Thirty SPF Kunming male mice aged 6-8 weeks were randomly divided into five groups:the control group,the model group,and the low,medium,and high dose groups of Dilong.Except for the control group,the other mice were continuously injected intraperitoneally with 80 mg/kg cyclophosphamide for 3 days to prepare an immunosuppressive model.After the model-ing was completed,the control group and the model group were given an equal amount of physio-logical saline by gavage,while the low,medium,and high dose Dilong groups were given 100,200,and 400 mg/kg of Dilong fermented material by gavage,respectively.The treatment lasted for 14 days,and thymus,spleen,blood,and intestinal contents were collected 2 hours after the last admin-istration.The results showed that cyclophosphamide caused a weight loss in mice during the exper-iment,reduced thymus index,white blood cells,lymphocytes,granulocytes,immunoglobulin levels in the blood,and disrupted gut microbiota.Dilong fermented material can effectively improve the decrease in body weight and thymus index of mice,increase the levels of immune cells and immu-noglobulin in the blood,improve the diversity and richness of intestinal microbiota,and increase the abundance of beneficial bacteria.Dilong fermented can effectively alleviate the immune suppres-sion and intestinal microbiota disorder caused by cyclophosphamide in mice.Considering all fac-tors,the optimal dosage is 200 mg/kg.

Thirty SPF Kunming male mice aged 6-8 weeks were randomly divided into five groups:the control group,the model group,and the low,medium,and high dose groups of Dilong.Except for the control group,the other mice were continuously injected intraperitoneally with 80 mg/kg cyclophosphamide for 3 days to prepare an immunosuppressive model.After the model-ing was completed,the control group and the model group were given an equal amount of physio-logical saline by gavage,while the low,medium,and high dose Dilong groups were given 100,200,and 400 mg/kg of Dilong fermented material by gavage,respectively.The treatment lasted for 14 days,and thymus,spleen,blood,and intestinal contents were collected 2 hours after the last admin-istration.The results showed that cyclophosphamide caused a weight loss in mice during the exper-iment,reduced thymus index,white blood cells,lymphocytes,granulocytes,immunoglobulin levels in the blood,and disrupted gut microbiota.Dilong fermented material can effectively improve the decrease in body weight and thymus index of mice,increase the levels of immune cells and immu-noglobulin in the blood,improve the diversity and richness of intestinal microbiota,and increase the abundance of beneficial bacteria.Dilong fermented can effectively alleviate the immune suppres-sion and intestinal microbiota disorder caused by cyclophosphamide in mice.Considering all fac-tors,the optimal dosage is 200 mg/kg.

Objective To explore the molecular genetic characteristics of primary and recurrent glioblastomas (GBMs) from the same patient in vivo, primary glioma stem cells cultured in vitro, and patient-derived xenograft (PDX). Methods (1) The primary and recurrent GBM specimens from one patient during surgical resection were collected; and the expressions of glial fibrillary acidic protein (GFAP), nestin and Ki-67 were detected by immunohistochemical staining; the methylation of O6-methylguanine DNA methyltransferase (MGMT) gene, mutation of isocitrate dehydrogenase (IDH) gene and amplification of epidermal growth factor receptor (EGFR) gene were analyzed. (2) The primary and recurrent GBM stem cells were cultured in vitro and named as SU5-1 and SU5-2 cells, respectively; the expressions of nestin and CD133 were detected by immunohistochemical staining; GFAP expression was detected by immunohistochemical staining after induced differentiation, and the growth curve was detected by CCK-8 assay; Transwell invasion assay was used to detect the invasion ability; cell resistance to temozolomide (TMZ), carboplatin (CBP), cisplatin (DDP) and adriamycin (ADM) was detected by CCK-8 assay; the protein expression of programmed death receptor-ligand 1 (PD-L1) was detected by Western blotting. The rate of PD-L1 positive cells was detected by flow cytometry; genetic testing analysis was as above. (3) The primary and recurrent in situ PDX models in nude mice were established, and the expressions of nestin, GFAP and Ki-67 were detected by immunohistochemical staining. Results (1) As compared with the primary GBM, the recurrent GBM had significantly higher percentages of Ki-67 and nestin positive cells, while statistically lower percentage of GFAP positive cells (P<0.05); genetic analysis showed that there was no mutation in IDH gene in the primary GBM tissues and recurrent GBM tissues; the MGMT gene in the primary GBM tissues was methylated and EGFR gene was not amplified, while the MGMT gene in recurrent GBM tissues was demethylated and EGFR gene amplification was positive. (2) Both SU5-1 and SU5-2 cells expressed nestin and CD133, and GFAP was expressed after induced differentiation; the growth curve showed that the proliferation of SU5-2 cells started earlier than that of SU5-1 cells, the two were equal on the 3rd, 4th, and 5th d, and the proliferation of SU5-1 cells was faster than that of SU5-2 cells from the 6th d; the invasion ability of SU5-2 cells was statistically stronger than that of SU5-1 cells (P<0.05); the inhibition rates of SU5-2 cells treated with 5, 10, and 15 mmol/L CBP, 0.3125, 1.25, and 5 mmol/L DDP, 0.5 and 2 mmol/L ADM, and 125 and 500 mmol/L TMZ were significantly lower than those of SU5-1 cells treated with the same concentrations and same drugs (P<0.05); the protein expression of PD-L1 in SU5-2 cells was higher than that in SU5-1 cells; the positive rate of PD-L1 in SU5-2 cells was statistically higher than that in SU5-1 cells (P<0.05); the results of genetic analysis were consistent with those of the primary and recurrent GBM samples. (3) As compared with those in the primary PDX model, the nestin and Ki-67 expressions were significantly higher and GFAP expression was significantly lower in the recurrent PDX model (P<0.05); the results of genetic analysis were consistent with those of the primary and recurrent GBM samples. Conclusions Genetic differences are detected between primary and recurrent GBMs; recurrent GBM has stronger invasive capacity and multi-drug resistance. The primary stem cells derived from surgical specimens and corresponding PDX models could replicate the molecular genetic characteristics of original tumors, which provide a reliable experimental platform for both tumor translation researches and screening of molecular therapeutic targets.

Various types of medical devices used in assisted reproductive technologies (ART) should be detected for their safety by strict biological assays. Mouse embryo assay(MEA)has been recognized as one of the most important and standardized methods with the threshold more than 80% of blastocyst formation rate (BR) after 96 h culture of fertilized eggs. The disadvantage using BR for embryonic quality control has been concerned as it is ubiquitously dependent of embryonic morphology and the detailed data including molecular and genetic information is obviously missing and incomplete. This leads to the urgent requirement for more sensitive and efficient assessments for the quality control of ART. This study evaluated the reliability of an immunofluorescent MEA by counting total cell and differential number of the cells in the inner cell mass (ICM) and trophectoderm (TE) in the blastocyst. This method improved the traditional MEA, provided a sensitive and powerful platform to assess embryonic developmental viability and should be suggested as a standard assay to be globally used for the quality control of medical devices and pre-clinical procedures in ART.
Animals ; Blastocyst ; Embryonic Development ; Equipment Safety ; Mice ; Reproducibility of Results ; Reproductive Techniques, Assisted ; instrumentation

Objective To compare the value of assessment with DWI and contrast-enhanced MRI (CE-MRI) in activity of sacroiliitis of patients with ankylosing spondylitis(AS).Methods Ninety-six patients conforming to modified New York criteria were prospectively collectedas the AS group, and twenty-one healthy volunteers were enrolled into the control group. According to the Bath AS disease activity index (BASDAI), erythrocyte sedimentation rate and C-reaction protein, AS patients were divided into the active AS group (n=60) and the chronic AS group (n=36) . All subjects were performed with conventional MRI, DWI and CE-MRI of bilateral sacroiliac joints. The MRI manifestations were reviewed and the ADC values and signal intensity enhancement rate (ΔSI) were measured.ANOVA was performed for the comparison ofΔSI and ADC values among active AS group, chronic AS group and control group with BASDAI and lab test results as the gold standards. ROC was analyzed with ΔSI and ADC values for activity of AS and paired
samples t test was obtained to comparethe areas under the ROC ofΔSI and ADC values.Results Among 96 cases of AS patients, MRI of sacroiliac jointsshowed that 62 cases had subchondral bone edema (57 cases of active group, 5 cases of chronic group), that 11 cases had bone surface erosion(4 cases of active group, 7 cases of chronic group), that 15 cases had bone sclerosis(6 cases of active group, 9 cases of chronic group) and that 58 cases had fat deposition on the sacroiliac joints (27 cases of active group, 31 cases of chronic group). The ΔSI values of the active group, the chronic group and control group were respectively (2.51 ± 1.69)%,(1.19 ± 0.67)%and(0.75 ± 0.21)%, and the ADCvalues were(1.33 ± 0.33)× 10-3,(1.00 ± 0.43)× 10-3 and(0.38±0.13)×10-3mm2/s. There were significant differences forΔSI and ADC values among three groups (F=18.375, 16.366. P<0.01), and statistical significance ofΔSI and ADC values were found between every two groups of three(P< 0.05).The area under the ROC between ΔSI and ADC to determine activity of AS patients were respectively 0.814 and 0.730, which had nostatistical significance(t=1.632, P=0.103). The sensitivity and specificity to determine activity of AS patients byΔSI=1.44%were 81.67%and 80.00%.The sensitivity and specificity to determine activity of AS patients by ADC=1.15 × 10-3/mm2 were 76.67% and 71.43%.Conclusion DWI and CE-MRI performed equally in detecting activity of AS patients.