Refine
Year of publication
Document Type
- Article (5423) (remove)
Has Fulltext
- yes (5423)
Keywords
- inflammation (80)
- COVID-19 (62)
- SARS-CoV-2 (47)
- cancer (38)
- glioblastoma (38)
- apoptosis (37)
- Inflammation (34)
- breast cancer (34)
- prostate cancer (30)
- autophagy (29)
Institute
- Medizin (5423) (remove)
We have investigated the role of reactive oxygen species and thiol-oxidizing agents in the induction of cell death and have shown that adenocarcinoma gastric (AGS) cells respond differently to the oxidative challenge according to the signaling pathways activated. In particular, apoptosis in AGS cells is induced via the mitochondrial pathway upon treatment with thiol-oxidizing agents, such as diamide. Apoptosis is associated with persistent oxidative damage, as evidenced by the increase in carbonylated proteins and the expression/activation of DNA damage-sensitive proteins histone H2A.X and DNA-dependent protein kinase. Resistance to hydrogen peroxide is instead associated with Keap1 oxidation and rapid translocation of Nrf2 into the nucleus. Sensitivity to diamide and resistance to hydrogen peroxide are correlated with GSH redox changes, with diamide severely increasing GSSG, and hydrogen peroxide transiently inducing protein-GSH mixed disulfides. We show that p53 is activated in response to diamide treatment by the oxidative induction of the Trx1/p38(MAPK) signaling pathway. Similar results were obtained with another carcinoma cell line, CaCo2, indicating that these findings are not limited to AGS cells. Our data suggest that thiol-oxidizing agents could be exploited as inducers of apoptosis in tumor histotypes resistant to ROS-producing chemotherapeutics.
Thioredoxin 1 and thioredoxin 2 have opposed regulatory functions on hypoxia-inducible factor-1α
(2007)
Hypoxia inducible factor 1 (HIF-1), a key regulator for adaptation to hypoxia, is composed of HIF-1alpha and HIF-1beta. In this study, we present evidence that overexpression of mitochondria-located thioredoxin 2 (Trx2) attenuated hypoxia-evoked HIF-1alpha accumulation, whereas cytosolic thioredoxin 1 (Trx1) enhanced HIF-1alpha protein amount. Transactivation of HIF-1 is decreased by overexpression of Trx2 but stimulated by Trx1. Inhibition of proteasomal degradation of HIF-1alpha in Trx2-overexpressing cells did not fully restore HIF-1alpha protein levels, while HIF-1alpha accumulation was enhanced in Trx1-overexpressing cells. Reporter assays showed that cap-dependent translation is increased by Trx1 and decreased by Trx2, whereas HIF-1alpha mRNA levels remained unaltered. These data suggest that thioredoxins affect the synthesis of HIF-1alpha. Trx1 facilitated synthesis of HIF-1alpha by activating Akt, p70S6K, and eIF-4E, known to control cap-dependent translation. In contrast, Trx2 attenuated activities of Akt, p70S6K, and eIF-4E and provoked an increase in mitochondrial reactive oxygen species production. MitoQ, a mitochondria specific antioxidant, reversed HIF-1alpha accumulation as well as Akt activation under hypoxia in Trx2 cells, supporting the notion of translation control mechanisms in affecting HIF-1alpha protein accumulation.
The activation of the transcription factor NF-E2-related factor 2 (Nrf2) maintains cellular homeostasis in response to oxidative stress by the regulation of multiple cytoprotective genes. Without stressors, the activity of Nrf2 is inhibited by its interaction with the Keap1 (kelch-like ECH-associated protein 1). Here, we describe (3S)-1-[4-[(2,3,5,6-tetramethylphenyl) sulfonylamino]-1-naphthyl]pyrrolidine-3-carboxylic acid (RA839), a small molecule that binds noncovalently to the Nrf2-interacting kelch domain of Keap1 with a Kd of ∼6 μm, as demonstrated by x-ray co-crystallization and isothermal titration calorimetry. Whole genome DNA arrays showed that at 10 μm RA839 significantly regulated 105 probe sets in bone marrow-derived macrophages. Canonical pathway mapping of these probe sets revealed an activation of pathways linked with Nrf2 signaling. These pathways were also activated after the activation of Nrf2 by the silencing of Keap1 expression. RA839 regulated only two genes in Nrf2 knock-out macrophages. Similar to the activation of Nrf2 by either silencing of Keap1 expression or by the reactive compound 2-cyano-3,12-dioxooleana-1,9-dien-28-oic acid methyl ester (CDDO-Me), RA839 prevented the induction of both inducible nitric-oxide synthase expression and nitric oxide release in response to lipopolysaccharides in macrophages. In mice, RA839 acutely induced Nrf2 target gene expression in liver. RA839 is a selective inhibitor of the Keap1/Nrf2 interaction and a useful tool compound to study the biology of Nrf2.
Resting state fMRI has been employed to identify alterations in functional connectivity within or between brain regions following acute and chronic exposure to Δ9-tetrahydrocannabinol (THC), the psychoactive component in cannabis. Most studies focused a priori on a limited number of local brain areas or circuits, without considering the impact of cannabis on whole-brain network organization. The present study attempted to identify changes in the whole-brain human functional connectome as assessed with ultra-high field (7T) resting state scans of cannabis users (N = 26) during placebo and following vaporization of cannabis. Two distinct data-driven methodologies, i.e. network-based statistics (NBS) and connICA, were used to identify changes in functional connectomes associated with acute cannabis intoxication and history of cannabis use. Both methodologies revealed a broad state of hyperconnectivity within the entire range of major brain networks in chronic cannabis users compared to occasional cannabis users, which might be reflective of an adaptive network reorganization following prolonged cannabis exposure. The connICA methodology also extracted a distinct spatial connectivity pattern of hypoconnectivity involving the dorsal attention, limbic, subcortical and cerebellum networks and of hyperconnectivity between the default mode and ventral attention network, that was associated with the feeling of subjective high during THC intoxication. Whole-brain network approaches identified spatial patterns in functional brain connectomes that distinguished acute from chronic cannabis use, and offer an important utility for probing the interplay between short and long-term alterations in functional brain dynamics when progressing from occasional to chronic use of cannabis.
Ataxia telangiectasia (A-T) is a progressive and life-limiting disease associated with cerebellar ataxia due to progressive cerebellar degeneration. In addition to ataxia, which is described in detail, the presence of chorea, dystonia, oculomotor apraxia, athetosis, parkinsonism, and myoclonia are typical manifestations of the disease. The study aimed to evaluate the specificity and sensitivity of neurofilament light chain (NfL) as a biomarker of neurodegeneration in relation to SARA score. In this prospective trial, one visit of 42 A-T patients aged 1.3–25.6 years (mean 11.6 ± 7.3 years) was performed, in which NfL was determined from serum by ELISA. Additionally, a neurological examination of the patients was performed. Blood was collected from 19 healthy volunteers ≥ 12 years of age. We found significantly increased levels of NfL in patients with A-T compared to healthy controls (21.5 ± 3.6 pg/mL vs. 9.3 ± 0.49 pg/mL, p ≤ 0.01). There was a significant correlation of NfL with age, AFP, and SARA. NfL is a new potential progression biomarker in blood for neurodegeneration in A-T which increases with age.
Background: The correct performance of a structured facial examination presents a fundamental clinical skill to detect facial pathologies. However, many students are not adequately prepared in this basic clinical skill. Many argue that the traditional ‘See One, Do One’ approach is not sufficient to fully master a clinical skill. ‘Mental Training’ has successfully been used to train psychomotor and technical skills in sports and other surgical fields, but its use in Oral and Maxillofacial Surgery is not described. We conducted a quasi-experimental to determine if ‘Mental Training’ was effective in teaching a structured facial examination.
Methods: Sixty-seven students were randomly assigned to a ‘Mental Training’ and ‘See One, Do One’ group. Both groups received standardized video instruction on how to perform a structured facial examination. The ‘See One, Do One’ group then received 60 min of guided physical practice while the ‘Mental Training’ group actively developed a detailed, stepwise sequence of the performance of a structured facial examination and visualized this sequence subvocally before practicing the skill. Student performance was measured shortly after (T1) and five to 10 weeks (T2) after the training by two blinded examiners (E1 and E2) using a validated checklist.
Results: Groups did not differ in gender, age or in experience. The ‘Mental Training’ group averaged significantly more points in T1 (pE1 = 0.00012; pE2 = 0.004; dE1 = 0.86; dE2 = 0.66) and T2 (pE1 = 0.04; pE2 = 0.008, dE1 = 0.37; dE2 = 0.64) than the ‘See One, Do One’ group. The intragroup comparison showed a significant (pE1 = 0.0002; pE2 = 0.06, dE1 = 1.07; dE2 = 0.50) increase in clinical examination skills in the ‘See One, Do One’ group, while the ‘Mental Training’ group maintained an already high level of clinical examination skills between T1 and T2.
Discussion: ‘Mental Training’ is an efficient tool to teach and maintain basic clinical skills. In this study ‘Mental Training’ was shown to be superior to the commonly used ‘See One, Do One’ approach in learning how to perform a structured facial examination and should therefore be considered more often to teach physical examination skills.
Assessment of individual therapeutic responses provides valuable information concerning treatment benefits in individual patients. We evaluated individual therapeutic responses as determined by the Disease Activity Score-28 joints critical difference for improvement (DAS28-dcrit) in rheumatoid arthritis (RA) patients treated with intravenous tocilizumab or comparator anti-tumor necrosis factor (TNF) agents. The previously published DAS28-dcrit value [DAS28 decrease (improvement) ≥ 1.8] was retrospectively applied to data from two studies of tocilizumab in RA, the 52-week ACT-iON observational study and the 24-week ADACTA randomized study. Data were compared within (not between) studies. DAS28 was calculated with erythrocyte sedimentation rate as the inflammatory marker. Stability of DAS28-dcrit responses and European League Against Rheumatism (EULAR) good responses was determined by evaluating repeated responses at subsequent timepoints. A logistic regression model was used to calculate p values for differences in response rates between active agents. Patient-reported outcomes (PROs; pain, global health, function, and fatigue) in DAS28-dcrit responder versus non-responder groups were compared with an ANCOVA model. DAS28-dcrit individual response rates were 78.2% in tocilizumab-treated patients and 58.2% in anti-TNF-treated patients at week 52 in the ACT-ion study (p = 0.0001) and 90.1% versus 59.1% at week 24 in the ADACTA study (p < 0.0001). DAS28-dcrit responses showed greater stability over time (up to 52 weeks) than EULAR good responses. For both active treatments, DAS28-dcrit responses were associated with statistically significant improvements in mean PRO values compared with non-responders. The DAS28-dcrit response criterion provides robust assessments of individual responses to RA therapy and may be useful for discriminating between active agents in clinical studies and guiding treat-to-target decisions in daily practice.
Human RNF213, which encodes the protein mysterin, is a known susceptibility gene for moyamoya disease (MMD), a cerebrovascular condition with occlusive lesions and compensatory angiogenesis. Mysterin mutations, together with exposure to environmental trigger factors, lead to an elevated stroke risk since childhood. Mysterin is induced during cell stress, to function as cytosolic AAA+ ATPase and ubiquitylation enzyme. Little knowledge exists, in which context mysterin is needed. Here, we found that genetic ablation of several mitochondrial matrix factors, such as the peptidase ClpP, the transcription factor Tfam, as well as the peptidase and AAA+ ATPase Lonp1, potently induces Rnf213 transcript expression in various organs, in parallel with other components of the innate immune system. Mostly in mouse fibroblasts and human endothelial cells, the Rnf213 levels showed prominent upregulation upon Poly(I:C)-triggered TLR3-mediated responses to dsRNA toxicity, as well as upon interferon gamma treatment. Only partial suppression of Rnf213 induction was achieved by C16 as an antagonist of PKR (dsRNA-dependent protein kinase). Since dysfunctional mitochondria were recently reported to release immune-stimulatory dsRNA into the cytosol, our results suggest that mysterin becomes relevant when mitochondrial dysfunction or infections have triggered RNA-dependent inflammation. Thus, MMD has similarities with vasculopathies that involve altered nucleotide processing, such as Aicardi-Goutières syndrome or systemic lupus erythematosus. Furthermore, in MMD, the low penetrance of RNF213 mutations might be modified by dysfunctions in mitochondria or the TLR3 pathway.
Purpose: Neonatal surgery for abdominal wall defects is not performed in a centralized manner in Germany. The aim of this study was to investigate whether treatment for abdominal wall defects in Germany is equally effective compared to international results despite the decentralized care.
Methods: All newborn patients who were clients of the major statutory health insurance company in Germany between 2009 and 2013 and who had a diagnosis of gastroschisis or omphalocele were included. Mortality during the first year of life was analysed.
Results: The 316 patients with gastroschisis were classified as simple (82%) or complex (18%) cases. The main associated anomalies in the 197 patients with omphalocele were trisomy 18/21 (8%), cardiac anomalies (32%) and anomalies of the urinary tract (10%). Overall mortality was 4% for gastroschisis and 16% for omphalocele. Significant factors for non-survival were birth weight below 1500 g for both groups, complex gastroschisis, volvulus and anomalies of the blood supply to the intestine in gastroschisis, and female gender, trisomy 18/21 and lung hypoplasia in omphalocele.
Conclusions: Despite the fact that paediatric surgical care is organized in a decentralized manner in Germany, the mortality rates for gastroschisis and omphalocele are equal to those reported in international data.
This study highlights the importance of insect evidence by evaluating 949 insect-associated cases, including 139 entomological reports, from 2001 to 2019 at the Institute of Legal Medicine Frankfurt/Germany. With a high number of cases in the summer months and a low number in the colder season, 78.5% of the bodies were found indoors, regardless of year or month. In more than 80% of the cases, where PMI information was available (n = 704), the presumed PMI ranged from 1 to 21 days, a period during which entomological evidence can provide a day-specific estimate of PMImin. In cases where insects have been identified to species level (n = 279), most bodies were infested by one or two species with a maximum of 10 different species. Overall, a total of 55 insect species were found. Information on biology, activity and distribution of the most abundant taxa is given and applied for 5 case histories estimating different PMImins of up to over 6 months. Despite proved importance and scientific development of forensic entomology, insects are still rarely considered as a tool in forensic case work. The main reasons are a lack of awareness and (too) late involvement of a forensic entomologist. Our work shows that forensic entomology is an independent discipline that requires specialist expertise.
Purpose of Review: To provide an overview of current surgical peri-implantitis treatment options.
Recent Findings: Surgical procedures for peri-implantitis treatment include two main approaches: non-augmentative and augmentative therapy. Open flap debridement (OFD) and resective treatment are non-augmentative techniques that are indicated in the presence of horizontal bone loss in aesthetically nondemanding areas. Implantoplasty performed adjunctively at supracrestally and buccally exposed rough implant surfaces has been shown to efficiently attenuate soft tissue inflammation compared to control sites. However, this was followed by more pronounced soft tissue recession. Adjunctive augmentative measures are recommended at peri-implantitis sites exhibiting intrabony defects with a minimum depth of 3 mm and in the presence of keratinized mucosa. In more advanced cases with combined defect configurations, a combination of augmentative therapy and implantoplasty at exposed rough implant surfaces beyond the bony envelope is feasible.
Summary: For the time being, no particular surgical protocol or material can be considered as superior in terms of long-term peri-implant tissue stability.
Purpose of Review: Attention deficit hyperactivity disorder (ADHD) shows high heritability in formal genetic studies. In our review article, we provide an overview on common and rare genetic risk variants for ADHD and their link to clinical practice.
Recent findings: The formal heritability of ADHD is about 80% and therefore higher than most other psychiatric diseases. However, recent studies estimate the proportion of heritability based on singlenucleotide variants (SNPs) at 22%. It is a matter of debate which genetic mechanisms explain this huge difference. While frequent variants in first mega-analyses of genome-wideassociation study data containing several thousand patients give the first genome-wide results, explaining only little variance, the methodologically more difficult analyses of rare variants are still in their infancy. Some rare genetic syndromes show higher prevalence for ADHD indicating a potential role for a small number of patients. In contrast, polygenic risk scores (PRS) could potentially be applied to every patient. We give an overview how PRS explain different behavioral phenotypes in ADHD and how they could be used for diagnosis and therapy prediction.
Summary: Knowledge about a patient’s genetic makeup is not yet mandatory for ADHD therapy or diagnosis. PRS however have been introduced successfully in other areas of clinical medicine, and their application in psychiatry will begin within the next years. In order to ensure competent advice for patients, knowledge of the current state of research is useful forpsychiatrists.
Despite the implementation of consolidative immune checkpoint inhibition after definitive chemoradiotherapy (CRT), the prognosis for locally advanced non-small-cell lung cancer (NSCLC) remains poor. We assessed the impact of the C-reactive protein (CRP) to albumin ratio (CAR) as an inflammation-based prognostic score in patients with locally advanced NSCLC treated with CRT. We retrospectively identified and analyzed 52 patients with primary unresectable NSCLC (UICC Stage III) treated with definitive/neoadjuvant CRT between 2014 and 2019. CAR was calculated by dividing baseline CRP by baseline albumin levels and correlated with clinicopathologic parameters to evaluate prognostic impact. After dichotomizing patients by the median, univariate and multivariate Cox regression analyses were performed. An increased CAR was associated with advanced T-stage (p = 0.018) and poor performance status (p = 0.004). Patients with pre-therapeutic elevated CAR had significantly lower hemoglobin and higher leukocyte levels (hemoglobin p = 0.001, leukocytes p = 0.018). High baseline CAR was shown to be associated with worse local control (LPFS, p = 0.006), shorter progression-free survival (PFS, p = 0.038) and overall survival (OS, p = 0.022), but not distant metastasis-free survival (DMFS). Multivariate analysis confirmed an impaired outcome in patients with high CAR (LPFS: HR 3.562, 95% CI 1.294–9.802, p = 0.011). CAR is an easily available and independent prognostic marker after CRT in locally advanced NSCLC. CAR may be a useful biomarker for patient stratification to individualize treatment concepts.
Calcium (Ca2+) elevation is an essential secondary messenger in many cellular processes, including disease progression and adaptation to external stimuli, e.g., gravitational load. Therefore, mapping and quantifying Ca2+ signaling with a high spatiotemporal resolution is a key challenge. However, particularly on microgravity platforms, experiment time is limited, allowing only a small number of replicates. Furthermore, experiment hardware is exposed to changes in gravity levels, causing experimental artifacts unless appropriately controlled. We introduce a new experimental setup based on the fluorescent Ca2+ reporter CaMPARI2, onboard LED arrays, and subsequent microscopic analysis on the ground. This setup allows for higher throughput and accuracy due to its retrograde nature. The excellent performance of CaMPARI2 was demonstrated with human chondrocytes during the 75th ESA parabolic flight campaign. CaMPARI2 revealed a strong Ca2+ response triggered by histamine but was not affected by the alternating gravitational load of a parabolic flight.
Acute respiratory distress syndrome (ARDS) is a major cause of patient mortality in intensive care units (ICUs) worldwide. Considering that no causative treatment but only symptomatic care is available, it is obvious that there is a high unmet medical need for a new therapeutic concept. One reason for a missing etiologic therapy strategy is the multifactorial origin of ARDS, which leads to a large heterogeneity of patients. This review summarizes the various kinds of ARDS onset with a special focus on the role of reactive oxygen species (ROS), which are generally linked to ARDS development and progression. Taking a closer look at the data which already have been established in mouse models, this review finally proposes the translation of these results on successful antioxidant use in a personalized approach to the ICU patient as a potential adjuvant to standard ARDS treatment.
Renal glomerular diseases such as glomerulosclerosis and diabetic nephropathy often result in the loss of glomerular function and consequently end-stage renal disease. The glomerulus consists of endothelial cells, mesangial cells and glomerular epithelial cells also referred to as podocytes. A fine-tuned crosstalk between glomerular cells warrants control of growth factor synthesis and of matrix production and degradation, preserving glomerular structure and function. Hydrogen sulfide (H2S) belongs together with nitric oxide (NO) and carbon monoxide (CO) to the group of gasotransmitters. During the last three decades, these higher concentration toxic gases have been found to be produced in mammalian cells in a well-coordinated manner. Recently, it became evident that H2S and the other gasotransmitters share common targets as signalling devices that trigger mainly protective pathways. In several animal models, H2S has been demonstrated as a protective factor in the context of kidney disorders, in particular of diabetic nephropathy. Here, we focus on the synthesis and action of H2S in glomerular cells, its beneficial effects in the glomerulus and its action in the context of the other gaseous signalling molecules NO and CO.
Macrophages constitute a major part of the tumor-infiltrating immune cells. Within the tumor microenvironment, they acquire an alternatively activated, tumor-supporting phenotype. Factors released by tumor cells are crucial for the recruitment of tumor-associated macrophages. In the present project, we aimed to understand the role of hsa-miR-200c-3p (miR-200c) in the interplay between tumor cells and macrophages. To this end, we employed a coculture system of MCF7 breast tumor cells and primary human macrophages and observed the transfer of miR-200c from apoptotic tumor cells to macrophages, which required intact CD36 receptor in macrophages. We further comprehensively determined miR-200c targets in macrophages by mRNA-sequencing and identified numerous migration-associated mRNAs to be downregulated by miR-200c. Consequently, miR-200c attenuated macrophage infiltration into 3-dimensional tumor spheroids. miR-200c-mediated reduction in infiltration further correlated with a miR-200c migration signature comprised of the four miR-200c-repressed, predicted targets PPM1F, RAB11FIB2, RDX, and MSN.
Physical activity and well-being during the second COVID19-related lockdown in Germany in 2021
(2021)
In the second wave of the COVID-19 pandemic in Germany, lockdown measures were reinstalled and were in place between November 2020 and April 2021, including the closure of physical activity facilities. The aim of the current online survey was to assess the lockdown effects on physical activity and well-being in the general population. Pre-lockdown vs. lockdown differences were tested with the Χ2 test and the Student’s t-test for paired data. Predictor variables to explain compliance with physical activity recommendations were identified using a fixed-effects binary logistic regression analysis. Data of 993 respondents were analyzed. Transport-related and leisure-time physical activity decreased (p < 0.001, d = 0.25, and p < 0.001, d = 0.33, respectively). Compliance with physical activity recommendations decreased from 42.2% to 29.4% (chi2 (1, 1986) = 35.335, p < 0.001, V = 0.13). Well-being decreased significantly (t (990) = 23.405, p < 0.001) by 16.3 points (d = 0.74). Physical activity and well-being declined in German adults during the second COVID-19-related lockdown. Physical activity should be promoted also in light of the emerging evidence on its protective effects against COVID-19.
Background and Objectives: We tested if a novel combination of predictors could improve the accuracy of outcome prediction after transfemoral transcatheter aortic valve implantation (TAVI). Materials and Methods: This prospective study recruited 169 participants (49% female; median age 81 years). The primary endpoint was midterm mortality; secondary endpoints were acute Valve Academic Research Consortium (VARC)-3 complication rate and post-TAVI in-hospital length of stay (LoS). EuroSCORE II (ESII), comorbidities (e.g., coronary artery disease), eGFR (estimated glomerular filtration rate; based on cystatin C), hemoglobin, creatinine, N-Terminal pro-Brain Natriuretic Peptide (NTproBNP) levels and patient-reported outcome measures (PROMs, namely EuroQol-5-Dimension-5-Levels, EQ5D5L; Kansas City Cardiomyopathy Questionnaire, KCCQ; clinical frailty scale, CFS) at baseline were tested as predictors. Regression (uni- and multi-variate Cox; linear; binary logistic) and receiver operating characteristic (ROC)-curve analysis were applied. Results: Within a median follow-up of 439 (318–585) days, 12 participants died (7.1%). Independent predictors of mortality using multivariate Cox regression were baseline eGFR (p = 0.001) and KCCQ (p = 0.037). Based on these predictors, a Linear Prediction Score (LPS1) was calculated. The LPS1-area under the curve (AUC)-value (0.761) was significantly higher than the ESII-AUC value (0.597; p = 0.035). Independent predictors for LoS > 6 days (the median LoS) were eGFR (p = 0.028), NTproBNP (p = 0.034), and EQ5D5L values (p = 0.002); a respective calculated LPS2 provided an AUC value of 0.677 (p < 0.001). Eighty participants (47.3%) experienced complications. Male sex predicted complications only in the univariate analysis. Conclusions: The combination of KCCQ and eGFR can better predict midterm mortality than ES II alone. Combining eGFR, NTproBNP, and EQ5D5L can reliably predict LoS after TAVI. This novel method improves personalized TAVI risk stratification and hence may help reduce post-TAVI risk.
Cellular response of human osteoblasts to different presentations of deproteinized bovine bone
(2022)
Objectives: This study evaluated the cellular response of primary osteoblasts exposed to two different presentations of a low-temperature non-sintered deproteinized bovine bone matrix (DBBM). Materials and methods: Six different baths of a commercially available DBBM block (Bonefill® Porous Block) and one of DBBM granule (Bonefill® Porous) were evaluated to identify the mineral structure and organic or cellular remnants. Samples of the same baths were processed in TRIZOL for RNA extraction and quantification. For the immunologic cell reaction assay, primary human osteoblasts (pOB) were exposed to DBMM block (pOB + B) or granules (pOB + G), or none (control) for 1, 3, or 7 days of cell cultivation. Expression of proinflammatory cytokines by pOB was evaluated by crosslinked ELISA assay. In addition, total DNA amount, as well as cell viability via LDH evaluation, was assessed. Results: Organic remnants were present in DBBM blocks; 45.55% (±7.12) of osteocytes lacunae presented cellular remnants in blocks compared to 17.31% (±1.31) in granules. In three of five batches of blocks, it was possible to isolate bovine RNA. The highest concentration of TGF-β1 was found in supernatants of pOB + G on day 7 (218.85 ± 234.62 pg/mL) (p < 0.05), whereas pOB + B presented the lowest amount of TGF-β1 secretion at the end of evaluation (30.22 ± 14.94 pg/mL, p < 0.05). For IL-6 and OPG, there was no statistical difference between groups, while pOB + G induced more IL-8 secretion than the control (3.03 ± 3.38 ng/mL, p < 0.05). Considering the kinetics of cytokine release during the study period, all groups presented a similar pattern of cytokines, estimated as an increasing concentration for IL-6, IL-8, and OPG during cultivation. Adherent cells were observed on both material surfaces on day 7, according to H&E and OPN staining. Conclusion: Neither tested material induced a pronounced inflammatory response upon osteoblast cultivation. However, further studies are needed to elucidate the potential influence of organic remnants in bone substitute materials on the regeneration process.
Aims: Patients with cardiovascular comorbidities have a significantly increased risk for a critical course of COVID-19. As the SARS-CoV2 virus enters cells via the angiotensin-converting enzyme receptor II (ACE2), drugs which interact with the renin angiotensin aldosterone system (RAAS) were suspected to influence disease severity.
Methods and results: We analyzed 1946 consecutive patients with cardiovascular comorbidities or hypertension enrolled in one of the largest European COVID-19 registries, the Lean European Open Survey on SARS-CoV-2 (LEOSS) registry. Here, we show that angiotensin II receptor blocker intake is associated with decreased mortality in patients with COVID-19 [OR 0.75 (95% CI 0,59–0.96; p = 0.013)]. This effect was mainly driven by patients, who presented in an early phase of COVID-19 at baseline [OR 0,64 (95% CI 0,43–0,96; p = 0.029)]. Kaplan-Meier analysis revealed a significantly lower incidence of death in patients on an angiotensin receptor blocker (ARB) (n = 33/318;10,4%) compared to patients using an angiotensin-converting enzyme inhibitor (ACEi) (n = 60/348;17,2%) or patients who received neither an ACE-inhibitor nor an ARB at baseline in the uncomplicated phase (n = 90/466; 19,3%; p<0.034). Patients taking an ARB were significantly less frequently reaching the mortality predicting threshold for leukocytes (p<0.001), neutrophils (p = 0.002) and the inflammatory markers CRP (p = 0.021), procalcitonin (p = 0.001) and IL-6 (p = 0.049). ACE2 expression levels in human lung samples were not altered in patients taking RAAS modulators.
Conclusion: These data suggest a beneficial effect of ARBs on disease severity in patients with cardiovascular comorbidities and COVID-19, which is linked to dampened systemic inflammatory activity.
Introduction Patients undergoing heart valve surgery are predominantly transferred postoperatively to the intensive care unit (ICU) under continuous sedation. Volatile anaesthetics are an increasingly used treatment alternative to intravenous substances in the ICU. As subject to inhalational uptake and elimination, the resulting pharmacological benefits have been repeatedly demonstrated. Therefore, volatile anaesthetics appear suitable to meet the growing demands of fast-track cardiac surgery. However, their use requires special preparation at the bedside and trained medical and nursing staff, which might limit the pharmacological benefits. The aim of our work is to assess whether the temporal advantages of recovery under volatile sedation outweigh the higher effort of special preparation.
Methods and analysis The study is designed to evaluate the differences between intravenous sedatives (n=48) and volatile sedatives (n=48) in continued intensive care sedation. This study will be conducted as a prospective, randomised, controlled, single-blinded, monocentre trial at a German university hospital in consenting adult patients undergoing heart valve surgery at a university hospital. This observational study will examine the necessary preparation time, staff consultation and overall feasibility of the chosen sedation method. For this purpose, the continuation of sedation in the ICU with volatile sedatives is considered as one study arm and with intravenous sedatives as the comparison group. Due to rapid elimination and quick awakening after the termination of sedation, closer consultation between the attending physician and the ICU nursing staff is required, in addition to a prolonged setup time. Study analysis will include the required setup time, time from admission to extubation as primary outcome and neurocognitive assessability. In addition, possible operation-specific (blood loss, complications), treatment parameters (catecholamine dosages, lung function) and laboratory results (acute kidney injury, acid base balance (lactataemia), liver failure) as influencing factors will be collected. The study-relevant data will be extracted from the continuous digital records of the patient data management system after the patient has been discharged from the ICU. For statistical evaluation, 95% CIs will be calculated for the median time to extubation and neurocognitive assessability, and the association will be assessed with a Cox regression model. In addition, secondary binary outcome measures will be evaluated using Fisher’s exact tests. Further descriptive and exploratory statistical analyses are also planned.
Ethics and dissemination The study was approved by the Institutional Ethics Board of the University of Frankfurt, Germany (#20-1050). Informed consent of all individual patients will be obtained before randomisation. Results will be disseminated via publication in peer-reviewed journals.
Hintergrund: Eine adäquate Anpassung der Glukokortikoidsubstitution an unterschiedliche Situationen ist essenziell für Leistungsfähigkeit und Lebensqualität von Patienten mit Nebennierenrindeninsuffizienz (NNRI). Sie dient darüber hinaus der Vermeidung lebensbedrohlicher adrenaler Krisen.
Ziel der Arbeit: Verbesserung der Versorgung von Patienten mit Nebennierenrindeninsuffizienz.
Material und Methoden: Selektive Literaturrecherche unter besonderer Berücksichtigung neuerer Studien.
Ergebnisse: Eine optimale Glukokortikoidsubstitution hat das Ziel, die physiologischen Kortisolschwankungen möglichst genau nachzuahmen. Hier haben in den letzten Jahren Präparate mit veränderter Pharmakokinetik das Therapiespektrum erweitert. Im Vordergrund stehen eine adäquate Anpassung der Substitution in Stresssituationen sowie die Vermeidung und adäquate Behandlung adrenaler Krisen, die mit einer Inzidenz von 4,8 bis 8,3 Krisen pro 100 Patientenjahre auftreten und mit einer Mortalität von etwa 0,5 pro 100 Patientenjahre weiterhin eine tödliche Bedrohung darstellen.
Schlussfolgerung: Zur Verhinderung lebensbedrohlicher Nebennierenkrisen ist eine Schulung von Patienten, Angehörigen und insbesondere auch medizinischem Personal notwendig.
Autophagy is an important survival mechanism that allows recycling of nutrients and removal of damaged organelles and has been shown to contribute to the proliferation of acute myeloid leukemia (AML) cells. However, little is known about the mechanism by which autophagy- dependent AML cells can overcome dysfunctional autophagy. In our study we identified autophagy related protein 3 (ATG3) as a crucial autophagy gene for AML cell proliferation by conducting a CRISPR/Cas9 dropout screen with a library targeting around 200 autophagy-related genes. shRNA-mediated loss of ATG3 impaired autophagy function in AML cells and increased their mitochondrial activity and energy metabolism, as shown by elevated mitochondrial ROS generation and mitochondrial respiration. Using tracer-based NMR metabolomics analysis we further demonstrate that the loss of ATG3 resulted in an upregulation of glycolysis, lactate production, and oxidative phosphorylation. Additionally, loss of ATG3 strongly sensitized AML cells to the inhibition of mitochondrial metabolism. These findings highlight the metabolic vulnerabilities that AML cells acquire from autophagy inhibition and support further exploration of combination therapies targeting autophagy and mitochondrial metabolism in AML.
Uterine cervical cancer is one of the leading causes of cancer-related mortality in women worldwide. Each year, over half a million new cases are estimated, resulting in more than 300,000 deaths. While less-invasive, fertility-preserving surgical procedures can be offered to women in early stages, treatment for locally advanced disease may include radical hysterectomy, primary chemoradiotherapy (CRT) or a combination of these modalities. Concurrent platinum-based chemoradiotherapy regimens remain the first-line treatments for locally advanced cervical cancer. Despite achievements such as the introduction of angiogenesis inhibitors, and more recently immunotherapies, the overall survival of women with persistent, recurrent or metastatic disease has not been extended significantly in the last decades. Furthermore, a broad spectrum of molecular markers to predict therapy response and survival and to identify patients with high- and low-risk constellations is missing. Implementation of these markers, however, may help to further improve treatment and to develop new targeted therapies. This review aims to provide comprehensive insights into the complex mechanisms of cervical cancer pathogenesis within the context of molecular markers for predicting treatment response and prognosis.
The microtubule (MT) cytoskeleton is crucial for cell motility and migration by regulating multiple cellular activities such as transport and endocytosis of key components of focal adhesions (FA). The kinesin-13 family is important in the regulation of MT dynamics and the best characterized member of this family is the mitotic centromere-associated kinesin (MCAK/KIF2C). Interestingly, its overexpression has been reported to be related to increased metastasis in various tumor entities. Moreover, MCAK is involved in the migration and invasion behavior of various cell types. However, the precise molecular mechanisms were not completely clarified. To address these issues, we generated CRISPR/dCas9 HeLa and retinal pigment epithelium (RPE) cell lines overexpressing or downregulating MCAK. Both up- or downregulation of MCAK led to reduced cell motility and poor migration in malignant as well as benign cells. Specifically, it’s up- or downregulation impaired FA protein composition and phosphorylation status, interfered with a proper spindle and chromosome segregation, disturbed the assembly and disassembly rate of FA, delayed cell adhesion, and compromised the plus-tip dynamics of MTs. In conclusion, our data suggest MCAK act as an important regulator for cell motility and migration by affecting the actin-MT cytoskeleton dynamics and the FA turnover, providing molecular mechanisms by which deregulated MCAK could promote malignant progression and metastasis of tumor cells.
Background: Inflammation, particularly cytokine release, contributes to epileptogenesis by influencing the cerebral tissue remodeling and neuronal excitability that occurs after a precipitating epileptogenic insult. While several cytokines have been explored in this process, release kinetics are less well investigated. Determining the time course of cytokine release in the epileptogenic zone is necessary for precisely timed preventive or therapeutic anti-inflammatory interventions. Methods: Hippocampal extracellular levels of six cytokines and chemokines (IL-1β, IL-6, IL-10, CCL2, CCL3, and CCL5) were quantified at various time points during epileptogenesis in a rat model of mesial temporal lobe epilepsy with hippocampal sclerosis (mTLE-HS) using microdialysis (MD). Results: The analysis of microdialysates demonstrated consistent elevation at all time points during epileptogenesis for IL-1β and IL-10. IL-10 release was maximal on day 1, IL-1β release peaked at day 8. No correlation between local hippocampal IL-1β concentrations and IL-1β blood levels was found. Conclusion: The release kinetics of IL-1β are consistent with its established pro-epileptogenic properties, while the kinetics of IL-10 suggest a counter-regulatory effect. This proof-of-concept study demonstrates the feasibility of intraindividual longitudinal monitoring of hippocampal molecular inflammatory processes via repetitive MD over several weeks and sheds light on the kinetics of hippocampal cytokine release during epileptogenesis.
In recent years, the infrapatellar fat pad (IFP) has gained increasing research interest. The contribution of the IFP to the development and progression of knee osteoarthritis (OA) through extensive interactions with the synovium, articular cartilage, and subchondral bone is being considered. As part of the initiation process of OA, IFP secretes abundant pro-inflammatory mediators among many other factors. Today, the IFP is (partially) resected in most total knee arthroplasties (TKA) allowing better visualization during surgical procedures. Currently, there is no clear guideline providing evidence in favor of or against IFP resection. With increasing numbers of TKAs, there is a focus on preventing adverse postoperative outcomes. Therefore, anatomic features, role in the development of knee OA, and consequences of resecting versus preserving the IFP during TKA are reviewed in the following article.
We proposed previously that closure of voltage-dependent anion channels (VDAC) in the mitochondrial outer membrane after ethanol exposure leads to suppression of mitochondrial metabolite exchange. Because ureagenesis requires extensive mitochondrial metabolite exchange, we characterized the effect of ethanol and its metabolite, acetaldehyde (AcAld), on total and ureagenic respiration in cultured rat hepatocytes. Ureagenic substrates increased cellular respiration from 15.8 ± 0.9 nmol O(2)/min/10(6) cells (base line) to 29.4 ± 1.7 nmol O(2)/min/10(6) cells in about 30 min. Ethanol (0-200 mM) suppressed extra respiration after ureagenic substrates (ureagenic respiration) by up to 51% but not base line respiration. Urea formation also declined proportionately. Inhibition of alcohol dehydrogenase, cytochrome P450 2E1, and catalase with 4-methylpyrazole, trans-1,2-dichloroethylene, and 3-amino-1,2,3-triazole restored ethanol-suppressed ureagenic respiration by 46, 37, and 66%, respectively. By contrast, inhibition of aldehyde dehydrogenase with phenethyl isothiocyanate increased the inhibitory effect of ethanol on ureagenic respiration by an additional 60%. AcAld, an intermediate product of ethanol oxidation, suppressed ureagenic respiration with an apparent IC(50) of 125 μM. AcAld also inhibited entry of 3-kDa rhodamine-conjugated dextran in the mitochondrial intermembrane space of digitonin-permeabilized hepatocytes, indicative of VDAC closure. In conclusion, AcAld, derived from ethanol metabolism, suppresses ureagenesis in hepatocytes mediated by closure of VDAC.
Several microRNAs (miRNAs) are associated with the molecular pathogenesis of hepatocellular carcinoma (HCC). However, previous studies analyzing the dysregulation of miRNAs in HCC show heterogeneous results. We hypothesized that part of this heterogeneity might be attributable to variations of miRNA expression deriving from the HCC capsule or the fibrotic septa within the peritumoral tissue used as controls. Tissue from surgically resected hepatitis C–associated HCC from six well-matched patients was microdissected using laser microdissection and pressure catapulting technique. Four distinct histologic compartments were isolated: tumor parenchyma (TP), fibrous capsule of the tumor (TC), tumor-adjacent liver parenchyma (LP), and cirrhotic septa of the tumor-adjacent liver (LC). MiRNA expression profiling analysis of 1105 mature miRNAs and precursors was performed using miRNA microarray. Principal component analysis and consecutive pairwise supervised comparisons demonstrated distinct patterns of expressed miRNAs not only for TP versus LP (e.g., intratumoral down-regulation of miR-214, miR-199a, miR-146a, and miR-125a; P< .05) but also for TC versus LC (including down-regulation within TC of miR-126, miR-99a/100, miR-26a, and miR-125b; P< .05). The tumor capsule therefore demonstrates a tumor-like phenotype with down-regulation of well-known tumor-suppressive miRNAs. Variations of co-analyzed fibrotic tissue within the tumor or in controls may have profound influence on miRNA expression analyses in HCC. Several miRNAs, which are proposed to be HCC specific, may indeed be rather associated to the tumor capsule. As miRNAs evolve to be important biomarkers in liver tumors, the presented data have important translational implications on diagnostics and treatment in patients with HCC.
Searching for new strategies to trigger apoptosis in rhabdomyosarcoma (RMS), we investigated the effect of two novel classes of apoptosis-targeting agents, i.e. monoclonal antibodies against TNF-related apoptosis-inducing ligand (TRAIL) receptor 1 (mapatumumab) and TRAIL receptor 2 (lexatumumab) and small-molecule inhibitors of inhibitor of apoptosis (IAP) proteins. Here, we report that IAP inhibitors synergized with lexatumumab, but not with mapatumumab, to reduce cell viability and to induce apoptosis in several RMS cell lines in a highly synergistic manner (combination index <0.1). Cotreatment-induced apoptosis was accompanied by enhanced activation of caspase-8, -9, and -3; loss of mitochondrial membrane potential; and caspase-dependent apoptosis. In addition, IAP inhibitor and lexatumumab cooperated to stimulate the assembly of a cytosolic complex containing RIP1, FADD, and caspase-8. Importantly, knockdown of RIP1 by RNA interference prevented the formation of the RIP1·FADD·caspase-8 complex and inhibited subsequent activation of caspase-8, -9, and -3; loss of mitochondrial membrane potential; and apoptosis upon treatment with IAP inhibitor and lexatumumab. In addition, RIP1 silencing rescued clonogenic survival of cells treated with the combination of lexatumumab and IAP inhibitor, thus underscoring the critical role of RIP1 in cotreatment-induced apoptosis. By comparison, the TNFα-blocking antibody Enbrel had no effect on IAP inhibitor/lexatumumab-induced apoptosis, indicating that an autocrine TNFα loop is dispensable. By demonstrating that IAP inhibitors and lexatumumab synergistically trigger apoptosis in a RIP1-dependent but TNFα-independent manner in RMS cells, our findings substantially advance our understanding of IAP inhibitor-mediated regulation of TRAIL-induced cell death.
Macrophages respond to the Th2 cytokine IL-4 with elevated expression of arachidonate 15-lipoxygenase (ALOX15). Although IL-4 signaling elicits anti-inflammatory responses, 15-lipoxygenase may either support or inhibit inflammatory processes in a context-dependent manner. AMP-activated protein kinase (AMPK) is a metabolic sensor/regulator that supports an anti-inflammatory macrophage phenotype. How AMPK activation is linked to IL-4-elicited gene signatures remains unexplored. Using primary human macrophages stimulated with IL-4, we observed elevated ALOX15 mRNA and protein expression, which was attenuated by AMPK activation. AMPK activators, e.g. phenformin and aminoimidazole-4-carboxamide 1-β-d-ribofuranoside inhibited IL-4-evoked activation of STAT3 while leaving activation of STAT6 and induction of typical IL-4-responsive genes intact. In addition, phenformin prevented IL-4-induced association of STAT6 and Lys-9 acetylation of histone H3 at the ALOX15 promoter. Activating AMPK abolished cellular production of 15-lipoxygenase arachidonic acid metabolites in IL-4-stimulated macrophages, which was mimicked by ALOX15 knockdown. Finally, pretreatment of macrophages with IL-4 for 48 h increased the mRNA expression of the proinflammatory cytokines IL-6, IL-12, CXCL9, and CXCL10 induced by subsequent stimulation with lipopolysaccharide. This response was attenuated by inhibition of ALOX15 or activation of AMPK during incubation with IL-4. In conclusion, limiting ALOX15 expression by AMPK may promote an anti-inflammatory phenotype of IL-4-stimulated human macrophages.
The Na+-F1F0-ATPase operon ofAcetobacterium woodii was recently shown to contain, among eleven atp genes, those genes that encode subunita and b, a gene encoding a 16-kDa proteolipid (subunit c 1), and two genes encoding 8-kDa proteolipids (subunits c 2 andc 3). Because subunits a,b, and c 1 were not found in previous enzyme preparations, we re-determined the subunit composition of the enzyme. The genes were overproduced, and specific antibodies were raised. Western blots revealed that subunits a,b, and c 1 are produced and localized in the cytoplasmic membrane. Membrane protein complexes were solubilized by dodecylmaltoside and separated by blue native-polyacrylamide gel electrophoresis, and the ATPase subunits were resolved by SDS-polyacrylamide gel electrophoresis. N-terminal sequence analyses revealed the presence of subunitsa, c 2, c 3,b, δ, α, γ, β, and ε. Biochemical and immunological analyses revealed that subunitsc 1, c 2, andc 3 are all part of the c-oligomer, the first of a F1F0-ATPase that contains 8- and 16-kDa proteolipids.
Mechanical stress is known to modulate fundamental events such as cell life and death. Mechanical stretch in particular has been identified as a positive regulator of proliferation in skin keratinocytes and other cell systems. In the present study it was investigated whether antiapoptotic signaling is also stimulated by mechanical stretch. It was demonstrated that mechanical stretch rapidly induced the phosphorylation of the proto-oncogene protein kinase B (PKB)/Akt at both phosphorylation sites (serine 473/threonine 308) in different epithelial cells (HaCaT, A-431, and human embryonic kidney-293). Blocking of phosphoinositide 3-OH kinase by selective inhibitors (LY-294002 and wortmannin) abrogated the stretch-induced PKB/Akt phosphorylation. Furthermore mechanical stretch stimulated phosphorylation of epidermal growth factor receptor (EGFR) and the formation of EGFR membrane clusters. Functional blocking of EGFR phosphorylation by either selective inhibitors (AG1478 and PD168393) or dominant-negative expression suppressed stretch-induced PKB/Akt phosphorylation. Finally, the angiotensin II type 1 receptor (AT1-R) was shown to induce positive transactivation of EGFR in response to cell stretch. These findings define a novel signaling pathway of mechanical stretch, namely the activation of PKB/Akt by transactivation of EGFR via angiotensin II type 1 receptor. Evidence is provided that stretch-induced activation of PKB/Akt protects cells against induced apoptosis.
Factor XI (FXI), the zymogen of the blood coagulation protease FXIa, and the structurally homologous protein plasma prekallikrein circulate in plasma in noncovalent complexes with H-kininogen (HK). HK binds to the heavy chains of FXI and of prekallikrein. Each chain contains four apple domains (F1-F4 for FXI and P1-P4 for prekallikrein). Previous studies indicated that the HK-binding site on FXI is located in F1, whereas the major HK-binding site on prekallikrein is in P2. To determine the contribution of each FXI apple domain to HK-FXI complex formation, we examined binding of recombinant single apple domain-tissue plasminogen activator fusion proteins to HK. The order of affinity from highest to lowest is F2 F4 > F1 F3. Monoclonal antibodies against F2 are superior to F4 or F1 antibodies as inhibitors of HK binding to FXI. Antibody alphaP2, raised against prekallikrein, cross-reacts with FXI F2 and inhibits FXI-HK binding with an IC(50) of 8 nm. HK binding to a platelet-specific FXI variant lacking the N-terminal half of F2 is reduced > 5-fold compared with full-length FXI. A chimeric FXI molecule in which F2 is replaced by P2 is cleaved within P2 during activation by factor XIIa, resulting in greatly reduced HK binding capacity. In contrast, wild-type FXI is not cleaved within F2, and its binding capacity for HK is unaffected by factor XIIa. Our data show that HK binding to FXI involves multiple apple domains, with F2 being most important. The findings demonstrate a similarity in mechanism for FXI and prekallikrein binding to HK.
Cholecystokinin (CCK) and related peptides are potent growth factors in the gastrointestinal tract and may be important for human cancer. CCK exerts its growth modulatory effects through G(q)-coupled receptors (CCK(A) and CCK(B)) and activation of extracellular signal-regulated protein kinase 1/2 (ERK1/2). In the present study, we investigated the different mechanisms participating in CCK-induced activation of ERK1/2 in pancreatic AR42J cells expressing both CCK(A) and CCK(B). CCK activated ERK1/2 and Raf-1 to a similar extent as epidermal growth factor (EGF). Inhibition of EGF receptor (EGFR) tyrosine kinase or expression of dominant-negative Ras reduced CCK-induced ERK1/2 activation, indicating participation of the EGFR and Ras in CCK-induced ERK1/2 activation. However, compared with EGF, CCK caused only small increases in tyrosine phosphorylation of the EGFR and Shc, Shc-Grb2 complex formation, and Ras activation. Signal amplification between Ras and Raf in a CCK-induced ERK cascade appears to be mediated by activation of protein kinase Cepsilon (PKCepsilon), because 1) down-modulation of phorbol ester-sensitive PKCs inhibited CCK-induced activation of Ras, Raf, and ERK1/2 without influencing Shc-Grb2 complex formation; 2) PKCepsilon, but not PKCalpha or PKCdelta, was detectable in Raf-1 immunoprecipitates, although CCK activated all three PKC isoenzymes. In addition, the present study provides evidence that the Src family tyrosine kinase Yes is activated by CCK and mediates CCK-induced tyrosine phosphorylation of Shc. Furthermore, we show that CCK-induced activation of the EGFR and Yes is achieved through the CCK(B) receptor. Together, our data show that different signals emanating from the CCK receptors mediate ERK1/2 activation; activation of Yes and the EGFR mediate Shc-Grb2 recruitment, and activation of PKC, most likely PKCepsilon, augments CCK-stimulated ERK1/2 activation at the Ras/Raf level.
In PC12 cells, a well studied model for neuronal differentiation, an elevation in the intracellular cAMP level increases cell survival, stimulates neurite outgrowth, and causes activation of extracellular signal-regulated protein kinase 1 and 2 (ERK1/2). Here we show that an increase in the intracellular cAMP concentration induces tyrosine phosphorylation of two receptor tyrosine kinases, i.e. the epidermal growth factor (EGF) receptor and the high affinity receptor for nerve growth factor (NGF), also termed Trk(A). cAMP-induced tyrosine phosphorylation of the EGF receptor is rapid and correlates with ERK1/2 activation. It occurs also in Panc-1, but not in human mesangial cells. cAMP-induced tyrosine phosphorylation of the NGF receptor is slower and correlates with Akt activation. Inhibition of EGF receptor tyrosine phosphorylation, but not of the NGF receptor, reduces cAMP-induced neurite outgrowth. Expression of dominant-negative Akt does not abolish cAMP-induced survival in serum-free media, but increases cAMP-induced ERK1/2 activation and neurite outgrowth. Together, our results demonstrate that cAMP induces dual signaling in PC12 cells: transactivation of the EGF receptor triggering the ERK1/2 pathway and neurite outgrowth; and transactivation of the NGF receptor promoting Akt activation and thereby modulating ERK1/2 activation and neurite outgrowth.
Erratum in: Correction: Cyclic AMP induces transactivation of the receptors for epidermal growth factor and nerve growth factor, thereby modulating activation of MAP kinase, Akt, and neurite outgrowth in PC12 cells. Journal of biological chemistry 2020 Oct 23;295(43):14792. doi: 10.1074/jbc.AAC120.016177.
Erratum for: Cyclic AMP induces transactivation of the receptors for epidermal growth factor and nerve growth factor, thereby modulating activation of MAP kinase, Akt, and neurite outgrowth in PC12 cells.Journal of biological chemistry, 2002 Nov 15;277(46):43623-30. doi: 10.1074/jbc.M203926200. Epub 2002 Sep 5.
Soluble guanylyl cyclase (sGC) is the major cytosolic receptor for nitric oxide (NO) that converts GTP into the second messenger cGMP in a NO-dependent manner. Other factors controlling this key enzyme are intracellular proteins such as Hsp90 and PSD95, which bind to sGC and modulate its activity, stability, and localization. To date little is known about the effects of posttranslational modifications of sGC, although circumstantial evidence suggests that reversible phosphorylation may contribute to sGC regulation. Here we demonstrate that inhibitors of protein-tyrosine phosphatases such as pervanadate and bisperoxo(1,10-phenanthroline)oxovanadate(V) as well as reactive oxygen species such as H2O2 induce specific tyrosine phosphorylation of the β1 but not of the α1 subunit of sGC. Tyrosine phosphorylation of sGCβ1 is also inducible by pervanadate and H2O2 in intact PC12 cells, rat aortic smooth muscle cells, and in rat aortic tissues, indicating that tyrosine phosphorylation of sGC may also occur in vivo. We have mapped the major tyrosine phosphorylation site to position 192 of β1, where it forms part of a highly acidic phospho-acceptor site for Src-like kinases. In the phosphorylated state Tyr(P)-192 exposes a docking site for SH2 domains and efficiently recruits Src and Fyn to sGCβ1, thereby promoting multiple phosphorylation of the enzyme. Our results demonstrate that sGC is subject to tyrosine phosphorylation and interaction with Src-like kinases, revealing an unexpected cross-talk between the NO/cGMP and tyrosine kinase signaling pathways at the level of sGC.
Nitric oxide (NO)-sensitive soluble guanylyl cyclase (sGC) is the major cytosolic receptor for NO, catalyzing the conversion of GTP to cGMP. In a search for proteins specifically interacting with human sGC, we have identified the multidomain protein AGAP1, the prototype of an ArfGAP protein with a GTPase-like domain, Ankyrin repeats, and a pleckstrin homology domain. AGAP1 binds through its carboxyl terminal portion to both the α1 and β1 subunits of sGC. We demonstrate that AGAP1 mRNA and protein are co-expressed with sGC in human, murine, and rat cells and tissues and that the two proteins interact in vitro and in vivo. We also show that AGAP1 is prone to tyrosine phosphorylation by Src-like kinases and that tyrosine phosphorylation potently increases the interaction between AGAP1 and sGC, indicating that complex formation is modulated by reversible phosphorylation. Our findings may hint to a potential role of AGAP1 in integrating signals from Arf, NO/cGMP, and tyrosine kinase signaling pathways.
Tensiomyography measures the radial displacement of a muscle during an electrically evoked twitch contraction. The rate of muscle displacement is increasingly reported to assess contractile properties. Several formulas currently exist to calculate the rate of displacement during the contraction phase of the maximal twitch response. However, information on the reproducibility of these formulas is scarce. Further, different rest intervals ranging from 10 s to 30 s are applied between consecutive stimuli during progressive electrical stimulation until the maximum twitch response. The effect of different rest intervals on the rate of displacement has not been investigated so far. The first aim of this study is to investigate the within and between-day reliability of the most frequently used formulas to calculate the rate of displacement. The second aim is to investigate the effect of changing the inter-stimulus interval on the rate of displacement. We will determine the rectus femoris and biceps femoris rate of displacement of twenty-four healthy subjects’ dominant leg on two consecutive days. The maximum displacement curve will be determined two times within three minutes on the first day and a third time 24 h later. On day two, we will also apply three blocks of ten consecutive stimuli at a constant intensity of 50 mA. Inter-stimuli intervals will be 10 s, 20 s or 30 s in each block, respectively, and three minutes between blocks. The order of inter-stimulus intervals will be randomized. This study will allow a direct comparison between the five most frequently used formulas to calculate the rate of displacement in terms of their reproducibility. Our data will also inform on the effect of different inter-stimulus intervals on the rate of displacement. These results will provide helpful information on methodical considerations to determine the rate of displacement and may thus contribute to a standardized approach.
Cervical cancer has caused substantial morbidity and mortality for millions of women over the past decades. While enormous progress has been made in diagnosis, prevention and therapy, the disease is still fatal for many women—especially in low-income countries. Since no detailed studies are available on the worldwide research landscape, we here investigated the global scientific output related to this cancer type by an established protocol. The “New Quality and Quantity Indices in Science” platform assessed all relevant cervical cancer research published in the Web of Science since 1900. A detailed analysis was conducted including country-specific research productivity, indicators for scientific quality, and relation of research activity to socioeconomic and epidemiologic figures. Visualization of data was generated by the use of density equalizing map projections. Our approach identified 22,185 articles specifically related to cervical cancer. From a global viewpoint, the United States of America was the dominating country in absolute numbers, being followed by China and Japan. By contrast, the European countries Sweden, Austria, and Norway were positioned first when the research activity was related to the population number. When the scientific productivity was related to annual cervical cancer cases, Scandinavian countries (Finland #1, Sweden #4, Norway #5, Denmark #7), the Alpine countries Austria (#2) and Switzerland (#6), and the Netherlands (#3) were leading the field. Density equalizing mapping visualized that large parts of Africa and South America were almost invisible regarding the global participation in cervical cancer research. Our data documented that worldwide cervical cancer research activity is continuously increasing but is imbalanced from a global viewpoint. Also, the study indicated that global and public health aspects should be strengthened in cervical carcinoma research in order to empower more countries to take part in international research activities.
Objective: After stratifying for age, sex and multimorbidity at baseline, our aim is to analyse time trends in incident multimorbidity and polypharmacy in the 15-year clinical trajectories of individual patients in a family medicine setting.
Methods: This study was carried out using data from the Registration Network Family Medicine in the South of the Netherlands. The clinical trajectories of 10037 subjects during the 15-year period (2000–2014) were analyzed in a repeated measurement of using a generalized estimating equations model as well as a multilevel random intercept model with repeated measurements to determine patterns of incident multimorbidity and polypharmacy. Hierarchical age-period-cohort models were used to generate age and cohort trajectories for comparison with prevalence trends in multimorbidity literature.
Results: Multimorbidity was more common in females than in males throughout the duration of the 15-year trajectory (females: 39.6%; males: 33.5%). With respective ratios of 11.7 and 5.9 between the end and the beginning of the 15-year period, the youngest female and male groups showed a substantial increase in multimorbidity prevalence. Ratios in the oldest female and male groups were 2.2 and 1.9 respectively. Females had higher levels of multimorbidity than males in the 0-24-year and 25-44-year age groups, but the levels converged to a prevalence of 92.2% in the oldest male and 90.7% in the oldest female group. Similar, albeit, moderate differences were found in polypharmacy patterns.
Conclusions: We sought to specify the progression of multimorbidity from an early age. As a result, our study adds to the multimorbidity literature by specifying changes in chronic disease accumulation with relation to polypharmacy, and by tracking differences in patient trajectories according to age and sex. Multimorbidity and polypharmacy are common and their prevalence is accelerating, with a relatively rapid increase in younger groups. From the point of view of family medicine, this underlines the need for a longitudinal approach and a life course perspective in patient care.
Purpose: Monocentric, prospective study to investigate whether concomitant support of cochlear implant (CI) patients by CI-trained otolaryngologists and application of a standardized head bandage can minimize potential complications during magnetic resonance imaging (MRI).
Methods: Thirty-seven patients with 46 CIs underwent MRI with a prophylactic head bandage. All participants and the otolaryngologist at the CI center completed pre- and post-MRI questionnaires documenting body region scanned, duration of MRI and bandage wear, field strength during the scan, and any complications. If pain was experienced, it was assessed using a visual analog scale (1–10).
Results: MRI was performed without adverse events in 37.8% of cases. Magnet dislocation requiring surgical revision occurred in 2% of cases. Pain was reported in 86% of cases, often due to the tightness of the dressing. Patients with rotating, MRI-compatible magnets reported significantly less pain than participants with older-generation implants. In 11% of cases, the MRI was discontinued.
Conclusion: Serious complications during MRI in cochlear implant patients are rare. Pain is the most common adverse event, probably mainly due to the tight bandage required by most implant types. With newer generations of magnets, these patients experience less pain, no dislocation of the magnets, and no need for bandaging. Although magnet dislocation cannot be completely prevented in older generations of implants, it appears to be reduced by good patient management, which recommends examination under the guidance of physicians trained in the use of hearing implants.
Bilateral simultaneous cochlear implantation is a safe method of hearing rehabilitation in adults
(2023)
Purpose: Bilateral cochlear implantation is an effective treatment for patients with bilateral profound hearing loss. In contrast to children, adults mostly choose a sequential surgery. This study addresses whether simultaneous bilateral CI is associated with higher rates of complications compared to sequential implantation.
Methods: 169 bilateral CI surgeries were analyzed retrospectively. 34 of the patients were implanted simultaneously (group 1), whereas 135 patients were implanted sequentially (group 2). The duration of surgery, the incidence of minor and major complications and the duration of hospitalization of both groups were compared.
Results: In group 1, the total operating room time was significantly shorter. The incidences of minor and major surgical complications showed no statistically significant differences. A fatal non-surgical complication in group 1 was particularly extensively reappraised without evidence of a causal relationship to the chosen mode of care. The duration of hospitalization was 0.7 days longer than in unilateral implantation but 2.8 days shorter than the combined two hospital stays in group 2.
Conclusion: In the synopsis of all considered complications and complication-relevant factors, equivalence of simultaneous and sequential cochlear implantation in adults in terms of safety was found. However, potential side effects related to longer surgical time in simultaneous surgery must be considered individually. Careful patient selection with special consideration to existing comorbidities and preoperative anesthesiologic evaluation is essential.
Untreated periodontal disease may influence general health. However, how may a physician, who is not trained in periodontal probing, detect untreated periodontitis?
Activated matrix metalloproteinase-8 (aMMP-
8) in saliva correlates with periodontal probing parameters. Thus, sensitivity and specificity of a chair-side test for aMMP-8 to detect periodontitis were evaluated. Thirty cases [untreated chronic periodontitis (ChP); 15 generalized moderate and 15 generalized severe] and 30 controls [probing depths (PD) ≤3 mm, vertical probing attachment level (PAL-V) ≤2 mm at <30 % of sites) were examined periodontally (PD, PAL-V, bleeding on probing). Subsequently, the aMMP-8 test was performed. The test kit
becomes positive with ≥25 ng/ml aMMP-8 in the sample.
The aMMP-8 test was positive in 87 % of ChP and in 40 % of controls. That corresponds to a sensitivity of 87 % and a specificity of 60 %. The sensitivity to detect generalized severe ChP was 93 % (60 % specificity). Backward
stepwise logistic regression analysis to explain positive
aMMP-8 tests identified exclusively ChP with an odds of 9.8 (p < 0.001). Positive results of the aMMP-8 test significantly correlate with generalized ChP. The aMMP-8 test may be used by physicians to detect periodontitis in their patients.
Integrin receptors contribute to hepatocellular carcinoma (HCC) invasion, while AKT-mTOR signaling controls mitosis. The present study was designed to explore the links between integrins and the AKT-mTOR pathway and the CDK-Cyclin axis. HCC cell lines (HepG2, Huh7, Hep3B) were stimulated with soluble collagen or Matrigel to activate integrins, or with insulin-like growth factor 1 (IGF1) to activate AKT-mTOR. HCC growth, proliferation, adhesion, and chemotaxis were evaluated. AKT/mTOR-related proteins, proteins of the CDK-Cyclin axis, focal adhesion kinase (FAK), and integrin-linked kinase (ILK) were determined following IGF1-stimulation or integrin knockdown. Stimulation with collagen or Matrigel increased tumor cell growth and proliferation. This was associated with significant alteration of the integrins α2, αV, and β1. Blockade of these integrins led to cell cycle arrest in G2/M and diminished the number of tumor cell clones. Knocking down the integrins α2 or β1 suppressed ILK, reduced FAK-phosphorylation and diminished AKT/mTOR, as well as the proteins of the CDK-Cyclin axis. Activating the cells with IGF1 enhanced the expression of the integrins α2, αV, β1, activated FAK, and increased tumor cell adhesion and chemotaxis. Blocking the AKT pathway canceled the enhancing effect of IGF on the integrins α2 and β1. These findings reveal that HCC growth, proliferation, and invasion are controlled by a fine-tuned network between α2/β1-FAK signaling, the AKT-mTOR pathway, and the CDK–Cyclin axis. Concerted blockade of the integrin α2/β1 complex along with AKT-mTOR signaling could, therefore, provide an option to prevent progressive dissemination of HCC.
We aimed to evaluate the factors associated with hemorrhage (HA) of melanoma brain metastases (MBM) after Cyberknife stereotactic radiosurgery (SRS) in the modern era of systemic therapy. A total of 55 patients with 279 MBM were treated in 93 fractions. The median age, SRS dose, radiological follow-up, and time to HA were 60.4 years, 20 Gy, 17.7 months, and 10.7 months, respectively. Radiologically evident HA was documented in 47 (16.8%) metastases. Of the 55 patients, 25 (45.4%) suffered an HA. Among those, HA caused grade 3 toxicity in 10 patients (40%) and grade 1 symptoms in 5 patients (20%). Ten patients (40%) with HA experienced no toxicity. Logistic regression revealed the use of anticoagulants and the administration of systemic therapy within 7/15 days from SRS to be predictive for HA. When considering the HA causing grade 3 symptomatology, only the use of anticoagulants was significant, with the delivery of whole brain radiation therapy (WBRT) before the HA narrowly missing statistical significance. Our retrospective analysis showed that the administration of modern systemic therapy within 7/15 days from SRS may contribute to HA of MBM, though it appears safe, at least concerning grade 3 toxicity. The use of anticoagulants by the time of SRS significantly increased the risk of HA.
The combination of atezolizumab and bevacizumab (A + B) is the new standard of care for the systemic first-line treatment of hepatocellular carcinoma (HCC). However, up to now there are only few data on the safety and efficacy of A + B in real life. We included patients with advanced HCC treated with A + B as first-line therapy at four cancer centers in Germany and Austria between December 2018 and August 2021. Demographics, overall survival (OS), and adverse events were assessed until 15 September 2021. We included 66 patients. Most patients had compensated cirrhosis (n = 34; 52%), while Child–Pugh class B cirrhosis was observed in 23 patients (35%), and class C cirrhosis in 5 patients (8%). The best responses included a complete response (CR) in 7 patients (11%), a partial response (PR) in 12 patients (18%), stable disease (SD) in 22 patients (33%), and progressive disease in 11 patients (17%). The median progression-free (PFS) survival was 6.5 months, while the median overall survival (OS) was not reached in this cohort (6-month OS: 69%, 12-month OS: 60%, 18-month OS: 58%). Patients with viral hepatitis seemed to have a better prognosis than patients with HCC of non-viral etiology. The real-world PFS and OS were comparable to those of the pivotal IMBRAVE trial, despite including patients with worse liver function in this study. We conclude that A + B is also highly effective in a real-life setting, with manageable toxicity, especially in patients with compensated liver disease. In patients with compromised liver function (Child B and C), the treatment showed low efficacy and, therefore, it should be well considered before administration to these patients.
DNA mismatch repair (MMR) deficiency plays an essential role in the development of colorectal cancer (CRC). We recently demonstrated in vitro that the serine/threonine casein kinase 2 alpha (CK2α) causes phosphorylation of the MMR protein MLH1 at position serine 477, which significantly inhibits the MMR. In the present study, CK2α-dependent MLH1 phosphorylation was analyzed in vivo. Using a cohort of 165 patients, we identified 88 CRCs showing significantly increased nuclear/cytoplasmic CK2α expression, 28 tumors with high nuclear CK2α expression and 49 cases showing a general low CK2α expression. Patients with high nuclear/cytoplasmic CK2α expression demonstrated significantly reduced 5-year survival outcome. By immunoprecipitation and Western blot analysis, we showed that high nuclear/cytoplasmic CK2α expression significantly correlates with increased MLH1 phosphorylation and enriched somatic tumor mutation rates. The CK2α mRNA levels tended to be enhanced in high nuclear/cytoplasmic and high nuclear CK2α-expressing tumors. Furthermore, we identified various SNPs in the promotor region of CK2α, which might cause differential CK2α expression. In summary, we demonstrated that high nuclear/cytoplasmic CK2α expression in CRCs correlates with enhanced MLH1 phosphorylation in vivo and seems to be causative for increased mutation rates, presumably induced by reduced MMR. These observations could provide important new therapeutic targets.