Objective To develop a novel approach of transfecting the exogenous gene safely and effectively mediated by receptor according to the cavitation effect of ultrasound-mediated microbubble destruction. Methods C-myc antisense oligonucleotide(ASODN) and galactosylation polylysine(G-PLL) which was fluoresce in-labeled specificity ligand and SonoVue microbbule were coupled by electrostatic interaction. SMMC-7721 cell with c-myc ASODN were transfected in vitro by ultrasound-mediated microbubble destruction to investigate the effect of c-myc ASODN on it. The c-myc mRNA gene expression of liver cancer SMMC-7721 cells was detected. Results Cultured with 10% SonoVue microbubble and under ultrasound at 1.5 MHz,1.0 W/cm2, 10% duty cycle,60 s respectively,the SMMC-7721 cells had not been changed significantly. Reverse transcription polymerase chain reaction(RT-PCR) assay showed that c-myc expression was decreased in transfected cells(P<0.01). Conclusions Receptor-mediated targeted microbubble destruction by ultrasound can enhance the exogenous gene transfection and expression. It is a promising method in gene therapy of liver cancer.

Administration, Intravaginal ; Adult ; Combined Modality Therapy ; Drugs, Chinese Herbal ; administration & dosage ; Female ; Humans ; Microwaves ; therapeutic use ; Middle Aged ; Phytotherapy ; Uterine Cervical Erosion ; therapy

Objective To investigate the value of clinical characteristics in diagnosis of vocal fold paralysis (VFP) and arytenoid dislocation .Methods Eighty - eight patients of VFP and 27 patients of arytenoid dislocation were studied , by comparing the causes , laryngeal morphologic characteristics and laryngeal electromyography (LEMG) .Results The causes of 88 VFP patients included surgery (45 cases) ,neck trauma(2 cases) ,idiopathic causes(16 cases) ,infection(16 cases) ,and tumor invasion - related(9 cases) .Of the 27 arytenoid dislocation pa‐tients ,24 had a history of endotracheal intubation and the others had a history of gastric tube insertion .The vocal folds were mostly fixed at the paramedian position ,followed by the abducent position and the median position .No significant differences were found in laryngeal morphologic characteristics between the two groups ,including vocal fold shape , glottis vertical symmetry , mucosal waves , supraglottic compensation , glottis closure and arytenoid movement .The LEMG of VFP patients appeared as denervation patterns ,reinnervation potentials ,or electrical si‐lence ;the recruitment patterns appeared as mix or simple patterns ;the evoked potentials were absent .Of the VFP patients ,54 cases(61 .36% % )were found synkinesis of involved posterior cricoarytenoid and two of them also in ‐volved thyroatenoid .The patients with synkinesis had lower percentage of vocal fold bowing and higher percentage of glottic vertical asymmetry compared to the ones without synkinesis .Of the VFP patients whose cause was surgery or neck trauma ,the median - position fixed vocal folds were mostly observed in the patients with duration of less than 1 month or with synkinesis .Of the 27 arytenoid dislocation patients ,20(74 .07% )showed normal LEMG pat‐terns and 7(25 .93% )showed apparent LEMG abnormality on the affected side .Conclusion The causes of vocal fold paralysis and arytenoid dislocation are different .Laryngeal morphologic characteristics have limitations in distinguis‐hing vocal fold paralysis from arytenoid dislocation .The shape and position of involved vocal folds of the VFP pa‐tients are correlated with duration ,nerve regeneration and synkinesis .

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Case discussion is an effective approach,which combines basic theory with clinical medicine.It can evoke students' interest and cultivate their creative thinking capacity.Moreover,it can improve teachers' general ability in teaching pathophysiolo- gy.In this article,we discuss the application of case discussion in pathophysiology teaching.

Glypican-3 (GPC3) is an oncofetal protein anchored on the plasma membrane and highly expressed in hepatocellular carcinoma (HCC).GPC3 could he used as a biomarker for the diagnosis of HCC and the serum levels of GPC3 in liver cancer patients has a significant role for their prognosis.Moreover, GPC3 in HCC cells is immunoreactive, rendering it a suitable target for the treatment of HCC.Nowadays, several clinical trials targeting GPC3 for HCC therapy have already been conducted: new anti- GPC3 antibodies are generated; the clinical trials about its combination administration with other targeted medicines are being in progress; (tP(3-targeted TRAB, GPC3 vaccines and GPC3-based chimeric antigen receptor T-cells (CAR-T) therapy are under investigation.In this review, we briefly discuss the structure of GPC3, its role in HCC pathogenesis and summarize the recent development in the clinical application of GPC3.We firmly believe that GPC3 would be a promising target for HCC therapy.And further studies focusing on GPC3 should provide us more solid evidence.

Abstract: Objective　To investigate the pollution of paralytic shellfish poisons (PSP) in shellfish sold in Hainan Province from 2018 to 2021. Methods　From 2018 to 2021, the content of 10 paralytic shellfish poisons including saxitoxin (STX), neosaxitoxin (neoSTX), gonyautoxins 1 (GTX1), gonyautoxins 2 (GTX2), gonyautoxins 3 (GTX3), gonyautoxins 4 (GTX4), gonyautoxins 5 (GTX5), decarbamoylsaxitoxin (dcSTX), decarbamoylgonyau toxins 2 (dcGTX2) and decarbamoylgonyau toxins 3 (dcGTX3) in 7 kinds of shellfish commonly sold in 13 cities and counties in Hainan province was analyzed. Results　The detection rate of PSP in 360 shellfish samples was 10.3%. Among them, the highest detection rate of STX was 5.83%, followed by GTX2 detection rate of 4.17%; the detection rate of neoSTX and GTX3 were both 1.67%; the detection rate of GTX1 was 1.39%. None of the five PSP, GTX4, GTX5, dcSTX, dcGTX2 and dcGTX3, were detected. Four types of PSP were detected in fanscallops, two were detected in oysters, mussels and Scapharca subcrenata, only one was detected in scallops, and no toxin contamination was detected in clams and razor clams. A single sample of fanscallops detected a maximum of 4 PSP, and a single sample of oysters, scallops, mussels and Scapharca subcrenata detected a maximum of 1 PSP. The equivalence of PSP in all samples was ND-155.6 μg/kg.The annual detection rate of PSP from high to low was: 20.0% in 2020, 15.6% in 2019, 5.3% in 2018, and 2.0% in 2021, and none of the samples tested exceeded the standard. Continuously detectable STX in 2018-2020, all PSP that could be detected in 2018 were STX. In 2019, in addition to STX detected in scallops and Scapharca subcrenata, neoSTX was also detected in oysters, mussels and Scapharca subcrenata. In 2020, PSP was only detected from scallops, and GTX2 could be detected in all positive specimens, while 5 STX, 5 GTX1 and 6 GTX3 were detected. Only GTX2 detected from scallops in 2021. STX was detected in shellfish sold in 12 cities and counties, GTX2 can be detected in 10 cities and counties, neoSTX can be detected in 5 cities and counties, GTX1 and GTX2 were detected in 4 cities and counties respectively. Shellfish sold in Wenchang and Lingshui markets can detect 5 types of PSP. Conclusion　Some types of shellfish on the market in Hainan are contaminated with some kind of PSP pollution risks, and it is necessary to strengthen the supervision of PSP in marketed shellfish.

Objective To explore change of peripheral blood CD5+B cells in the systemic lupus ery- thematosus(SLE)patients and the association of this change with the disease activity,Methods The percent- age of CD5~+B ceils in peripheral blood in 57 patients with SLE and 35 healthy controls was determined by flow eytometry.Levels of anti-dsDNA,complement C3 and C4,anti-nuclear antibody(ANA),anticardiolipin anti- body(ACL),were also measured.Results The percentage of CD5~+B cell in SLE patients(2.1?0.4)% was high- er than that in normal controls [(1.5?0.4)%](P＜0.05).In active disease group,the percentage of CD5~+B cells [(2.5?0.5)%] was significantly higher than that in inactive SLE patients [(1.4?0.5)% ](P＜0.01).The percent- age of CD5~+B cell associated with anti-dsDNA,ANA,ALA positively,with C3 negatively and had no associa- tion with C4.Conclusion The percentage of CD5~+B cells in the SLE is significantly high and has correlation and some relationship with the SLE.

OBJECTIVETo investigate the effects of hypoxia on lipid metabolism in the normal human hepatic cell line L02 and to investigate the underlying molecular mechanisms.

METHODSL02 cells were exposed to hypoxic conditions (experimental groups: at 1% O2, 5% CO2, 94% N2 for 3, 6, 12, 24, or 48 hours) or normoxic conditions (control group: at 21% O2). Lipid droplet accumulation and triglyceride content were measured in each group by oil red O staining and biochemical assay, respectively. The mRNA expression levels of hypoxia-inducible factor (HIF)-2a and sterol regulatory element binding protein (SREBP)-1c were detected by reverse transcription-polymerase chain reaction. Protein expression levels of HIF-2a, adipose differentiation-related protein (ADRP), and Fas were tested by Western blot analysis.

RESULTSLipid droplet accumulation and the triglyceride content were significantly higher in the hypoxia group than the normoxia group. In addition, the hypoxia groups had significantly down-regulated mRNA expression of SREBP-1c (12h: 0.236+/-0.043, 24 h: 0.287+/-0.044, 48 h: 0.342+/-0.049 vs. normoxia: 0.503+/-0.037; F = 28.37, P less than 0.01) and FAS protein (12 h: 0.562+/-0.054, 24 h: 0.674+/-0.062, 48 h: 0.682+/-0.057 vs normoxia: 0.857+/-0.069; F = 16.08, P less than 0.01). In normoxic cells, little or no expression of HIF-2a protein was detected by Western blot. In hypoxic cells, HIF-2a protein expression peaked at 6h (0.973+/-0.067). ADRP protein expression was significantly higher in hypoxia groups than in the normoxia group (12 h: 0.319+/-0.043, 24 h: 0.732+/-0.056 and 48 h: 0.873+/-0.066 vs. 0.211+/-0.019; all, P less than 0.05.

CONCLUSIONExposure to hypoxic conditions might induce lipidosis in normal human hepatic cells by stimulating HIF-2a and ADRP expression.

Basic Helix-Loop-Helix Transcription Factors ; metabolism ; Cell Hypoxia ; Cell Line ; Down-Regulation ; Hepatocytes ; metabolism ; Humans ; Lipid Metabolism ; Membrane Proteins ; metabolism ; Oxygen ; metabolism ; Perilipin-2 ; Sterol Regulatory Element Binding Protein 1 ; metabolism

OBJECTIVETo investigate the effect of Hepatitis B Virus X Protein (HBx) on the expression of lipid metabolism-related genes and its role in pathogenesis of hepatocyte fatty degeneration.

METHODSHepatitis B Virus X gene eukaryon expression vector pIRES2-eGFP-HBx was transfected into HepG2 cells to establish HepG2/HBx cell model for HBx expression. HepG2 cells transfected with pIRES2-eGFP (HepG2/pIRES2 cell) and non-transfected were used as controls. At 24, 48 and 72 hours after transfection, the expression of green fluorescent protein (GFP) was observed by fluorescence microscope and the triglyceride(TG) content was detected. RT-PCR and Western blot were applied to detect the levels of sterol regulatory element binding protein-1 (SREBP-1), liver x receptor alpha (LXRalpha) mRNA and the levels of HBx, LXRalpha and fatty acid synthase (FAS) protein. At 24, 48 and 72 hours after transfection, the expression of GFP was found in HepG2/HBx and HepG2/pIRES2 cells, and increased gradually. The expression of HBx was detected only in HepG2/HBx cells, and was increased with time after transfection (F = 32.21, P less than 0.01). These suggested successful obtaining of HepG2-HBx cell model for HBx expression.

RESULTSAt 24h, 48h and 72h after transfection, the expression levels of LXRalpha mRNA (0.386+/-0.055, 0.505+/-0.071, 0.649+/-0.058 ) and SREBP-1 mRNA (0.395+/-0.055, 0.548+/-0.047, 0.795+/-0.058), as well as the levels of LXRalpha protein(0.178+/-0.036, 0.263+/-0.047, 0.347+/-0.058) and FAS protein(0.436+/-0.055, 0.608+/-0.053, 0.827+/-0.046) in HepG2-HBx group were dramatically higher than those in the controls at the same time points (all P less than 0.05/0.01), and were gradually increased with time (all P less than 0.05/0.01). A positive correlationship was observed between HBX protein level and the LXRalpha, SREbP-1 mRNA and LXRalpha, FAS protein levels. The difference of TG content between HepG2/HBx group and control groups was not statistically significant (P more than 0.05).

CONCLUSIONSHBx-LXRalpha-SREBP-1/FAS pathway suggested regulating transcription and expression of lipid metabolism-related genes, which might be one of the important molecular mechanism causing hepatocyte fatty degeneration.

Carcinoma, Hepatocellular ; metabolism ; Fatty Acid Synthase, Type I ; metabolism ; Hep G2 Cells ; Humans ; Lipid Metabolism ; genetics ; Liver Neoplasms ; metabolism ; Liver X Receptors ; Orphan Nuclear Receptors ; metabolism ; Sterol Regulatory Element Binding Protein 1 ; metabolism ; Trans-Activators ; genetics ; Transfection