This research is to explore the perfusion time-intensity curve parameters of a lung adenocarcinoma xenograft into nude mouse model with contrast enhanced ultrasonography (CEUS); and to investigate the angiogenesis features of tumor at different growth time. Twenty one lung adenocarcinoma xenografted nude mice were divided into three groups and inculcated with human lung adenocarcinoa. Time window for examining CEUS were respectively in 7-day, 14-day and 28-day. The perfusion parameters including rise time (RT), peak intensity (PI), area under the curve (AUC) of lung tumor were obtained on CEUS images by using off-line software Q lab. Immunohistochemically staining for CD34 was used to observe the microvessel density (MVD).The 7-day group had the highest AUC and PI; AUC and PI of 14-day and 28-day group decreased gradually (P < 0.05). RT was increased as tumor growth. In tumor with necrosis, AUC and PI of non-necrosis part were also larger than necrosis part (P < 0.05). Immunohistochemically staining for CD34 of all tumors reflected that the density of microvessels in necrosis tumor was significantly higher than those without necrosis (7.50±3.44 vs.12.44±5.74, P=0.034). Pearson correlation indicated that PI was positively related with MVD (r=0.668, P=0.008). Lung adenocarcinoma perfusion characteristic can be accessed from time-intensity curve parameters by using noninvasively and non-radiative contrast enhanced ultrasonography. Time-intensity curve parameters including AUC, PI and RT may reflect tumor angiogenesis.
ObjectiveTo observe the expression of miR-142-5p and forkhead transcription protein O subgroup 3 (FOXO3) in CD4+ T cells of experimental autoimmune uveitis (EAU) model rats, and preliminarily explore the targeting relationship between the two and the effect on EAU impact.MethodsTen Lewis rats were randomly divided into model group and control group. Rats in the model group wree induced an EAU animal model by adoptive immunization. Twenty days after immunization, CD4+ T cells were extracted from the eyeballs and draining lymph nodes of rats in the control group and model group, and divided into control group, model group, mimic-negative control (NC) group, miR-142-5p-mimic group, and small interference (si)-NC group, si-FOXO3 group for in vitro experiments. The miR-142-5p-mimic group and si-FOXO3 group were transfected with miR-142-5p-mimic and si-FOXO3, respectively. Twenty-five Lewis rats were randomly divided into model group, mimic-NC transfected group, miR-142-5p-mimic transfected group, si-NC transfected group, and si-FOXO3 transfected group. The above-mentioned in vitro experimental groups were injected with cells respectively. Slit lamp microscopy and EAU score were performed on 4, 8, 12, 16, 20 days after immunization; on 20 days after immunization, hematoxylin-eosin staining was performed for histopathological grading. Real-time fluorescence quantitative polymerase chain reaction was used to detect the relative expression of miR-142-5p and FOXO3 mRNA in CD4+ T cells and eye tissues of rats in each group, and helper T cell 17 (Th17) marker interleukin (IL)-17, IL-22, retinoic acid-related orphan receptor gamma (ROR gamma) relative expression level in the supernatant. Bioinformatics website and dual luciferase was used to predict the targeting relationship between miR-142-5p and FOXO3. One-way analysis of variance or t test was used for comparison between groups.ResultsAll rats in the model group showed symptoms of EAU to varying degrees, and the symptoms became worse with time. Compared with the control group, the relative expression of miR-142-5p mRNA in CD4+ T cells of the model group increased, and the relative expression of FOXO3 mRNA decreased. The differences were statistically significant (t=7.374, 10.423; P=0.002, 0.001). Compared with the mimic-NC group, the relative expression of miR-142-5p mRNA in the CD4+ T cells of the miR-142-5p-mimic group increased, and the difference was statistically significant (t=6.540, P=0.003). Compared with the model group, mimic-NC group, and si-NC group, the relative expression of IL-17, IL-22, and RORγ mRNA in CD4+ T cells in the miR-142-5p-mimic group and si-FOCO3 group increased significantly. The difference was statistically significant (F=26.110, 6.292, 5.269, 55.660, 10.490, 11.430; P＜0.05). Compared with the mimic-NC transfected group, the relative expression of miR-142-5p mRNA in the ocular tissues of the miR-142-5p-mimic transfected rats increased significantly, and the difference was statistically significant (t=6.690, P＜0.05). Compared with the transfected si-NC group, the relative expression of FOXO3 mRNA in the eye tissue of the transfected si-FOXO3 group was significantly decreased, and the difference was statistically significant (t=17.751, P＜0.05). Rats in the mimic-NC transfected group, miR-142-5p-mimic transfected group, si-NC transfected group, and si-FOXO3 transfected group prolonged with time after immunization, and the EAU scores showed an upward trend. The EAU score and histopathological grade of rats in the miR-142-5p-mimic transfected group were higher than those in the mimic-NC transfected group, and the difference was statistically significant (t=5.633, 6.286; P＜0.05). The EAU score and histopathological grade of the rats in the transfected si-FOXO3 group were higher than those in the transfected si-NC group, and the difference was statistically significant (t=6.852, 6.635; P＜0.05). FOXO3 has a targeting relationship with miR-142-5p.ConclusionsIn EAU rat CD4+ T cells, the expression of miR-142-5p is up-regulated, while the expression of FOXO3 is down-regulated. miR-142-5p targets the expression of FOXO3 to promote the development of Th17 cell-related inflammatory factors.
ObjectiveTo observe the expressions of miR-183 and retinal dehydrogenase 11 (RDH11) in exosomes derived from bone marrow mesenchymal stem cells (BMSC), and to preliminarily explore their targeting relationship and their effects on retinal pigment epithelial (RPE) cells. MethodsBMSC from C57BL/6 (C57) mice were isolated and cultured, and BMSC-derived exosomes were identified. BMSC were divided into blank group, simulation blank control group (mimic-NC group), miR-183 simulation group (miR-183-mimic group). C57 mice and retinal degeneration 10 (rd10) mouse RPE cells were cultured with reference to literature methods. RPE cells from rd10 mice were transfected with BMSC exosomes and co-cultured and divided into control group, exosome group, mimic-NC-exosome group (mimic-NC-exo group), miR-183-mimic-exosome group (miR-183-mimic-exo group). The relative expression levels of miR-183, RDH11 mRNA and protein in C57 mice, rd10 mice and RPE cells in each group were detected by real-time quantitative polymerase chain reaction and western blotting. The targeting relationship between miR-183 and RDH11 was analyzed by bioinformatics website and dual luciferase reporter. Cell counting kit 8 was used to detect the effect of miR-183 on BMSC exosomes on RPE cell proliferation; in situ labeling end labeling method was used to detect RPE cells apoptosis. One-way ANOVA was used to compare multiple groups. ResultsCompared with C57 mouse RPE cells, the relative expression of miR-183 in rd10 mouse RPE cells was down-regulated, and the relative expression of RDH11 mRNA was up-regulated, and the differences were statistically significant (t=5.230, 8.548; P=0.006, 0.001). Compared with the blank group and the mimic-NC group, the relative expression of miR-183 mRNA in the exosomes of the miR-183-mimics group was significantly increased (F=60.130, P＜0.05). After 24 h of co-culture, exosomes entered RPE cells. Compared with the mimic-NC-exo group, the relative expression of miR-183 mRNA in RPE cells in the miR-183-mimic-exo group was significantly increased, the proliferation ability was enhanced (t=7.311, P=0.002), and the number of apoptotic cells was decreased (F=10.949, P=0.012), and the differences were statistically significant (t=4.571, P=0.002). Bioinformatics website and dual-luciferase report confirmed that miR-183 has a targeting relationship with RDH11. Compared with the mimic-NC group, the relative expression of RDH11 mRNA and protein in the exosomes of the miR-183-mimic group was decreased, and the difference was statistically significant (t=5.361, 6.591; P=0.006, 0.003). After co-culture, compared with the control group, there was no significant difference in the relative expression of RDH11 mRNA and protein in RPE cells in the exosome group (t=0.169, 1.134; P=0.874, 0.320); The relative expressions of RDH11 mRNA and protein in RPE cells in -183-mimic-exo group were decreased, and the difference was statistically significant (t=5.554, 5.546; P=0.005, 0.005). ConclusionUp-regulation of BMSC-derived exosomal miR-183 promote the proliferation of RPE cells in vitro by targeting the expression of RDH11 and reduce the number of apoptosis.
【Abstract】 Objective Using 16-slice multi-detector row helical CT (16-slice MDCT) to investigate the value of multiplanar reformation technique (MPR) in the diagnosis of bowel obstruction. Methods Thirty patients with surgically (27 cases) or clinically (3 cases) proofed diagnosis of bowel obstruction underwent 16-slice MDCT examination of the entire abdomen. All cases had plain CT scan, while 20 cases had additional contrast-enhanced CT scan at portal venous phase. In addition to the conventional axial images, the original CT raw data were then reconstructed into both coronal and sagittal images using MPR technique. Imaging findings were analyzed on axial, MPR coronal and sagittal images. Results Among the 30 patients with bowel obstruction, there were 8 cases caused by adhesion, 7 by simple intestinal tumor, 5 by intussusception (including caused by instestinal tumor), 4 by abdominal hernia, 2 by volvulus, 1 by ileocecal abscess, 1 by stenosis of mesenteric artery，1 by retroperitoneal cyst, and 1 by carcinoma in pancreatic tail. Six patients developed intestinal ischemia or strangulation. Both axial and MPR images correctly depicted the presence of bowel obstruction. Based on CT axial view （AV）, the site and the underlying etiology of bowel obstruction were determined in 26 (86.7%) and 22 (73.3%) patients respectively, while the combination with MPR coronal and sagittal images improved the diagnostic performance to 29 (96.7%) and 27 (90.0%) patients respectively. Both axial and MPR images correctly revealed the presence of intestinal ischemia or strangulation in 5 （83.3%）patients. Conclusion MPR technique of MDCT is very useful for evaluating the site and etiology of bowel obstruction, as well as the circulation status of involved bowel loop.
ObjectiveTo investigate regulation mechanism of glypican-3 (GPC3) on Hippo signaling pathway and its effects on biological behavior of hepatocellular carcinoma Huh7 cells. MethodsShort hairpin RNAs (shRNA) targeting GPC3 and Yes-associated protein 1 (YAP1) genes were constructed. All of the shRNAs were transfected into Huh7 cells by liposome transfection in order to screen out the stable expression cell lines. The expressions of GPC3 and YAP1 in Huh7 cells were detected by PCR and Western blot in order to screen out the effective shRNAs. The proliferation, invasion, and apoptosis of Huh7 cells were detected by Edu cell proliferation assay, transwell, and flow cytometry. GPC3 shRNA transfection experiments were divided into 6 groups:non-transfection group, empty vector group, GPC3-714-shRNA group, GPC3-647-shRNA group, GPC3-1718-shRNA group, and GPC3-2134-shRNA group. YAP1 shRNA transfection experiments were divided into 6 groups:non-transfection group, empty vector group, YAP1-906-shRNA group, YAP1-1363-shRNA group, YAP1-1666-shRNA group, and YAP1-2895-shRNA group. GPC3 regulation experiments were divided into 5 groups:non-transfection group, empty vector group, GPC3-1718-shRNA group, GPC3-1718-shRNA+ rhYAP1 group, and YAP1-1666-shRNA group. Results① GPC3-1718-shRNA and YAP1-1666-shRNA plasmids were successfully constructed to silence the expressions of GPC3 and YAP1. ② The expressions of GPC3 mRNA and protein in each transfection group were significantly lower than those in the non-transfection group (P<0.05) and the empty vector group (P<0.05), while which in the GPC3-1718-shRNA group was significantly lower than those in all the other transfection groups (P<0.05). The expressions of YAP1 mRNA and protein in each transfection group were significantly lower than those in the non-transfection group and empty vector group (P<0.05), while which in the YAP1-1666-shRNA group was significantly lower than those in all the other transfection groups (P<0.05). ③ The expressions of YAP1 mRNA and protein in the GPC3-1718-shRNA group and the YAP1-1666-shRNA group were significantly lower than those in the non-transfection group (P<0.05) and the empty vector group (P<0.05), while which in the GPC3-1718-shRNA+rhYAP1 group were significantly higher than those in the GPC3-1718-shRNA group (P<0.05) and the YAP1-1666-shRNA group (P<0.05). ④ Compared with the non-transfection group and the empty vector group, the abilities of cell proliferation and invasion in the GPC3-1718-shRNA group and the YAP1-1666-shRNA group were significantly decreased, and the cell apoptosis was significantly increased (P<0.05); The cell proliferation, invasion, and apoptosis in the GPC3-1718-shRNA+rhYAP1 group were significantly improved (P<0.05). ConclusionGPC3 is likely to affect biological behavior of hepatocellular carcinoma Huh7 cells through regulation of YAP1 in Hippo signaling pathway.
ObjectiveTo investigate the high-resolution computed tomography (HRCT) signs of patients diagnosed with the coronavirus disease 2019 (COVID-19) and explore its evolution features during hospitalization.MethodsFrom January 17, 2020 to February 26, 2020, HRCT images from 15 COVID-19 patients were analyzed. All the patients had positive nucleic acid test results of SARS-CoV-2. The imaging features of initial and follow-up of each patient were reviewed and graded based on the severity of lung lesions.ResultsAmong the 15 COVID-19 patients, ground-glass opacity (GGO) was found in 14 cases. Six patients presented with consolidation and 3 with fibrosis. Five patients had multi-lobe involvement. Subpleural distribution pattern was present in 12 patients (80.0%) and peribronchovascular distribution pattern was present in 2 patients (13.3%). The severity score on HRCT images at the follow-up was significantly higher than that at the initial (4.6±3.4 vs. 3.5±2.5, P=0.018 2). Increase of random distribution pattern (5 cases) were also noted at the follow-up.ConclusionsChest HRCT of COVID-19 patients is characterized with GGO mainly distributed in subpleural areas and a rapid progression within a short time interval. HRCT could provide a sensitive monitor to observe disease progression for COVID-19 patients.
ObjectiveTo explore the dynamic changes of immune cell populations in Panc02 pancreatic cancer bearing immunocompetent mice. MethodsThe C57BL6/J mice syngeneic pancreatic cancer cell line Panc02 were subcutaneously implanted to establish the immunocompetent murine pancreatic cancer bearing model.According to the tumor size, the tumor was classified into 4 stages, named T1-T4, respectively.The flow cytometry was performed to identify the dynamic changes of different cell populations, such as inflammatory cells (CD45+), T helper (Th) lymphocytes, cytotoxic T lymphocytes (CTL), B lymphocytes, granulocytes, macrophages, dendritic cells (DC), natural killer (NK) cells, and natural killer T (NKT) cells in the peripheral blood and tumor tissue. ResultsThree dynamic types of immune cells with the tumor progression were identified:consistent increase, consistent decrease, increase and then decrease.①In peripheral blood:The proportion of the Th lymphocytes, CTL, and B lymphocytes consistently decreased; The proportion of granulocytes consistently increased; The proportion of the DC, macrophages, NK cells, and NKT cells increased from T1 to T3 stage but sharply decreased at T4 stage.②In tumor tissue:The intratumoral CD45+ cells consis-tently increased; The proportion of the granucolyte, macrophages, and DC consistently increased; The proportion of the Th lymphocytes and the CTL consistently decreased; The proportion of the B lymphocytes did not change significantly; The proportion of the NK cells or NKT cells increased from T1 to T3 stage but sharply decreased at T4 stage. ConclusionWith pancreatic cancer progression, the immune cell populations show different dynamic change models, which imply their important roles in predicting the prognosis and the integrated treatments of pancreatic cancer.