A phytoplasma effector destabilizes chloroplastic glutamine synthetase inducing chlorotic leaves that attract leafhopper vectors.

Leaf yellowing is a well-known phenotype that attracts phloem-feeding insects. However, it remains unclear how insect-vectored plant pathogens induce host leaf yellowing to facilitate their own transmission by insect vectors. Here, we report that an effector protein secreted by rice orange leaf phytoplasma (ROLP) inhibits chlorophyll biosynthesis and induces leaf yellowing to attract leafhopper vectors, thereby presumably promoting pathogen transmission. This effector, designated secreted ROLP protein 1 (SRP1), first secreted into rice phloem by ROLP, was subsequently translocated to chloroplasts by interacting with the chloroplastic glutamine synthetase (GS2). The direct interaction between SRP1 and GS2 disrupts the decamer formation of the GS2 holoenzyme, attenuating its enzymatic activity, thereby suppressing the synthesis of chlorophyll precursors glutamate and glutamine. Transgenic expression of SRP1 in rice plants decreased GS2 activity and chlorophyll precursor accumulation, finally inducing leaf yellowing. This process is correlated with the previous evidence that the knockout of GS2 expression in rice plants causes a similar yellow chlorosis phenotype. Consistently, these yellowing leaves attracted higher numbers of leafhopper vectors, caused the vectors to probe more frequently, and presumably facilitate more efficient phytoplasma transmission. Together, these results uncover the mechanism used by phytoplasmas to manipulate the leaf color of infected plants for the purpose of enhancing attractiveness to insect vectors.

Review on recent progress in on-line monitoring technology for atmospheric pollution source emissions in China.

Emissions from mobile sources and stationary sources contribute to atmospheric pollution in China, and its components, which include ultrafine particles (UFPs), volatile organic compounds (VOCs), and other reactive gases, such as NH3 and NOx, are the most harmful to human health. China has released various regulations and standards to address pollution from mobile and stationary sources. Thus, it is urgent to develop online monitoring technology for atmospheric pollution source emissions. This study provides an overview of the main progress in mobile and stationary source monitoring technology in China and describes the comprehensive application of some typical instruments in vital areas in recent years. These instruments have been applied to monitor emissions from motor vehicles, ships, airports, the chemical industry, and electric power generation. Not only has the level of atmospheric environment monitoring technology and equipment been improving, but relevant regulations and standards have also been constantly updated. Meanwhile, the developed instruments can provide scientific assistance for the successful implementation of regulations. According to the potential problem areas in atmospheric pollution in China, some research hotspots and future trends of atmospheric online monitoring technology are summarized. Furthermore, more advanced atmospheric online monitoring technology will contribute to a comprehensive understanding of atmospheric pollution and improve environmental monitoring capacity.

Intracranial complete response to toripalimab and anlotinib in a patient with recurrent brain metastases of small cell lung cancer after failure of second-line maintenance therapy: a case report.

Background: Approximately 10-25% of patients with small cell lung cancer (SCLC) have brain metastases at the time of diagnosis. Radiotherapy is a common treatment for brain metastases, but the relapse rates are high. Accumulating evidence suggests that immunotherapy may have a better therapeutic effect for brain metastases. Here, we reported a patient with limited-stage SCLC and relapsed brain metastases who achieved sustained intracranial complete response (CR) to programmed cell death-1 (PD-1) inhibitor toripalimab and multikinase inhibitor anlotinib. Case Description: A 59-year-old female patient developed brain metastases after initial treatment for limited stage SCLC. CR of brain lesions was achieved after intensity-modulated radiation therapy followed by chemotherapy with irinotecan plus lobaplatin and concurrent anlotinib. PD-1 inhibitor sintilimab combined with anlotinib were given as maintenance therapy. Small and asymptomatic brain lesions relapsed 2.5 months after achieving CR. Another three cycles of sintilimab combined with anlotinib failed to control the relapsed brain lesions. Following two cycles of another PD-1 inhibitor toripalimab combined with anlotinib, the relapsed brain metastases disappeared. Then the patient received another seven cycles of this regimen with sustained CR, and no serious adverse reactions occurred. Interestingly, the primary lung tumor achieved sustained CR from the end of initial treatment to the last follow-up. Conclusions: This case suggests that toripalimab in combination with anlotinib may be a promising treatment option for patients with brain metastases from SCLC.

Effects of micro-nano bubble with CO2 treated water on the growth of Amaranth green (Amaranthus viridis).

The micro and nano bubble (MNB) technology, due to its promising features and advantages, has become increasingly popular in agriculture. MNB-treated water positively impacts plant growth, especially when it is treated with a combination of gas-like carbon dioxide (CO2), injected through the MNB generator. Therefore, this study used MNB water with CO2 that are small bubbles of nanometer and micrometer diameters having several unique physical properties that make them useful for water treatments. This research evaluates the effect of MNBs and CO2-treated water on leafy vegetable Amaranth green (Amaranthus viridis). The experiment divided the Amaranth plants into three major groups, G1, G2, and G3, irrigated by MNB water with dissolved CO2, MNBs with only Air, and simple tap water, respectively. The first treatment group (G1) (MNBs with CO2) was further divided into three sub-divisions, i.e., G1A, G1B, G1C, and the second treatment group G2 (MNBs with Air) was divided into three sub-groups, i.e., G2A, G2B, and G2C, while the third group G3 with only one category as only controlled group. These sub-divisions of treatment groups G1 and G2 were done to investigate the impact of MNBs and CO2 treated water with different time durations. For example, in G1A, the water treatment with MNBs and CO2 was kept five minutes, for G1B 10 minutes, and G1C 15 minutes. Similar method was adopted for G2 as well. According to the results, water treated with MNB and CO2 has a significant (90%) impact on the Amaranth germination rate and plant growth. Specifically, pots irrigated with the MNBs + CO2-treated water showed better germination and plant growth rate than the MNBs + Air treated water. Overall, both treatment groups, G1 and G2, showed significantly higher impacts than the CK groups (simple water). Further, this experiment showed that the 10 and 15 minutes treatment of water (G1B, G1C and G2B, G2C) increased the stem height and root size compared to the 5 minutes treated water (G1A, G2A). This study concludes that the water with MNBs has a positive impact on the vegetables and can be an effective technology to improve crop yield.

A Co-Nanoparticles Modified Electrode for On-Site and Rapid Phosphate Detection in Hydroponic Solutions.

Conventional strategies for determining phosphate concentration is limited in efficiency due to the cost, time, and labor that is required in laboratory analysis. Therefore, an on-site and rapid detection sensor for phosphate is urgently needed to characterize phosphate variability in a hydroponic system. Cobalt (Co) is a highly sensitive metal that has shown a selectivity towards phosphate to a certain extent. A disposable phosphate sensor based on the screen-printed electrode (SPE) was developed to exploit the advantages of Co-nanoparticles. A support vector machine regression model was established to predict the concentration of phosphate in the hydroponic solutions. The results showed that Co-nanoparticles improve the detection limit of the sensor in the initial state. Meanwhile, the corrosion of Co-nanoparticles leads to a serious time-drift and instability of the electrodes. On the other hand, the coefficient of variation of the disposable phosphate detection chip is 0.4992%, the sensitivity is 33 mV/decade, and the linear range is 10-1-10-4.56 mol/L. The R2 and mean square error of the buffer-free sensor in the hydroponic solution are 0.9792 and 0.4936, respectively. In summary, the SPE modified by the Co-nanoparticles is a promising low-cost sensor for on-site and rapid measurement of the phosphate concentration in hydroponic solutions.

Rice yellow stunt virus activates polyamine biosynthesis to promote viral propagation in insect vectors by disrupting ornithine decarboxylase antienzyme function.

Intracellular polyamines (putrescine, spermidine, and spermine) have emerged as important molecules for viral infection; however, how viruses activate polyamines biosynthesis to promote viral infection remains unclear. Ornithine decarboxylase 1 (ODC1) and its antienzyme 1 (OAZ1) are major regulators of polyamine biosynthesis in animal cells. Here, we report that rice yellow stunt virus (RYSV), a plant rhabdovirus, could activate putrescine biosynthesis in leafhoppers to promote viral propagation by inhibiting OAZ1 expression. We observed that the reduction of putrescine biosynthesis by treatment with difluormethylornithine (DFMO), a specific nontoxic inhibitor of ODC1, or with in vitro synthesized dsRNAs targeting ODC1 mRNA could inhibit viral infection. In contrast, the supplement of putrescine or the increase of putrescine biosynthesis by treatment with dsRNAs targeting OAZ1 mRNA could facilitate viral infection. We further determined that both RYSV matrix protein M and ODC1 directly bind to the ODC-binding domain at the C-terminus of OAZ1. Thus, viral propagation in leafhoppers would decrease the ability of OAZ1 to target and mediate the degradation of ODC1, which finally activates putrescine production to benefit viral propagation. This work reveals that polyamine-metabolizing enzymes are directly exploited by a vector-borne virus to increase polyamine production, thereby facilitating viral infection in insect vectors.

Simulation-Based Design and Optimization of Rectangular Micro-Cantilever-Based Aerosols Mass Sensor.

Micro-Cantilever (MCL) is a thin film structure that is applied for aerosol particle mass sensing. Several modifications to the rectangular MCL (length-to-width ratio, slots at the anchor, serrations at its side edges) are made to deduce the role and influence of the shape of rectangular MCL-based aerosol mass sensors and reduce gas damping. A finite element fluid-structure interaction model was used to investigate the performance of MCL. It is found that (I) the mass sensitivity and quality factor decline with the increasing of length-to-width ratio which alters the resonant frequency of the MCL. The optimum conditions, including the length-to-width ratio (σlw = 5) and resonant frequency (f0 = 540.7 kHz) of the MCL, are obtained with the constant surface area (S = 45,000 µm2) in the frequency domain ranging from 0 to 600 kHz. (II) The slots can enhance the read-out signal and bring a small Q factor drop. (III) The edge serrations on MCL significantly reduce the gas damping. The results provide a reference for the design of aerosol mass sensor, which makes it possible to develop aerosol mass sensor with high frequency, sensitivity, and quality.

ZEB1-AS1 is associated with poor prognosis in non-small-cell lung cancer and influences cell migration and apoptosis by repressing ID1.

Long non-coding RNAs (lncRNAs) have been reported to play a vital role in non-small-cell lung cancer (NSCLC). ZEB1-AS1 overexpression predicts a poor prognosis in osteosarcoma and colorectal cancers. In the current study, we determined the clinical significance and prognostic value of ZEB1-AS1 in patients with NSCLC. The expression of ZEB1-AS1 and inhibitor of differentiation-1 (ID1) was measured using qRT-PCR and Western blot. Cell growth, migration, and invasion were determined using colony formation assays, Transwell assay, and flow cytometry, respectively. Tumor growth was determined with a xenograft model. ZEB1-AS1 was significantly up-regulated in NSCLC tissues compared with normal samples. ZEB1-AS1 overexpression was significantly associated with advanced tumor, lymph node, and metastases (TNM) stage and tumor size, as well as poorer overall survival. Moreover, ZEB1-AS1 knockdown inhibited NSCLC cell proliferation and migration, and promoted cell apoptosis. In addition, a chromatin immunoprecipitation assay revealed that ZEB1-AS1 interacted with STAT3, thereby repressing ID1 expression. Furthermore, rescue experiments indicated that ZEB1-AS1 functioned as an oncogene partly by repressing ID1 in NSCLC cells. Taken together, our findings indicate that ZEB1-AS1 serves as a promising therapeutic target to predict the prognosis of NSCLC.

Elevated serum soluble programmed cell death ligand 1 concentration as a potential marker for poor prognosis in small cell lung cancer patients with chemotherapy.

BACKGROUND: Potential relationship between serum soluble programmed cell death ligand 1 and prognosis of small cell lung cancer is not well explored. The aim of the study was to reveal the prognostic significance of serum soluble programmed cell death ligand 1 in patients with small cell lung cancer. METHODS: A total of 250 small cell lung cancer patients and 250 controls were included. Research information was obtained from their medical records. Blood samples were collected on admission. Serum concentration of programmed cell death ligand 1 was measured using Enzyme-Linked Immunosorbent Assay. The patients underwent cisplatin-etoposide chemotherapy with a maximum of six cycles. Subsequently, they were followed-up for 12 months, and therapeutic response and cancer death were recorded. RESULTS: Serum concentration of programmed cell death ligand 1 was higher in the patients than in the controls on admission (P < 0.001). After chemotherapy, 112 patients had no response to this therapy. In the 12-month follow up period, 118 patients died due to this cancer. Multivariate Cox regression model revealed that the higher serum concentration of programmed cell death ligand 1 on admission was associated with the higher risk of no response to chemotherapy or cancer caused death (HR: 1.40, 95% CI: 1.05 ~ 1.87; HR: 1.43, 95% CI: 1.08 ~ 1.87). CONCLUSION: Elevated serum concentration of soluble programmed cell death ligand 1 might be an independent risk factor for non-response to chemotherapy and cancer caused death in small cell lung cancer patients.

Fibroblast activation protein-α in tumor cells promotes colorectal cancer angiogenesis via the Akt and ERK signaling pathways.

Fibroblast activation protein-α (FAP-α) is a cell surface serine protease of the post-prolyl peptidase family, and stromal FAP-α expression may serve important functions in tumor occurrence and progression. In recent years, FAP-α expression in tumor cells has been detected in a number of types of tumor, and its roles in tumor growth and metastasis have been reported. However, the presence of FAP-α in colorectal cancer (CRC) cells lacks sufficient evidence and its role in angiogenesis remains unknown. The present study confirmed FAP-α expression in CRC cells at the tissue and cellular level, using immunohistochemistry and western blot analysis, respectively; it additionally identified that FAP-α in CRC cells was positively associated with vascular endothelial growth factor (VEGF)-A expression and microvessel density in stained tissue samples for the first time. In addition, western blotting identified that FAP-α overexpression in SW1116 cells significantly upregulated VEGF-A expression, and silencing of FAP-α in HT29 cells markedly inhibited VEGF-A expression. Survival analysis demonstrated that patients with high expression of FAP-α and VEGF-A had the shortest survival time. To detect the effects of FAP-α on human umbilical vein endothelial cells (HUVECs), conditioned medium (CM) from CRC cell lines was used and it was identified that CM from SW1116 cells with overexpressed FAP-α exhibited significantly increased VEGF-R2, phosphorylated extracellular signal-regulated kinase (p-ERK) and p-RAC-α serine/threonine-protein kinase (Akt) in HUVECs, in addition to the proliferation rate. Conversely, CM from HT29 cells with FAP-α silenced exhibited a significantly inhibited proliferation rate. Molecular mechanism analysis demonstrated that p-ERK and p-Akt in SW1116 and HT29 cells were affected by alterations in FAP-α expression, and treatment with a p-ERK inhibitor (U0126) and p-Akt inhibitor (LY294002) ameliorated VEGF-A upregulation induced by FAP-α overexpression. All the results confirmed the presence of FAP-α in CRC cells and suggested that FAP-α may effectively promote angiogenesis in CRC via the Akt and ERK signaling pathways.

Comparative study of cylindrical and parallel-plate electrophoretic separations for the removal of ions and sub-23 nm particles.

Cylindrical and parallel-plate electrophoretic separations for the removal of ions and sub-23 nm particles were compared in this study. First, COMSOL Multiphysics® software was utilized to simulate the ion and particle trajectories inside both electrophoretic separations. The results show that ions and sub-23 nm particles are removed simultaneously and that all particles can pass through both electrophoretic separations smoothly at a trap voltage of 25 V. The experimental results show that ion losses become smaller with increasing ion flow rates, and ion losses of the cylindrical and parallel-plate electrophoretic separations range from 56.2 to 71.6% and from 43.8 to 59.6%, respectively, at ion flow rates ranging from 1-3 L/min. For the removal of ions and sub-23 nm particles, the collection efficiency of both electrophoretic separations can reach 100%, but the parallel-plate electrophoretic separation requires a lower trap voltage. The particle loss of the parallel-plate electrophoretic separation is under approximately 10%, which is lower than that of the cylindrical electrophoretic separation. In particular, for large particles (800-2500 nm), the particle losses inside the cylindrical electrophoretic separation are approximately two times higher than those inside the parallel-plate electrophoretic separation. The parallel-plate electrophoretic separation is beneficial for the removal of ions and sub-23 nm particles.

All-Fiber Configuration Laser Self-Mixing Doppler Velocimeter Based on Distributed Feedback Fiber Laser.

In this paper, a novel velocimeter based on laser self-mixing Doppler technology has been developed for speed measurement. The laser employed in our experiment is a distributed feedback (DFB) fiber laser, which is an all-fiber structure using only one Fiber Bragg Grating to realize optical feedback and wavelength selection. Self-mixing interference for optical velocity sensing is experimentally investigated in this novel system, and the experimental results show that the Doppler frequency is linearly proportional to the velocity of a moving target, which agrees with the theoretical analysis commendably. In our experimental system, the velocity measurement can be achieved in the range of 3.58 mm/s-2216 mm/s with a relative error under one percent, demonstrating that our novel all-fiber configuration velocimeter can implement wide-range velocity measurements with high accuracy.

Matrine induces mitochondrial apoptosis in cisplatin-resistant non-small cell lung cancer cells via suppression of ß-catenin/survivin signaling.

Matrine is an alkaloid isolated from Sophora flavescens and shows anticancer activities. The present study was carried out to determine the cytotoxic effects of matrine on cisplatin-resistant non-small cell lung cancer (NSCLC) cells and the associated molecular mechanisms. Parental and cisplatin-resistant A549 and H460 NSCLC cells were treated with 1 or 2 g/l of matrine for 48 h, and cell viability and apoptosis were assessed. ß-catenin-mediated transcriptional activity, mitochondrial membrane potential (ΔΨm) changes, activation of caspases, and survivin expression were examined. The effect of overexpression of survivin on the anticancer activity of matrine was investigated. Compared to the parental cells, cisplatin-resistant NSCLC cells showed increased ß-catenin transcriptional activity. Matrine treatment resulted in a significant reduction in ß-catenin activation and survivin expression in the cisplatin-resistant cells. Matrine caused apoptotic death in the cisplatin-resistant NSCLC cells, coupled with loss of ΔΨm and activation of caspase-9 and -3. Matrine-induced apoptosis of the cisplatin-resistant NSCLC cells was significantly reversed by overexpression of survivin. In conclusion, matrine exposure induces mitochondrial apoptosis in cisplatin-resistant NSCLC cells, which is largely mediated through inactivation of ß-catenin/survivin signaling. Further investigation of the therapeutic benefit of matrine in overcoming cisplatin resistance in NSCLC is warranted.

Isolation of a new lycodine alkaloid from Lycopodium japonicum.

A new lycodine alkaloid, N-methylhydroxypropyllycodine (1), was isolated from the club moss Lycopodium japonicum Thunb, together with five known compounds, N-methyllycodine (2), huperzinine (3), ß-obscurine (4), α-obscurine (5) and des-N-methyl-α-obscurine (6). Their structures were elucidated by spectroscopic analyses, including 2D NMR techniques.

A substitution in nsP1 combined with a double substitution in E2 glycoprotein renders Sindbis-like virus XJ-160 fully neurovirulent for adult mice.

Sindbis virus (SV) can be rendered neurovirulent for adult mice by a double substitution within the E2 glycoprotein, including replacing Gln at position 55 of E2 with a His (E2-55: Gln-His) and E2-70: Glu to Lys. However, the mutant Sindbis-like virus XJ-160 with the double substitution (BR-E5570) does not show neurovirulence for adult mice, although the mutant apparently reduced the average survival time of neonatal mice. To produce an XJ-160 virus neurovirulent for adult mice, the BR-E5570 virus containing the double substitution was provided with another substitution in the nsP1 region (nsP1-173: Thr-Ile), which could enhance viral infectivity and neurovirulence for neonatal mice. The mutant containing these three substitutions was accordingly designated as BR-5570-ns173. Like the BR-XJ160 virus derived from the wild-type clone, BR-E5570 and BR-E5570-ns173 were both virulent for newborn mice, between which BR-E5570-ns173 virus showed the greatest neurovirulence. Furthermore, only BR-E5570-ns173 virus was fully neurovirulent for 14-day-old mice, and this fatal adult mouse-virulence was dependent on the E2 double substitutions at positions 55 and 70. Compared with BR-XJ160, both the mutants showed a higher capacity for propagation both in cultured cells and in the mouse brain. In particular, BR-E5570-ns173 virus showed a more persistent existence and higher titer in the brains of 7-day-old mice. These findings indicate that the substitution at nsP1-173 combination with a double substitution in the E2 region renders the XJ-160 virus fully neurovirulent for adult mice, and this neurovirulence may be related to the increased efficiency and persistence of propagation of this virus.

[Development and Application of An Assay for High-throughput Antiviral Compounds Screening against Alphaviruses].

To establish a cell-based rapid luciferase suppression assay for high-throughput screening (HTS) anti-alphaviruses compounds screening, which could cause viral encephalitis, raise the social issues associated directly with public health and huge economic burden to the society. The Gaussia luciferase assay system was used for HTS model for identifying inhibitors of labeled virus XJ160-GLUC. The decreased 50% GLUC activity inhibition ratio was deemed to be the screening positive index. The reaction system in this model was optimized, and the reliability of the model was evaluated. For HTS model's optimization, cells were infected with XJ160-GLUC at an MOI of 0.025 PFU/cell. The supernatant treated with compounds 48h were collected for GLUC expression detection. In the model, Z' factor was up to 0.71, demonstrating that HTS assay for identifying inhibitors that target all aspects of the viral life cycle of XJ160-GLUC was stable and reliable. After screening 8080 compounds (five-in-one), 341 positive samples were selected, and the positive rate was 4.2% with a cutoff at 50% inhibition. Then 1705 compounds were screened subsequently and the positive rate was 1.1% with obtaining 19 positive compounds. These results will lay the foundation for finding the anti-alphaviruses' drug targets.

Arctigenin enhances chemosensitivity to cisplatin in human nonsmall lung cancer H460 cells through downregulation of survivin expression.

Arctigenin, a dibenzylbutyrolactone lignan, enhances cisplatin-mediated cell apoptosis in cancer cells. Here, we sought to investigate the effects of arctigenin on cisplatin-treated non-small-cell lung cancer (NSCLC) H460 cells. The 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay and annexin-V/propidium iodide staining were performed to analyze the proliferation and apoptosis of H460 cells. Arctigenin dose-dependently suppressed cell proliferation and potentiated cell apoptosis, coupled with increased cleavage of caspase-3 and poly(ADP-ribose) polymerase. Moreover, arctigenin sensitized H460 cells to cisplatin-induced proliferation inhibition and apoptosis. Arctigenin alone or in combination with cisplatin had a significantly lower amount of survivin. Ectopic expression of survivin decreased cell apoptosis induced by arctigenin (P < 0.05) or in combination with cisplatin (P < 0.01). Moreover, arctigenin (P < 0.05) or in combination with cisplatin (P < 0.01) induced G1/G0 cell-cycle arrest. Our data provide evidence that arctigenin has a therapeutic potential in combina-tion with chemotherapeutic agents for NSLC.

[Construction and characterization of EGFP reporter gene labeled Sindbis virus].

OBJECTIVE: To construct and characterize EGFP reporter gene labeled Sindbis virus (SINV). METHODS: The reporter gene EGFP was inserted into the genome of infectious clone pBR-XJ160 by using multi-fusion long fragment PCR method. Then apply reverse genetic manipulation technique to rescue and obtain EGFP labeled SINV. RESULTS: We successively obtained labeled SINV, which has good fluorescent expression characteristics and genetic stability. CONCLUSION: The labeled virus can be seen in living cells and living body, and this serves as a good tool for cell and tissue tropism and biological function study of viruses. This study laid a foundation for further studying the cell tropism, biological functions and infection mechanism of SINV.

Development of EGFP/GLUC-tagged Sindbis-like virus XJ-160.

Based on an infectious clone of Sindbis-like virus XJ-160, recombinant vectors containing a reporter gene (enhanced green fluorescence protein [EGFP] or Gaussia luciferase [GLUC]) were constructed by placing the reporter gene cassette containing the subgenomic promoter behind the 3' terminus of the viral structural protein gene. EGFP/GLUC-tagged Sindbis-like viruses were rescued in BHK-21 cells transfected with transcripts produced from the recombinant vectors. EGFP expression and strong luciferase activity were detected in BHK-21 cells infected with repeated passages of the EGFP/GLUC-tagged viruses, revealing the genetic stability of the chimeric viruses. The EGFP/GLUC-tagged Sindbis viruses reported will contribute to the assessment of viral replication and proliferation, tracking and elucidating Alphavirus-host interactions, and screening for antiviral compounds.

Induction of humoral and cellular immune responses against hepatitis C virus by vaccination with replicon particles derived from Sindbis-like virus XJ-160.

A replication-defective, recombinant Sindbis virus vector was utilized in a novel immunization strategy to induce humoral and cellular responses against hepatitis C virus (HCV). The recombinant vector, pVaXJ-E1E2, expressing the gene for HCV glycoproteins E2 and E1, was constructed by inserting the E1E2 gene into the replicon pVaXJ, a DNA vector derived from Sindbis-like virus XJ-160. The defective replicon particles, XJ-E1E2, were produced by transfecting BHK-21(E+Capsid) cells, the packaging cell lines for the vector from XJ-160 virus, with pVaXJ-E1E2. Both glycoproteins, E2 and E1, were stably expressed, as indicated by immunofluorescence assay (IFA) and Western blotting. Mice were vaccinated using a prime-boost strategy with XJ-E1E2 particles combined with Freund's incomplete adjuvant via intramuscular injection at 0 and 2 weeks. HCV-specific IgG antibody levels and cellular immune responses were evaluated by IFA and IFN-γ ELISPOT, respectively. The results showed that the defective XJ-E1E2 particles in combination with Freund's incomplete adjuvant induced effective humoral and cellular immune responses against HCV glycoprotein E1 or E2, suggesting that a defective Sindbis particle vaccine is capable of eliciting an effective immune response. These findings have important implications for the development of HCV vaccine candidates.