Assessment of lumbar paraspinal muscle morphology using mDixon Quant magnetic resonance imaging (MRI): a cross-sectional study in healthy subjects.

Background: Lumbar paraspinal muscles (LPM) are a part of the deep spinal stabilisation system and play an important role in stabilising the lumbar spine and trunk. Inadequate function of these muscles is thought to be an essential aetiological factor in low back pain, and several neuromuscular diseases are characterised by dysfunction of LPM. The main aims of our study were to develop a methodology for LPM assessment using advanced magnetic resonance imaging (MRI) methods, including a manual segmentation process, to confirm the measurement reliability, to evaluate the LPM morphological parameters [fat fraction (FF), total muscle volume (TMV) and functional muscle volume (FMV)] in a healthy population, to study the influence of physiological factors on muscle morphology, and to build equations to predict LPM morphological parameters in a healthy population. Methods: This prospective cross-sectional observational comparative single-centre study was conducted at the University Hospital in Brno, enrolling healthy volunteers from April 2021 to March 2023. MRI of the lumbar spine and LPM (erector spinae muscle and multifidus muscle) were performed using a 6-point Dixon gradient echo sequence. The segmentation of the LPM and the control muscle (psoas muscle) was done manually to obtain FF and TMV in a range from Th12/L1 to L5/S1. Intra-rater and inter-rater reliability were evaluated. Linear regression models were constructed to assess the effect of physiological factors on muscle FF, TMV and FMV. Results: We enrolled 90 healthy volunteers (median age 38 years, 45 men). The creation of segmentation masks and the assessment of FF and TMV proved reliable (Dice coefficient 84% to 99%, intraclass correlation coefficient ≥0.97). The univariable models showed that FF of LPM was influenced the most by age (39.6% to 44.8% of variability, P<0.001); TMV and FMV by subject weight (34.9% to 67.6% of variability, P<0.001) and sex (24.7% to 64.1% of variability, P<0.001). Multivariable linear regression models for FF of LPM included age, body mass index and sex, with R-squared values ranging from 45.4% to 51.1%. Models for volumes of LPM included weight, age and sex, with R-squared values ranged from 37.4% to 76.8%. Equations were developed to calculate predicted FF, TMV and FMV for each muscle. Conclusions: A reliable methodology has been developed to assess the morphological parameters (biomarkers) of the LPM. The morphological parameters of the LPM are significantly influenced by physiological factors. Equations were constructed to calculate the predicted FF, TMV and FMV of individual muscles in relation to anthropometric parameters, age, and sex. This study, which presented LPM assessment methodology and predicted values of LPM morphological parameters in a healthy population, could improve our understanding of diseases involving LPM (low back pain and some neuromuscular diseases).

Adherence to the ISHLT Protocol for the Referral of Patients with Idiopathic Pulmonary Fibrosis to the Transplantation Center among of Czech Centers for Interstitial Lung Diseases.

There are limited data on referral rates and the number of patients with idiopathic pulmonary fibrosis (IPF) who are eligible for lung transplantation. The aim of the present study was to assess adherence to the consensus of the International Society for Heart and Lung Transplantation (ISHLT) for the referral of patients with IPF among Czech interstitial lung disease (ILD) centers. Czech patients who were diagnosed with IPF between 1999 and 2021 (n = 1584) and who were less than 65 years old at the time of diagnosis were retrospectively selected from the Czech Republic of the European Multipartner Idiopathic Pulmonary Fibrosis Registry (EMPIRE). Nonsmokers and ex-smokers with a body mass index (BMI) of <32 kg/m2 (n = 404) were included for further analyses. Patients with a history of cancer <5 years from the time of IPF diagnosis, patients with alcohol abuse, and patients with an accumulation of vascular comorbidities were excluded. The trajectory of individual patients was verified at the relevant ILD center. From the database of transplant patients (1999-12/2021, n = 541), all patients who underwent transplantation for pulmonary fibrosis (n = 186) were selected, and the diagnosis of IPF was subsequently verified from the patient's medical records (n = 67). A total of 304 IPF patients were eligible for lung transplantation. Ninety-six patients were referred to the transplant center, 50% (n = 49) of whom were referred for lung transplantation. Thirty percent of potentially eligible patients not referred to the transplant center were considered to have too many comorbidities by the reporting physician, 19% of IPF patients denied lung transplantation, and 17% were not referred due to age. Among Czech patients with IPF, there may be a larger pool of potential lung transplant candidates than has been reported to the transplant center to date.

Single-cell profiling of surface glycosphingolipids opens a new dimension for deconvolution of breast cancer intratumoral heterogeneity and phenotypic plasticity.

Glycosylated sphingolipids (GSLs) are a diverse group of cellular lipids, typically reported as rare in normal mammary tissue. In breast cancer (BCa), GSLs have emerged as noteworthy markers associated with breast cancer stem cells, mediators of phenotypic plasticity, and contributors to cancer cell chemoresistance. GSLs are potential surface markers that can uniquely characterize the heterogeneity of the tumor microenvironment, including cancer cell subpopulations and epithelial-mesenchymal plasticity (EMP). In this study, mass spectrometry analyses of the total sphingolipidome in breast epithelial cells and their mesenchymal counterparts revealed increased levels of Gb3 in epithelial cells and significantly elevated GD2 levels in the mesenchymal phenotype. To elucidate whether GSL-related epitopes on BCa cell surfaces reflect EMP and cancer status, we developed and rigorously validated a 12-color spectral flow cytometry panel. This panel enables the simultaneous detection of native GSL epitopes (Gb3, SSEA1, SSEA3, SSEA4, GD2), epithelial-mesenchymal transition (EMT) markers (EpCAM, TROP2, CD9), and lineage markers (CD45, CD31, CD90) at the single-cell level. As a next step, the established panel was used for the analysis of BCa primary tumors and revealed surface heterogeneity in SSEA1, SSEA3, SSEA4, GD2, and Gb3, indicative of native epitope presence also on non-tumor cells. These findings further highlighted the phenotype-dependent alterations in GSL surface profiles, with differences between epithelial and stromal cells in the tumor. This study provides novel insights into BCa heterogeneity, shedding light on the potential of native GSL-related epitopes as markers for EMP and cancer status in fresh clinical samples. The developed single-cell approach offers promising avenues for further exploration.

Orthotopic model for the analysis of melanoma circulating tumor cells.

Metastatic melanoma, a highly lethal form of skin cancer, presents significant clinical challenges due to limited therapeutic options and high metastatic capacity. Recent studies have demonstrated that cancer dissemination can occur earlier, before the diagnosis of the primary tumor. The progress in understanding the kinetics of cancer dissemination is limited by the lack of animal models that accurately mimic disease progression. We have established a xenograft model of human melanoma that spontaneously metastasizes to lymph nodes and lungs. This model allows precise monitoring of melanoma progression and is suitable for the quantitative and qualitative analysis of circulating tumor cells (CTCs). We have validated a flow cytometry-based protocol for CTCs enumeration and isolation. We could demonstrate that (i) CTCs were detectable in the bloodstream from the fourth week after tumor initiation, coinciding with the lymph node metastases appearance, (ii) excision of the primary tumor accelerated the formation of metastases in lymph nodes and lungs as early as one-week post-surgery, accompanied by the increased numbers of CTCs, and (iii) CTCs change their surface protein signature. In summary, we present a model of human melanoma that can be effectively utilized for future drug efficacy studies.

Inhibition of Chk1 stimulates cytotoxic action of platinum-based drugs and TRAIL combination in human prostate cancer cells.

Checkpoint kinase 1 (Chk1) plays an important role in regulation of the cell cycle, DNA damage response and cell death, and represents an attractive target in anticancer therapy. Small-molecule inhibitors of Chk1 have been intensively investigated either as single agents or in combination with various chemotherapeutic drugs and they can enhance the chemosensitivity of numerous tumor types. Here we newly demonstrate that pharmacological inhibition of Chk1 using potent and selective inhibitor SCH900776, currently profiled in phase II clinical trials, significantly enhances cytotoxic effects of the combination of platinum-based drugs (cisplatin or LA-12) and TRAIL (tumor necrosis factor-related apoptosis inducing ligand) in human prostate cancer cells. The specific role of Chk1 in the drug combination-induced cytotoxicity was confirmed by siRNA-mediated silencing of this kinase. Using RNAi-based methods we also showed the importance of Bak-dependent mitochondrial apoptotic pathway in the combined anticancer action of SCH900776, cisplatin and TRAIL. The triple drug combination-induced cytotoxicity was partially enhanced by siRNA-mediated Mcl-1 silencing. Our findings suggest that targeting Chk1 may be used as an efficient strategy for sensitization of prostate cancer cells to killing action of platinum-based chemotherapeutic drugs and TRAIL.

The Impact of Switching to a Second Antifibrotic in Patients With Idiopathic Pulmonary Fibrosis: A Retrospective Multicentre Study From the EMPIRE Registry.

INTRODUCTION: Most patients with idiopathic pulmonary fibrosis (IPF) treated with antifibrotics (AF) have progressive disease despite treatment. A switch of AF may improve survival, but evidence from randomised controlled trials is missing. We aimed to evaluate the efficacy of an AF switch on survival and FVC decline in patients from the European MultiPartner IPF registry (EMPIRE). METHODS: The study included 612 patients who discontinued the first antifibrotic therapy. Patients were grouped and analysed from two perspectives: (1) whether they had received a second antifibrotic treatment after the discontinuation of the first therapy, and (2) a reason for discontinuation of the first AF - "lack of efficacy" (LE) and "intolerance" (INT). RESULTS: While 263 (43%) of 612 patients received no second AF ("non-switched"), 349 (57%) patients switched. Overall survival was higher in patients who received a second AF (median 50 vs. 29 months; adjusted HR 0.64, P=0.023). Similarly, the annual FVC decline was significantly reduced in switched patients: -98ml/y in switched and -172ml/y in non-switched patients (P=0.023), respectively. The switched patients had similar risk for mortality in both LE and INT groups (adjusted HR 0.95, P=0.85). The high impact of switching on survival was demonstrated in LE patients (adjusted HR 0.27, P<0.001). CONCLUSION: The patients without a second AF had significantly shorter overall survival. Our analysis suggests the importance of switching patients with an ineffective first AF therapy to a second AF therapy.