Objective To investigate the changes in eosinophil counts and the activation of microglial cells in the brain tissues of mice at different stages of Angiostrongylus cantonensis infection, and to examine the role of microglia in regulating the progression of angiostrongyliasis and unravel the possible molecular mechanisms. Methods Fifty BALB/c mice were randomly divided into the control group and the 7-d, 14-d, 21-day and 25-d infection groups, of 10 mice in each group. All mice in infection groups were infected with 30 stage III A. cantonensis larvae by gavage, and animals in the control group was given an equal amount of physiological saline. Five mice were collected from each of infection groups on days 7, 14, 21 d and 25 d post-infection, and 5 mice were collected from the control group on the day of oral gavage. The general and focal functional impairment was scored using the Clark scoring method to assess the degree of mouse neurological impairment. Five mice from each of infection groups were sacrificed on days 7, 14, 21 d and 25 d post-infection, and 5 mice from the control group were sacrificed on the day of oral gavage. Mouse brain tissues were sampled, and the pathological changes of brain tissues were dynamically observed using hematoxylin and eosin (HE) staining. Immunofluorescence staining with eosinophilic cationic protein (ECP) and ionized calcium binding adaptor molecule 1 (Iba1) was used to assess the degree of eosinophil infiltration and the counts of microglial cells in mouse brain tissues in each group, and the morphological parameters of microglial cells (skeleton analysis and fractal analysis) were quantified by using Image J software to determine the morphological changes of microglial cells. In addition, the expression of M1 microglia markers Fcγ receptor III (Fcgr3), Fcγ receptor IIb (Fcgr2b) and CD86 antigen (Cd86), M2 microglia markers Arginase 1 (Arg1), macrophage mannose receptor C-type 1 (Mrc1), chitinase-like 3 (Chil3), and phagocytosis genes myeloid cell triggering receptor expressed on myeloid cells 2 (Trem2), CD68 antigen (Cd68), and apolipoprotein E (Apoe) was quantified using real-time quantitative reverse transcription PCR (RT-qPCR) assay in the mouse cerebral cortex of mice post-infection. Results A large number of A. cantonensis larvae were seen on the mouse meninges surface post-infection, and many neuronal nuclei were crumpled and deeply stained, with a large number of bleeding points in the meninges. The median Clark scores of mouse general functional impairment were 0 (interquartile range, 0), 0 (interquartile range, 0.5), 6 (interquartile range, 1.0), 14 (interquartile range, 8.5) points and 20 (interquartile range, 9.0) points in the control group and the 7-d, 14-d, 21-d and 25-d groups, respectively (H = 22.45, P < 0.01), and the median Clark scores of mouse focal functional impairment were 0 (interquartile range, 0), 2 (interquartile range, 2.5), 7 (interquartile range, 3.0), 18 (interquartile range, 5.0) points and 25 (interquartile range, 6.5) points in the control group and the 7-d, 14-d, 21-d and 25-d groups, respectively (H = 22.72, P < 0.01). The mean scores of mice general and focal functional impairment were all higher in the infection groups than in the control group (all P values < 0.05). Immunofluorescence staining showed a significant difference in the eosinophil counts in mouse brain tissues among the five groups (F = 40.05, P < 0.000 1), and the eosinophil counts were significantly higher in mouse brain tissues in the 14-d (3.08 ± 0.78) and 21-d infection groups (5.97 ± 1.37) than in the control group (1.00 ± 0.28) (both P values < 0.05). Semi-quantitative analysis of microglia immunofluorescence showed a significant difference in the counts of microglial cells among the five groups (F = 17.66, P < 0.000 1), and higher Iba1 levels were detected in mouse brain tissues in 14-d (5.75 ± 1.28), 21-d (6.23 ± 1.89) and 25-d infection groups (3.70 ± 1.30) than in the control group (1.00 ± 0.30) (all P values < 0.05). Skeleton and fractal analyses showed that the branch length [(162.04 ± 34.10) μm vs. (395.37 ± 64.11) μm; t = 5.566, P < 0.05] and fractal dimension of microglial cells (1.30 ± 0.01 vs. 1.41 ± 0.03; t = 5.266, P < 0.05) were reduced in mouse brain tissues in the 21-d infection group relative to the control group. In addition, there were significant differences among the 5 groups in terms of M1 and M2 microglia markers Fcgr3 (F = 48.34, P < 0.05), Fcgr2b (F = 55.46, P < 0.05), Cd86 (F = 24.44, P < 0.05), Arg1 (F = 31.18, P < 0.05), Mrc1 (F = 15.42, P < 0.05) and Chil3 (F = 24.41, P < 0.05), as well as phagocytosis markers Trem2 (F = 21.19, P < 0.05), Cd68 (F = 43.95, P < 0.05) and Apoe (F = 7.12, P < 0.05) in mice brain tissues. Conclusions A. cantonensis infections may induce severe pathological injuries in mouse brain tissues that are characterized by massive eosinophil infiltration and persistent activation of microglia cells, thereby resulting in progressive deterioration of neurological functions.

Objective To investigate the capillarization of liver sinusoidal endothelial cells (LSECs) and its association with hepatic fibrosis during the development of alveolar echinococcosis, so as to provide the basis for unraveling the mechanisms underlying the role of LSEC in the development and prognosis of hepatic injuries and hepatic fibrosis caused by alveolar echinococcosis. Methods Forty C57BL/6 mice at ages of 6 to 8 weeks were randomly divided into a control group and 1-, 2- and 4-week infection groups, of 10 mice in each group. Each mouse in the infection groups was intraperitoneally injected with 2 000 Echinococcus multilocularis protoscoleces, while each mouse in the control group was given an equal volume of phosphate-buffered saline using the same method. All mice were sacrificed 1, 2 and 4 weeks post-infection and mouse livers were collected. The pathological changes of livers were observed using hematoxylin-eosin (HE) staining, and hepatic fibrosis was evaluated through semi-quantitative analysis of Masson’s trichrome staining-positive areas. The activation of hepatic stellate cells (HSCs) and extracellular matrix (ECM) deposition were examined using immunohistochemical staining of α-smooth muscle actin (α-SMA) and collagen type I alpha 1 (COL1A1), and the fenestrations on the surface of LSECs were observed using scanning electron microscopy. Primary LSECs were isolated from mouse livers, and the mRNA expression of LSEC marker genes Stabilin-1, Stabilin-2, Ehd3, CD209b, GATA4 and Maf was quantified using real-time fluorescence quantitative PCR (qPCR) assay. Results Destruction of local liver lobular structure was observed in mice 2 weeks post-infection with E. multilocularis protoscoleces, and hydatid cysts, which were surrounded by granulomatous tissues, were found in mouse livers 4 weeks post-infection. Semi-quantitative analysis of Masson’s trichrome staining showed a significant difference in the proportion of collagen fiber contents in mouse livers among the four groups (F = 26.060, P < 0.001), and a higher proportion of collagen fiber contents was detected in mouse livers in the 4-week infection group [(11.29 ± 2.58)%] than in the control group (P < 0.001). Immunohistochemical staining revealed activation of a few HSCs and ECM deposition in mouse livers 1 and 2 weeks post-infection, and abundant brown-yellow stained α-SMA and COL1A1 were deposited in the lesion areas in mouse livers 4 weeks post-infection, which spread to surrounding tissues. Semi-quantitative analysis revealed significant differences in α-SMA (F = 7.667, P < 0.05) and COL1A1 expression (F = 6.530, P < 0.05) in mouse levers among the four groups, with higher α-SMA [(7.13 ± 3.68)%] and COL1A1 expression [(13.18 ± 7.20)%] quantified in mouse livers in the 4-week infection group than in the control group (both P values < 0.05). Scanning electron microscopy revealed significant differences in the fenestration frequency (F = 37.730, P < 0.001) and porosity (F = 16.010, P < 0.001) on the surface of mouse LSECs among the four groups, and reduced fenestration frequency and porosity were observed in the 1-[(1.22 ± 0.48)/μm² and [(3.05 ± 0.91)%] and 2-week infection groups [(3.47 ± 0.10)/μm² and (7.57 ± 0.23)%] groups than in the control group (all P values < 0.001). There was a significant difference in the average fenestration diameter on the surface of mouse LSECs among the four groups (F = 15.330, P < 0.001), and larger average fenestration diameters were measured in the 1-[(180.80 ± 16.42) nm] and 2-week infection groups [(161.70 ± 3.85) nm] than in the control group (both P values < 0.05). In addition, there were significant differences among the four groups in terms of Stabilin-1 (F = 153.100, P < 0.001), Stabilin-2 (F = 57.010, P < 0.001), Ehd3 (F = 31.700, P < 0.001), CD209b (F = 177.400, P < 0.001), GATA4 (F = 17.740, P < 0.001), and Maf mRNA expression (F = 72.710, P < 0.001), and reduced mRNA expression of Stabilin-1, Stabilin-2, Ehd3, CD209b, GATA4 and Maf genes was quantified in three infection groups than in the control group (all P values < 0.001). Conclusions E. multilocularis infections may induce capillarization of LSECs in mice, and result in a reduction in the expression of functional and phenotypic marker genes of LSECs, and capillarization of LSECs occurs earlier than activation of HSC and development of hepatic fibrosis.

This article explores the standardized management of colorectal polyps, including classification, treatment, follow-up, and preventive control. Corresponding treatment strategies, including endoscopic resection and surgical intervention, are employed for different types of polyps. Currently, there is debate over whether to choose endoscopic resection or surgical intervention for malignant polyps at pT1 stage. Drawing on the latest literature and guidelines, the article elaborates on polyp classification, treatment modalities, follow-up, and preventive measures. Standardized management of colorectal polyps is important for reducing the incidence of colorectal cancer and improving the cure rate of early-stage colorectal cancer.

This article explores the standardized management of colorectal polyps, including classification, treatment, follow-up, and preventive control. Corresponding treatment strategies, including endoscopic resection and surgical intervention, are employed for different types of polyps. Currently, there is debate over whether to choose endoscopic resection or surgical intervention for malignant polyps at pT1 stage. Drawing on the latest literature and guidelines, the article elaborates on polyp classification, treatment modalities, follow-up, and preventive measures. Standardized management of colorectal polyps is important for reducing the incidence of colorectal cancer and improving the cure rate of early-stage colorectal cancer.

Objective·To investigate the impact of KH-type splicing regulatory protein(KHSRP)on the proliferation of prostate cancer cells and the regulation of downstream gene expression,and explore the potential role and mechanism of KHSRP in the transition of prostate cancer from androgen-dependent to androgen-independent.Methods·Recombinant lentivirus was used to infect androgen-dependent prostate cancer LNCaP cells and androgen-independent prostate cancer DU 145 cells to establish stable cell lines with functional deficiency/acquisition of KHSRP,and the functional differences of KHSRP between the two cell types were compared.Western blotting was used to detect the expression levels of KHSRP,androgen receptor(AR),and ankyrin 3(ANK3)in the stable cell lines.Cell proliferation assay,colony formation assay,and mouse xenograft assay were performed to assess the impact of KHSRP on the proliferation ability of LNCaP cells.RNA sequencing(RNA-seq)was performed to identify downstream genes affected by KHSRP,and the mRNA expression levels of these genes were measured by quantitative real-time PCR(RT-qPCR).The expression of ANK3 and KHSRP in prostate tissue samples and the difference of ANK3 expression in different prostate cancer cell lines were analyzed by combining Cancer Cell Line Encyclopedia(CCLE),The Cancer Genome Atlas(TCGA)and Gene Expression Omnibus(GEO)databases.Results·GEO data analysis showed that KHSRP was upregulated in prostate cancer tissues compared to benign prostate tissues,suggesting its association with prostate tumorigenesis.Cell proliferation assay,colony formation assay,and mouse xenograft assay demonstrated a negative correlation between KHSRP expression and the proliferation ability of LNCaP cells,indicating that KHSRP can inhibit the proliferation of prostate cancer cells,with a stronger effect on LNCaP cells than on DU 145 cells.Western blotting and RT-qPCR analysis of the stable LNCaP cell lines showed that KHSRP overexpression led to a decrease in AR protein level without affecting its mRNA level,suggesting that KHSRP can indirectly regulate AR protein level.RNA-seq and RT-qPCR results showed KHSRP was positively correlated with the expression of ANK3,a regulatory factor affecting AR protein stability,and subsequent Western blotting confirmed an increase in ANK3 protein expression after KHSRP overexpression.TCGA data analysis further supported the correlation between KHSRP and ANK3 mRNA expression.Interestingly,according to CCLE and GEO data,the expression of ANK3 was closely related to prostate cancer malignancy.Conclusion·KHSRP may indirectly regulate AR protein stability through ANK3,thereby influencing the proliferation of androgen-dependent prostate cancer cells and mediating the altered responsiveness to androgen in prostate cancer cells.

AIM:To explore the mechanism by which over-expression of enhancer of zeste homolog 2 (EZH2) in a panel of gastric cancer cell lines is involved in tumorigenesis of gastric cancer .METHODS: Real-time PCR and Western blot were employed to examine the mRNA and protein levels of EZH 2, respectively.MTS assay, cell migration and soft agar assay were performed to investigate the role of EZH 2 in the regulation of stomach cancer behaviors .The effect of EZH2 on NF-κB target gene expression was determined by Luciferase reporter and real-time PCR.Co-immunoprecipitati-on was used to analyze the interaction of EZH 2 and p65 in HEK293T cells.RESULTS: The expression levels of EZH2 were significantly increased in the gastric cancer cells compared with normal gastric epithelial cells .Pharmacological inhibi-tion by DZNep or knockdown of EZH2 significantly compromised AGS and SNU-16 cell activity , cell migration and anchor-age-independent cell growth.Moreover, siRNA knockdown of EZH2 impaired NF-κB downstream targets, such as IL-8, CXCL5 and CCL20.In addition, the interaction of EZH2 and p65 was detected.CONCLUSION: EZH2 mediates the growth of gastric cancer cells through the regulation of NF-κB downstream gene expression .