On the basis of introducing FDA's regulatory measure and relevant requirement for life-cycle management of combination product, this paper aims to discuss corresponding countermeasure for supervision system construction in consideration of domestic drug-device combination product's current situation, in order to promote innovative development of relevant industries.
Device Approval ; Drug Approval ; United States ; United States Food and Drug Administration

Objective:To investigate the level of mumps virus (MuV) IgG antibody of healthy population and the genotyping of MuV in Fujian province in 2018.Methods:MuV IgG antibody of healthy population was detected by enzyme-linked immunosorbent assay (ELISA). Small hydrophobin (SH) gene of MuV was genotyped in pharyngeal swab and cell cultures of mumps patients using reverse transcription polymerase chain reaction (RT-PCR). The phylogenetic tree by SH sequences was constructed to identify MuV genotype.Results:A total of 4 925 people completed antibody testing, the positive rate of MuV IgG antibody was 78.58%(3780/4925), and the geometric mean concentration (GMC) was 245.83 IU/ml. There was no statistically significant difference in the positive rate of antibodies between people of different genders, while GMC had statistically significant differences ( χ2=4.295, P=0.117; Z=-2.220, P=0.026). There were significant differences in the positive rate of antibodies and GMC between people in different regions and age groups. Especially in infants and 12-15 years old group of people, the MuV IgG antibody positive rate and GMC were at low levels. The antibody positive rate and GMC of people with a history of mumps-containing vaccine (MuCV) immunization were higher than those without a history of MuCV immunization and those with unknown MuCV immunization history ( χ2=259.315, P<0.001; Z=-16.319, P<0.001). Eight strains of MuV were isolated from the mumps outbreak, which were all F genotypes. Conclusion:The immune level of mumps in infants and young children in Fujian province was low, and the 12-15 age group is the focus of attention. The epidemic strains of MuV in Fujian province in 2018 were mainly F genotype strains.

More efficient drug delivery system and formulation with less adverse effects are needed for the clinical application of broad-spectrum antineoplastic agent doxorubicin (DOX). Here we obtained outer-membrane vesicles (OMVs), a nano-sized proteoliposomes naturally released by Gram-negative bacteria, from attenuated and prepared doxorubicin-loaded O0MVs (DOX-OMV). Confocal microscopy and distribution study observed that DOX encapsulated in OMVs was efficiently transported into NSCLC A549 cells. DOX-OMV resulted in intensive cytotoxic effects and cell apoptosis as evident from MTT assay, Western blotting and flow cytometry due to the rapid cellular uptake of DOX. In A549 tumor-bearing BALB/c nude mice, DOX-OMV presented a substantial tumor growth inhibition with favorable tolerability and pharmacokinetic profile, and TUNEL assay and H&E staining displayed extensive apoptotic cells and necrosis in tumor tissues. More importantly, OMVs' appropriate immunogenicity enabled the recruitment of macrophages in tumor microenvironment which might synergize with their cargo DOX . Our results suggest that OMVs can not only function as biological nanocarriers for chemotherapeutic agents but also elicit suitable immune responses, thus having a great potential for the tumor chemoimmunotherapy.

Objective:We isolated and identified the genotypes and molecular characteristics of the imported B3 measles virus (MeV) in Fujian province in 2018.Methods:Throat swab specimens were collected from clinically diagnosed measles patients and tested for viral RNA, using the real-time reverse transcription-polymerase chain reaction after the RNA extraction. Reverse transcription-polymerase chain reaction method was undertaken to amplify the 634 nucleotide acids of 3-terminal of the nucleoprotein gene. A phylogenetic tree was constructed and similarities in homology assessed.Results:We successfully isolated and obtained two measles virus strains and eighteen viral nucleic acid sequences. The Fujian strains were clustered within the same genotype group of WHO genotype B3 reference strains. Compared to the major circulating measles strain genotype B3 in the world, two Fujian strains MV18-41 and MV18-42 showed 100.0 % nucleic acid homology to HongKong.CHN/35.18 strain which was isolated from Hong Kong in 2018. The remaining 16 Fujian strains showed the highest homology (99.9 %) with the Mvs/Osaka.JPN/38.18/B3 strain isolated from Japan in 2018. Compared with other 23 WHO genotype reference strains, homology on both nucleotide and amino acid of the Fujian strain and the B1 genotype reference strain were the smallest, as 95.1 %-95.4 % and 95.3 %, respectively. The differences of homology between the Fujian strain and H1 genotype reference strain were the largest, as 88.7 %-89.0 % and 87.3 %, respectively. In addition, there were 13 mutation sites between the Fujian strain and the vaccine strain (Shanghai-191) at the 150 amino acid position of carboxy terminus on N protein, However, these sites did not cause functional changes in the protein region. Conclusions:In Fujian province, two strains of B3 genotype measles virus were obtained successfully, which were considered to be new genotype measles virus found in 2018. These findings showed it is necessary to strengthening the monitoring program on imported cases for better control and eliminate the measles virus.

Objective To explore the role of forkhead box F2 (FoxF2) in the extracellular matrix of trabecular meshwork.Methods The cultured human trabecular meshwork cells (HTMCs) were divided into Scramble control group and FoxF2 small hairpin RNA (shRNA) group,then FoxF2 shRNA,the FoxF2 restructuring interference carrier was built,HTMCs were infected with FoxF2 shRNA lentivirus.Western blot assay was used to detect the expression of FoxF2 protein and extracellular matrix.Furthermore,Transwell counting experiment was used to analyze the migration ability of HTMCs.Results The cultured HTMCs grew well and showed a long spindle shape.The growth status of HTMCs was well,and their morphological characteristics were consistent with the HTMCs in vivo.The relative expression level of FoxF2 protein in the FoxF2 shRNA group was lower than that in the Scramble control group,with a significant difference between them (0.72 ± 0.02 vs.1.27 ± 0.05;t =16.68,P ＜ 0.01).The relative expression level of fibronectin (FN),collagen type Ⅰ (COL Ⅰ) and α-smooth muscle actin (α-SMA) were 0.43±0.03,0.53 ±0.08 and O.86±0.15 in the FoxF2 shRNA group,and 0.87±0.04,1.66±0.06 and 1.73 ±0.13 in the Scramble control group,respectively,the relative expression levels of FN,COL Ⅰ and α-SMA in the FoxF2 shRNA group were significantly lower than those in the Scramble control group (t =15.08,18.81,7.50,all at P＜0.01).The migration number of HTMCs in the FoxF2 shRNA group was significantly lower than that in the Scramble control group (117.30±11.41 vs.251.00±10.37;t =8.72,P＜0.01).Conclusions The FoxF2 shRNA lentivirus are successfully constructed,which can decrease the expression of FoxF2 in HTMCs.Low expression of FoxF2 can reduce the expression level of extracellular matrix protein in HTMCs and inhibit the migration ability of HTMCs.

Objective To observe the inhibitory effect of lentiviral vector miR-191 (LV-191) on retinal neovascularization (RNV) in mice model of oxygen-induced retinopathy (OIR).Methods Eighty healthy 7-day-old C57BL/6J mice were randomly divided into 5 groups including normal group, non-intervention group, normal saline (NS) group, LV-191 group and LV-green fluorescent protein (GFP) group, 16 mice in each group. The OIR model was established in the non-intervention group, NS group, LV-191 group and LV-GFP group. NS group, LV-191 group and LV-GFP group were given an intravitreal injection of 1 μl of NS, LV-191 and LV-GFP at the age of 12 days. No injection was performed in the non-intervention group. In normal group,newborn mouse were maintained in room air form P0 to P17, and no treatment was performed. Mice in all five groups were euthanized at P17. Retinal neovasculation (RNV) was evaluated by counting the number of pre-retinal neovascular cells and analysis of non-perfusion area area by immunofluorescent staining of the mouse retina. Real-time quantitative PCR (RT-PCR) to detect miR -191 and P21 expression of retinal tissue.Results In the LV-191 group, the non-perfusion area were both significantly smaller than those in non-intervention group, NS group and LV-GFP group (F=127.20,P<0.001). The number of pre-retinal neovascular cell nuclei in retinas from LV-191 group were obviously lower than those in the retinas from non-intervention group, NS group and LV-GFP group (F=31.71,P<0.05). RT-PCR showed that the LV-191 and P21 level of LV-191 group increased significantly than other groups (F=10.95, 15.60;P<0.05).Conclusion Intravitreal injection of LV-191 inhibits RNV in mice model of OIR possibly through up-regulating p21.

Objective To observe the effect ofpolypyramidine tract binding protein-associated splicing factor (PSF) towards advanced glycation end products (AGEs) induced the apoptosis of Müller cells in vitro.Methods Experimental study.Müller cells were cultured and divided into groups according to the project design,plasmid enhanced green fluorescent protein-PSF were transfected into the cells to achieve the overexpression of PSF Müller cells in vitro,then cells were exposed to AGEs and the Morphological changes were observed by HE staining and Hoechst 33258 staining while the survival rate of cells were detected by MTT assay.The effects of PSF on AGEs-induced Müller apoptosis was measured by Cell Death Detection ELISA kit.Meanwhile,2',7'-diehlorofluorescin diaeetate staining was performed to monitor the protective effects of PSF on AGEs-induced Müller cells ROS.Results The morphology of cells in normal group was full and the cytoplasm staining was uniform.In N+AGEs group and Vec+AGEs group,cell volume decreased,cytoplasm was dense and concentrated,and eosinophilic staining was enhanced.The cell morphology of PSF+AGEs group was still full,with uniform cytoplasm staining and uniform nucleus staining.The viability of N+AGEs group,Vec+AGEs group and PSF+AGEs group were 0.42±0.11,0.35±0.12 and 0.68±0.12.The apoptosis values were 1.08 ± 0.16,0.96± 0.20 and 0.44± 0.08.The intracellular ROS levels were 28 833.67± 3 550.06,28 356.67±4 854.81,186 163.00±382.54.Compared with N+AGEs group and Vec+AGEs group,the cell viability of PSF+AGEs group was significantly improved (F=20.65,P=0.000),ce11 apoptosis value (F=43.43,P=0.000) and intracellular ROS level (F=1 8.86,P=0.000).Conclusion PSF overexpression play a protective role in AGEs-induced apoptosis by inhibiting the production of ROS in Müiller cells.

Objective To observe RNA-Seq analysis ofgene expression profiling in retinal vascular endothelial cells after anti-vascular endothecial growth factor (VEGF) treatment.Methods Retinal vascular endothelial cells were cultured in vitro,and the logarithmic growth phase cells were used for experiments.The cells were divided into the control group and high glucose group.The cells of two groups were cultured for 5 hours with 5,25 mmol/L glucose,respectively.And then,whole transcriptome sequencing approach was applied to the above two groups of cells through RNA-Seq.Now with biological big data obtained as a basis,to analyze the differentially expressed genes (DEGs).And through enrichment analysis to explain the differential functions of DEGs and their signal pathways.Results The gene expression profiles of the two groups of cells were obtained.Through analysis,449 DEGs were found,including 297 upregulated and 152 downregulated ones.The functions of DEGs were influenced by regulations over molecular biological process,cellular energy metabolism and protein synthesis,etc.Among these genes,ITGB 1BP2,NCF 1 and UNC5C were related to production of inflammation;AKR1C4,ATP 1A3,CHST5,LCTL were related to energy metabolism of cells;DAB 1 and PRSS55 were related to protein synthesis;SMAD9 and BMP4 were related to the metabolism of extracellular matrix.GO enrichment analysis showed that DEGs mainly act in three ways:regulating biological behavior,organizing cellular component and performing molecular function,which were mainly concentrated in the system generation of biological process part and regulation of multicellular organisms.Pathway enrichment analysis showed that gene expressions of the two cell groups were differentiated in transforming growth factor-β (TGF-β) signaling pathway,complement pathway and amino acid metabolism-related pathways have also been affected,such as tryptophan,serine and cyanide.Among them,leukocyte inhibitory factor 9 and bone morphogenetic protein 4 play a role through the TGF-β signaling pathway.Conclusions High glucose affects the function of retinal vascular endothelial cells by destroying transmembrane conduction of retinal vascular endothelial cells,metabolism of extracellular matrix,and transcription and translation of proteins.

Objective To observe the synergistic effect of metformin and anti-vascular endothelial growth factor (VEGF) in the treatment of diabetic retinopathy.Methods This study was composed of clinical data review and in vitro cell experiment.Ten patients (12 eyes) with diabetic macular edema treated with antiVEGF drugs were included in the study.Patients were randomly divided into the VEGF group (anti-VEGF drug therapy) and the combined treatment group (anti-VEGF drug combined with metformin).The changes of visual acuity and central retinal thickness (CRT) were compared between the two groups.As far as the in vitro experiment was concerned,vascular endothelial cells were divided into the control group (normal cells),the VEGF group (50 ng/ml VEGF),the anti-VEGF group (50 ng/ml VEGF+2.5 μg/ml of conbercept),and the combined group (50 ng/ml VEGF +2.5 μg/ml of conbercept +2.0 mmol/L of metforrnin).And then MTT cell viability assay,scratch assay and real-time quantitative polymerase chain reaction assay were performed to analyze the cell viability,cell migration and mRNA level of VEGFR2,protein kinase C (PKC)-α and PKC-β successively.Results Review of clinical trial shows that the CRT recovery rates in the combined treatment group were much higher than that in the VEGF group at 3 month after the operation,while the difference was statistically significant (t=-2.462,P＜0.05).In vitro cell experiment results showed that VEGF induction upregulated the viability and mobility of vascular endothelial cells obviously compared with control group,at the same time,the use of anti VEGF drugs can effectively reverse the trend,in contrast,combination of metformin and anti-VEGF showed a more superior effect to some extent (P＜0.05).In the VEGF group,the mRNA expression of VEGFR2,PKC-αand PKC-[β were significantly increased compared with the control group (P＜ 0.01);while the mRNA expression of VEGFR2,PKC-αand PKC-β in the combination group decreased significantly compared with the VEGF group and the control group (P＜0.05).However,in the anti-VEGF group,the mRNA expression of VEGFR2,PKC-αand PKC-β were decreased,but has failed to reach the level of statistical learn the difference.Conclusions The combination ofmetformin and anti-VEGF drugs can reduce the CRT of diabetic retinopathy patients and inhibit the proliferation and migration of retinal vascular endothelial cells which induced by VEGF.The synergistic mechanism may be related to the inhibitory effect of metformin on the expression of VEGFR and PKC.

Objective To observe the protective effect of dl-3-n-Butylphthalide (NBP) on apoptosis of retinal Müller cells induced by hydrogen peroxide (H2O2).Methods Human retinal Müller cells cultured in vitro were divided into normal control group,model group (H2O2 group) and experimental group (H2O2+NBP group).The cells in the H2O2 group and H2O2+NBP group were cultured with 200 μ mol/L H2O2 for 2 h.Then the culture solution of the H2O2 group replace with complete medium and the H2O2+NBP group replace with complete medium containing 1 tmol/L NBP.The normal control group was a conventional cultured cells.Müller cells were identified by immunofluorescence staining.Hematoxylin-eosin (HE) staining was used to observe the apoptosis morphological changes.MTT assay was used to detect the activity of of retinal Müller cells after after 24 h and 48 h of NBP intervention.Hoechst33258 staining was used to observe the apoptosis.LIVE/DEAD (R)cell activity/cytotoxicity kit was used to detect cell viability.Dichlorofluorescein diacetate (DCFH-DA) + endoplasmic reticulum (ER) red fluorescent probe (ER-Tracker Red) double staining was used to observe the expression level of reactive oxygen species (ROS) in ER of cells.One-way ANOVA combined with Dunnett statistical method were used for data analysis.Results HE staining showed that the number of cells in H2O2+NBP group was higher than that in H2O2 group.MTT assay showed that after 24 h and 48 h of NBP intervention,the differences in cell viability between the normal control group and the H2O2 group,the H2O2 group and the H2O2+NBP group were statistically significant (t=28.96,3.658,47.58,20.33;P＜0.001,0.022).The results of Hoechst33258 showed that the nuclear nucleus of a few cells in the H2O2+NBP group was crescent-shaped and the nuclear fragmentation was reduced,and the blue fluorescence of the remaining cells was uniform.The LIVE/DEAD ~ cell activity/cytotoxicity kit showed that the number of dead cells with red fluorescence in the H2O2 group increased significantly,and the number of viable cells with green fluorescence decreased significantly.In the H2O2+NBP group,the number of viable cells with green fluorescence increased,and the number of dead cells with red fluorescence decreased.The double staining results of DCFH-DA+ER-Tracker Red showed that the green fluorescence intensity of H2O2 group was significantly enhanced;the green fluorescence intensity of H2O2+NBP group was lower than that of H2O2 group.Conclusion NBP alleviates H2O2-induced apoptosis of human retinal Müller cells by inhibiting ROS production.