European Journal of Histochemistry最新文献

Stroke is a brain system disease with a high fatality rate and disability rate. About 80% of strokes are ischemic strokes. Cerebral ischemia-reperfusion injury (CIRI) caused by ischemic stroke seriously affects the prognosis of stroke patients. The purpose of this study is to investigate the effect of sufentanil (SUF) on CIRI model rats. We used middle cerebral artery occlusion (MCAO) to make the CIRI model in rats and monitored region cerebral blood flow (rCBF) to ensure that blood flow was blocked and recanalized. We used ELISA and RT-PCR to detect the expression of inflammatory factors in rat serum and brain tissue. In addition, we detected the expression of metalloproteinase (MMP) 2, MMP9 and collagen IV in brain tissues and performed Evans blue (EB) assay to determine the permeability of the blood-brain barrier (BBB). Finally, we clarified the apoptosis of brain tissue through the TUNEL staining and the detection of caspase3, Bcl2 and Bax. Various concentrations of SUF, especially 5, 10 and 25 μg/kg of SUF, all alleviated the infarct size, neurological function and brain edema of MCAO rats. SUF pretreatment also effectively reduced the expression of inflammatory cytokines in MCAO rats, including interleukin (IL)-1β, IL-4, IL-6, IL-8, IL-10 and tumor necrosis factor (TNF)-α. In addition, SUF also inhibited MMP2 and MMP9 and promoted the expression of collagen IV, indicating that SUF attenuated the destruction of the BBB. SUF also inhibited caspase3 and Bax rats and promoted Bcl2 in MCAO rats, thus inhibiting cell apoptosis. SUF pretreatment effectively improved the neurological function and cerebral infarction of MCAO rats, inhibited excessive inflammation in rats, protected the BBB, and inhibited cell apoptosis in brain tissue.

Psoriasis is a chronic inflammatory skin disease. Although miRNAs are reported to be associated with the pathogenesis of psoriasis, the contribution of individual microRNAs toward psoriasis remains unclear. The miR-17-92 cluster regulates cell growth and immune functions that are associated with psoriasis. miR-17-3p is a member of miR-17-92 cluster; however, its role in dermatological diseases remains unclear. Our study aims at investigating the effects of miR-17-3p and its potential target gene on keratinocytes proliferation and secretion of pro-inflammatory cytokine and their involvement in psoriasis. Initially, we found that miR-17-3p was upregulated in psoriatic skin lesions, and bioinformatic analyses suggested that CTR9 is likely to be a target gene of miR-17-3p. Quantitative reverse-transcriptase PCR and immunohistochemical analysis revealed that CTR9 expression was downregulated in psoriatic lesions. Using dual-luciferase reporter assays, we identified CTR9 as a direct target of miR-17-3p. Further functional experiments demonstrated that miR-17-3p promoted the proliferation and pro-inflammatory cytokine secretion of keratinocytes, whereas CTR9 exerted the opposite effects. Gain-of-function studies confirmed that CTR9 suppression partially accounted for the effects of miR-17-3p in keratinocytes. Furthermore, Western blot revealed that miR-17-3p activates the downstream STAT3 signaling pathway while CTR9 inactivates the STAT3 signaling pathway. Together, these findings indicate that miR-17-3p regulates keratinocyte proliferation and pro-inflammatory cytokine secretion partially by targeting the CTR9, which inactivates the downstream STAT3 protein, implying that miR-17-3p might be a novel therapeutic target for psoriasis.

The purpose of this study was to investigate the effect of Ghrelin on the polarization of microglia/ macrophages after cerebral ischemia (CI) in rats. 60 wild-type SD rats were randomly divided into sham group, CI group, CI+Ghrelin group, 20 rats in each group. The modified Longa suture method was used to establish the middle cerebral artery occlusion (MCAO) model in rats. Before surgery, Ghrelin was injected subcutaneously (100μg/kg, twice a day) for 4 consecutive weeks. After modeling, neurological function scores were performed with three behavioral experiments: mNSS score, Corner test, and Rotarod test, to evaluate the recovery of neurological function after Ghrelin treatment. At the same time, the brain tissues were collected and stained with 2,3,5-triphenyltetrazolium chloride (TTC) to detect the cerebral infarct volume. RT-qPCR was used to detect the expression of TNF-α and IL-1β in the ischemic brain tissue, and the TUNEL staining was used to detect the apoptosis of brain tissue. Flow cytometry was used to detect the percentage of M1 type microglia/macrophages which were isolated by trypsin digestion of fresh cerebral cortex. Then, the Western blotting and immunofluorescence method were used to detect the phosphorylation level of AKT (P-AKT) and AKT. Compared with the CI group, the neurological function of the rats in the CI+Ghrelin group was dramatically improved, and the cerebral infarction area was dramatically reduced. At the same time, the expression of TNF-α and IL-1β in the ischemic brain tissue of rats in the CI+Ghrelin group decreased, and the apoptotic cells in the brain tissue also decreased. Compared with the CI treatment group, the activation of M1 microglia/macrophages in the cortex of the ischemic side of the infarct and the peri-infarct area in the CI+Ghrelin group was dramatically inhibited. At the same time, the ratio of P-AKT/AKT of the brain tissue in the CI+Ghrelin group was dramatically higher than that of the CI group. In the rat cerebral ischemia model, Ghrelin can promote the repair of brain damage and the recovery of neurological function after ischemia. Its mechanism may be related to activating AKT to selectively reduce M1 microglia/macrophages, reducing inflammation and cell apoptosis in brain tissue.

During the last two centuries, histochemistry has provided significant advancements in many fields of life sciences. After a period of neglect due to the great development of biomolecular techniques, the histochemical approach has been reappraised and is now widely applied in the field of nanomedicine. In fact, the novel nanoconstructs intended for biomedical purposes must be visualized to test their interaction with tissue and cell components. To this aim, several long-established staining methods have been re-discovered and re-interpreted in an unconventional way for unequivocal identification of nanoparticulates at both light and transmission electron microscopy.

The purpose of this work was to prove that oxidative stress is the main mechanism responsible for retinal neurodegenerative changes, subsequent apoptosis, and inflammatory cytokine release in rats fed with a high cholesterol diet (HCD) and determine the role of garlic in alleviating these changes. Forty rats were equally divided into four groups: control, garlic-treated (positive control), HCD, and HCD + garlic-treated (HCD + G). By the end of the experiment (24 weeks) blood samples were collected for assessment of serum lipid profile, oxidative stress parameters, and plasma levels of IL-6 and TNF-α. Both eyes of the rats were enucleated; one was used for light microscopic examination and the other for electron microscopic examination. There was a significant increase in the levels of serum lipids, oxidative stress parameters, IL-6 and TNF-α, and area of expression of caspase-3 in the HCD group compared to both the control and HCD + G groups. Histological examination revealed degenerative changes in all layers of the neural retina in the HCD group. Garlic administration resulted in a significant improvement in the biochemical, immunohistochemical, and histological characteristics of hypercholesterolemic rats. These findings support the hypotheses that garlic has strong antioxidant, anti-apoptotic, and anti-inflammatory properties. Garlic ameliorates the neurodegenerative changes in the neural retina of hypercholesteremic rats.

Autofluorescent granules of various sizes were observed in primary human liver endothelial cells (LSECs) upon laser irradiation using a wide range of wavelengths. Autofluorescence was detected in LAMP-1 positive vesicles, suggesting lysosomal location. Confocal imaging of freshly prepared cultures and imaging flow cytometry of non-cultured cells revealed fluorescence in all channels used. Treatment with a lipofuscin autofluorescence quencher reduced autofluorescence, most efficiently in the near UV-area. These results, combined with the knowledge of the very active blood clearance function of LSECs support the notion that lysosomally located autofluorescent material reflected accumulation of lipofuscin in the intact liver. These results illustrate the importance of careful selection of fluorophores, especially when labelling of live cells where the quencher is not compatible.

Spastin is one of the proteins which lead to hereditary spastic paraplegia (HSP), whose dysfunction towards microtubule severing and membrane transporting is critically important. The present study is to elucidate the mechanisms of the protein stability regulation of spastin. The ubiquitin encoding plasmids are transfected into COS-7 cells with different fusion tags including Green Fluorescent Protein (GFP), mCherry and Flag. The expression level of spastin was detected, microtubule severing activity and neurite outgrowth were quantified. The data showed that ubiquitin overexpression significantly induced the decreased expression of spastin, suppressed the activity of microtubule severing in COS-7 cells and inhibited the promoting effect on neurite outgrowth in cultured hippocampal neurons. Furthermore, when modulating the overexpression experiments of ubiquitin, it was found that relatively small tag like Flag, but not large tags such as GFP or mCherry fused with ubiquitin, retained the activity on spastin stability. The present study investigated the effects of small/large tags addition to ubiquitin and the novel mechanisms of post-transcriptional modifications of spastin on regulating neurite outgrowth, in the attempt to experimentally elucidate the mechanisms that control the level or stability of spastin in hereditary spastic paraplegia.

Osteosarcoma (OS), characterized by high morbidity and mortality, is the most common bone malignancy worldwide. MicroRNAs (miRNAs) play a crucial role in the initiation and development of OS. The purpose of this study was to investigate the roles of miR-1270 in OS. RT-qPCR and Western blot were applied to detect the mRNA and protein level, respectively. CCK-8, colony formation, and TUNEL assays were conducted to determine the cell viability, proliferation, and apoptosis of OS cells. Wound healing and transwell assay were performed to detect the migration and invasion ability of OS cells. Bioinformatics analysis and dual-luciferase reporter assay were used to predict the target genes of miR-1270. Tumor xenograft in vivo assay was carried out to determine miR-1270 effect on the tumor size, volume, and weight. In this study, miR-1270 was overexpressed in OS tissues and cells. However, miR-1270 knockdown inhibited the proliferation, migration and invasion, and promoted the OS cells' apoptosis. Mechanistically, cingulin (CGN) was predicted and proved to be a target of miR-1270 and partially alleviated the effects of miR-1270 on the proliferation, migration and invasion ability of OS cells. Taken together, knockdown of miR-1270 may inhibit the development of OS via targeting CGN. This finding may provide a novel therapeutic strategy for OS.

Acute lung injury/acute respiratory distress syndrome (ALI/ARDS) is a kind of diffuse inflammatory injury caused by various factors, characterized by respiratory distress and progressive hypoxemia. It is a common clinical critical illness. The aim of this study was to investigate the effect and mechanism of the Mucin1 (MUC1) gene and its recombinant protein on lipopolysaccharide (LPS)-induced ALI/ARDS. We cultured human alveolar epithelial cell line (BEAS-2B) and used MUC1 overexpression lentivirus to detect the effect of MUC1 gene on BEAS-2B cells. In addition, we used LPS to induce ALI/ARDS in C57/BL6 mice and use hematoxylin and eosin (H&E) staining to verify the effect of their modeling. Recombinant MUC1 protein was injected subcutaneously into mice. We examined the effect of MUC1 on ALI/ARDS in mice by detecting the expression of inflammatory factors and oxidative stress molecules in mouse lung tissue, bronchoalveolar lavage fluid (BALF) and serum. Overexpression of MUC1 effectively ameliorated LPS-induced damage to BEAS-2B cells. Results of H&E staining indicate that LPS successfully induced ALI/ARDS in mice and MUC1 attenuated lung injury. MUC1 also reduced the expression of inflammatory factors (IL-1β, TNF-α, IL-6 and IL-8) and oxidative stress levels in mice. In addition, LPS results in an increase in the activity of the TLR4/NF-κB signaling pathway in mice, whereas MUC1 decreased the expression of the TLR4/NF-κB signaling pathway. MUC1 inhibited the activity of TLR4/NF-κB signaling pathway and reduced the level of inflammation and oxidative stress in lung tissue of ALI mice.

Preeclampsia (PE) is one of the leading causes of maternal morbidity and mortality in pregnant women. This study aimed to investigate the potential impact and regulatory mechanisms of bone morphogenetic protein receptor 2 (BMPR2) on the progression of PE. We obtained placental tissues from pregnant women with PE and normal pregnant women, and the results showed that BMPR2 was expressed at low levels in the tissue from PE women. Genetic knockdown of BMPR2 increased the proliferation and invasion of cultured trophoblast cells, whereas its overexpression reduced these characteristics. Bioinformatics analysis and luciferase reporter gene assays confirmed that BMPR2 is a direct target of miR-21. Overexpression of a miR-21 inhibitor promoted the growth and invasiveness of trophoblast cells, whereas the opposite results were observed for the miR-21 mimic. Furthermore, miR-21 was sponged by the lncRNA MEG3, and shRNA inhibition of MEG3 reduced trophoblast cell growth and invasiveness. miR-21 was upregulated in the tissues from PE women, whereas MEG3 was downregulated, and the two were negatively correlated. Collectively, this study demonstrates that the lncRNA MEG3 acts as a sponge for miR-21, which regulates BMPR2 expression and promotes trophoblast cell proliferation and invasiveness, thereby preventing the development of PE. These findings provide novel insight into a targeted therapy that could be used to treat or prevent the development of PE.