Acta histochemica最新文献

Central giant cell granuloma (CGCG) is a benign jaw lesion with variable clinical behavior. Cell cannibalism is a cellular process associated with aggressiveness and invasion in malignant neoplasms. Here, we morphologically investigated cell cannibalism as an auxiliary method to predict CGCG clinical behavior. Cell cannibalism was quantitatively evaluated in 19 cases of peripheral giant cell granuloma (PGCG), 38 cases of CGCG (non-aggressive and aggressive), and 19 cases of giant cell tumor of bone (GCT) stained with hematoxylin and eosin. T-test was performed to assess the differences between the variables analyzed (p ≤ 0.05). Cell cannibalism was identified in 21% of non-aggressive CGCGs and 68.4% of aggressive CGCGs. A significantly higher amount of cannibal multinucleated giant cells (CMGC) was observed in aggressive CGCG compared to PGCG and non-aggressive CGCG (p = 0.042; p = 0.044, respectively). There were no significant differences in the CMGC index between non-aggressive CGCG and PGCG (p = 0.858) and between aggressive CGCG and GCT (p = 0.069). CGGC cases that exhibited rapid growth and tooth displacement and/or root resorption had a higher amount of CMGC (p = 0.035; p = 0.041, respectively). Cell cannibalism can be identified in CGCG through routine anatomopathological examination. The quantification of CMGC can help to predict the clinical behavior of central giant cell granuloma.

Even after more than two years of intensive research, not all of the pathophysiological processes of Coronavirus Disease 2019 (COVID-19), induced by severe acute respiratory syndrome coronavirus type 2 (SARS-CoV-2) infection, have been fully elucidated. The initial virus-host interaction at the respiratory epithelium plays a crucial role in the course and progression of the infection, and is highly dependent on the glycosylation pattern of the host cell and of the secreted mucins. Glycans are polysaccharides that can be attached to proteins and thereby add to their stability and functionality. Lectins are glycan-binding proteins that recognize specific glycan motifs, and lectin histochemistry is a suitable tool to visualize and examine glycosylation pattern changes in tissues. In this study we used lectins with different glycan-specificities for the visualization of glycosylation pattern changes in the respiratory tract of SARS-CoV-2 infected Golden Syrian hamsters. While some lectins (LEL, STL) enable the visualization of the damage to alveolar type 1 pneumocytes, other lectins, e.g., GSLI, visualized the loss and subsequent hyperplasia of type 2 pneumocytes. UEAI staining was co-localized with KI67, a proliferation marker. Double staining of lectins LEL, STL and WGA with specific immune cell markers (Iba1, CD68) showed co-localization and the dominant infiltration of monocyte-derived macrophages into infected alveolar tissue. The elucidation of the glycosylation pattern of the respiratory tract cells in uninfected and infected Golden Syrian hamsters revealed physiological and pathological aspects of the disease that may open new possibilities for therapeutic development.

Topiramate [2,3:4,5-bis-o-(1-methylethylidene) β-D-fructo-pyranose sulfamate; TPM] is one of the most used new-generation antiepileptic drugs. It has been reported to regulate the differentiation of human bone cells. However, the molecular mechanism of TPM in osteoblast differentiation is not fully elucidated. In the present study, we examined the effect of TPM on osteogenic differentiation of C3H10T1/2, MC3T3-E1, primary mouse calvarial cells, and primary bone marrow stem cells (BMSCs). Primary cells were isolated from mice calvaria and bone marrow respectively. Expression of the osteogenic gene was determined by RT-PCR. The osteogenic protein levels were measured by Western blot analysis. Alkaline phosphatase (ALP) staining experiment was performed to evaluate ALP activity. Alizarin red s (ARS) staining was performed to measure zebrafish caudal fin regeneration. Treatment of TPM up-regulated the osteogenic genes including distal-less homeobox 5 (Dlx5) and runt-related transcription factor 2 (Runx2). In addition, TPM also increased the Dlx5 and Runx2 protein levels, Smad1/5/9 phosphorylation, and alkaline phosphatase (ALP) activity. Furthermore, TPM activated AMPK, and inhibition of AMPK decreased TPM-induced osteogenic differentiation. In the zebrafish model, osteogenic effect of TPM was identified. TPM was increased amputated caudal fin rays of zebrafish. These results demonstrate that TPM enhances osteogenic differentiation via AMPK-mediated Smad1/5/9 phosphorylation.

HSCs (hepatic stellate cells) contribute to the excessive extracellular matrix (ECM) deposition plays a key role in the progression of hepatic fibrosis. The present study focused on the hepatoprotective effect of Ginsenoside Rc (Rc), one of the protopanaxadiol type ginsenoside, which has contributed to reverse activated HSCs to improve hepatic fibrosis via regulating Nur77-TLR4/MyD88 signaling pathway. We established the hepatic fibrosis model by intraperitoneal injection of carbon tetrachloride (CCl₄). And HSCs were stimulated with TGF-β, followed by silencing of Nur77, and then incubated in Rc. Rc significantly alleviated histopathological changes, reduced serum alanine aminotransferase (ALT) and aspartate aminotransferase (AST) levels. Rc could upregulate the Nur77 and downregulate fibrosis markers in the liver of mice, including decreasing the expressions of α-SMA, Collagen-I, the ratio of TIMP-1/MMP-13. Rc significantly increased the expression of Nur77 and suppressed the production of ECM in HSCs. Rc inhibited TLR4 signaling pathway, consequently reversing the inflammatory response, including the production of MyD88, IRAK1, IRAK4 and IL-23. When Nur77 was knocked in TGF-β-stimulated HSCs, TLR4 and α-SMA production were increased. Rc suppressed these activatory effects in Nur77 knockdown HSCs. Rc reduced inflammatory reaction by regulating the Nur77-TLR4 signaling pathway while suppressing the fibrogenesis suggesting, underscoring a promising approach of Rc for the treatment in hepatic fibrosis. Targeting Nur77-TLR4 signaling in HSCs would be the potential strategy for Rc against hepatic fibrosis.

Background

Fibroblasts (FBs) have been widely used as a typical in vitro cell model for investigating the biological processes and cell pathophysiological mechanisms. However, FBs are prone to senescence in cell culture process after several passages. Thus, a new approach to cell culture is quite required to enhance the viability of cells.

Objective

To explore a novel method of cell culture based on skin FBs.

Methods

Dermal tissue blocks were obtained from BALB/c neonatal mice and randomly divided into experimental group and control group. The experimental group received the newly improved culture method, namely, continuous adherence subculture of tissue block (CASTB) method; while the traditional subculture method was applied in the control group. Cells at 1st, 5th and 10th passages were collected and identified by using histological/immunohistochemical and western blot analysis. Cellular viability, proliferation, senescence and apoptosis were analyzed through application of cell growth curve, CCK-8 assay, Ki67 assay, PCNA protein analysis, β-galactosidase staining, flow cytometry and western blot analysis.

Results

Cells under two culture patterns exhibited spindle/irregular shape and vimentin positive expression. With the increase of passage times, the cellular growth rate in the control group gradually decreased, but no alterations emerged from the experimental group. CASTB method remarkably promoted cell growth and proliferation. Moreover, a greatly lower apoptosis and senescence tendency appeared in the experimental group than the control group with passages increasing.

Conclusion

The method of CASTB is superior to traditional subculture, offering a large number of primary FBs with higher efficiency and success rate and being worth of further popularization and application.

Background

Each eccrine sweat gland (ESG) is a single-tubular structure with a central lumen, and the formation of hollow lumen in the initial solid cell mass is a key developmental process. To date, there are no reports on the mechanism of native ESG lumen formation.

Methods

To investigate the lumen morphogenesis and the lumen formation mechanisms of Sprague-Dawley (SD) rat ESGs, SD rat hind-footpads at E20.5, P1–P5, P7, P9, P12, P21, P28 and P56 were obtained. The lumen morphogenesis of ESGs was examined by HE staining and immunofluorescence staining for polarity markers. The possible mechanisms of lumen formation were detected by terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) apoptosis assay and autophagy marker LC3B immunofluorescence staining, and further explored by ouabain intervention experiment.

Results

In SD rat ESGs, the microlumen was formed at P1, and the small intact lumen with apical-basal polarity appeared at P3. The expression of apical marker F-actin, basal marker Laminin, basolateral marker E-cadherin was consistent with the timing of lumen formation of SD rat ESGs. During rat ESG development, apoptosis and autophagy were not detected. However, inhibition of Na⁺-K⁺-ATPase (NKA) with ouabain resulted in decreased lumen size, although neither the timing of lumen formation nor the expression of polarity proteins was altered.

Conclusions

Epithelial polarity-driven membrane separation but not cavitation regulates lumen formation of SD rat ESGs. NKA-regulated fluid accumulation drives lumen expansion.

The vomeronasal organ is an olfactory organ found in amphibians and higher vertebrates. Type 1 vomeronasal receptors, one of the major olfactory receptors in vertebrates, are expressed in the vomeronasal organ in mammals. In amphibians and fish, they are expressed in the olfactory epithelium. The lungfish, which is the species of fish most closely related to amphibians, has a primitive vomeronasal organ: the recess epithelium. Expression of type 1 vomeronasal receptors has been reported in both the olfactory epithelium and the recess epithelium in three species of African lungfish and one species of South American lungfish. However, a previous study suggested that in the African lungfish Protopterus dolloi these receptors are expressed only in the olfactory epithelium. In this study, we identified 21 type 1 vomeronasal receptor genes in P. dolloi and examined the expression sites in the olfactory organ. In P. dolloi, most cells expressing the type 1 vomeronasal receptor were distributed in the olfactory epithelium, but a few were also found in the recess epithelium. This implies that the functions of the olfactory epithelium and the primitive vomeronasal organ are incompletely separated, and that all extant African and South American lungfish share this trait.

ITGB3, an osteoclast marker, is involved in osteoclast formation. Nevertheless, its related mechanism remains poorly characterized. Herein, this study examines the mechanisms affecting osteoclast formation with the involvement of ITGB3. Osteoclast formation was induced with macrophage colony-stimulating factor (M-CSF) and receptor activator of nuclear factor-kappa B ligand (RANKL), followed by measurement of the mRNA and protein expression of ITGB3 and LSD1. After gain- and loss-of-function assays, cell viability and the expression of osteoclast marker genes (NFATc1, ACP5, and CTSK) were assessed, and osteoclast formation was evaluated with TRAP staining. ChIP assays were used to examine histone 3 lysine 9 (H3K9) monomethylation (H3K9me1) and H3K9 dimethylation (H3K9me2) modifications and LSD1 protein enrichment in the ITGB3 promoter. During osteoclast formation, ITGB3 and LSD1 were gradually augmented. Knockdown of LSD1 or ITGB3 curbed cell viability, the expression of osteoclast marker genes, and osteoclast formation. Moreover, overexpression of ITGB3 nullified the suppressive impact of LSD1 knockdown on osteoclast formation. Mechanistically, LSD1 promoted ITGB3 expression by reducing H3K9 levels in the ITGB3 promoter. LSD1 enhanced ITGB3 expression by decreasing H3K9me1 and H3K9me2 levels in ITGB3 promoter to boost osteoclast formation.

Situated in the oral cavity, the rabbit palatine tonsils are part of the mucosal immune system and help to defend the body against foreign pathogens. Expressed as two oval protrusions in the wall of the oropharynx, the rabbit palatine tonsils are characterized by excretory ducts and trabeculae. We here compare paraffin embedded and cryosections of the healthy rabbit tonsils. This analysis centers on evaluating the differential outcomes resulting from the application of these fixation methodologies in conjunction with immunohistochemical assays targeting collagen I, collagen III, fibronectin, α-smooth muscle actin (α-SMA), and ki67. Subsequent recommendations are provided based on our findings. Furthermore, we demonstrate the advantage of an antigen retrieval step in immunohistochemical labeling of paraffin sections. Basic classical histological stainings as HE, GT and elastin were also performed. Comparison of different stainings and labelings was furthermore performed in serial sections, showing that adjacent to the excretory ducts, the tonsillar tissue was particularly composed of collagen I and fibronectin, while the vessel walls were predominantly α-SMA positive. Moreover, PAR-2 immunohistochemical staining was performed, where a small fraction of the cells found in the tonsillar connective tissue were PAR-2 positive (probably a subpopulation of mast cells), as well as the lumen of some excretory ducts and trabeculae. Collagen III on the other hand was only weakly expressed in the tonsils. Proliferating ki67 positive cells were rare. This endeavor serves to furnish the scientific community with reference imagery pertinent to researchers opting for the rabbit palatine tonsil model. The diversity of staining techniques employed herein establishes a foundational repository of images, primed for comparative analysis against pathological conditions. Furthermore, these images hold the potential to illustrate inter-species variations. For instance, they can be juxtaposed against murine or rodent tonsils, or even offer insights into the human context.

Since their relatively recent discovery, telocytes (TCs) have been described as peculiar cells strategically positioned in the stromal tissue component of multiple organ systems of the mammalian body including female reproductive organs (i.e., ovary, uterine tube, and uterus). Nevertheless, current knowledge of TCs in the vagina is very limited. The present study was therefore undertaken to investigate the existence and characteristics of TCs in the stromal tissue of human vaginal mucosa by means of immunohistochemistry, immunofluorescence confocal microscopy, and transmission electron microscopy. In the vaginal lamina propria, TCs were first identified by CD34 immunohistochemistry that revealed the presence of CD34⁺ stromal cells arranged in networks, especially around blood vessels. Double immunofluorescence confocal microscopy allowed to precisely distinguish the perivascular networks of CD34⁺ stromal cells lacking CD31 immunoreactivity from adjacent CD31⁺ microvessels. All the perivascular networks of TCs/CD34⁺ stromal cells situated in the vaginal lamina propria coexpressed platelet-derived growth factor receptor α, which strengthened their identification as TCs. Instead, vaginal mucosal TCs were immunophenotypically negative for c-kit/CD117. The ultrastructural examination confirmed the presence of TCs, namely stromal cells with characteristic cytoplasmic processes (i.e., telopodes) forming labyrinthine networks around blood vessels and releasing extracellular vesicles. Together, our morphological findings provide the first comprehensive demonstration that TCs reside in the human vaginal lamina propria, thus paving the way for further investigation of their putative functions in vaginal mucosal homeostasis and pathophysiology.