Journal of Histochemistry & Cytochemistry最新文献

Several types of cytotoxic insults disrupt endoplasmic reticulum (ER) homeostasis, cause ER stress, and activate the unfolded protein response (UPR). The role of ER stress and UPR activation in hypersensitivity pneumonitis (HP) has not been described. HP is an immune-mediated interstitial lung disease that develops following repeated inhalation of various antigens in susceptible and sensitized individuals. The aim of this study was to investigate the lung expression and localization of the key effectors of the UPR, BiP/GRP78, CHOP, and sXBP1 in HP patients compared with control subjects. Furthermore, we developed a mouse model of HP to determine whether ER stress and UPR pathway are induced during this pathogenesis. In human control lungs, we observed weak positive staining for BiP in some epithelial cells and macrophages, while sXBP1 and CHOP were negative. Conversely, strong BiP, sXBP1- and CHOP-positive alveolar and bronchial epithelial, and inflammatory cells were identified in HP lungs. We also found apoptosis and autophagy markers colocalization with UPR proteins in HP lungs. Similar results were obtained in lungs from an HP mouse model. Our findings suggest that the UPR pathway is associated with the pathogenesis of HP.

Lactate and ATP formation by aerobic glycolysis, the Warburg effect, is considered a hallmark of cancer. During angiogenesis in non-cancerous tissue, proliferating stalk endothelial cells (ECs) also produce lactate and ATP by aerobic glycolysis. In fact, all proliferating cells, both non-cancer and cancer cells, need lactate for the biosynthesis of building blocks for cell growth and tissue expansion. Moreover, both non-proliferating cancer stem cells in tumors and leader tip ECs during angiogenesis rely on glycolysis for pyruvate production, which is used for ATP synthesis in mitochondria through oxidative phosphorylation (OXPHOS). Therefore, aerobic glycolysis is not a specific hallmark of cancer but rather a hallmark of proliferating cells and limits its utility in cancer therapy. However, local treatment of angiogenic eye conditions with inhibitors of glycolysis may be a safe therapeutic option that warrants experimental investigation. Most types of cells in the eye such as photoreceptors and pericytes use OXPHOS for ATP production, whereas proliferating angiogenic stalk ECs rely on glycolysis for lactate and ATP production. (J Histochem Cytochem XX.XXX-XXX, XXXX).

The TRPA1 ion channel is a sensitive detector of reactive chemicals, found primarily on sensory neurons. The phenotype exhibited by mice lacking TRPA1 suggests its potential as a target for pharmacological intervention. Antibody-based detection for distribution analysis is a standard technique. In the case of TRPA1, however, there is no antibody with a plausible validation in knockout animals or functional studies, but many that have failed in this regard. To this end we employed the single molecule in situ hybridization technique RNAscope on sensory neurons immediately after detection of calcium responses to the TRPA1 agonist allyl isothiocyanate. There is a clearly positive correlation between TRPA1 calcium imaging and RNAscope detection (R = 0.43), although less than what might have been expected. Thus, the technique of choice should be carefully considered to suit the research question. The marginal correlation between TRPV1 RNAscope and the specific agonist capsaicin indicates that such validation is advisable for every RNAscope target. Given the recent description of a long-awaited TRPA1 reporter mouse, TRPA1 RNAscope detection might still have its use cases, for detection of RNA at particular sites, for example, defined structurally or by other molecular markers.

To clarify the cellular mechanism of cortical porosity induced by intermittent parathyroid hormone (PTH) administration, we examined the femoral cortical bone of mice that received 40 µg/kg/day (four times a day) human PTH (hPTH) (1-34). The PTH-driven cortical porosity initiated from the metaphyseal region and chronologically expanded toward the diaphysis. Alkaline phosphatase (ALP)-positive osteoblasts in the control mice covered the cortical surface, and endomucin-positive blood vessels were distant from these osteoblasts. In PTH-administered mice, endomucin-reactive blood vessels with TRAP-positive penetrated the ALP-positive osteoblast layer, invading the cortical bone. Statistically, the distance between endomucin-positive blood vessels and the cortical bone surface abated after PTH administration. Transmission electron microscopic observation demonstrated that vascular endothelial cells often pass through the flattened osteoblast layer and accompanied osteoclasts in the deep region of the cortical bone. The cell layers covering mature osteoblasts thickened with PTH administration and exhibited ALP, α-smooth muscle actin (αSMA), vascular cell adhesion molecule-1 (VCAM1), and receptor activator of NF-κB ligand (RANKL). Within these cell layers, osteoclasts were found near endomucin-reactive blood vessels. In PTH-administered femora, osteocytes secreted Dkk1, a Wnt inhibitor that affects angiogenesis, and blood vessels exhibited plasmalemma vesicle-associated protein, an angiogenic molecule. In summary, endomucin-positive blood vessels, when accompanied by osteoclasts in the ALP/αSMA/VCAM1/RANKL-reactive osteoblastic cell layers, invade the cortical bone, potentially due to the action of osteocyte-derived molecules such as DKK1.

The large family of transient receptor potential (TRP) channels are integral membrane proteins that function as environmental sensors and act as ion channels after activation by mechanical (touch), physical (heat, pain), and chemical stimuli (pungent compounds such as capsaicin). Most TRP channels are localized in the plasma membrane of cells but some of them are localized in membranes of organelles and function as intracellular Ca²⁺-ion channels. TRP channels are involved in neurological disorders but their precise role(s) and relevance in these disorders are not clear. Endothelial cells of the blood-brain barrier (BBB) express TRP channels such as TRP vanilloid 1-4 and are involved in thermal detection by regulating BBB permeability. In neurological disorders, TRP channels in the BBB are responsible for edema formation in the brain. Therefore, drug design to modulate locally activity of TRP channels in the BBB is a hot topic. Today, the application of TRP channel antagonists against neurological disorders is still limited.

Liver regeneration is a well-orchestrated compensatory process that is regulated by multiple factors. We recently reported the importance of the chromatin protein, a high-mobility group box 2 (HMGB2) in mouse liver regeneration. However, the molecular mechanism remains unclear. In this study, we aimed to study how HMGB2 regulates hepatocyte proliferation during liver regeneration. Seventy-percent partial hepatectomy (PHx) was performed in wild-type (WT) and HMGB2-knockout (KO) mice, and the liver tissues were used for microarray, immunohistochemistry, quantitative polymerase chain reaction (qPCR), and Western blotting analyses. In the WT mice, HMGB2-positive hepatocytes colocalized with cell proliferation markers. In the HMGB2-KO mice, hepatocyte proliferation was significantly decreased. Oil Red O staining revealed the transient accumulation of lipid droplets at 12-24 hr after PHx in the WT mouse livers. In contrast, decreased amount of lipid droplets were found in HMGB2-KO mouse livers, and it was preserved until 36 hr. The microarray, immunohistochemistry, and qPCR results demonstrated that the expression of lipid metabolism-related genes was significantly decreased in the HMGB2-KO mouse livers. The in vitro experiments demonstrated that a decrease in the amount of lipid droplets correlated with decreased cell proliferation activity in HMGB2-knockdown cells. HMGB2 promotes de novo lipogenesis to accelerate hepatocyte proliferation during liver regeneration.

Xylene is the commonest clearing agent even though it is hazardous and costly. This study evaluated the clearing properties of coconut oil as an alternative cost-effective clearing agent for histological processes. Ten (10) prostate samples fixed in formalin were taken and each one was cut into 4 before randomly separating them into four groups (A, B, C and D). Tissues were subjected to ascending grades of alcohol for dehydration. Group A was cleared in xylene and Groups B, C, and D were cleared at varying times of 1hr 30mins, 3hrs, and 4hrs in coconut oil respectively before embedding, sectioning, and staining were carried out. Gross and histological features were compared. Results indicated a significant shrinkage in coconut oil-treated specimen compared with the xylene-treated specimen and only the tissues cleared in coconut oil for 4hrs were as rigid as the tissues cleared in xylene (p > 0.05). No significant difference was found in either of the sections when checked for cellular details and staining quality (p > 0.999). Coconut oil is an efficient substitute for xylene in prostate tissues with a minimum clearing time of 4hrs, as it is environmentally friendly and less expensive, but causes significant shrinkage to prostate tissue.

Diabetes is not only an endocrine but also a vascular disease. Vascular defects are usually seen as consequence of diabetes. However, at the level of the pancreatic islet, vascular alterations have been described before symptom onset. Importantly, the cellular and molecular mechanisms underlying these early vascular defects have not been identified, neither how these could impact the function of islet endocrine cells. In this review, we will discuss the possibility that dysfunction of the mural cells of the microvasculature-known as pericytes-underlies vascular defects observed in islets in pre-symptomatic stages. Pericytes are crucial for vascular homeostasis throughout the body, but their physiological and pathophysiological functions in islets have only recently started to be explored. A previous study had already raised interest in the "microvascular" approach to this disease. With our increased understanding of the crucial role of the islet microvasculature for glucose homeostasis, here we will revisit the vascular aspects of islet function and how their deregulation could contribute to diabetes pathogenesis, focusing in particular on type 1 diabetes (T1D).

Correlative microscopy is a sophisticated imaging technique that combines optical and electron microscopes, with the most common approach being the integration of light microscopy and electron microscopy, known as correlative light and electron microscopy (CLEM). While CLEM provides a comprehensive view of biological samples, it presents a significant challenge in sample preparation due to the distinct processes involved in each technique. Striking a balance between these methods is crucial. Despite numerous approaches, achieving seamless imaging with CLEM remains a complex task. Exosomes, nanovesicles ranging from 30 to 150 nm in size, are enclosed by a lipid bilayer and released by various cell types. Visualizing exosomes poses difficulties due to their small size and minimal electric charge. However, imaging exosomes at high resolution offers a direct method to understand their morphology and functions. In this study, we evaluated exosome imaging with CLEM using a combination of confocal, transmission electron microscope, and scanning electron microscope (SEM). In addition, we conducted a comparative analysis of these two techniques, evaluating their suitability and efficiency in imaging nanoscale structures. In this study, we found that confocal-SEM correlation is more applicable for imaging exosomes. Moreover, we observed that exosomes were found in clusters in confocal-SEM correlation.