European Journal of Histochemistry最新文献

Prolactin (PRL) is a hormone crucial for normal reproduction, functioning as an autocrine, paracrine, and endocrine factor. This study aimed to examine the immunolocalization and expression patterns of PRL, prolactin receptor (PRLR), and signal transducer and activator of transcription 5 (STAT5) in the ovaries of wild ground squirrels during both breeding and non-breeding periods. Significant seasonal variations were observed in ovarian weights, with higher values during the breeding season and relatively lower values during the nonbreeding season. PRL, PRLR, STAT5, and p-STAT5 were immunolocalized in granulosa cells and luteal cells during the breeding season, whereas they were exclusively found in granulosa cells during the non-breeding season. The mRNA expression levels of Prl, Prlr, and Stat5 were increased in ovarian tissues during the breeding season compared to the non-breeding season. Moreover, the mean mRNA levels of Prl, Prlr, and Stat5 exhibited a positive correlation with ovarian weights. Both circulating PRL and ovarian PRL concentrations were significantly elevated during the breeding season. Additionally, transcriptomic analysis of ovarian tissues revealed differentially expressed genes possibly associated with ovarian function and mammary gland development, including ovarian follicle development, steroid synthesis, and regulation of reproductive process. These findings suggest that PRL might play an essential endocrine, autocrine, or paracrine role in the regulation of seasonal changes in the ovarian functions in wild ground squirrels.

Autofluorescence (AF) poses challenges for detecting proteins of interest in situ when employing immunofluorescence (IF) microscopy. This interference is particularly pronounced in strongly autofluorescent tissues such as myocardium, where tissue AF can be comparable to IF. Although various histochemical methods have been developed to achieve effective AF suppression in different types of tissue, their applications on myocardial  samples have not been well validated. Due to inconsistency across different autofluorescent structures in sometypes of tissue, it is unclear if these methods can effectively suppress AF across all autofluorescent structures within the myocardium. Here, we quantitatively evaluated the performance of several commonly used quenching treatments on formaldehyde-fixed myocardial samples, including 0.3 M glycine, 0.3% Sudan Black B (SBB), 0.1% and 1% sodium borohydride (NaBH4), TrueVIEW® and TrueBlack®. We further assessed their quenching performance by employing the pre-treatment and post-treatment protocols, designed to cover two common IF staining scenarios where buffers contained detergents or not. The results suggest that SBB and TrueBlack® outperform other reagents in AF suppression on formaldehyde-fixed myocardial samples in both protocols. Furthermore, we inspected the quenching performance of SBB and TrueBlack® on major autofluorescent myocardial structures and evaluated their influence on IF imaging. The results suggest that SBB outperforms TrueBlack® in quenching major autofluorescent structures, while TrueBlack® excels in preserving IF labeling signal. Surprisingly, we found the treatment of NaBH4 increased AF signal and enhanced the AF contrast of major autofluorescent structures. This finding suggests that NaBH4 has the potential to act as an AF enhancer and may facilitate the interpretation of myocardial structures without the need for counterstaining.

Pulmonary fibrosis is a progressive lung disorder. Evidence has shown that hsa_circular (circ)RNA_0001861 is dysregulated in pulmonary fibrosis. However, the detailed function of hsa_circRNA_0001861 in pulmonary fibrosis remains unexplored. To investigate the function of hsa_circRNA_0001861 in pulmonary fibrosis, human pulmonary fibroblasts in vitro were used, and cell counting kit-8 (CCK-8) and 5-ethynyl-2'-deoxyuridine (EdU) staining were performed to assess cell viability and proliferation, respectively. Western blot analysis and reverse transcription-quantitative PCR (RT-qPCR) were used to evaluate protein and mRNA levels. Meanwhile, the relationship among hsa_circRNA_0001861, miR-296-5p and BCL-2 binding component 3 (BBC3) was investigated by RNA pull-down assays. Furthermore, an in vivo model of lung fibrosis was constructed to assess the function of hsa_circRNA_0001861 in lung fibrosis. The data revealed that TGF‑β1 significantly increased the proliferation of pulmonary fibroblasts, while this phenomenon was markedly abolished by hsa_circRNA_0001861 overexpression. hsa_circRNA_0001861 overexpression markedly inhibited TGF‑β1‑induced fibrosis in pulmonary fibroblasts through the mediation of α-smooth muscle actin, E-cadherin, collagen III and fibronectin 1. Meanwhile, hsa_circRNA_0001861 could bind with miR-296-5p, and BBC3 was identified to be the downstream mRNA of miR-296-5p. In addition, the upregulation of hsa_circRNA_0001861 clearly reversed TGF‑β1‑induced fibrosis and proliferation in pulmonary fibroblasts through the upregulation of BBC3. Furthermore, hsa_circRNA_0001861 upregulation markedly alleviated pulmonary fibrosis in vivo. Hsa_circRNA_0001861 upregulation attenuated pulmonary fibrosis by modulating the miR-296-5p/BBC3 axis. Hence, the present study may provide some insights for the discovery of new methods against pulmonary fibrosis.

Crocin has been reported to have therapeutic effects on multiple cancers including colon cancer, but its specific mechanism is still ambiguous and needs to be further explored. Human colorectal adenocarcinoma cells (HCT-116) and human normal colonic epithelial cells (CCD841) were first treated with increasing concentrations of crocin. Subsequently, with 150 and 200 μM of crocin, the cell vitality was examined by cell counting kit 8. Cell apoptosis and proliferation were tested by TUNEL staining and colony formation assay, respectively. The expression of Ki-67 was assessed by immunofluorescence. Enzyme-linked immunosorbent assay was used to evaluate the level of inflammation- and oxidative-related factors. The reactive oxygen species (ROS) production and mitochondrial membrane potential (MMP) were examined by flow cytometer. Janus kinase (JAK), signal transducer and activator of transcription 3 (STAT3), and extracellular regulated protein kinases (ERK) in HCT-116 cells were tested by Western blot. Different concentrations of crocin barely affected the CCD841 cell vitality, while crocin restrained the HCT-116 cells vitality, proliferation and the expression of Ki-67, while inducing apoptosis in a concentration-dependent manner. Moreover, the contents of inflammation- and oxidative-related factors in HCT-116 cells were largely blunted by crocin that enhanced ROS and restrained the MMP and suppressed p-JAK2/JAK2, p-STAT3/STAT3, and p-ERK/ERK expression in HCT-116 cells. Crocin induced apoptosis and restored mitochondrial function in HCT-116 cells via repressing the JAK pathway. If the threptic effect works in patients, it could herald a new, effective treatment for colon cancer, improving the patients' prognosis and quality of life.

Geniposide (GEN), a medical herb, is known for its therapeutic applications in cardiovascular diseases, though its efficacy in treating myocardial ischemia/reperfusion injury (MI/RI) is yet to be fully elucidated. This study is an endeavor to explore the potential protective mechanism of GEN against MI/RI. To simulate the MI/RI condition, the left anterior descending artery was occluded for 30 min, followed by a reperfusion period of 120 min in a rat model. Three dosages (50, 100, or 150 mg/kg) of GEN were intraperitoneally injected to the Sprague-Dawley rats once a day, for seven days before the ligation of the artery. The rats were categorized into sham group, MI/RI group, and three different dosages GEN-treated groups. As the results showed, the pretreatment with GEN mitigated myocardial injury, reduced infarct volume, inhibited apoptosis, enhanced superoxide dismutase activity, and decreased malondialdehyde and myeloperoxidase activity, as well as serum creatine kinase-MB and lactate dehydrogenase levels. Moreover, GEN ameliorated MI/RI by downregulating protein expression of toll-like receptor 4, myeloid differentiation primary response 88, and p-nuclear factor-κB. In conclusion, the pretreatment of GEN may be considered as a potential therapeutic option for MI/RI.

The expression changes of high-mobility group box 1 (HMGB1) in the mouse cochlea have recently been implicated in noise-induced hearing loss, suggesting that HMGB1 participates in regulating cochlear function. However, the precise role of HMGB1 in the auditory system remains largely unclear. This study aimed to investigate its function in the developing mouse cochlea by examining the expression pattern of HMGB1 in the mouse cochlea from embryonic day (E) 18.5 to postnatal day (P) 28 using double immunofluorescence on frozen sections. Our findings revealed that HMGB1 was extensively expressed in the cell nucleus across various regions of the mouse cochlea, including the organ of Corti. Furthermore, its expression underwent developmental regulation during mouse cochlear development. Specifically, HMGB1 was found to be localized in the tympanic border cells at each developmental stage, coinciding with the gradual anatomical in this region during development. In addition, HMGB1 was expressed in the greater epithelial ridge (GER) and supporting cells of the organ of Corti, as validated by the supporting cell marker Sox2 at P1 and P8. However, at P14, the expression of HMGB1 disappeared from the GER, coinciding with the degeneration of the GER into the inner sulcus cells. Moreover, we observed that HMGB1 co-localized with Ki-67-positive proliferating cells in several cochlear regions during late embryonic and early postnatal stages, including the GER, the tympanic border cells, cochlear lateral wall, and cochlear nerves. Furthermore, by dual-staining Ki-67 with neuronal marker TUJ1 and glial marker Sox10, we determined the expression of Ki-67 in the neonatal glial cells. Our spatial-temporal analysis demonstrated that HMGB1 exhibited distinct expression patterns during mouse cochlear development. The co-localization of HMGB1 with Ki-67-positive proliferating cells suggested that HMGB1 may play a role in cochlear development.

This study aimed at exploring the expression and clinical significance of aromatase P450, adhesion molecule CD24 and HER2/neu in endometrial cancer. The expression of aromatase P450, adhesion molecule CD24 and HER2/neu was detected by immunohistochemistry in 15 cases of endometrial hyperplasia group, 50 cases of endometrial adenocarcinoma and 3 cases of uterine papillary adenocarcinoma, with 15 cases of normal endometrium as control group. We detected no expression of aromatase P450, adhesion molecule CD24 or HER2/neu in control group. Aromatase P450 positive expression rate was 66.7% in endometrial hyperplasia group and 70.3% in endometrial carcinoma group, without significant difference (p>0.05). There was no significant difference (p>0.05) in the positive expression rate of aromatase P450 between different myometrial invasion groups of endometrial adenocarcinomas. CD24 positive expression rate was 40.0% in endometrial hyperplasia group and 79.6% in endometrial carcinoma group, with significant difference (p<0.05). HER2/neu positive expression rate was 26.7% in the endometrial hyperplasia group and 57% in endometrial carcinoma group, with significant difference (p<0.05). In conclusion, aromatase P450 may be one factor associated with endometrial cancer cell proliferation, while CD24 and HER2/neu may be important factors associated with the invasion and metastasis of endometrial cancer.

Lots of adrenergic receptors (ARs) are widely present across the auditory pathways and are positioned to affect auditory and vestibular functions. However, noradrenergic regulation in the cochlea has not been well characterized. In this study, a rat model of noise-induced hearing loss was developed to investigate the expression of α2A-adrenergic receptor (AR) after acoustic trauma, then, we investigated the expression of α2A-AR in the developing rat cochlea using immunofluorescence, qRT-PCR, and Western blotting. We found that the expression of α2A-AR significantly increased in rats exposed to noise compared with controls. Immunofluorescence analysis demonstrated that α2A-AR is localized on hair cells (HCs), spiral ganglion neurons (SGNs), and the stria vascularis (SV) in the postnatal developing cochlea from post-natal day (P) 0 to P28. Furthermore, we observed α2A-AR mRNA reached a maximum level at P14 and P28 when compared with P0, while no significant differences in α2A-AR protein levels at the various stages when compared with P0. This study provides direct evidence for the expression of α2A-AR in HCs, SGNs, and the SV of the cochlea, indicating that norepinephrine might play a vital role in hearing function within the cochlea through α2A-AR.

Quercetin (Que) has been proven to enhance the chemosensitivity of multiple cancers, including colon cancer (CC). However, whether the combination of Que and 5-fluorouracil (5-FU) has a synergistic effect on drug-resistant CC cells has not previously been reported. The effect of Que (5 and 10 μg/mL) on cell vitality and apoptosis of CC and CC drug-resistant cells was examined using a cell counting kit-8 (CCK-8) and flow cytometry. After cells were treated with 5-FU (10, 40 μg/mL), Que (10 μM, 40 μM), or 5-FU in combination with Que, cell proliferation, apoptosis, oxidative stress-related factors, reactive oxygen species (ROS), and nuclear factor erythroid 2-related factor (Nrf2)/heme oxygenase-1 (HO-1) pathway-related factors were examined by colony formation assay, flow cytometry, ELISA, ROS kit, immunofluorescence assay, and Western blot. The results showed that 5-FU reduced cell viability and induced apoptosis of CC as well as 5-FU-resistant CC cells. Que further restrained the proliferation, oxidative stress-related factors (SOD, CAT, GPx, and GR), ROS production, and induced apoptosis in CC cells and 5-FU-resistant CC cells induced by 5-FU. Moreover, the combination of Que and 5-FU attenuated the Nrf2/HO-1 pathway-related marker levels in CC cells and 5-FU-resistant CC cells. Therefore, our results suggest that Que reverses 5-FU resistance in CC cells via modulating the Nrf2/HO-1 pathway.

Lung cancer originating from the bronchial epithelium is the most common lung malignancy. It has been reported that programmed cell death 1 ligand 1 (PD-L1) and tumor-associated macrophages are closely related to the development of lung cancer. However, whether tumor-derived exosomal PD-L1 could mediate the regulation of macrophage polarization in lung cancer remains unclear. For this research, the level of PD-L1 in normal tissues and lung cancer tissues was evaluated using RT-qPCR. Next, the apoptosis of lung cancer cells was evaluated using flow cytometry assay. Then, the structure and morphology of vesicles were observed using transmission electron microscopy and nanoparticle tracking analysis. Later on, the internalization of exosomes by macrophage was observed using fluorescence microscopy. Our results showed that the level of PD-L1 was upregulated in tumor tissues and lung cancer cells. Knockdown of PD-L1 notably inhibited the viability, migration and invasion of lung cancer cells. In addition, lung cancer cells-derived exosomal PD-L1 could be absorbed by macrophages. Meanwhile, exosomal PD-L1 was able to promote macrophages M2 polarization. Moreover, macrophages M2 polarization induced by exosomal PD-L1 further remarkably promoted the viability, migration, invasion, and epithelial-mesenchymal transition process of lung cancer cells. Collectively, knockdown of PD-L1 notably inhibited the viability, migration and invasion of lung cancer cells. Tumor cell-derived exosomal PD-L1 could promote the growth of lung cancer cells by mediating macrophages M2 polarization. Thus, inhibiting macrophages M2 polarization might be a promoting therapy for the treatment of lung cancer.