Acta Histochemica Et Cytochemica最新文献

Pathological hallmark of Alzheimer’s disease (AD) is characterized by the accumulation and aggregation of amyloid β (Aβ) peptides into extracellular plaques of the brain. Clarification of the process of how soluble Aβ starts to assemble into amyloid fibrils is an essential step in elucidating the pathogenesis of AD. In our previous study, Aβ proteoforms including full-length Aβ40 and Aβ42/43 with N- and C-terminal truncated forms were visualized in postmortem brains from AD patients with matrix-assisted laser desorption/ionization-based mass spectrometry imaging (MALDI-MSI). In this study, Aβ proteoforms were consistently visualized by an updated protocol, and uncharacterized peptides such as Aβ1-29 and Aβ10-40 in AD brains were also visualized. To decipher neurotoxic effects of Aβ in patients’ brains, here we integrate liquid chromatography tandem mass spectrometry (LC-MS/MS) based shotgun proteomics with laser microdissection (LMD) excised tissue samples as well as direct tissue imaging with MALDI-MSI. With this approach, we have highlighted dynamic alterations of microtubule associating proteins (MAPs) including MAP1A, MAP1B and MAP2 as well as AD dominant proteins including APP, UCHL1, SNCA, and APOE. Of note, as lipid dysregulation has been implicated with AD pathology, we have challenged to integrate proteomics and lipid imaging for AD and control brain tissue. Spatial multi-omics is also valid to uncover molecular pathology of white matter as well as grey matter and leptomeningeal area, for example, by visualizing heme in patients’ postmortem brains.

Mitochondrial ferritin (FtMt) is a novel ferritin that sequesters iron and plays a protective role against oxidative stress. FtMt shares a high homology with H-ferritin but is expressed only in the brain, heart, and testis. In the midbrain, FtMt expression is observed in the substantia nigra. FtMt plays a neuroprotective role in the pathology of neurodegenerative diseases such as Parkinson’s disease, where excessive iron induces oxidative stress, causing cell death. Herein, we investigated FtMt immunoreactivity in the brains of patients with subarachnoid hemorrhage (SAH). Double immunofluorescence labeling of tyrosine hydroxylase (TH) and FtMt showed high colocalization in the substantia nigra pars compacta (SNc) in control and SAH cases. However, in SAH cases, FtMt immunoreactivity was observed in some TH-negative neurons. Double immunofluorescence labeling of glial cell markers and FtMt showed no apparent colocalization. The number and ratio of FtMt-positive but TH-negative neurons significantly differed between the control and SAH groups. Prussian blue staining in SAH cases showed positive iron staining over a wide surface range and the substantia nigra. Thus, FtMt may be related to iron dynamics in the substantia nigra following subarachnoid hemorrhage.

High-mobility group box 1 (HMGB1) functions as damage-associated molecular pattern (DAMPs), released into extracellular space during cellular stress. Extracellular HMGB1 act as signal molecules through Toll-like receptor (TLR) 2 or TLR4, exerting diverse functions in both normal cells and malignant cells including breast cancer. However, their comprehensive examination in breast cancer tissues is lacking. Thus, we immunolocalized them in 112 breast cancer tissues, correlating their immunoreactivity with clinicopathological parameters and clinical outcomes to clarify their significance in breast cancer. We demonstrated that nuclear HMGB1 immunoreactivity was correlated with tumor progression and longer disease-free survival. In contrast, TLR2 immunoreactivity was correlated with increased cell proliferation and shorter disease-free survival, dependent on cytoplasmic HMGB1 immunoreactivity. Additionally, TLR4 immunoreactivity correlated with chemoresistance, regardless of cytoplasmic HMGB1 immunoreactivity. It was therefore considered that TLR2 collaboratively contributed to breast cancer progression with HMGB1-DAMPs to become a worse prognostic factor. Meanwhile, TLR4 served as a worse prognostic factor associated with chemoresistance, irrespective of HMGB1.

Recent advances in viral vector technology, specifically using adeno-associated virus (AAV) vectors, have significantly expanded possibilities in neuronal tracing. We have utilized the Cre/loxP system in combination with AAV techniques in rats to explore the subcellular localization of palmitoylation signal-tagged GFP (palGFP) in oxytocin-producing neurosecretory neurons. A distinctive branching pattern of single axons was observed at the level of the terminals in the posterior pituitary. Despite challenges in detecting palGFP signals by fluorescent microscopy, immunoelectron microscopy demonstrated predominant localization on the plasma membrane, with a minor presence on the neurosecretory vesicle membrane. These findings suggest that membrane-anchored palGFP may undergo exocytosis, translocating from the plasma membrane to the neurosecretory vesicle membrane. In this study, we observed characteristic axon terminal structures in the posterior pituitary of oxytocin neurons. This study indicates the importance of understanding the plasma membrane-specific sorting system in neuronal membrane migration and encourages future studies on the underlying mechanisms.

Protein lactylation is a post-translational modification associated with glycolysis. Although recent evidence indicates that protein lactylation is involved in epigenetic gene regulation, its pathophysiological significance remains unclear, particularly in neoplasms. Herein, we investigated the potential involvement of protein lactylation in the molecular mechanisms underlying benign and malignant pancreatic epithelial tumors, as well as its role in the response of pancreatic cancer (PC) cells to gemcitabine. Increased lactylation was observed in the nuclei of intraductal papillary mucinous adenoma, non-invasive intraductal papillary mucinous carcinoma, and invasive carcinoma, in parallel to the upregulation of hypoxia-inducible factor-1α. This observation indicated that a hypoxia-associated increase in nuclear protein lactylation could be a biochemical hallmark in pancreatic epithelial tumors. The standard PC chemotherapy drug gemcitabine suppressed histone lactylation in vitro, suggesting that histone lactylation might be relevant to its mechanism of action. Taken together, our findings suggest that protein lactylation may be involved in the development of pancreatic epithelial tumors and could represent a potential therapeutic target for PC.

Cancer tissue generally possesses an immunosuppressive microenvironment. However, some cancers are associated with lymphoid stroma (i.e., a widely developed tertiary lymphoid structure). The T-cell zone (paracortex) of secondary lymphoid organs, particularly lymph nodes, is characterized by an abundance of T-cell zone fibroblastic reticular cells (TCZ-FRCs) that express C-C motif chemokine ligand 21 (CCL21) and smooth muscle actin (SMA). We analyzed the presence of TCZ-FRCs in 30 cases of carcinomas with lymphoid stroma of the breast, stomach, colon, tongue, and skin. Immunohistochemistry corroborated the abundance of CCL21⁺ SMA⁺ TCZ-FRCs in the normal lymph nodes. In sharp contrast, all 30 carcinomas with lymphoid stroma displayed no CCL21⁺ SMA⁺ TCZ-FRCs despite the affluence of T cells. Real-time reverse transcription polymerase chain reaction confirmed a marked decrease in the messenger ribonucleic acid expression of CCL21 and its receptor C-C motif chemokine receptor 7 in cancer lymphoid stroma compared to that in lymph nodes. Next, we analyzed the T cell phenotypes. The cancer lymphoid stroma demonstrated an abundance of CD3⁺ CD62L⁻ memory-type T cells, in contrast to the presence of CD3⁺ CD62L⁺ naïve- and central memory T cells in the T cell zone of lymphoid tissues. Our data demonstrated the following: 1) Cancer lymphoid stroma lacked TCZ-FRCs with abundance of more activated T cells than in lymph nodes and 2) these were common phenomena in cancer lymphoid stroma irrespective of the histological types and organs involved.

We aimed to investigate the effects of short-term corticosteroid administration after anterior cruciate ligament (ACL) reconstruction on marrow adipose tissue (MAT) and trabecular bone mass, as well as to examine whether treadmill exercise can mitigate MAT increase and trabecular bone deterioration caused by corticosteroid. ACL-reconstructed rats were divided into groups: no intervention, daily treadmill exercise (60 min/day), administration of the steroidal drug dexamethasone (250 μg/kg on days 0–5, 7, and 9 post-operatively), or dexamethasone administration combined with treadmill exercise. Untreated rats were served as controls. At day 10 or 30 post-operatively, histological assessments were performed in the proximal tibial epiphysis. MAT accumulation and trabecular bone loss were observed after ACL reconstruction. Dexamethasone promoted MAT accumulation at day 10 post-operatively but did not affect the trabecular bone loss. The MAT accumulation caused by dexamethasone reversed within 21 days after discontinuation. Treadmill exercise did not influence the changes in the MAT and trabecular bone areas. Short-term corticosteroid administration after ACL reconstruction promoted MAT accumulation while not affecting trabecular bone area. The MAT accumulation resulting from corticosteroid administration was reversible after discontinuation. Treadmill exercise could not mitigate the accumulation of MAT caused by corticosteroid administration and did not affect trabecular bone area.

Peritoneal dialysis (PD) fluid, which contains a high concentration of glucose, is involved in peritoneal damage after long-term use. The mechanisms through which glucose induces damage to the mesothelium have not been clearly elucidated. Although, endoplasmic reticulum (ER) stress response is associated with several diseases, the involvement of ER stress in peritoneal damage has not yet been demonstrated. Primary-cultured rat peritoneal mesothelial cells (RPMCs) and rat PD model were used to investigate the influence of glucose on the peritoneum. Cells treated with glucose were examined for cytotoxicity, induction of apoptosis, and activation of the ER stress pathway. Glucose treatment of RPMCs induced cell death at concentrations higher than 3%. Annexin V positive, that is a feature of apoptosis, occurred in dead cells. Treatment with glucose led to the activation of protein kinase R-like ER kinase (PERK) and eukaryotic translation initiation factor-2α (eIF-2α). Glucose also induced the expression and nuclear translocation of homologous protein C/EBP. Cell death was rescued by the integrated stress response inhibitor, ISRIB, which suppresses the integrated stress response pathway, including ER stress. Glucose in PD fluid induces PERK/eIF-2α-mediated ER stress in RPMCs, resulting in apoptosis. This cellular stress may cause peritoneal damage in patients receiving PD.

Multiple sclerosis (MS) is an inflammatory demyelinating disease of the central nervous system, characterized by remyelination failure and axonal dysfunction. Remyelination by oligodendrocytes is critical for improvement of neurological deficits associated with demyelination. Rodent models of demyelination are frequently used to develop and evaluate therapies for MS. However, a suitable mouse model for assessing remyelination-associated recovery of motor functions is currently unavailable. In this review, we describe the development of the mouse model of internal capsule (IC) demyelination by focal injection of lysolecithin into brain and its application in the evaluation of drugs for demyelinating diseases. This mouse model exhibits motor deficits and subsequent functional recovery accompanying IC remyelination. Notably, this model shows enhancement of functional recovery as well as tissue regeneration when treated with clemastine, a drug that promotes remyelination. The IC demyelination mouse model should contribute to the development of novel drugs that promote remyelination and ameliorate neurological deficits in demyelinating diseases.

Aquaporin-5 (AQP5) water channel, transmembrane protein 16A (TMEM16A) Ca²⁺-activated Cl⁻ channel, and Na⁺-K⁺-2Cl⁻ cotransporter (NKCC1) are membrane proteins on salivary gland acinar cells that function in watery saliva secretion. We examined their expression changes in rat parotid glands under reduced mastication. Rats were either fed regular chow as a control group, fasted for 48 hr or fed a liquid diet for 48 hr or 1 week to reduce mastication. The parotid glands were then resected to analyze the protein and mRNA levels by immunofluorescence, immunoblotting, and reverse-transcription quantitative PCR (RT-qPCR). AQP5 protein was significantly decreased in both liquid diet groups and the fasting group but its mRNA levels showed no apparent changes compared with the control group. The protein and mRNA levels of TMEM16A and NKCC1 showed no significant changes between any of the groups other than an increase in NKCC1 mRNA in the 1-week liquid diet group. These results suggest that reduced mastication may increase the AQP5 protein degradation, but not that of other membrane proteins necessary for saliva secretion.