (74%) of which were taking metformin concurrent with SBRT. Median PFS for metformin-users vs. metformin non-users was 36.4 months vs 48.9 months, respectively (p = 0.29). Among diabetic patients, median PFS for metformin users was 36.4 months and was unobserved for non-users (p= 0.40). On univariable analysis, male sex (p = 0.03) and tumor size (p = 0.01) were associated with the risk of progression or death; use of metformin was not significant (p = 0.34). There was no difference in grade ≥2 radiation pneumonitis between metformin users vs non-users (p = 0.51) Conclusion In this retrospective sample of lung SBRT patients, we did not detect a meaningful effect of concurrent metformin use on PFS. Since SBRT and conventional RT may have different cell kill mechanisms, the previously described beneficial effects of metformin may not apply in a hypofractionated setting. These results should be validated in an independent dataset, and we await the results of ongoing clinical trials., Competing Interests: The authors have declared that no competing interests exist., (Copyright © 2021, Stang et al.)

Aim: The study aims to explore the views of children with cerebral palsy (CP) regarding their experiences being bullied and their perceptions of their primary bully., Method: Forty-three children aged 10-18 with CP were given the California Bullying Victimization Scale. Fourteen of the children shared a bullying experience orally or written after the survey. Using a mixed methods analysis, the survey answers were tabulated with descriptive statistics and analysed by Gross Motor Function Classification System (GMFCS) level. The comments were coded to determine qualitative responses., Results: Most bullied children viewed the bully as less than or at least equal to them regarding popularity (73%), performance in schoolwork (85%), and physical strength (56%). Bullied children preferentially told an adult at home. Code words and common categories of bullying were determined from their experiences., Interpretation: Children with CP experience bullying but do not view themselves as inferior to their bully in popularity, intelligence, or physical strength. Most bullied participants confided in an adult at home. Children with GMFCS Levels 1, 2, and 3 seem to be at greater risk for bullying than children with GMFCS Levels 4 and 5. Children with CP demonstrate individualized strategies for resilience with reliance on adults for resources., (© 2020 John Wiley & Sons Ltd.)

Objective Radiation pneumonitis (RP) is a dose-limiting toxicity that affects the treatment of lung cancer. Data on factors predictive of developing symptomatic RP after stereotactic body radiation therapy (SBRT) are limited. We reviewed data to identify pretreatment factors predictive of the development of symptomatic RP in patients' lung cancer treated with SBRT. Methods Data were collected on 296 patients treated with SBRT for lung cancer. Factors available at time of consultation were analyzed for the development of symptomatic RP, defined as CTCAE v. 4.0 ≥ Grade 2. The factors analyzed included patient demographic, tumor-specific, and pretreatment pulmonary function data. Univariate and multivariate analyses were performed to assess for predictive factors. Results Median follow-up was 22 months. The rate of symptomatic RP was 16%. Univariate analysis showed an increased rate of symptomatic RP with treatments to the right lung (22% vs. 9%, p = 0.007), driven primarily by an increased rate of symptomatic RP when treating the right lower lobe (RLL) vs. other lobes (31 vs. 13%, p = 0.03). Patients with a history of prior lung directed therapy were also more likely to develop symptomatic RP (12% vs. 24%, p = 0.008). These statistical differences were retained on multivariate analysis. Conclusion SBRT to the right lung, especially the RLL, and to patients with a history of prior lung-directed therapy increases the risk of developing symptomatic RP after SBRT. Further studies on ways to predict and prevent symptomatic RP are needed., Competing Interests: The authors have declared that no competing interests exist., (Copyright © 2020, Harris et al.)

Cotton-based surgical invasive devices with their desired hemostyptic properties have been used for decades in the surgical field. However, in cardiac surgery using the heart-lung machine with direct retransfusion of suction blood, activated blood may re-enter the circulation without filtration and may trigger a cascade reaction leading to systemic inflammation and thrombosis. We therefore set out to evaluate the inflammatory potential of untreated and pyrogen-impregnated cotton-based surgical invasive medical devices. After incubation of the swabs with whole blood or PBMC, the cell-free supernatant was investigated for IL1β and IL6. While the reaction of human whole blood toward cotton swabs could not be influenced by any sterilization technique, dry heat and gamma-irradiation were able to diminish the inflammatory reaction of PBMC toward the material and the used pyrogens. In conclusion, using PBMC in direct contact to cotton we are the first to establish a suitable test method for quantification of the pyrogenic/inflammatory activity of this material. The unaltered reaction of whole blood, however, suggests a crosstalk of cells and plasma proteins in the inflammation activation that is not prevented by sterilization of the swabs. This new in vitro testing methodology may help to better display the clinical situation during development of new materials. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater 107B: 1877-1888, 2019., (© 2018 Wiley Periodicals, Inc.)

Purpose: The use of stereotactic body radiation therapy (SBRT) has emerged as an effective treatment modality for patients with early-stage non-small-cell lung cancer (NSCLC), with excellent local control rates. Despite this, there is a predominant pattern of distant failure. We sought to identify factors that help predict which patients with stages I to IIA NSCLC treated with SBRT are at highest risk of distant failure, so that we may utilize these factors in the future to help determine which patients may benefit from the addition of systemic therapies., Patients and Methods: We retrospectively reviewed 292 patients treated with SBRT for early stage NSCLC from 2006 to 2016 at 2 institutions. Patients were classified by T stage, tumor size, location and histology, pretreatment positron emission tomography/computed tomography (PET/CT) standardized uptake value (SUV), smoking status, and age. The primary endpoint of the study was distant failure. We aimed to analyze if patient characteristics could be identified that predicted for distant failure through the use of competing risk analysis., Results: The median follow-up was 21.9 months. The median dose of radiation and fractionation delivered was 50 Gy (range, 45-65 Gy) in 5 fractions (range, 3-13 fractions). The median patient age was 72.8 years (interquartile range, 65.4-79.7 years). The 2-year distant failure was 22.0%, and overall survival at 2 years was found to be 61.0%. For every 1-year increase in patient age, the hazard of distant failure at any given time was 3% lower (hazard ratio, 0.97; 95% confidence interval, 0.94-0.99; P = .04). None of the remaining characteristics emerged as significant risk factors for distant failure on univariable or multivariable analysis., Conclusions: Overall, our cohort had distant failure and survival rates comparable with what has been described in the literature. Although we were unable to identify factors outside of age that correlated to risk of distant failure, this topic warrants further investigation, as distant failure is the primary pattern of failure with SBRT when used as the primary management for early-stage NSCLC. Additional molecular studies are needed to further inform on the role of systemic therapy in patients with early-stage NSCLC to improve clinical outcomes., (Published by Elsevier Inc.)

Hemocompatibility and cytocompatibility of biomaterials codetermine the success of tissue engineering applications. DNA, the natural component of our cells, is an auspicious biomaterial for the generation of designable scaffolds with tailorable characteristics. In this study, a combination of rolling circle amplification and multiprimed chain amplification is used to generate hydrogels at centimeter scale consisting solely of DNA. Using an in vitro rotation model and fresh human blood, the reaction of the hemostatic system on DNA hydrogels is analyzed. The measurements of hemolysis, platelets activation, and the activation of the complement, coagulation, and neutrophils using enzyme-linked immunosorbent assays demonstrate excellent hemocompatibility. In addition, the cytocompatibility of the DNA hydrogels is tested by indirect contact (agar diffusion tests) and material extract experiments with L929 murine fibroblasts according to the ISO 10993-5 specifications and no negative impact on the cell viability is detected. These results indicate the promising potential of DNA hydrogels as biomaterials for versatile applications in the field of regenerative medicine., (© 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.)

Pyrogen testing represents a crucial safety measure for parental drugs and medical devices, especially in direct contact with blood or liquor. The European Pharmacopoeia regulates these quality control measures for parenterals. Since 2010, the monocyte activation test (MAT) has been an accepted pyrogen test that can be performed with different human monocytic cell sources: whole blood, isolated monocytic cells or monocytic cell lines with IL1β, IL6, or TNFα as readout cytokines. In the present study, we examined the three different cell sources and cytokine readout parameters with the scope of accelerating the assay time. We could show that despite all cell types being able to detect pyrogens, primary cells were more sensitive than the monocytic cell line. Quantitative real-time PCR revealed IL6 mRNA transcripts having the largest change in Ct-values upon LPS-stimulation compared to IL1β and TNFα, but quantification was unreliable. IL6 protein secretion from whole blood or peripheral blood mononuclear cells (PBMC) was also best suited for an accelerated assay with a larger linear range and higher signal-to-noise ratios upon LPS-stimulation. The unique combination with propan-2-ol or a temperature increase could additionally increase the cytokine production for earlier detection in PBMC. The increased incubation temperature could finally increase not only responses to lipopolysaccharides (LPS) but also other pyrogens by up to 13-fold. Therefore, pyrogen detection can be accelerated considerably by using isolated primary blood cells with an increased incubation temperature and IL6 as readout. These results could expedite assay time and thus help to promote further acceptance of the MAT. Copyright © 2016 John Wiley & Sons, Ltd., (Copyright © 2016 John Wiley & Sons, Ltd.)

Background: Recent reports demonstrate impaired tumor re-oxygenation 24-48h after stereotactic body radiation therapy (SBRT), suggesting that non-consecutive treatment delivery may be advantageous. To test this hypothesis clinically, we compared local control in patients treated in consecutive daily fractions vs. nonconsecutive fractions., Methods: We retrospectively reviewed 107 lung SBRT patients (117 tumors) treated for T1-T2N0 NSCLC with LINAC based SBRT (50 or 60Gy/5fractions). Patients were characterized as having been treated in consecutive daily fractions vs. in non-consecutive fractions. Local control, survival and toxicity end points (CTCAE V4.0) were compared. Propensity score matching and Cox regression analyses were performed in order to determine the effect of fractionation on local control., Results: With a median follow up of 23.7months, 3-year local control was superior at 93.3% vs. 63.6% in the non-consecutive and consecutive group, respectively (p=0.001). Multivariate analysis and propensity score matching showed that consecutive fractionation was an independent predictor of local failure. Overall survival trended toward improvement in the non-consecutive group, but this was not statistically significant (p=0.188). Development of any grade 2 toxicity was not significantly different between the two groups (p=0.75)., Conclusion: Five-fraction SBRT delivered over non-consecutive days imparts superior LC and similar toxicity compared to consecutive fractionation. These results should be validated in independent datasets and in a prospective fashion., (Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.)

Extracorporeal membrane oxygenation (ECMO) is a pivotal bridge to recovery for cardiopulmonary failure in children. Besides its life-saving quality, it is often associated with severe system-related complications, such as hemolysis, inflammation, and thromboembolism. Novel oxygenator and pump systems may reduce such ECMO-related complications. The ExMeTrA oxygenator is a newly designed pediatric oxygenator with an integrated pulsatile pump minimizing the priming volume and reducing the surface area of blood contact. The aim of our study was to investigate the feasibility and safety of this new ExMeTrA (expansion mediated transport and accumulation) oxygenator in an animal model. During 6 h of extracorporeal circulation (ECC) in pigs, parameters of the hemostatic system including coagulation, platelets and complement activation, and flow rates were investigated. A nonsignificant trend in C3 consumption, thrombin-antithrombin-III (TAT) complex formation and a slight trend in hemolysis were detected. During the ECC, the blood flow was constantly at 500 ml/min using only flexible silicone tubes inside the oxygenator as pulsatile pump. Our data clearly indicate that the hemostatic markers were only slightly influenced by the ExMeTrA oxygenator. Additionally, the oxygenator showed a constant quality of blood flow. Therefore, this novel pediatric oxygenator shows the potential to be used in pediatric and neonatal support with ECMO.

Systemic inflammatory response syndrome (SIRS) is a common complication after cardiovascular surgery that in severe cases can lead to multiple organ dysfunction syndrome and even death. We therefore set out to identify reliable early biomarkers for SIRS in a prospective small patient study for timely intervention. 21 Patients scheduled for planned cardiovascular surgery were recruited in the study, monitored for signs of SIRS and blood samples were taken to investigate biomarkers at pre-assigned time points: day of admission, start of surgery, end of surgery, days 1, 2, 3, 5 and 8 post surgery. Stored plasma and cryopreserved blood samples were analyzed for cytokine expression (IL1β, IL2, IL6, IL8, IL10, TNFα, IFNγ), other pro-inflammatory markers (sCD163, sTREM-1, ESM-1) and response to endotoxin. Acute phase proteins CRP, PCT and pro-inflammatory cytokines IL6 and IL8 were significantly increased (p<0.001) at the end of surgery in all patients but could not distinguish between groups. Normalization of samples revealed significant increases in IL1β changes (p<0.05) and decreased responses to endotoxin (p<0.01) in the SIRS group at the end of surgery. Soluble TREM-1 plasma concentrations were significantly increased in patients with SIRS (p<0.01). This small scale patient study could show that common sepsis markers PCT, CRP, IL6 and TNFα had low predictive value for early diagnosis of SIRS after cardiovascular surgery. A combination of normalized IL1β plasma levels, responses to endotoxin and soluble TREM-1 plasma concentrations at the end of surgery are predictive markers of SIRS development in this small scale study and could act as an indicator for starting early therapeutic interventions.

Aptamers are synthetic single-stranded DNA (ssDNA) molecules with the ability to fold into complex three-dimensional structures. They can bind their targets with a high selectivity and affinity, thus they have an enormous potential as therapeutic agents. However, since aptamers are synthetic and especially since certain sequences can increasingly bind to the pattern recognition receptors of the immune cells when applied in vivo, they can induce an immune activation. Here, we established a real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) based assay to evaluate aptamers-induced immune activation prior to in vivo studies. Human whole blood or plasmacytoid dendritic cell line (PMDC05) were incubated with CpG, R10-60 aptamer, start library, or a CpG containing aptamer. After 2 and 4 h, cytokine expression was measured using qRT-PCR to determine immune reaction against different aptamers. CpG containing a phosphorothioate backbone led to a significant up-regulation of CCL-7, IFN-1α, IFN-1β in whole blood after 4 h. Compared to the samples without ssDNA, significantly higher TNF-α expression was detected after the R10-60 aptamer incubation for 4 h. The stimulation of PMDC05 cells with different ssDNA enabled more sensitive detection of aptamer sequence specific immune activation. After 4 h, CpG led to a significantly higher expression of CCL-8, CXCL-10, IL-1β, IL-6, IL-8, IFN-1β, and TNF-α. R10-60 aptamer caused a significant up-regulation of IL-1β, IFN-1β, and TNF-α. Negative control aptamers did not induce an immune activation. The use of this assay before starting with in vivo studies will facilitate the in vitro prediction of immune activation potential of aptamers., (Copyright © 2014 John Wiley & Sons, Ltd.)

Next to good hemocompatibility performance of new medical devices, which has to be tested according to the ISO 10993-4, the detection of pyrogen-contaminated devices plays a pivotal role for safe device application. During blood contact with pyrogen-contaminated devices, intense inflammatory and hemostatic reactions are feared. The aim of our study was to investigate the influence of pyrogenic contaminations on stents according to the ISO 10993-4. The pyrogens of different origins like lipopolysaccharides (LPS), purified lipoteichoic acid (LTA) or zymosan were used. These pyrogens were dried on stents or dissolved and circulated in a Chandler-loop model for 90 min at 37°C with human blood. Before and after circulation, parameters of the hemostatic system including coagulation, platelets, complement and leukocyte activation were investigated. The complement system was activated by LPS isolated from Klebsiella pneumoniae and Pseudomonas aeruginosa and by LTA. Leukocyte activation was triggered by LPS isolated from K. pneumoniae, LTA and zymosan, whereas coagulation and platelet activation were only slightly influenced. Our data indicate that pyrogen-contaminated devices lead to an alteration in the hemostatic response when compared to depyrogenized devices. Therefore, pyrogenicity testing should be performed prior to hemocompatibility tests according to ISO 10993-4 in order to exclude hemostatic activation induced by pyrogen contaminations., (Copyright © 2014 Elsevier B.V. All rights reserved.)

Pyrogens are components of microorganisms, like bacteria, viruses or fungi, which can induce a complex inflammatory response in the human body. Pyrogen contamination on medical devices prior operation is still critical and associated with severe complications for the patients. The aim of our study was to develop a reliable test, which allows detection of pyrogen contamination on the surface of medical devices. After in vitro pyrogen contamination of different medical devices and incubation in a rotation model, the human whole blood monocyte activation test (MAT), which is based on an IL-1β-specific ELISA, was employed. Our results show that when combining a modified MAT protocol and a dynamic incubation system, even smallest amounts of pyrogens can be directly detected on the surface of medical devices. Therefore, screening of medical devices prior clinical application using our novel assay, has the potential to significantly reduce complications associated with pyrogen-contaminated medical devices.

Age-associated changes in tongue musculature may contribute to dysphagia. One possible treatment is tongue exercise. Exercise induces synaptic plasticity by increasing neurotrophic factors in spinal cord and limb musculature. However, effects of exercise on neurotrophic factors in the cranial sensorimotor system are unknown. Our purpose was to examine the effects of age and exercise on brain-derived neurotrophic factor (BDNF) and its receptor TrkB in the rat hypoglossal nucleus. Young, middle-aged, and old rats were assigned to exercise or no-exercise control conditions. Exercise animals were trained to perform a tongue press task for 8 weeks. Samples from the hypoglossal nucleus were analyzed for BDNF and TrkB immunoreactivity. Baseline maximum tongue forces were similar in all age groups and increased significantly following exercise. BDNF immunoreactivity did not show a significant decrease with age in control group. However, in the exercise group, BDNF was significantly increased in young animals. TrkB immunoreactivity decreased significantly with age in control group, but did not change with exercise. BDNF and TrkB immunoreactivity levels were positively correlated with exercise in young and middle aged animals, but were negatively or weakly correlated with exercise in old animals and with a lack of exercise in no-exercise controls. Tongue exercise was associated with increased tongue forces in rats at all ages. While increases in BDNF and TrkB levels associated with exercise may play a role in mechanisms contributing to increased tongue forces in young and middle-aged rats, other mechanisms may be involved in increased tongue forces observed in old rats., (Copyright © 2011 Elsevier B.V. All rights reserved.)

During hibernation in the 13-lined ground squirrel, Ictidomys tridecemlineatus, the cerebral cortex is electrically silent, yet the brainstem continues to regulate cardiorespiratory function. Previous work showed that neurons in slices through the medullary ventral respiratory column (VRC) but not the cortex are insensitive to high doses of pentobarbital during hibernation, leading to the hypothesis that GABA(A) receptors (GABA(A)R) in the VRC undergo a seasonal modification in subunit composition. To test whether alteration of GABA(A)R subunits are responsible for hibernation-associated pentobarbital insensitivity, we examined an array of subunits using RT-PCR and Western blots and identified changes in ε- and δ-subunits in the medulla but not the cortex. Using immunohistochemistry, we confirmed that during hibernation, the expression of ε-subunit-containing GABA(A)Rs nearly doubles in the VRC. We also identified a population of δ-subunit-containing GABA(A)Rs adjacent to the VRC that were differentially expressed during hibernation. As δ-subunit-containing GABA(A)Rs are particularly sensitive to ethanol (EtOH), multichannel electrodes were inserted in slices of medulla and cortex from hibernating squirrels and EtOH was applied. EtOH, which normally inhibits neuronal activity, excited VRC but not cortical neurons during hibernation. This excitation was prevented by bicuculline pretreatment, indicating the involvement of GABA(A)Rs. We propose that neuronal activity in the VRC during hibernation is unaffected by pentobarbital due to upregulation of ε-subunit-containing GABA(A)Rs on VRC neurons. Synaptic input from adjacent inhibitory interneurons that express δ-subunit-containing GABA(A)Rs is responsible for the excitatory effects of EtOH on VRC neurons during hibernation.