This paper introduces the development of the MEMS at home and abroad,besides the key technology,the application in medicine and military about the MEMS also expounded in detail.

Guanylate binding protein-1(GBP-1) is an interferon-induced protein. To observe its antiviral effect against Hepatitis B virus (HBV) and Coxsackie virus B3 (CVB3), we constructed an eukaryotic expression vector of human GBP-1(hGBP-1). Full-length encoding sequence of hGBP-1 was amplified by long chain RT-PCR and inserted into a pCR2.1 vector, then subcloned into a pCDNA3.1(-) vector. Recombinant hGBP-1 plasmids and pHBV1.3 carrying 1.3-fold genome of HBV were contransfected into HepG2 cells, and inhibition effect of hGBP-1 against HBV replication was observed. Hela cells transfected with recombinant hGBP-1 plasmids were challenged with CVB3, and viral yield in cultures were detected. The results indicated that recombinant eukaryotic expression plasmid of hGBP-1 was constructed successfully and the hGBP-1 gene carried in this plasmid could be efficiently expressed in HepG2 cells and Hela cells. hGBP-1 inhibit CVB3 but not HBV replication in vitro. These results demonstrate that hGBP-1 mediates an antiviral effect against CVB3 but not HBV and perhaps plays an important role in the interferon-mediated antiviral response against CVB3.

To establish a replication cellular model of hepatitis B virus (HBV) and determine its application in antiviral drug evaluation,we constructed an expression plasmid which contained 1.3 copies of the HBV genome,and measured the level of viral replication after transient transfection in Huh7 cells.We then observed the effect of antiviral drug administration.1.3 fold of the HBV(ayw) gene fragment was cloned into pCR2.1 by PCR and restriction endonuclease digestion.The recombinant plasmid was trans ient transfected into Huh7 cells,HBsAg,HBeAg and HBV DNA in supernatant of Huh7 cells were measured by ELISA and real-time PCR respectively; intracellular HBV replicative intermediates and intracellular HBV transcripts were detected by Southern blot and Northern blot respectively.The antiviral effect of adefovir,a novel anti-HBV nucleotide analogue,was evaluated in this cellular model system.The results indicated that a recombinant plasmid of HBV replicon was constructed successfully; the HBV genome carried in plasmid pHBV1.3 could efficiently replicate and be expressed in Huh 7 cells,adefovir could inhibit HBV replication in this cellular model,and the inhibition was dosage-dependent.The conclusion is HBV replicon,which can initiate viral replication efficiently in hepatoma cells,may be a useful tool in the study of HBV replication and antiviral drug.

OBJECTIVETo investigate the inhibitory effects of CaMKIIN on acute myeloid leukemia cell line HL-60 to explore a novel therapeutic target of leukemia.

METHODSHuman CaMK II N gene expression vector pcDNA3.1/hCaMKIIN or empty vector pcDNA3.1/myc-His (-) B was transfected into HL-60 cells by Lipofectamine 2000. Human CaMK II N proteins of transfected cells were detected by Western blot. Cell proliferation affected by human CaMKIIN was determined by MTT. Colony-forming assay was performed by soft agar growth system. The cells transfected with CaMKIIN were stained with Hoechst 33342 to detect the apoptotic proportion under fluorescence microscopy. Cell cycle was analyzed by flow cytometry.

RESULTSHuman CaMKIIN was stably transfected into HL-60 cells, and overexpression of human CaMKIIN inhibited the proliferation of HL-60/CaMKIIN cells compared to HL-60/mock cells and HL-60 cells [(0.44 ± 0.03) vs (0.94 ± 0.05) vs (0.94 ± 0.04), P<0.01]. The colony formation of HL-60/CaMKIIN was also markedly smaller[(21.00 ± 3.05)/500] than that of mock-transfected [(111.00±4.58)/500]] and control cells [(119.00±6.09)/500] (P<0.01). After 72 hrs-culture, the apoptotic proportion in cells transfected with CaMK II N was obviously higher than of cells transfected with mock DNA or control [(22.49 ± 2.15)% vs (7.17 ± 0.72)% vs (6.40 ± 0.55)%, P<0.01]. Up to (82.97 ± 2.90)% human CaMKIIN/HL-60 cells were arrested at G0/G1 phase, which was more than mock-transfected [(40.53 ± 2.38)%] and control cells [(41.63 ± 2.27)%] (P<0.05). Human CaMKIIN could down-regulate expression of Bcl-2 in transfected cells.

CONCLUSIONCaMK IIN up-regulation could inhibit proliferation and induce apoptosis of human acute myeloid leukemia cell HL-60.

Apoptosis ; Cell Proliferation ; Genetic Vectors ; HL-60 Cells ; Humans ; Proteins ; genetics ; metabolism ; Transfection ; Up-Regulation

OBJECTIVETo explore the mechanism of Alpha-fetoprotein (AFP) effects on hepatocellular carcinoma cells (HCC) resistances apoptosis induced by tumor necrosis factor-related apoptosis inducing-ligand (TRAIL).

METHODSThe expressed alteration of TRAIL receptor-2 (DR5) after the human hepatoma cells line Bel 7402 (AFP-producing) and HLE cells (non-AFP producing) were treated with all trans retinoic acid (ATRA) were determined by Western blot; Interaction of AFP with RAR-beta was analyzed by co-immunoprecipitation (Co-IP); Laser confocal microscopy was used to observe co-localization of AFP and RAR-beta; Short small RNA interfering (RNAi) was applied to knock down the expression of AFP in Bel 7402 cells; The full AFP gene cDNA was inserted into pcDNA3.1 vector and constructed the expressed vector of AFP (named pcDNA3.1-afp); The growth of hepatoma cells was analyzed by MTT.

RESULTSBel 7402 and HLE cells expressed DR5, lowed dosage of ATRA (40mumol/L) had no influence on the expression of DR5 in Bel 7402 cells, but ATRA (160mumol/L) could inhibit the expression of AFP and promote the expression of DR5 significantly; Co-IP indicated that AFP had a property for interacting with RAR-beta; The results also demonstrated AFP co-localization with RAR-beta in cytoplasm of Bel 7202 cells; The expression of DR5 was enhanced while the expression of AFP was knocked down by RNAi. pcDNA3.1-afp vector was transfected into HLE cells, the growth of HLE cells were stimulated and TRAIL cytotoxicity of HLE cells were reduced. But when the expression of AFP was knocked down the sensitivity of Bel 7402 cells to TRAIL was enhanced.

CONCLUSIONSThese data provided that AFP had a capability to interact with RAR-beta and suppressed the expression of DR5. AFP could play pivotal role on hepatoma cells resistance-induced apoptosis by TRAIL.

Apoptosis ; Cell Line, Tumor ; metabolism ; Humans ; Receptors, Retinoic Acid ; metabolism ; Receptors, TNF-Related Apoptosis-Inducing Ligand ; metabolism ; TNF-Related Apoptosis-Inducing Ligand ; metabolism ; Tretinoin ; pharmacology ; alpha-Fetoproteins ; metabolism

The paper reports the systematic study on felodipine and its impurities in tablets, to improve its quality standards for the control of the related substances. HPLC-DAD, UPLC-MS, IR and NMR methods were used for the isolation of felodipine and its impurities in tablets, their identification and the zebrafish animal model was used for the analysis of the toxic impurities. In felodipine material and its tablets, three impurities are isolated and identified. They are impurity 1 [dimethyl 4-(2, 3-dichlorophenyl)-2, 6-dimethyl-1, 4-dihydropyridine-3, 5-dicarboxylate], impurity 2 [ethyl methyl 4-(2, 3-dichlorophenyl)-2, 6-dimethylpyridine-3, 5-dicarboxylate] and impurity 3 [diethyl 4-(2, 3-dichlorophenyl)-2, 6-dimethyl-1, 4-dihydropyridine-3, 5-dicarboxylate], separately. The result of zebrafish animal model analysis showed that the teratogenic effects of four compounds were: impurity 3 > or = felodipine > impurity 1 > impurity 2, lethal effects were as follows: impurity 2 = impurity 3 > felodipine > or = impurity 1. This study confirmed the toxicity of three impurities in felodipine. According to the results, the paper suggested the amendments to the standard of the medicine and provided the support to the control of impurities in the manufacturing process.
Abnormalities, Drug-Induced ; Animals ; Antihypertensive Agents ; administration & dosage ; chemistry ; toxicity ; Calcium Channel Blockers ; administration & dosage ; chemistry ; toxicity ; Chromatography, High Pressure Liquid ; methods ; Drug Contamination ; Felodipine ; administration & dosage ; chemistry ; toxicity ; Magnetic Resonance Spectroscopy ; Molecular Structure ; Pharmaceutical Preparations ; analysis ; chemistry ; Quality Control ; Spectrophotometry, Infrared ; Tablets ; Tandem Mass Spectrometry ; Zebrafish

Objective To learn the changes of blood lead levels and serum biochemical parameters of the school agechildren from different areas. Methods All research objects, the school age children, were from three different areasrespectively, including a mountainous area (L area), an island area (H area) where there is not history of Pb pollution,and an industry area (N area) in relation to Pb pollution. The morning urine and peripheral venous blood samples werecollected from the school age children. Pb in blood (PbB), δ-aminoaevulinic acid in urine (ALA), Ca2+, BUN, Cr inserum, and Thyroid Stimulating Hormone (TSH), thyroxin (T4), free thyroxin (FT4) levels were detected. ResultsPbB levels [M was 36.0 ppb] of the school age children from N area were significantly higher than that of L area [22.0 ppb] andH area [23.8 ppb]. On the contrary, serum Ca2+ levels of the school age children from N area were significantly lower than thatof L area and H area. Serum T4 of N area was significantly lower than that of L area and H area. Serum FT4 of H area wassignificantly higher than that of L area and N area. And TSH of N area and H area were both obviously lower than that of L area.But all of these thyroxin indexes were in the range of normal values. Conclusion It should be widely concerned that thesignificant elevation of PbB levels may have a negative impact on school age children in the future.

OBJECTIVE: To carry out prenatal diagnosis for a fetus with mosaicism Yq deletion. METHODS: A fetus with high risk of sex chromosomes indicated by non-invasive prenatal testing (NIPT) at Tianjin Medical University General Hospital in July 2021 was selected as the study subject. Prenatal diagnosis of the fetus was performed with combined G-banded chromosomal karyotyping, fluorescence in situ hybridization (FISH), copy number variation sequencing (CNV-seq), real-time fluorescence PCR (QF-PCR), and ultrasound examination. RESULTS: Analysis of the amniocytes at 23 gestational weeks had yielded a 45,X karyotype. However, FISH had shown signals of Y chromosome. Re-examination by cordocentesis had shown a mosaicism of 46,X,+mar[33]/45,X[17]. FISH showed that 69% of the cells had contained Y chromosome signals. The result of CNV-seq was seq[19]del(Y)(q11.1q12)(mos) chrY: g.13200001_ 28820000del (mosaicism rate = 64%), which suggested mosaicism for a Yq deletion, which encompassed the azoospermia factor (AZF) region. Deletion of the AZF region was verified by QF-PCR. The fetal karyotype was ultimately determined as mos46,X,del(Y)(q11.1)[33]/45,X[17]. Although ultrasound examination had shown no abnormality in the fetus, the couple had opted to terminate the pregnancy, and the induced fetus had a normal male appearance. CONCLUSION The combined use of multiple techniques is beneficial for accurate and rapid prenatal diagnosis. For fetuses with mosaicism chromosomal abnormalities, it may be difficult to accurately predict the postnatal phenotype. It is therefore necessary to further explore their genotype-phenotype correlation in order to provide better guidance upon genetic counseling.
Female ; Pregnancy ; Male ; Humans ; Mosaicism ; DNA Copy Number Variations ; In Situ Hybridization, Fluorescence ; Y Chromosome ; Fetus

OBJECTIVETo study the effects of benzo(a)pyrene (BaP) on the cell cycle distribution and activities of mitogen-activated protein kinase (MAPK) signal molecules (ERK1/2, JNK1/2 and p38) in human embryo lung cells (HELF), and to investigate the relationship between alterations of MAPK protein phosphorylation and the cell cycle distributions.

METHODSThe phosphorylation of MAPK were induced by exposing HELF cells to BaP at 0.1, 0.5, 2.5 and 12.5 micromol/L. The phosphorylation and protein expression levels of ERK1/2, JNK1/2 and p38 were determined through western-blotting assay. And the flow cytometry assay was used to measure the cell cycle effects in HELF cells after treatment with 2.5 micromol/L BaP for 24 h.

RESULTSThe phosphorylation levels of ERK1/2, JNK1/2 and p38 were significantly increased through BaP exposure. In addition, the phosphorylation of these three MAPKs has similar alteration pattern. We found that exposure of cells to 2.5 microM of BaP for 24 h resulted in a decrease of G(0) and G(1) population by 11.9% (F = 41.38, P < 0.01) and an increase of S population by 17.2% (F = 68.13, P < 0.01). Three chemical inhibitors of MAPK (AG126, SP600125 and SB203580) could significantly inhibit the cell cycle alteration because of BaP treatment.

CONCLUSIONERK1/2, JNK1/2 and p38 could positively regulate the BaP independently induced cell cycle alterations.

Benzo(a)pyrene ; toxicity ; Cell Cycle ; drug effects ; Cells, Cultured ; Fibroblasts ; drug effects ; metabolism ; Humans ; JNK Mitogen-Activated Protein Kinases ; metabolism ; Lung ; cytology ; embryology ; MAP Kinase Kinase 4 ; metabolism ; MAP Kinase Signaling System ; drug effects ; Mitogen-Activated Protein Kinase 1 ; metabolism ; Mitogen-Activated Protein Kinase 3 ; metabolism ; Mitogen-Activated Protein Kinase 8 ; metabolism ; Mitogen-Activated Protein Kinase 9 ; metabolism ; Signal Transduction ; drug effects ; p38 Mitogen-Activated Protein Kinases ; metabolism

OBJECTIVETo investigate the roles of cyclin D1/CDK4-E2F-1/4 pathway in cell cycle changes of human embryo lung fibroblasts (HELF) induced by two different benzo(a)pyrene [B(a)P] treatment models.

METHODSTwo B(a)P treatment models: HELF were treated by 2 micromol/L B(a)P for 24 hours; HELF were treated by 100 micromol/L B(a)P three times 24 hours each and provide with some characteristics of transformed cells (T-HELF). Changes of cell cycle and the expression of cyclin D1, CDK4 and E2F-1/4 were checked using the flow cytometry and Western bolt analysis.

RESULTSAfter 24 hours 2 microml/L B(a)P treatment, the HELFs in the G(1) phase was decreased. In HELF transfected with antisense cyclin Dl (A-Dl) and antisense CDK4 (A-K4), the expression of cyclin Dl and CDK4 blocked the cell cycle changes from the G(1) phase to the S phase induced by B(a)P. The overexpression of cyclin Dl and E2F-1 in HELF was induced by B(a)P. The E2F-1 overexpression in A-D1 induced B(a)P was inhibited. The E2F-4 expression was decreased and the CDK4 expression was increased significantly in A-K4 after B(a)P treatment. Most of T-HELF transfected with antisense cyclin Dl (T-A-Dl) and antisense CDK4 (T-A-K4) were retained in G(1) phase. The cyclin Dl expression in T-HELF was increased significantly compared with that in HELF. The E2F-4 expression in T-A-Dl and T-A-K4 was increased significantly compared with that in T-HELF.

CONCLUSIONB(a)P induces the cell cycle changes through cyclin D1/CDK4-E2F-1/4 pathway in HELF treated by 2 micromol/L B(a)P while it induces cell cycle changes through cyclin D1/CDK4-E2F-4 pathway in T-HELF.

Benzo(a)pyrene ; pharmacology ; Blotting, Western ; Cell Cycle ; drug effects ; Cells, Cultured ; Cyclin D1 ; biosynthesis ; Cyclin-Dependent Kinase 4 ; biosynthesis ; Dose-Response Relationship, Drug ; E2F1 Transcription Factor ; biosynthesis ; E2F4 Transcription Factor ; biosynthesis ; Fibroblasts ; cytology ; drug effects ; metabolism ; Flow Cytometry ; Humans ; Lung ; cytology ; embryology