While the cellular mechanisms of liver regeneration have been thoroughly studied, the role of extracellular matrix (ECM) in liver regeneration is still poorly understood. We utilized a ...proteomics-based approach to identify the shifts in ECM composition after CCl4 or DDC treatment and studied their effect on the proliferation of liver cells by combining biophysical and cell culture methods. We identified notable alterations in the ECM structural components (eg collagens I, IV, V, fibronectin, elastin) as well as in non-structural proteins (eg olfactomedin-4, thrombospondin-4, armadillo repeat-containing x-linked protein 2 (Armcx2)). Comparable alterations in ECM composition were seen in damaged human livers. The increase in collagen content and decrease in elastic fibers resulted in rearrangement and increased stiffness of damaged liver ECM. Interestingly, the alterations in ECM components were nonhomogenous and differed between periportal and pericentral areas and thus our experiments demonstrated the differential ability of selected ECM components to regulate the proliferation of hepatocytes and biliary cells. We define for the first time the alterations in the ECM composition of livers recovering from damage and present functional evidence for a coordinated ECM remodelling that ensures an efficient restoration of liver tissue.
Infertility affects around 7% of men worldwide. Idiopathic non-obstructive azoospermia (NOA) is defined as the absence of spermatozoa in the ejaculate due to failed spermatogenesis. There is a high ...probability that NOA is caused by rare genetic defects. In this study, whole-exome sequencing (WES) was applied to two Estonian brothers diagnosed with NOA and Sertoli cell-only syndrome (SCOS). Compound heterozygous loss-of-function (LoF) variants in FANCM (Fanconi anemia complementation group M) were detected as the most likely cause for their condition. A rare maternally inherited frameshift variant p.Gln498Thrfs∗7 (rs761250416) and a previously undescribed splicing variant (c.4387−10A>G) derived from the father introduce a premature STOP codon leading to a truncated protein. FANCM exhibits enhanced testicular expression. In control subjects, immunohistochemical staining localized FANCM to the Sertoli and spermatogenic cells of seminiferous tubules with increasing intensity through germ cell development. This is consistent with its role in maintaining genomic stability in meiosis and mitosis. In the individual with SCOS carrying bi-allelic FANCM LoF variants, none or only faint expression was detected in the Sertoli cells. As further evidence, we detected two additional NOA-affected case subjects with independent FANCM homozygous nonsense variants, one from Estonia (p.Gln1701∗; rs147021911) and another from Portugal (p.Arg1931∗; rs144567652). The study convincingly demonstrates that bi-allelic recessive LoF variants in FANCM cause azoospermia. FANCM pathogenic variants have also been linked with doubled risk of familial breast and ovarian cancer, providing an example mechanism for the association between infertility and cancer risk, supported by published data on Fancm mutant mouse models.
Prostate adenocarcinoma (PCa) stromal markers have recently gained attention as complementary diagnostic tools. The DNA reparation complex protein FANCM has been shown to express in the normal ...prostate stroma and FANCM gene alterations to be associated with PCa susceptibility; this has led to the hypothesis that an insufficient level of FANCM expression may provide additional information for the evaluation of PCa. The study cohort comprised 60 radical prostatectomy specimens. The controls involved 11 autopsies (CTRL) and non‐cancerous tissue (NCT) areas from the prostatectomy specimen. The samples were stained with the FANCM antibody. The quantification of the stromal staining index (SSI) was made using ImageJ and QuPath. Overall, 655 regions of interest (ROI) were analyzed. FANCM expression appeared equally intense and stroma specific in both CTRL and NCT, indicating the absence of underlying baseline alterations. Within the age span of the cohort 47–89 years, no significant effect of the age of the patients on the FANCM expression was seen. FANCM demonstrated Gleason grade (G) dependent decline in PCa, being statistically significant in controls versus G1 and G2 versus G3. In other adjacent International Society of Urological Pathology (ISUP) groups, it remained insignificant, still being meaningful between high and low‐grade cancers.
This paper presents a technique that can potentially help to determine the receptivity stage of the endometrium from histology images by automatically measuring the stromal nuclear changes. The ...presented technique is composed of an image segmentation model and the statistical evolution of segmented areas in hematoxylin and eosin (HE)-stained histology images. Three different endometrium receptivity stages, namely pre-receptive, post-receptive, and receptive, were compared. An ensemble-based AI model was proposed for histology image segmentation, which is based on individual UNet++, UNet, and ResNet34-UNet segmentation models. The performance of the ensemble-based technique was assessed using the Dice score and intersection over unit (IoU) values. In comparison to alternative segmentation architectures that were applied singly, the current ensemble-based method obtained higher Dice score (0.95) and IoU (0.90) values. The statistical comparison highlighted a noticeable difference in the number of nuclei and the size of the stroma tissue. The proposed technique demonstrated the positive potential for practical implementation for automatic endometrial tissue analysis.
Despite the importance of placenta in mediating rapid physiological changes in pregnancy, data on temporal dynamics of placental gene expression are limited. We completed the first transcriptome ...profiling of human placental gene expression dynamics (GeneChips, Affymetrix®; ~47,000 transcripts) from early to mid-gestation (n = 10; gestational weeks 5-18) and report 154 genes with significant transcriptional changes (ANOVA, FDR P<0.1). TaqMan RT-qPCR analysis (n = 43; gestational weeks 5-41) confirmed a significant (ANOVA and t-test, FDR P<0.05) mid-gestational peak of placental gene expression for BMP5, CCNG2, CDH11, FST, GATM, GPR183, ITGBL1, PLAGL1, SLC16A10 and STC1, followed by sharp decrease in mRNA levels at term (t-test, FDR P<0.05). We hypothesized that normal course of late pregnancy may be affected when genes characteristic to mid-gestation placenta remain highly expressed until term, and analyzed their expression in term placentas from normal and complicated pregnancies preeclampsia (PE), n = 12; gestational diabetes mellitus (GDM), n = 12; small- and large-for-gestational-age newborns (SGA, LGA), n = 12+12. STC1 (stanniocalcin 1) exhibited increased mRNA levels in all studied complications, with the most significant effect in PE- and SGA-groups (t-test, FDR P<0.05). In post-partum maternal plasma, the highest STC1 hormone levels (ELISA, n = 129) were found in women who had developed PE and delivered a SGA newborn (median 731 vs 418 pg/ml in controls; ANCOVA, P = 0.00048). Significantly higher expression (t-test, FDR P<0.05) of CCNG2 and LYPD6 accompanied with enhanced immunostaining of the protein was detected in placental sections of PE and GDM cases (n = 15). Our study demonstrates the importance of temporal dynamics of placental transcriptional regulation across three trimesters of gestation. Interestingly, many genes with high expression in mid-gestation placenta have also been implicated in adult complex disease, promoting the discussion on the role of placenta in developmental programming. The discovery of elevated maternal plasma STC1 in pregnancy complications warrants further investigations of its potential as a biomarker.
Celotno besedilo
Dostopno za:
DOBA, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
Parkinson's disease (PD) is the second-most common neurodegenerative disorder worldwide and is diagnosed based on motor impairments. Non-motor symptoms are also well-recognised in this disorder, and ...peripheral neuropathy is a frequent but poorly appreciated non-motor sign. Studying how central and peripheral sensory systems are affected can contribute to the development of targeted therapies and deepen our understanding of the pathophysiology of PD. Although the cause of sporadic PD is unknown, chronic exposure to the pesticide rotenone in humans increases the risk of developing the disease. Here, we aimed to investigate whether peripheral neuropathy is present in a traditional model of PD. Mice receiving intrastriatal rotenone showed greatly reduced dopamine terminals in the striatum and a reduction in tyrosine hydroxylase-positive neurons in the
and developed progressive motor impairments in hindlimb stepping and rotarod but no change in spontaneous activity. Interestingly, repeated testing using gold-standard protocols showed no change in gut motility, a well-known non-motor symptom of PD. Importantly, we did not observe any change in heat, cold, or touch sensitivity, again based upon repeated testing with well-validated protocols that were statistically well powered. Therefore, this traditional model fails to replicate PD, and our data again reiterate the importance of the periphery to the disorder.
The elastic section of the giant muscle protein titin contains many immunoglobulin-like domains, which have been shown by single-molecule mechanical studies to unfold and refold upon stretch-release. ...Here we asked whether the mechanical properties of Ig domains and/or other titin regions could be responsible for the viscoelasticity of nonactivated skeletal-muscle sarcomeres, particularly for stress relaxation and force hysteresis. We show that isolated psoas myofibrils respond to a stretch-hold protocol with a characteristic force decay that becomes more pronounced following stretch to above 2.6-
μm sarcomere length. The force decay was readily reproducible by a Monte Carlo simulation taking into account both the kinetics of Ig-domain unfolding and the worm-like-chain model of entropic elasticity used to describe titin’s elastic behavior. The modeling indicated that the force decay is explainable by the unfolding of only a very small number of Ig domains per titin molecule. The simulation also predicted that a unique sequence in titin, the PEVK domain, may undergo minor structural changes during sarcomere extension. Myofibrils subjected to 1-Hz cycles of stretch-release exhibited distinct hysteresis that persisted during repetitive measurements. Quick stretch-release protocols, in which variable pauses were introduced after the release, revealed a two-exponential time course of hysteresis recovery. The rate constants of recovery compared well with the refolding rates of Ig-like or fibronectin-like domains measured by single-protein mechanical analysis. These findings suggest that in the sarcomere, titin’s Ig-domain regions may act as entropic springs capable of adjusting their contour length in response to a stretch.
Glioblastoma multiforme (GBM) is one of the most angiogenic tumors. However, antiangiogenic therapy has not shown significant clinical efficacy. The aim of our study was to evaluate the impact of ...inflammatory tumor microenvironment on the expression of vascular endothelial growth factor receptor 2 (VEGFR-2). Surgically excised primary GBM tissues were histologically examined for overall extent of inflammation (score 1–3). After immunohistochemistry, the tissue expression of ICAM-1 (optical density), the number of VEGFR-2 positive (VEGFR-2+) blood vessels (per microscopic field), and the endothelial staining intensity of VEGFR-2 (score 0–3) were determined. In GBM, the extent of inflammation was 1.9 ± 0.7 (group mean ± SD). Mean optical density of inflammatory mediator ICAM-1 was 57.0 ± 27.1 (pixel values). The number of VEGFR-2+ blood vessels and endothelial VEGFR-2 staining intensity were 6.2 ± 2.4 and 1.2 ± 0.8, respectively. A positive association was found between endothelial VEGFR-2 staining intensity and the extent of inflammation (p=0.005). Moreover, VEGFR-2 staining intensity correlated with the expression level of ICAM-1 (p=0.026). The expression of VEGFR-2, one of the main targets of antiangiogenic therapy, depends on GBM microenvironment. Higher endothelial VEGFR-2 levels were seen in the presence of more pronounced inflammation. Target dependence on inflammatory tumor microenvironment has to be taken into consideration when treatment approaches that block VEGFR-2 signaling are designed.
Purpose
Lumbar disc degeneration may be associated with intensity of neovascularization in disc herniations. Our study was designed to evaluate how much the severity of histodegeneration is related ...to the development of neovascularization and to the level of pleiotrophin in the herniated lumbar discs.
Methods
Surgically excised lumbar disc specimens were obtained from 29 patients with noncontained (i.e., extruding through the posterior longitudinal ligament) and 21 patients with contained disc herniations. The histodegeneration scores and levels of neovascularization were estimated according to semiquantitative analysis in lumbar disc and endplate samples. Immunohistochemical staining were performed to identify the newly formed blood vessels and to detect the presence of pleiotrophin in the specimens.
Results
Higher levels of disc and endplate neovascularity were registered in noncontained herniations. The level of neovascularization was significantly related to the score of histodegeneration in the herniated disc tissues but not in the endplate specimens. Both contained and noncontained herniations had the highest values of histodegeneration in conjunction with the highest level of neovascularization but the relations between neovascularity and degenerative changes remained to be significant only in the group of noncontained herniations. Registration or frequency of pleiotrophin positive cells did not correlate significantly with histodegeneration or level of neovascularization in the disc samples.
Conclusion
Severe histodegeneration of the lumbar disc herniations is associated with enhanced neovascularization and potentially also spontaneous regression of the herniated tissue.
Pathadin (https://gitlab.com/Digipathology/Pathadin) was designed as a WSI oriented open-source set of tools for beginners to experience the advantages of computer-assisted image analysis and cover ...essential features, frequently strenuous to access with the alternative programs. It is mainly oriented to work with histology slides but also includes a significant part of modern image formats. Introducing Pathadin, the manuscript aims to improve understanding of contemporary image analysis components, resolve technophobia and misbeliefs in the computational field, simplifying pathology research work, and shifting it into a quantitative paradigm.
Despite being easy to use, Pathadin includes both basic and advanced analytical algorithms, as the application of machine learning.
The functionality of Pathadin is demonstrated by AI-enhanced quantification of epithelial and stromal changes in prostate carcinoma, and their dependence on ISUP grade. The material included 5 radical prostatectomy samples for training and 83 (including 11 autopsies) samples for analysis. The analytical material was stained with Masson’s trichrome and Ki67, as widely available and potentially prognostic markers. An integrated solution by HistomicsTK for Ki67 evaluation was used. A U-net model for separating glands and stroma was trained, simplifying the independent analysis of these components.
During the process, the model successfully highlighted glands and stroma. Masson’s trichrome stain demonstrated a gradual increase in collagen expression, being statistically significant between controls vs. G1, and G3 vs. G4. Although there was considerable overlap between adjacent groups, there was only a minor overlap in collagen amount between high- and low-grade carcinomas, affirming that with further research, stroma could be an additional diagnostic marker in prostate adenocarcinoma. Ki67 showed a statistically significant gradual increase in all groups except G1 vs. G2 and G4 vs. G5.
Pathadin demonstrates that there is no need for significant resources to experience the advantages of modern computer-assisted analysis.
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