In this randomized, placebo-controlled cross-over trial we aimed to investigate if radon spa therapy exerts more pain relief than exposure to warm water alone. In addition, immunological parameters ...were assessed in both treatment groups. In the RAD-ON02 trial, 116 patients suffering from musculoskeletal disorders (MSDs) received either serial radon spa or solely warm water baths. Pain intensity was assessed by determination of different pain parameters on a visual analogue scale and by pressure point dolorimetry at baseline and at weeks 4, 12 and 24. The longitudinal immune status of the patients was analyzed by a flow cytometry-based assay from peripheral blood at the time points of pain assessments. There were no side effects attributable to radon exposure observed. However, radon spa was superior to warm water applications at week 4 in terms of pain reduction. Pain and morning stiffness at the time of assessment were significantly reduced after radon spa (p<0.001, p<0.01) but not after warm water baths. The dolorimetry resulted in a significantly higher exerted pressure strength in patients after radon spa (p<0.001), but not after warm water applications. During the long-term follow-up, both treatment modalities reduced pain to a similar degree and pain modulation was not distorted by the participants' intake of analgesics. No significant changes in the immune status attributable specifically to radon were found, even though the increase in regulatory T cell counts occurs earlier after radon baths than after sole warm water baths and a higher level of significance is reached after radon spa at week 24. Serial radon spa has additive pain-relieving effects. The immunological parameters assessed in our study appear not to be directly linked to the pain reduction caused by radon exposure, at least in MSD patients with predominantly degenerative diseases.
https://www.clinicaltrialsregister.eu/ctr-search/search?query=rad-on02, identifier 2016-002085-31; https://drks.de/search/de/trial, identifier DRKS00016019.
The human oral microbiota consists of over 700 widespread taxa colonizing the oral cavity in several anatomically diverse oral niches. Lately, sequencing of the 16S rRNA genes has become an ...acknowledged, culture-independent method to characterize the oral microbiota. However, only a small amount of data are available concerning microbial differences between oral niches in periodontal health and disease. In the context of periodontitis, the cytokine expression in the gingival crevicular fluid has been studied in detail, whereas little is known about the cytokine profile in hard and soft tissue biofilms. In order to characterize oral niches in periodontal health, the oral microbiota and cytokine pattern were analyzed at seven different sites (plaque (P), gingival crevicular fluid (GCF), saliva (S), tongue (T), hard palate (HP), cheek (C) and sublingual area (U)) of 20 young adults using next-generation sequencing and multiplex immunoassays. Site-specific microbial compositions were detected, which clustered into three distinct metaniches ("P-GCF", "S-T-HP" and "C-U") and were associated with niche-/metaniche-specific cytokine profiles. Our findings allow the definition of distinct metaniches according to their microbial composition, partly reflected by their cytokine profile, and provide new insights into microenvironmental similarities between anatomical diverse oral niches.
BackgroundThe predictive power of novel biological markers for treatment response to immune checkpoint inhibitors (ICI) is still not satisfactory for the majority of patients with cancer. One should ...identify valid predictive markers in the peripheral blood, as this is easily available before and during treatment. The current interim analysis of patients of the ST-ICI cohort therefore focuses on the development and validation of a liquid immune profile-based signature (LIPS) to predict response of patients with metastatic cancer to ICI targeting the programmed cell death protein 1 (PD-1)/programmed cell death-ligand 1 (PD-L1) axis.MethodsA total of 104 patients were prospectively enrolled. 54 immune cell subsets were prospectively analyzed in patients’ peripheral blood by multicolor flow cytometry before treatment with ICI (pre-ICI; n=89), and after the first application of ICI (n=65). Pre-ICI, patients were randomly allocated to a training (n=56) and a validation cohort (n=33). Univariate Cox proportional hazards regression analysis and least absolute shrinkage and selection operator Cox model were used to create a predictive immune signature, which was also checked after the first ICI, to consider the dynamics of changes in the immune status.ResultsWhole blood samples were provided by 89 patients pre-ICI and by 65 patients after the first ICI. We identified a LIPS which is based on five immune cell subtypes: CD14high monocytes, CD8+/PD-1+ T cells, plasmacytoid dendritic cells, neutrophils, and CD3+/CD56+/CD16+ natural killer (NK)T cells. The signature achieved a high accuracy (C-index 0.74 vs 0.71) for predicting overall survival (OS) benefit in both the training and the validation cohort. In both cohorts, the low-risk group had significantly longer OS than the high-risk group (HR 0.26, 95% CI 0.12 to 0.56, p=0.00025; HR 0.30, 95% CI 0.10 to 0.91, p=0.024, respectively). Regarding the whole cohort, LIPS also predicted progression-free survival (PFS). The identified LIPS was not affected by clinicopathological features with the exception of brain metastases. NKT cells and neutrophils of the LIPS can be used as dynamic predictive biomarkers for OS and PFS after first administration of the ICI.ConclusionOur study identified a predictive LIPS for survival of patients with cancer treated with PD-1/PD-L1 ICI, which is based on immune cell subsets in the peripheral whole blood.Trial registration numberNCT03453892.
Deep learning-based head and neck lymph node level (HN_LNL) autodelineation is of high relevance to radiotherapy research and clinical treatment planning but still underinvestigated in academic ...literature. In particular, there is no publicly available open-source solution for large-scale autosegmentation of HN_LNL in the research setting.
An expert-delineated cohort of 35 planning CTs was used for training of an nnU-net 3D-fullres/2D-ensemble model for autosegmentation of 20 different HN_LNL. A second cohort acquired at the same institution later in time served as the test set (n = 20). In a completely blinded evaluation, 3 clinical experts rated the quality of deep learning autosegmentations in a head-to-head comparison with expert-created contours. For a subgroup of 10 cases, intraobserver variability was compared to the average deep learning autosegmentation accuracy on the original and recontoured set of expert segmentations. A postprocessing step to adjust craniocaudal boundaries of level autosegmentations to the CT slice plane was introduced and the effect of autocontour consistency with CT slice plane orientation on geometric accuracy and expert rating was investigated.
Blinded expert ratings for deep learning segmentations and expert-created contours were not significantly different. Deep learning segmentations with slice plane adjustment were rated numerically higher (mean, 81.0 vs. 79.6, p = 0.185) and deep learning segmentations without slice plane adjustment were rated numerically lower (77.2 vs. 79.6, p = 0.167) than manually drawn contours. In a head-to-head comparison, deep learning segmentations with CT slice plane adjustment were rated significantly better than deep learning contours without slice plane adjustment (81.0 vs. 77.2, p = 0.004). Geometric accuracy of deep learning segmentations was not different from intraobserver variability (mean Dice per level, 0.76 vs. 0.77, p = 0.307). Clinical significance of contour consistency with CT slice plane orientation was not represented by geometric accuracy metrics (volumetric Dice, 0.78 vs. 0.78, p = 0.703).
We show that a nnU-net 3D-fullres/2D-ensemble model can be used for highly accurate autodelineation of HN_LNL using only a limited training dataset that is ideally suited for large-scale standardized autodelineation of HN_LNL in the research setting. Geometric accuracy metrics are only an imperfect surrogate for blinded expert rating.
EF5 Is the High-Affinity Mg2+ Site in ALG‑2 Tanner, John J; Frey, Benjamin B; Pemberton, Travis ...
Biochemistry (Easton),
09/2016, Letnik:
55, Številka:
36
Journal Article
Recenzirano
The penta-EF-hand (PEF) protein ALG-2 (apoptosis-linked gene 2) has been implicated in several important physiological processes, including endoplasmic reticulum–Golgi vesicular transport and ...endosomal biogenesis/transport. ALG-2 was recently shown to harbor a metal ion-binding site with a high affinity for Mg2+ and a low affinity for Ca2+. We herein present the X-ray structure of Mg2+-bound ALG-2des23wt. Although the Cα trace is nearly indistinguishable from that of the Ca2+-free protein, the orientation of the C-terminal helix differs in the two structures. Consistent with that observation, replacement of the +x ligand in EF5, D169, with alanine eliminates high-affinity Mg2+ binding. It also eliminates the low-affinity Ca2+ site and lowers the affinity of the remaining Ca2+-binding sites, EF3 and EF1. The coordination environment in EF5 approaches ideal Mg2+ octahedral geometry. The ligand array, consisting of three carboxylates (+x, +y, +z), a backbone carbonyl (−y), and two water molecules (−x, –z), may offer a recipe for a high-affinity, high-selectivity Mg2+-binding site. Sequence data for other PEF proteins indicate that select calpain large subunits, notably CAPN1 and CAPN8, may also possess a high-affinity Mg2+-binding site. In Mg2+-bound ALG-2, the carbonyl of F188 and the C-terminal carboxylate of V191 interact with the ε-ammonium group of K137 in the opposing subunit, suggesting that Mg2+ binding could have an impact on dimerization. Interestingly, EF1 and EF3 are also occupied in the crystal, despite having modest affinity for Mg2+. The results of a calorimetry-based analysis indicate that their Mg2+ binding constants are 2 orders of magnitude lower than that determined for EF5.
•Peripheral blood immune parameters predict efficacy of chemoimmunotherapy in HNSCC.•Various differences of immune parameters before and after treatment allowed the best prediction.•Cells of the ...innate immune system are prominent predictors.•Polymorphonuclear cells, monocytes, and plasmacytoid dendritic cells serve as best predictors.
Individual prediction of treatment response is crucial for personalized treatment in multimodal approaches against head-and-neck squamous cell carcinoma (HNSCC). So far, no reliable predictive parameters for treatment schemes containing immunotherapy have been identified. This study aims to predict treatment response to induction chemo-immunotherapy based on the peripheral blood immune status in patients with locally advanced HNSCC.
The peripheral blood immune phenotype was assessed in whole blood samples in patients treated in the phase II CheckRad-CD8 trial as part of the pre-planned translational research program. Blood samples were analyzed by multicolor flow cytometry before (T1) and after (T2) induction chemo-immunotherapy with cisplatin/docetaxel/durvalumab/tremelimumab. Machine Learning techniques were used to predict pathological complete response (pCR) after induction therapy.
The tested classifier methods (LDA, SVM, LR, RF, DT, and XGBoost) allowed a distinct prediction of pCR. Highest accuracy was achieved with a low number of features represented as principal components. Immune parameters obtained from the absolute difference (lT2-T1l) allowed the best prediction of pCR. In general, less than 30 parameters and at most 10 principal components were needed for highly accurate predictions. Across several datasets, cells of the innate immune system such as polymorphonuclear cells, monocytes, and plasmacytoid dendritic cells are most prominent.
Our analyses imply that alterations of the innate immune cell distribution in the peripheral blood following induction chemo-immuno-therapy is highly predictive for pCR in HNSCC.
Hyperthermia (HT) is an accepted treatment for recurrent breast cancer which locally heats the tumor to 39-44 °C, and it is a very potent sensitizer for radiotherapy (RT) and chemotherapy. However, ...currently little is known about how HT with a distinct temperature, and particularly, how the sequence of HT and RT changes the immune phenotype of breast cancer cells. Therefore, human MDA-MB-231 and MCF-7 breast cancer cells were treated with HT of different temperatures (39, 41 and 44 °C), alone and in combination with RT (2 × 5 Gy) in different sequences, with either RT or HT first, followed by the other. Tumor cell death forms and the expression of immune checkpoint molecules (ICMs) were analyzed by multicolor flow cytometry. Human monocyte-derived dendritic cells (moDCs) were differentiated and co-cultured with the treated cancer cells. In both cell lines, RT was the main stressor for cell death induction, with apoptosis being the prominent cell death form in MCF-7 cells and both apoptosis and necrosis in MDA-MB-231 cells. Here, the sequence of the combined treatments, either RT or HT, did not have a significant impact on the final outcome. The expression of all of the three examined immune suppressive ICMs, namely PD-L1, PD-L2 and HVEM, was significantly increased on MCF-7 cells 120 h after the treatment of RT with HT of any temperature. Of special interest for MDA-MB-231 cells is that only combinations of RT with HT of both 41 and 44 °C induced a significantly increased expression of PD-L2 at all examined time points (24, 48, 72, and 120 h). Generally, high dynamics of ICM expression can be observed after combined RT and HT treatments. There was no significant difference between the different sequences of treatments (either HT + RT or RT + HT) in case of the upregulation of ICMs. Furthermore, the co-culture of moDCs with tumor cells of any treatment had no impact on the expression of activation markers. We conclude that the sequence of HT and RT does not strongly affect the immune phenotype of breast cancer cells. However, when HT is combined with RT, it results in an increased expression of distinct immune suppressive ICMs that should be considered by including immune checkpoint inhibitors in multimodal tumor treatments with RT and HT. Further, combined RT and HT affects the immune system in the effector phase rather than in the priming phase.
Radiotherapy (RT) is commonly applied for the treatment of glioblastoma multiforme (GBM). Following the planning target volume (PTV) definition procedure standardized in guidelines, a 20% risk of ...missing non-local recurrences is present. Purpose of this study was to evaluate whether diffusion tensor imaging (DTI)-based fiber tracking may be beneficial for PTV definition taking into account the prediction of distant recurrences. 56 GBM patients were examined with magnetic resonance imaging (MRI) including DTI performed before RT after resection of the primary tumor. Follow-up MRIs were acquired in three month intervals. For the seven patients with a distant recurrence, fiber tracking was performed with three algorithms and it was evaluated whether connections existed from the primary tumor region to the distant recurrence. It depended strongly on the used tracking algorithm and the used tracking parameters whether a connection was observed. Most of the connections were weak and thus not usable for PTV definition. Only in one of the seven patients with a recurring tumor, a clear connection was present. It seems unlikely that DTI-based fiber tracking can be beneficial for predicting distant recurrences in the planning of PTVs for glioblastoma multiforme.
Rheumatoid arthritis (RA) is a multifactorial autoimmune disease whose main hallmark is inflammation and destruction of the joints. Two cell types within the synovium that play an important role in ...RA are fibroblast-like synoviocytes (FLS) and macrophages. The latter innate immune cells show a high plasticity in their phenotype and are central in inflammatory processes. Low-dose radiotherapy (LD-RT) with particularly a single dose of 0.5 Gy has been demonstrated to have a positive impact on pain, inflammation, and bone in inflamed joints. We now examined for the first time how LD-RT influences FLS and bone marrow-derived macrophages in co-culture systems of an experimental model of RA to reveal further mechanisms of immune modulatory effects of low and intermediate dose of ionizing radiation. For this, the bone marrow of hTNF-α tg mice was differentiated either with cytokines to obtain key macrophage phenotypes (M0, M1, and M2) or with supernatants (SN) of untreated or irradiated FLS. Flow cytometry analyses were used to analyse the impact of radiation (0.1, 0.5, 1.0, and 2.0 Gy) on the phenotype of macrophages in the presence or absence of SN of FLS. LD-RT had no impact on cytokine-mediated macrophage polarization in M0, M1, or M2 macrophages. However, SN of irradiated FLS particularly reduced CD206 expression on macrophages. Macrophage phenotype was stable when being in contact with SN of nonirradiated FLS, but significantly increased surface expression of CD206 and slightly decreased CD80 and CD86 expression were observed when macrophage themselves were irradiated with 0.5 Gy under these microenvironmental conditions, again highlighting discontinuous dose dependencies in the low and intermediate dose range. One can conclude that FLS-dependent microenvironmental conditions have a slight influence on the modulation of macrophage phenotype under radiation exposure conditions. Future studies are needed to reveal the impact of radiation exposure on the functions of treated macrophages under such microenvironmental conditions.
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