ABSTRACT
Abstract Objectives Accumulating research have reported that microRNAs (miRNAs) play important roles in Retinoblastoma (RB). Nonetheless, the function and underlying mechanism of miR-181a-5p in RB remain ambiguous. Methods The relative expression levels of miR-181a-5p and NRAS mRNA were detected by quantitative Reverse Transcription-Polymerase Chain Reaction (qRT-PCR). RB cell proliferation was measured using the Cell Counting Kit-8 (CCK-8) and 5′-Bromo-2′-deoxyuridine (BrdU) assays. Transwell assays and flow cytometry were performed to detect the migration, invasion, and apoptosis of RB cells. The interaction between miR-181a-5p and NRAS was explored using luciferase experiments, western blotting, and qRT-PCR. Results miR-181a-5p expression was found to be decreased in RB tissues and cell lines, and its expression was correlated with unfavorable pathological features of the patients. In vitro experiments revealed that miR-181a-5p reduced RB cell proliferation, migration, and invasion while enhancing apoptosis. Further research confirmed that NRAS is a direct target of miR-181a-5p. miR-181a-5p inhibited NRAS expression at both the mRNA and protein levels. Co-transfection of pcDNA-NRAS or NRAS small interfering RNA (siRNA) reversed the effects of miR-181a-5p mimics or miR-181a-5p inhibitors on RB cells. Conclusion miR-181a-5p was significantly downregulated during the development of RB, and it suppressed the malignant behaviors of RB cells by targeting NRAS.
ABSTRACT
Huperzine A is a potent,reversible,and blood-brain barrier permeable acetylcholinesterase irhibitor.The aim of this study was to compare the pharmacokinetics,tolerability,and bioavailability of two formulations with the established reference formulation of huperzine A in a fasting,healthy Chinese male population.This was a randomized,single-dose,3-period,6-sequence crossover study.The plasma concentrations of huperzine A were determined by liquid chromatography tandem mass spectrometry.Tolerability was assessed based on subject interview,vital sign monitoring,physical examination,and routine blood and urine tests.The mean (SD) pharmacokinetic parameters of the reference drug were Cmax,1.550 (0.528) ng/mL;t1/2,12.092 (1.898) h;AUC0-72h,17.550 (3.794) ng.h/mL.Those of the test formulation A and test formulation B were Cmax,1.412 (0.467),1.521 (0.608) ng/mL;t1/2,12.073 (2.068),12.271 (1.678) h;AUC0-72h,15.286 (3.434) ng.h/mL,15.673 (3.586) ng.h/mL.The 90％ confidence intervals for the AUC0-72h and Cmax were between 0.80 and 1.25.No adverse events were reported by the subjects or found with results of clinical laboratory test.The test and reference products met the regulatory criteria for bioequivalence in these fasting,healthy Chinese male volunteers.All three formulations appeared to be well tolerated.
ABSTRACT
To construct a lentiviral shRNA vector targeting human protein phosphatase 1D magnesium-dependent (PPM1D) gene and detect its effectiveness of gene silencing in human gliomas,specific siRNA targets with short hairpin frame were designed and synthesized.DNA oligo was cloned into the pFU-GW-iRNA lentiviral expression vector,and then PCR and sequencing analyses were conducted to verify the constructs.After the verified plasmids were transfected into 293T cells,the lentivirus was produced and the titer of virus was determined.Real-time quantitative PCR and Western blot were performed to detect the PPM1D expression level in the infected glioma cells.PCR and Western blot analyses revealed the optimal interfering target,and the virus with a titer of 6×108 TU/mL was successfully packaged.The PPM1D expression in human glioma cells was knocked down at both mRNA and protein levels by virus infection.The expression of PPM1D mRNA and protein was decreased by 76.3％ and 87.0％ respectively as compared with control group.The multiple functions of human glioma cells after PPM1D RNA interference were detected by flow cytometry and cell counting kit-8 (CCK-8).Efficient down-regulation of PPM1D resulted in significantly increased cell apoptosis and reduced cell proliferation and invasion potential in U87-MG cells.We have successfully constructed the lentiviral shRNA expression vector capable of stable PPM1D gene silencing at both mRNA and protein levels in glioma cells.And our data gave evidence that the reduced cell growth observed after PPM1D silencing in glioma cells was at least partly due to increased apoptotic cell death.
ABSTRACT
The expression of paired immunoglobulin-like receptor B(PirB)in normal and injured spinal cord of rats was investigated.The SD rat hemi-sectioned spinal cord injury(SCI)model was established.Before and 1,3,7,10 days after SCI,the spinal cord tissues were harvested,and Western blot and immunohistochemistry were used to examine the expression and location of PirB.The results showed that the expression level of PirB in the normal spinal cord of SD rats was low.At the first day after SCI,the expression of PirB was obviously increased,and that in the injured spinal cord from the first day to the 10th day was significantly higher than in the normal spinal cord.The positive expression of PirB in neurons from different regions of gray matter of the injured spinal cord was seen.It was concluded that the expression of PirB in the normal spinal cord of rats was low.The expression of PirB in SCI was significantly increased till at least the 10th day.