Acta histochemica最新文献

Pterygium is a common eye surface disease with high recurrence and unclear pathogenesis. In current study, RNA sequencing was conducted in 6 pairs of human pterygium and conjunctival tissues, and Matr3 as a novel candidate gene was significantly reduced in pterygium compared to control tissues. Moreover, immunoprecipitation was performed to pull down MATR3, and WTAP specially interacting with MATR3 in control but not pterygium was identified by mass spectrum. Immunoprecipitation was performed to validate the interaction between MATR3 and WTAP/METTL3/METTL14 complex. (Methylated) RNA immunoprecipitation was performed to further reveal that the binding affinity of WTAP and MATR3 was lost at 3′ UTR of RNA molecules of down-regulated genes in pterygium. Overall, we figured out the loss of intercrossing between MATR3 and N6-methyladenosine methyltransferase complex, as well as indicated the potential impact on transcription of target genes in pterygium.

Background

Adipose mesenchymal stem cell (AMSC)-derived exosomes are promising novel factors for wound repair and regeneration. This study aimed to explore the potential roles and underlying mechanisms of specific miRNA in wound healing using AMSC-derived exosomes as carriers.

Methods

The expression profiles of GSE197840 were downloaded to screen for differentially expressed miRNAs (DEmiRNAs), and the corresponding genes of the identified miRNAs were predicted. Next, miRNA-mRNA co-expression networks were constructed and the genes in these networks were subjected to functional analysis. miR-223–3p overexpressed AMSCs were then established to isolate exosomes, and the effects of AMSC-derived exosomes carrying miR-223–3p on wound healing and the related potential mechanisms were further investigated in vivo.

Results

35 DEmiRNAs were identified and a co-expression network containing 22 miRNAs and 91 target genes was constructed. Based on the network, miR-223–3p was the hub node and the genes were significantly enriched in 15 GO terms of biological processes and 14 KEGG pathways, including cAMP, PI3K-Akt, cGMP-PKG, neurotrophin signaling pathway, and dopaminergic synapse. Then, miR-223–3p overexpressed AMSCs-derived exosomes were successfully extracted, and miR-223–3p was found to directly bind with MAPK10. In vivo experiments validated that AMSCs-derived exosomal miR-223–3p could promote wound healing, and up-regulated α-SMA, CD31, COL1A1, COL2A1, COL3A1, and down-regulated MAPK10, TNF-α, IL-β, and IL-6.

Conclusions

AMSC-derived exosomal miR-223–3p may accelerate wound healing by targeting MAPK10.

Objective

Idiopathic pulmonary fibrosis (IPF) is considered as a chronic interstitial lung disease with underlying mechanism of IPF remaining unclear, while there are no definitive treatment options. In recent years, scientists have gradually paid attention to the influence of angiogenesis on IPF. Because IPF is a progressive with microvascular remodeling disorder, scientists have postulated that angiogenesis may also be one of the initiating and contributing factors of the disease. Bupleurum is a common natural Chinese herbal medicine with antibacterial, anti-inflammatory, anti-tumor and other pharmacological effects. As the most important active monomer of Bupleurum, Saikosaponin-d (SSd) is a new discovery with anti-pulmonary fibrosis (PF) activity. This study attempts to investigate the role of SSd in the interference of PF through regulation of angiogenesis in IPF through Angiopoietin (Angpt) /Tie receptor 2 (Tie2) pathway.

Methods

Randomly, we allocated C57BL/6 mice into four groups (n = 20 in each group). Afterwards, establishment of IPF model was accomplished via intratracheal administration of bleomycin (BLM, 5 mg/kg), while corresponding drug intervention was given accordingly. On 3rd, 7th, 14th and 28th days after modeling, we performed histopathological examination through staining. Meanwhile, immunohistochemistry (IHC) of PF and the expression of related factors were observed, while Ang/Tie2 pathway was assessed by ELISA with the effect of SSd on angiogenesis related proteins in IPF being explored with IHC and Western Blot technique.

Results

Our results showed that SSd could reduce inflammation and PF levels in lung tissue of experimental mice, while levels of angiogenesis-related factors, namely Tie-2, Ang-1 and ANGPT2 (Ang-2), fibrosis- associated factors like Alpha-smooth muscle actin (α-SMA), collagen-I and hydroxyproline in SSd and dexamethasone (DXM) mice were significantly reduced at each time point compared to BLM (p < 0.01). Additionally, we discovered substantial decreased expressions of Ang-1, Ang-2, Tie-2, α-SMA and collagen-I at protein level in SSd and DXM mice at each time point compared to BLM (p < 0.05). Besides, insignificant differences were observed between SSd and DXM groups (p > 0.05).

Conclusion

This study has demonstrated that SSd could down-regulate the expression of ANG-1, Ang-2 and Tie2 in the Ang/Tie2 pathway, and may reduce lung inflammation and PF in BLM-induced mice via inhibition of angiogenesis.

Purpose

Telocytes (TCs), a novel type of stromal cells found in tissues, induce macrophage differentiation into classically activated macrophages (M1) types and enhance their phagocytic function. The purpose of this study was to investigate the inhibitory effects of TC-induced M1 macrophages on endometriosis (EMs).

Methods

mouse uterine primary TCs and endometrial stromal cells (ESCs) were isolated and identified using double immunofluorescence staining. For the in vitro study, ESCs were treated with TC-induced M1 macrophages, and the vascular endothelial growth factor (VEGF), matrix metalloproteinase 9 (MMP9), and nuclear factor kappa B (NF-κb) genes were identified by quantitative real-time PCR (qRT-PCR) or western blotting (WB). For the in vivo study, an EMs mouse model received TC-conditioned medium (TCM) via abdominal administration, and characterized the inhibitory effects on growth (lesion weight, volume, and pathology), tissue-resident macrophages differentiation by immunostaining, angiogenic capacity (CD31 and VEGF), invasive capacity (MMP9), and NF-κb expression within EMs lesions.

Results

immunofluorescent staining showed that uterine TCs expressed CD34+ and vimentin+, whereas ESCs expressed vimentin+ and cytokeratin-. At the cellular level, TC-induced M1 macrophages can significantly inhibit the expression of VEGF and MMP9 in ESCs through WB or qRT-PCR, possibly by suppressing the NF-κb pathway. The in vivo study showed that macrophages switch from the alternatively activated macrophages (M2) in untreated EMs lesions to the M1 subtype after TCM exposure. Thereby, TC-induced M1 macrophages contributed to the inhibition of EMs lesions. More importantly, this effect may be achieved by suppressing the expression of NF-κb to inhibit angiogenesis (CD31 and VEGF) and invasion (MMP9) in the tissue.

Conclusion

TC-induced M1 macrophages play a prevailing role in suppressing EMs by inhibiting angiogenic and invasive capacity through the NF-κb pathway, which provides a promising therapeutic approach for EMs.

The ability to differentiate into cells of different lineages, such as bone cells, is the principal value of adult mesenchymal stem cells (MSCs), which can be used with the final aim of regenerating damaged tissue. Due to its potential use and importance in regenerative medicine and tissue engineering, several questions have been raised regarding the molecular mechanisms of MSC differentiation. As one of the crucial mediators in organism development, the transforming growth factor-beta (TGF-β) superfamily directs MSCs’ commitment to selecting differentiation pathways. This review aims to give an overview of the current knowledge on the mechanisms of the TGF-β superfamily in MSCs bone differentiation, with additional insight into the mutual regulation of microRNAs and TGF-β in osteogenesis.

Objectives

Cholestatic liver diseases are characterized by hepatocellular damage, cholangiocyte proliferation, and progressive fibrosis. Bile duct ligation (BDL) is widely used to resemble liver injuries induced by cholestasis. Peroxisome proliferator-activated receptor-gamma coactivator 1 alpha (PGC1α) was reported to play a critical role in multiple biological responses. Nevertheless, whether PGC1α is involved in bile acid metabolism and biliary disorders remains unclear. This study aimed to investigate the effect of PGC1α on hepatic responses after cholestatic injury.

Materials and methods

Wild-type mice were subjected to BDL or sham surgery for 14 days and human liver specimens from patients with primary biliary cholangitis (PBC) were collected to detect the expression of PGC1α. Hepatic-specific PGC1α knockout mice (HKO) were constructed and subjected to BDL, in which the effects of PGC1α on cholestatic liver injury were demonstrated by biochemical and histopathological assessments, immunoblotting, and metabolomics.

Results

The expression of PGC1α was upregulated in the liver of PBC patients and murine models. Both in vivo and in vitro experiments supported the protective effects of PGC1α on cholestasis-induced hepatocyte injury. Infiltrated inflammatory cells after BDL were decreased in HKO mice. Inhibited Wnt/β-Catenin pathway and enhanced Notch signaling promoted transdifferentiation of hepatic progenitor cells (HPC)/ hepatocytes into cholangiocytes, leading to the greater ductular reaction observed in the HKO mice. But bile acids metabolism and mitochondrial function were not affected due to hepatic PGC1α deficiency in cholestasis.

Conclusions

Hepatic-specific deletion of PGC1α regulated liver regeneration by promoting ductular reactions, thereby exerting protective effects against BDL-induced liver injury, which could be a new potential therapeutic target.

Germinal centers (GCs) are some of the most important structures in the human immune system. As such, their cell types and functions have been thoroughly investigated. B cells, T cells, follicular dendritic cells (FDCs), and macrophages have widely been found to typically be aggregated in GCs. However, the amount of space occupied by each of these cell types has yet to be investigated. In this study, we conducted confocal laser-based 3D cell-volume quantification of typical GC cells under reactive conditions in lymphadenitis and investigated how volume proportions change during GC development. For this investigation, we used anti-CD3 (T cells), anti-CD20 and anti-Pax5 (B cells), anti-CD23 (FDCs), anti-CD68 (macrophages), and DAPI (nuclear staining). We detected average proportions of about 11% CD3, 9% CD20, 6% CD23, and 2% CD68 in the largest possible regions of interest within GCs. Interestingly, these values remained steady relatively independent of GC size. The remarkably low B cell proportion can be attributed to technical constraints given the use of the CD20 antibody in 3D. Applying the B cell marker Pax5, we found that about 44% of the volume was occupied by B cells after extrapolating the volume of B cell nuclei to that of whole B cells. We concluded that Pax5 is more suitable than anti-CD20 for 3D B cell quantification in GCs. The substantial unstained volume in GCs raises the question of whether other cell types fill these open spaces. Our 3D investigation enabled a unique morphological and volumetric evaluation of GC cells that balance their overall volumes in GCs.

We demonstrate herein a refined method to evaluate the migration capacity of monolayer cells using the CellProfiler pipeline. We used MDA-MB-231 cells, a triple-negative breast cancer cell line, as a model to perform the wound healing assay and proceeded with the pipeline analysis. In order to see a contrast in our analysis of cell migration, we treated the cells with 10 µM kartogenin for 48 h and compared the result to the control cells treated with 0.1 % dimethyl sulfoxide (DMSO). The migration rate of MDA-MB-231 cells could be measured precisely using this method whereby in the presence of 10 µM kartogenin, the cells migrated at 6.3 ± 1.7µmh⁻¹ whilst the vehicle control migrated at 9.1 ± 3.2 µmh⁻¹ (p < 0.05). The small changes in the rate of migration could be significantly differentiated and we believe that this method is accurate in analyzing data of scratch assays as it is of high precision and therefore can be used for high-throughput screenings.

In maize, immunoprecipitation assays have shown that CycD2;2 interacts with KRPs. However, evidence on CycD2;2 or KRPs localization and their possible interaction in specific tissues is lacking and its physiological consequence is still unknown. This work explores the spatiotemporal presence of CyclinD2s and KRPs, cell cycle regulators, during maize seed germination (18 and 36 h) after soaking on glucose or sucrose (120 mM). CyclinD2s are positive actors driving proliferation; KRPs are inhibitors of the main kinase controlling proliferation (a negative signal that slows down the cell cycle). Cell cycle proteins were analyzed by immunolocalization on longitudinal sections of maize embryo axis in seven different tissues or zones (with different proliferation or differentiation potential) and in the nucleus of their cells. Results showed a prevalence of these cell cycle proteins on embryo axes from dry seeds, particularly, their accumulation in nuclei of radicle cells. The absence of sugar caused the accumulation of these regulators in different proliferating zones. CyclinD2 abundance was reduced during germination in the presence of sucrose along the embryo axis, while there was an increase at 36 h on glucose. KRP proteins showed a slight increase at 18 h and a decrease at 36 h on both sugars. There was no correlation between cell cycle regulators/DNA co-localization on both sugars. Results suggest glucose induced a specific accumulation of each cell cycle regulator depending on the proliferation zone as well as nuclear localization which may reflect the differential morphogenetic program regarding the proliferation potential in each zone, while sucrose has a mild influence on both cell cycle proteins accumulation during germination. Whenever CycD2s were present in the nucleus, KRPs were absent after treatment with either sugar and at the two imbibition times analyzed, along the different embryo axe zones.