We investigated the relationship between neutrophil apoptosis and endoplasmic reticulum stress (ERS) in sepsis and its mechanism. A prospective cohort study was conducted by recruiting a total of 58 patients with sepsis. Peripheral blood samples were collected on 1, 3, 5 and 7 days after admission to the ICU. The expressions of endoplasmic reticulum specific glucose regulatory protein 78 (GRP78), C/EBP homologous protein (CHOP), apoptosis signal-regulating kinase 1 (ASK1), Bcl-2-like 11 (BIM), death receptor 5 (DR5), c-Jun N-terminal kinases (JNK) and p38 were detected by Western blot and PCR. The subcellular location of CHOP and GRP78 was observed by immunofluorescence analysis. Spearman correlation was used to analyze the correlation between the expression of chop protein and the apoptosis rate of peripheral blood neutrophils. Healthy volunteers in the same period were selected as the healthy control group. The expression of GRP78 protein was significantly elevated on the first day of ICU admission and showed a decreasing trend on the third, fifth and seventh day, but was significantly higher than the corresponding healthy control group. The expression of CHOP protein reached the highest level on the third day. The expression of chop protein in each group was significantly higher than that in the corresponding healthy control group. Immunofluorescence staining clearly showed that the CHOP protein accumulated in the nucleus, with an elevation in the intensity of GRP78. The neutrophil apoptosis rate of sepsis patients on the 1st, 3rd, 5th and 7th day of ICU stay was significantly higher than that of the healthy control group, with the highest apoptosis rate on the 3rd day, and then decreased gradually. CHOP protein expression level was significantly positively correlated with neutrophil apoptosis rate in sepsis patients. Endoplasmic reticulum stress occurs in neutrophils during the development of sepsis. GRP78 protein and CHOP protein may be involved in the pathological process of neutrophil apoptosis in sepsis.

Altered metabolism of lipids is a key factor in many diseases including cancer. Therefore, investigations into the impact of unsaturated and saturated fatty acids (FAs) on human body homeostasis are crucial for understanding the development of lifestyle diseases. In this paper, we focus on the impact of palmitic (PA), linoleic (LA), and eicosapentaenoic (EPA) acids on human colon normal (CCD-18 Co) and cancer (Caco-2) single cells using Raman imaging and spectroscopy. The label-free nature of Raman imaging allowed us to evaluate FAs dynamics without modifying endogenous cellular metabolism. Thanks to the ability of Raman imaging to visualize single-cell substructures, we have analyzed the changes in chemical composition of endoplasmic reticulum (ER), mitochondria, lipid droplets (LDs), and nucleus upon FA supplementation. Analysis of Raman band intensity ratios typical for lipids, proteins, and nucleic acids (I1656/I1444, I1444/I1256, I1444/I750, I1304/I1256) proved that, using Raman mapping, we can observe the metabolic pathways of FAs in ER, which is responsible for the uptake of exogenous FAs, de novo synthesis, elongation, and desaturation of FAs, in mitochondria responsible for energy production via FA oxidation, in LDs specialized in cellular fat storage, and in the nucleus, where FAs are transported via fatty-acid-binding proteins, biomarkers of human colon cancerogenesis. Analysis for membranes showed that the uptake of FAs effectively changed the chemical composition of this organelle, and the strongest effect was noticed for LA. The spectroscopy studies have been completed using XTT tests, which showed that the addition of LA or EPA for Caco-2 cells decreases their viability with a stronger effect observed for LA and the opposite effect observed for PA. For normal cells, CCD-18 Co supplementation using LA or EPA stimulated cells for growing, while PA had the opposite impact.

The endoplasmic reticulum（ER）is the largest membranous network serving as a region for protein, lipid and steroid synthesis, transport and storage. Detailed information about ER-cisternae, ER-tubules and rough endoplasmic reticulum (rER) is scarce in human blood cells. This study describes a series of giant inclusions and Auer bodies in promyeloblasts in six patients with acute promyelocytic leukemia (APL), by light microscopy, transmission electron microscopy (TEM) and cytochemical stains. TEM revealed that giant inclusions and pro-Auer bodies were associated with rER and surrounded by tubular structures composed of degenerated or redundant membrane in promyeloblasts, which corresponded with elements of the ER system. This paper reveals that in the promyeloblasts of APL, ER is the source of and transforms progressively into giant inclusions and Auer bodies.

Photoactivation is a paradigm consisting in local molecular fluorescent activation by laser illumination in a chosen region (source) while measuring the concentration at a target region. Data-driven modeling is concerned with the following questions: how from the measurement in these two regions is it possible to infer the properties of molecular propagation? How is it possible to use such responses to infer motions occurring in networks such as the endoplasmic reticulum? In this book chapter, we shall review the data-driven analysis based on diffusion-transport models and numerical simulations to interpret the photoactivation dynamics and extract biophysical parameters. We will discuss modeling approaches to reconstruct local network properties from photoactivation transients.

In this chapter, approaches to the image analysis of the choreography of the plant endoplasmic reticulum (ER) labeled with fluorescent fusion proteins (\"stars,\" if you wish) are presented. The approaches include the analyses of those parts of the ER that are attached through membrane contact sites to moving or non-moving partners (other \"stars\"). Image analysis is also used to understand the nature of the tubular polygonal network, the hallmark of this organelle, and how the polygons change over time due to tubule sliding or motion. Furthermore, the remodeling polygons of the ER interact with regions of fundamentally different topologies, the ER cisternae, and image analysis can be used to separate the tubules from the cisternae. ER cisternae, like polygons and tubules, can be motile or stationary. To study which parts are attached to non-moving partners, such as domains of the ER that form membrane contact sites with the plasma membrane/cell wall, an image analysis approach called persistency mapping has been used. To study the domains of the ER that move rapidly and stream through the cell, image analysis of optic flow has been used. However, optic flow approaches confuse the movement of the ER itself with the movement of proteins within the ER. As an overall measure of ER dynamics, optic flow approaches are of value, but their limitation as to what exactly is \"flowing\" needs to be specified. Finally, there are important imaging approaches that directly address the movement of fluorescent proteins within the ER lumen or in the membrane of the ER. Of these, fluorescence recovery after photobleaching (FRAP), inverse FRAP (iFRAP), and single particle tracking approaches are described.

S100B, a homodimeric Ca2+-binding protein, is produced and secreted by astrocytes, and its extracellular levels have been used as a glial marker in brain damage and neurodegenerative and psychiatric diseases; however, its mechanism of secretion is elusive. We used primary astrocyte cultures and calcium measurements from real-time fluorescence microscopy to investigate the role of intracellular calcium in S100B secretion. In addition, the dimethyl sulfoxide (DMSO) effect on S100B was investigated in vitro and in vivo using Wistar rats. We found that DMSO, a widely used vehicle in biological assays, is a powerful S100B secretagogue, which caused a biphasic response of Ca2+ mobilization. Our data show that astroglial S100B secretion is triggered by the increase in intracellular Ca2+ and indicate that this increase is due to Ca2+ mobilization from the endoplasmic reticulum. Also, blocking plasma membrane Ca2+ channels involved in the Ca2+ replenishment of internal stores decreased S100B secretion. The DMSO-induced S100B secretion was confirmed in vivo and in ex vivo hippocampal slices. Our data support a nonclassic vesicular export of S100B modulated by Ca2+, and the results might contribute to understanding the mechanism underlying the astroglial release of S100B.

Cytosolic-free calcium ions play an important role in various physical and physiological processes. A vital component of neural signaling is the free calcium ion concentration often known as the second messenger. There are many parameters that effect the cytosolic free calcium concentration like buffer, voltage-gated ion channels, Endoplasmic reticulum, Mitochondria, etc. Mitochondria are small organelles located within the nervous system that are involved in processes within cells such as calcium homeostasis management, energy generation, response to stress, and cell demise pathways. In this work, a mathematical model with fuzzy boundary values has been developed to study the effect of Mitochondria and ER fluxes on free Calcium ions. The intended findings are displayed utilizing the physiological understanding that amyloid beta plaques and tangles of neurofibrillary fibers have been identified as the two main causes of AD. The key conclusion of the work is the investigation of [Formula: see text] for healthy cells and cells affected by Alzheimer\'s disease, which may aid in the study of such processes for computational scientists and medical practitioners. Also, it has been shown that when a unique solution is found for a specific precise problem, it also successfully deals with any underlying ambiguity within the problem by utilizing a technique based on the principles of linear transformation. Furthermore, the comparison between the analytical approach and the generalized hukuhara derivative approach is shown here, which illustrates the benefits of the analytical approach. The simulation is carried out in MATLAB.

It is known that the unfavorable outcome in patients infected with SARS-CoV-2 may be associated with the development of complications caused by heart damage due to the direct virus action. The mechanism of these cardiovascular injuries caused by SARS-CoV-2 infection has not been fully understood; however, the study of COVID-19-associated myocardial microcirculatory dysfunction can represent the useful strategy to solving this challenge. Thus, here we aimed to study the ultrastructural organization of endothelial cells of myocardial capillaries in patients with COVID-19. The morphology of endotheliocytes of the myocardial blood capillaries in patients with COVID-19 was studied on cardiac autopsy material using transmission electron microscopy. The endotheliocytes of myocardial capillaries in patients with COVID-19 were characterized by the abundant rough endoplasmic reticulum (ER) membranes, the Golgi complex, and free polysomal complexes of ribosomes and lipids. The presence of double membrane vesicles with virions and zippered ER was detected in the cytoplasm of endotheliocytes. The revealed endothelial ultrastructural changes indicate the remodeling of intracellular membranes during SARS-CoV-2 infection. Our findings confirm the formation of virus-induced structures in myocardial endothelial cells considered critical for viral replication and assembly. The data may elucidate the mechanisms of endothelial dysfunction development in patients with COVID-19 to provide potential targets for drug therapy.

Wilson disease (WD) is caused by mutations in the ATP7B gene that encodes a copper (Cu) transporting ATPase whose trafficking from the Golgi to endo-lysosomal compartments drives sequestration of excess Cu and its further excretion from hepatocytes into the bile. Loss of ATP7B function leads to toxic Cu overload in the liver and subsequently in the brain, causing fatal hepatic and neurological abnormalities. The limitations of existing WD therapies call for the development of new therapeutic approaches, which require an amenable animal model system for screening and validation of drugs and molecular targets. To achieve this objective, we generated a mutant Caenorhabditis elegans strain with a substitution of a conserved histidine (H828Q) in the ATP7B ortholog cua-1 corresponding to the most common ATP7B variant (H1069Q) that causes WD. cua-1 mutant animals exhibited very poor resistance to Cu compared to the wild-type strain. This manifested in a strong delay in larval development, a shorter lifespan, impaired motility, oxidative stress pathway activation, and mitochondrial damage. In addition, morphological analysis revealed several neuronal abnormalities in cua-1 mutant animals exposed to Cu. Further investigation suggested that mutant CUA-1 is retained and degraded in the endoplasmic reticulum, similarly to human ATP7B-H1069Q. As a consequence, the mutant protein does not allow animals to counteract Cu toxicity. Notably, pharmacological correctors of ATP7B-H1069Q reduced Cu toxicity in cua-1 mutants indicating that similar pathogenic molecular pathways might be activated by the H/Q substitution and, therefore, targeted for rescue of ATP7B/CUA-1 function. Taken together, our findings suggest that the newly generated cua-1 mutant strain represents an excellent model for Cu toxicity studies in WD.

Organelles are dynamic entities whose functions are essential for the optimum functioning of cells. It is now known that the juxtaposition of organellar membranes is essential for the exchange of metabolites and their communication. These functional apposition sites are termed membrane contact sites. Dynamic membrane contact sites between various sub-cellular structures such as mitochondria, endoplasmic reticulum, peroxisomes, Golgi apparatus, lysosomes, lipid droplets, plasma membrane, endosomes, etc. have been reported in various model systems. The burgeoning area of research on membrane contact sites has witnessed several manuscripts in recent years that identified the contact sites and components involved. Several methods have been developed to identify, measure and analyze the membrane contact sites. In this manuscript, we aim to discuss important methods developed to date that are used to study membrane contact sites.