Room Temperature Ionic Liquid Capping Layer for High Efficiency FAPbI3 Perovskite Solar Cells with Long-Term Stability.

Ionic liquid salts (ILs) are generally recognized as additives in perovskite precursor solutions to enhance the efficiency and stability of solar cells. However, the success of ILs incorporation as additives is highly dependent on the precursor formulation and perovskite crystallization process, posing challenges for industrial-scale implementation. In this study, a room-temperature spin-coated IL, n-butylamine acetate (BAAc), is identified as an ideal passivation agent for formamidinium lead iodide (FAPbI3) films. Compared with other passivation methods, the room-temperature BAAc capping layer (BAAc RT) demonstrates more uniform and thorough passivation of surface defects in the FAPbI3 perovskite. Additionally, it provides better energy level alignment for hole extraction. As a result, the champion n-i-p perovskite solar cell with a BAAc capping layer exhibits a power conversion efficiency (PCE) of 24.76%, with an open-circuit voltage (Voc) of 1.19 V, and a Voc loss of ≈330 mV. The PCE of the perovskite mini-module with BAAc RT reaches 20.47%, showcasing the effectiveness and viability of this method for manufacturing large-area perovskite solar cells. Moreover, the BAAc passivation layer also improves the long-term stability of unencapsulated FAPbI3 perovskite solar cells, enabling a T80 lifetime of  3500 h when stored at 35% relative humidity at room temperature in an air atmosphere.

High effects of climate oscillations on population diversity and structure of endangered Myricaria laxiflora.

Exploring the effects of climate oscillations on the population diversity and structure of endangered organisms in the Three Gorges Reservoir (TGR) area is essential for hydrological environment changes on endangered organism evolution. Myricaria laxiflora is an endemic and endangered shrub restricted to the TGR along the banks of Yangtze River, China. Recently, six natural populations of this species were newly found upstream and downstream of the TGR, whose habitats have been dramatically changed by the summer flooding regulated by large dams. To study the water level fluctuations and climatic shifts on the genetic diversity and genetic differentiation of the six natural populations, 303 individuals from six populations were analyzed based on one nuclear DNA (ITS) and four chloroplast fragments (trnL-F, psbA-trnH, rps16, and rpl16). The phylogenetic tree and significant genetic divergence identified in the cpDNA and ITS with genetic isolation and limited gene flow among regions suggested that the six populations separated well to two groups distributed upstream and downstream. The MaxEnt modeling results indicated that obvious unidirectional eastward migration via Yangtze River gorges watercourse mediated from Last Interglacial to Last Glacial Maximum were showed with the narrow scale distributions of six remnant populations and nine extirpated populations. The initial habitat fragmentation could be triggered by the accumulation of local habitat loss of the impoundment of the TGR during the Present period and might remain stable restoration with bidirectional diffusion in the Future. Divergences among M. laxiflora populations might have been induced by the drastic changes of the external environment and limited seed/pollen dispersal capacity, as the results of long-term ecological adaptability of summer flooding stress. The haplotypes of nuclear gene could be used for population's differentiation and germplasm protection. This identified gene flow and range dynamics have provided support for the gene-flow and geology hypothesis. It is also crucial for rescuing conservation to understand the impact of environmental dynamics on endangered organism evolution.

Correction: An interfacial toughening strategy for high stability 2D/3D perovskite X-ray detectors with a carbon nanotube thin film electrode.

Correction for 'An interfacial toughening strategy for high stability 2D/3D perovskite X-ray detectors with a carbon nanotube thin film electrode' by Liwen Qiu et al., Nanoscale, 2023, 15, 14574-14583, https://doi.org/10.1039/D3NR02801A.

An interfacial toughening strategy for high stability 2D/3D perovskite X-ray detectors with a carbon nanotube thin film electrode.

Single-crystalline metal halide perovskite materials hold great promise for developing next-generation low-dose X-ray detection. To bring this new technology into reality, it is important to improve the durability of perovksite detectors by suppressing the well-known corrosion and ion diffusion problems at the perovskite/electrode interface. For imaging application, it is also imperative to develop new assembling approaches to realise non-planar interconnection between thick perovskite crystals and thin-film transistor (TFT) backplanes. Herein, a flexible and mechanically robust carbon nanotube (CNT) film was proposed to replace noble metal electrodes. The proposed CNT film, whose binder contains a carboxyl group, can form solid contact with a phenethylamine-based two-dimensional (2D) perovskite via amide coupling, thus toughening the perovskite-electrode interface. The resulting CNT/2D-3D perovskite detector shows an applaudable low dark current, high sensitivity, a low dose detection limit and excellent stability, retaining 98% of its initial sensitivity after storage for three months. Moreover, the flexible CNT films are also beneficial for making non-planar interconnection between thick perovskite crystals and TFT backplanes. The proposed flexible CNT thin film electrode thus provides a facile route towards realising a low-dose, high-resolution and highly stable perovskite X-ray detector.

Summer dormancy of Myricaria laxiflora to escape flooding stress: Changes in phytohormones and enzymes induced by environmental factors.

Dormancy is an adaptation mechanism of plants to environmental stress. Myricaria laxiflora undergoes a long period of flooding stress every year. In order to determine whether this species escapes flooding stress through dormancy, young branches and leaves were collected at different time points before the onset of flooding, and changes in the content/activity of hormones/enzymes that are closely involved in plant growth were monitored. The inducing environmental factors of summer dormancy were identified. The branches and leaves of M. laxiflora showed the following trends as summer flooding approached: (1) gradual increase in the abscisic acid content; (2) gradual decrease in the gibberellin and cytokinin contents; and (3) a continuous decrease in the activities of malate dehydrogenase (MDH), ribulose diphosphate carboxylase (RuBisCo), and glycolate oxidase (GLO). Pearson correlation analysis revealed (1) daylight duration was highly correlated with the hormone content and enzyme activity; (2) the daily mean air temperature (DMAT) was significantly correlated with the cytokinin content. These findings suggest that daylight duration was the main environmental factor leading to changes in the phytohormone content and enzyme activity as well as leading to summer dormancy. M. laxiflora undergoes dormancy before the onset of summer flooding to escape summer flooding stress. Our data indicate that summer flooding does not impede the survival and growth of M. laxiflora.

[Preparation and identification of neutralizing antibodies to envelope protein domain III(ED III) of the four dengue virus serotypes].

Objective To prepare neutralizing monoclonal antibodies (mAbs) against envelope protein domain III of the four dengue virus serotypes (DENV-EDIII) and identify its specificity. Methods BALB/c mice were immunized with recombinant EDIII protein (rDENV-EDIII) of the four DENV serotypes. Hybridoma cells secreting DENV-EDIII antibodies were screened by indirect ELISA. The specificity of positive hybridoma cells were further tested by indirect immunofluorescence assay (IFA). The neutralizing activities of DENV-EDIII mAbs in vitro were determined by the enzyme-linked immunospot microneutralization test (ELISPOT-MNT). Results 6 mAbs specific for the EDIII of the four DENV serotypes and 11 mAbs specific for only one serotype of DENV-EDIII protein were obtained, of which one monoclonal antibody had a balanced strong neutralizing activity against the four DENV serotypes in vitro, and its 50% inhibitory concentrations (IC50) for the four DENV serotypes was 0.05 µg/mL, 1.89 µg/mL, 0.02 µg/mL, 3.91 µg/mL, respectively. Conclusion A specific monoclonal antibody against DENV-EDIII is successfully screened and obtained to neutralize the four DENV serotypes.

Morphological structures and histochemistry of roots and shoots in Myricaria laxiflora (Tamaricaceae).

Myricaria laxiflora (Tamaricaceae) is an endangered plant that is narrowly distributed in the riparian zone of the Three Gorges, along the Yangtze River, China. Using bright-field and epifluorescence microscopy, we investigated the anatomical and histochemical features that allow this species to tolerate both submerged and terrestrial environments. The adventitious roots of Myr. laxiflora had an endodermis with Casparian bands and suberin lamellae; the cortex and hypodermal walls had lignified thickenings in the primary structure. In the mature roots, the secondary structure had cork. The apoplastic barriers in stems consisted of a lignified fiber ring and a cuticle at the young stage and cork at the mature stage. The leaves had two layers of palisade tissue, a hyaline epidermis, sunken stomata, and a thick, papillose cuticle. Aerenchyma presented in the roots and shoots. Several Myr. laxiflora structures, including aerenchyma, apoplastic barriers in the roots and shoots, were adapted to riparian habitats. In addition, shoots had typical xerophyte features, including small leaves, bilayer palisade tissues, sunken stomata, a thick, papillose cuticle, and a hyaline epidermis. Thus, our study identified several anatomical features that may permit Myr. laxiflora to thrive in the riparian zone of the Three Gorges, China.

Structural and Functional Characterization of a Cross-Reactive Dengue Virus Neutralizing Antibody that Recognizes a Cryptic Epitope.

Understanding the molecular basis of the neutralizing antibody response to dengue virus (DENV) is an essential component in the design and development of effective vaccines and immunotherapeutics. Here we present the structure of a cross-reactive, neutralizing antibody, 3E31, in complex with domain III (DIII) of the DENV envelope (E) protein and reveal a conserved, temperature-sensitive, cryptic epitope on DIII that is not available in any of the known conformations of E on the dengue virion. We observed that 3E31 inhibits E-mediated membrane fusion, suggesting that the antibody is able to neutralize virus through binding an as-yet uncharacterized intermediate conformation of DENV E and sterically block trimer formation. Finally, we show that, unlike cross-reactive fusion peptide-specific antibodies, 3E31 does not promote antibody-dependent enhancement of infection at sub-neutralizing concentrations. Our results highlight the 3E31 epitope on the E protein DIII as a promising target for immunotherapeutics or vaccine design.

[Mapping of the B Cell Neutralizing Epitopes on ED III of Envelope Protein from Dengue Virus].

Dengue virus (DENV) envelope [E] protein is the major surface protein of the virions that indued neutralizing antibodies. The domain III of envelope protein (EDIII) is an immunogenic region that holds potential for the development of vaccines; however, the epitopes of DENV EDIII, especially neutralizing B-cell linear epitopes, have not been comprehensively mapped. We mapped neutralizing B-cell linear epitopes on DENV-1 EDIII using 27 monoclonal antibodies against DENV-1 EDIII proteins from mice immunized with the DENV-1 EDIII. Epitope recognition analysis was performed using two set of sequential overlapping peptides (16m and 12m) that spanned the entire EDIII protein from DENV-1, respectively. This strategy identified a DENV-1 type- specific and a group-specific neutralizing epitope, which were highly conserved among isolates of DENV-1 and the four DENV serotypes and located at two regions from DENV-1 E, namely amino acid residues 309-320 and 381-392(aa 309-320 and 381-392), respectively. aa310 -319(310KEVAETQHGT319)was similar among the four DENV serotypes and contact residues on aa 309 -320 from E protein were defined and found that substitution of residues E309 , V312, A313 and V320 in DENV-2, -3, -4 isolates were antigenically silent. We also identified a DENV-1 type-specific strain-restricted neutralizing epitope, which was located at the region from DENV-1 E, namely amino acid residues 329-348 . These novel type- and group-specific B-cell epitopes of DENV EDIII may aid help us elucidate the dengue pathogenesis and accelerate vaccine design.

[Establishment and preliminary application of dengue virus envelope domain III IgG antibody capture enzyme-linked immuno-absorbent assay].

OBJECTIVE: To establish a highly sensitive and specific assay to detect dengue virus (DENV) envelope protein domain III (EDIII) IgG antibody, and to explore its value in the diagnosis and seroepidemiological survey of dengue. METHODS: The DENV EDIII IgG antibody capture ELISA was developed using the recombinant full-length DENV EDIII, which was prepared by Pichia yeast expression system as the capture antigen. The serum samples were collected from the same group of 35 DENV-1 patients of primary infection during disease period in 2006 and their follow-up phase in 2010; and the sensitivity of the assay was compared to that of the commercial Panbio DENV IgG ELISA. RESULTS: The sensitivity of DENV EDIII IgG ELISA in detecting the serum samples from disease period and follow-up phase was 87% (20/23) and 94% (33/35), respectively; whereas the sensitivity of Panbio DENV IgG ELISA was 71% (25/35) and 0, respectively. The sensitivity of DENV EDIII IgG ELISA in detecting the serum samples from both periods was similar, without statistical significance (χ(2) = 0.946, P = 0.331). For serum samples from disease period, the sensitivity of DENV EDIII IgG ELISA was comparable with that of Panbio DENV IgG ELISA (χ(2) = 1.924, P = 0.165). However, DENV EDIII IgG ELISA demonstrated a significantly higher sensitivity than Panbio DENV IgG ELISA in detecting the serum samples from follow-up phase (χ(2) = 62.432, P = 0.000). CONCLUSION: DENV EDIII IgG capture ELISA is highly sensitive in detecting IgG in the serum samples from either disease period or follow-up phase. This method might be a promising alternative for diagnosis and seroepidemiologic survey of dengue.

Dengue virus envelope domain III immunization elicits predominantly cross-reactive, poorly neutralizing antibodies localized to the AB loop: implications for dengue vaccine design.

Dengue virus (DENV) is a mosquito-borne virus that causes severe health problems. An effective tetravalent dengue vaccine candidate that can provide life-long protection simultaneously against all four DENV serotypes is highly anticipated. A better understanding of the antibody response to DENV envelope protein domain III (EDIII) may offer insights into vaccine development. Here, we identified 25 DENV cross-reactive mAbs from immunization with Pichia pastoris-expressed EDIII of a single or all four serotype(s) using a prime-boost protocol, and through pepscan analysis found that 60 % of them (15/25) specifically recognized the same highly conserved linear epitope aa 309-320 of EDIII. All 15 complex-reactive mAbs exhibited significant cross-reactivity with recombinant EDIII from all DENV serotypes and also with C6/36 cells infected with DENV-1, -2, -3 and -4. However, neutralization assays indicated that the majority of these 15 mAbs were either moderately or weakly neutralizing. Through further epitope mapping by yeast surface display, two residues in the AB loop, Q316 and H317, were discovered to be critical. Three-dimensional modelling analysis suggests that this epitope is surface exposed on EDIII but less accessible on the surface of the E protein dimer and trimer, especially on the surface of the mature virion. It is concluded that EDIII as an immunogen may elicit cross-reactive mAbs toward an epitope that is not exposed on the virion surface, therefore contributing inefficiently to the mAbs neutralization potency. Therefore, the prime-boost strategy of EDIII from a single serotype or four serotypes mainly elicited a poorly neutralizing, cross-reactive antibody response to the conserved AB loop of EDIII.

Evaluation and analysis of dengue virus enhancing and neutralizing activities using simple high-throughput assays.

The risk of antibody-dependent enhancement (ADE) of dengue virus (DENV) infection is a major obstacle for the development of dengue vaccine candidates. Here, we described a novel approach for assessment of ADE by measuring DENV nonstructural protein 1 (NS1) production in culture supernatants with Fcγ receptor-expressing K562 cells in ELISA format (ELISA-ADE). Enhancing activities quantified by measurement of kinetics of NS1 production were in a good agreement with the results of the virus titration assay. In conjunction with the previously established enzyme-linked immunospot-based micro-neutralization test (ELISPOT-MNT) in 96-well format, the observable dose-response profiles of enhancing and neutralizing activities against all four DENV serotypes were produced with two flaviviral envelope cross-reactive monoclonal antibodies and four primary DENV-1-infected human sera. The simple high-throughput ELISA-ADE assay offers advantages for quantitative measurement of infection enhancement that can potentially be applied to large-scale seroepidemiological studies of DENV infection and vaccination.

Comparison of surface plasmon resonance, resonant waveguide grating biosensing and enzyme linked immunosorbent assay (ELISA) in the evaluation of a dengue virus immunoassay.

Two label-free biosensor platforms, Resonance Waveguide Grating (RWG) and Surface Plasmon Resonance (SPR), were used to rank a large panel of anti-dengue virus NS1 antibodies. Dengue non-structural 1 (NS1) protein is an established serological marker for the early detection of dengue infection. A variety of commercial dengue NS1 antigen capture immunoassays are available in both ELISA and lateral flow format. However, there is a significant scope to improve both the sensitivity and the specificity of those tests. The interactions of antibody (Ab)-antigen (Ag) were profiled, with weak interactions (KD = 1-0.1 µM) able to be detected under static equilibrium conditions by RWG, but not observed to under more rigorous flow conditions using SPR. There were significant differences in the absolute affinities determined by the two technologies, and there was a poor correlation between antibodies best ranked by RWG and the lower limit of detection (LLOD) found by ELISA. Hence, whilst high-throughput RWG can be useful as preliminary screening for higher affinity antibodies, care should be exercised in the assignation of quantitative values for affinity between different assay formats.

[Analysis of serum neutralizing antibody response in patients with primary dengue virus type 1 infection].

OBJECTIVE: To investigate the characteristics and dynamic changes of serum neutralizing antibody response in patients with primary infection of dengue virus type 1 (DENV-1). METHODS: Serum samples were obtained from the same patients with primary infection of DENV-1 within 2 weeks after symptom onset in 2006 and in 2010. A group-specific DENV NS1 capture ELISA-based micro-neutralizing test (ELISA-MNT) capable of detecting neutralizing antibodies against all the 4 serotypes of DENV was used to test the neutralizing antibody titers against DENV in the serum samples. The neutralizing antibody titers against a standard strain and 2 clinically isolated strains of DENV-1 were detected in serum samples collected in 2010. RESULTS: Cross-reactive neutralizing antibody response against all the 4 serotypes of DENV was found in both of the serum samples collected in 2006 and 2010, but the samples collected in 2006 showed stronger cross-reactive neutralizing antibody responses. The neutralizing antibody against DENV-2, rather than the anticipated DENV-1 antibody, had the highest titer in the samples collected in 2006, whereas the antibody against homologous DENV-1 had the highest titer in the samples obtained in 2010. The neutralizing antibody titers against the homologous DENV-1 was significantly higher in samples collected in 2010 (U=86.500, P=0.000), which also demonstrated significantly different neutralizing antibody titers against the 3 different strains of DENV-1 (Χ(2)=12.123, P=0.002). CONCLUSION: The production of cross-reactive neutralizing antibodies between the 4 serotypes of DENV is a characteristic of DENV infection, particularly during early infection, but only the homologous neutralizing antibody increases obviously over time. The titers of the neutralizing antibodies against different strains, even of the same serotype, may differ distinctly.

[Effect of lentiviral vector-mediated short hairpin RNA targeting survivin against endometriosis-like lesions in chick embryo chorioallantoic membrane].

OBJECTIVE: To investigate the inhibitory effect of lentiviral vector-mediated short hairpin RNA targeting survivin (LV-survivin shRNA) on angiogenesis and growth of endometriosis-like lesions in chick en embryo chorioallantoic membrane. METHODS: Eutopic endometrium from women with endometriosis was transplanted onto the non-vascular region of (CAM), where LV-survivin shRNA was delivered subsequently. The angiogenesis and the growth of endometriosis-like lesions in the CAM model were evaluated. RESULTS: The angiogenesis and formation of endometriosis-like lesions were significantly suppressed in the CAM model by treatment with LV-survivin shRNA in comparison with those in the untreated CAM models and models treated with empty LV or DMEM (P<0.001). LV-survivin shRNA also caused a significantly higher cell apoptotic rate in the endometriosis-like lesions than the other treatments (P<0.001) and induced necrosis in the lesions. CONCLUSION: LV-survivin shRNA can effectively inhibit angiogenesis induced by the eutopic endometrium and markedly suppress the formation of endometriosis-like lesions in the CAM model.

Prognostic implications of microRNA-100 and its functional roles in human epithelial ovarian cancer.

Dysregulation of microRNAs (miRNAs) has been found to be associated with a variety of diseases, including epithelial ovarian cancer (EOC). Recently, miR-100 was reported to be downregulated in human ovarian carcinoma, however, the clinical significance and functional roles of miR-100 expression in human EOC are unclear. TaqMan real-time quantitative RT-PCR assay was performed to detect the expression of miR-100 in 98 EOC tissues and 15 adjacent normal epithelial tissues. The relationship between miR-100 expression and clinicopathological factors in 98 EOC patients was statistically analyzed. The effect of miR-100 expression on patient survival was determined. Finally, the role of miR-100 in EOC cell growth and its possible mechanisms were analyzed with miR-100 precursor or inhibitor-transfected cells. We showed that the level of miR-100 was significantly lower in EOC tissues compared to adjacent normal tissues. Low miR-100 expression was found to be closely correlated with advanced FIGO stage, higher serum CA125 expression level and lymph node involvement. Also, low miR-100 expression was correlated with shorter overall survival of EOC patients, and multivariate analysis showed that the status of miR-100 expression was an independent predictor of overall survival in EOC. Additionally, miR-100 could affect the growth of EOC cells by post-transcriptionally regulating polo-like kinase 1 (PLK1) expression. Together, these results suggest that low miR-100 expression may be an independent poor prognostic factor and miR-100 can function as a tumor suppressor by targeting PLK1 in human EOCs.

Comparison of plaque- and enzyme-linked immunospot-based assays to measure the neutralizing activities of monoclonal antibodies specific to domain III of dengue virus envelope protein.

The plaque reduction neutralization test (PRNT) is used widely to measure the neutralization activity of anti-dengue virus (DENV) antibodies, but it is time-consuming and labor-intensive and has low sample throughput. For fast and convenient measurement of neutralizing antibodies, especially in evaluating the efficiency of the DENV vaccines on a large scale, a new method is needed to replace PRNT. In recent decades, several microneutralization assays have been developed to overcome the limitations of PRNT. In the present study, we evaluated one of these, the enzyme-linked immunospot microneutralization test (ELISPOT-MNT), in comparison with PRNT. ELISPOT-MNT is performed in 96-well format, and the plaques are developed after 2 to 4 days using an ELISA to transform them into spots, which are detected automatically with an ELISPOT instrument. The assay is faster than PRNT, has a high throughput, and is more objective. We used 10 monoclonal antibodies (MAbs) against domain III of the DENV envelope protein (EDIII) to evaluate the two assays; all of these MAbs cross-react with all four serotypes of DENV as measured by immunofluorescence assay. The two neutralization assays were performed simultaneously to measure the 50% inhibitory concentration (IC(50)) of these MAbs. Using PRNT as the reference and treating IC(50) values higher than 50 µg/ml of MAbs as negative, ELISPOT-MNT showed a sensitivity of 95.6% and specificity of 88.24% when 10 MAbs were tested against four DENV serotype strains. A good correlation (R(2) = 0.672; P = 0.000) was observed between the two assays, making ELISPOT-MNT a potentially valuable method for measure of neutralizing antibodies against DENV.

Enzyme-linked immunosorbent assay-format tissue culture infectious dose-50 test for titrating dengue virus.

A dengue nonstructural protein 1 (NS1) antigen capture enzyme-linked immunosorbent assay (ELISA)-based tissue culture infectious dose-50 (TCID(50)) test (TCID(50)-ELISA) was developed as an alternative to the standard plaque assay for titrating dengue virus. Virus titers obtained by TCID(50)-ELISA were comparable to those obtained by the plaque assay and by the traditional TCID(50)-cytopathic effect (CPE) test (TCID(50)-CPE), with a better reproducibility and a lower coefficient of variation. Quantitative comparison of TCID(50)-ELISA and TCID(50)-CPE resulted in a correlation coefficient of 0.976. Moreover, this new method showed a wider application to C6/36, Vero E6, BHK-21, and Vero cells compared with other titration methods. In summary, the novel TCID(50)-ELISA method described here provides a more reliable and more accurate alternative compared to the plaque assay and TCID(50)-CPE for titration of dengue virus.

Full serotype- and group-specific NS1 capture enzyme-linked immunosorbent assay for rapid differential diagnosis of dengue virus infection.

Dengue virus (DENV), a member of the Flavivirus family, has four distinct serotypes (DENV serotype 1 [DENV1], DENV2, DENV3, and DENV4) that require differentiation for the effective prevention of morbid disease. Early and rapid differentiation between flaviviruses remains challenging. Full assays combining four individual, serotype-specific and one group-specific nonstructural protein 1 (NS1) antigen capture enzyme-linked immunosorbent assays (ELISAs) based on monoclonal antibodies (MAbs) against DENV NS1 were developed and validated. The sensitivities and specificities of the full NS1 ELISAs were evaluated with viral cultures and dengue acute-phase sera. Four serotype-specific NS1 ELISAs displayed high specificities for the detection and differentiation of appropriate serotypes. The group-specific NS1 ELISA was broadly reactive with the four dengue virus serotypes. None of the NS1 ELISAs displayed cross-reactivity with the other flaviviruses or samples from febrile patients with non-dengue virus infections. The full serotype- and group-specific MAb-based NS1 capture ELISAs may provide tools for the early detection and typing of dengue infection, which is preferable to reverse transcriptase PCR (RT-PCR) for the rapid differential diagnosis of dengue virus infection in the field.

[Inhibitory effect of short hairpin RNA targeting survivin gene on human ectopic endometrial cells].

OBJECTIVE: To construct a lentiviral expression vector for short hairpin RNA (shRNA) of human survivin gene, and assess its gene silencing effect in human ectopic endometrial cells. METHODS: Human survivin gene shRNA sequence was designed using a software available on-line. The synthesized shRNA sequence was cloned into the pGCL-GFP vector to construct LV-survivin shRNA, which was confirmed by PCR and DNA sequence analysis. The packaging 293T cells were cotransfected with LV-survivin shRNA, pHelper 1.0 and pHelper 2.0, and the titer of the lentivirus was determined. The recombinant lentivirus was injected into human ectopic endometrial cells and the survivin mRNA expression was examined by semi-quantitative reverse transcription-polymerase chain reaction (RT-PCR) in comparison with that in the non-transfected and blank vector-transfected human ectopic endometrial cells. RESULTS: PCR analysis and DNA sequencing confirmed correct insertion of the shRNA sequence into the lentiviral vector. The titer of virus after packaging was 8x10(8) U/ml. Survivin mRNA expression in human ectopic endometrial cells transfected by LV-survivin shRNA was significantly inhibited compared with those in the non-transfected and empty vector transfected human ectopic endometrial cells (P<0.01), and no significant difference was found between the latter two groups. CONCLUSION: The lentiviral shRNA vector of survivin gene constructed can effectively inhibit the expression of survivin gene in human ectopic endometrial cells in vitro. This vector provides a tool for investigating the role of survivin gene in the occurrence and progression of endometriosis and for searching new therapeutic targets.