Recent advances in nanocarrier-based drug delivery systems in treatment of rheumatoid arthritis / 中国中药杂志

Rheumatoid arthritis(RA) is a widely prevalent autoimmune inflammatory disease that severely affects patients' quality of life. Currently, conventional formulations against RA have several limitations, such as nonspecificity, poor efficacy, large drug dosages, frequent administration, and systemic side effects. Nanotechnology-based drug delivery systems have emerged as a promising stra-tegy for the diagnosis and treatment of RA since nanotechnology can overcome the limitations of traditional treatments and simplify the complexity of the disease. These systems enable targeted delivery of anti-inflammatory drugs to the inflamed areas through active and passive targeting, achieving specificity to the joints, overcoming the need for increased dosage and administration frequency, and reducing associated adverse reactions. This article aimed to review nanocarrier-based drug delivery systems in the field of RA and elucidate how nanosystems can be utilized to deliver therapeutic drugs to inflamed joints for controlling RA progression. By discussing the current issues and challenges faced by nanodrug delivery systems and highlighting the urgent need for solutions, this article offers theoretical support for further research on nanotechnology-based co-delivery systems in the future.

Advance on nitric oxide combined photothermal therapy for tumor treatment / 药学学报

Photothermal therapy (PTT) has attracted significant attention due to minimal side effects and high treatment specificity. However, it often requires very high temperature to achieve complete tumor ablation under a single PTT. Such high temperature brings obvious thermal damage and inflammatory response to the body, affecting the therapeutic effect. In recent years, nitric oxide (NO) has been used to significantly inhibit tumor growth and enhance the sensitivity of tumor cells of temperature and drugs, thus enhancing the therapeutic effect. However, compounds as NO donors often have some disadvantages such as poor biocompatibility and untargeted delivery, etc., therefore, this medical application based on NO therapy is limited. In conclusion, the organic combination of NO donors and photothermal agents (PTAs) is expected to overcome the shortcomings of single therapy and achieve the antitumor effect of "1 + 1 > 2". In view of the rapid development of NO combining with PTT in tumor therapy, this review firstly introduces the antitumor mechanisms of different types of NO donors. Then the treatment strategy based on NO combined with PTT is discussed. Finally, the prospects and challenges of this combination therapy strategy in the clinical treatment of cancer are discussed.

Treatment of rheumatoid arthritis by injection of sinomenine solid lipid nanoparticles under a fluorescence endoscopic laser confocal microscope / 中国中药杂志

A fluorescence endoscopic laser confocal microscope(FELCM) was used to direct the injection of sinomenine solid lipid nanoparticles(Sin-SLN) into the joint, and the in vitro effectiveness of Sin-SLN in the treatment of rheumatoid arthritis(RA) was evaluated. Sin-SLN was prepared with the emulsion evaporation-low temperature curing method. The Sin-SLN prepared under the optimal conditions showed the encapsulation efficiency of 64.79%±3.12%, the drug loading of 3.84%±0.28%, the average particle size of(215.27±4.21) nm, and the Zeta potential of(-32.67±0.84) mV. Moreover, the Sin-SLN demonstrated good stability after sto-rage for 30 days. The rabbit model of RA was established by the subcutaneous injection of ovalbumin and complete Freund's adjuvant. Five groups were designed, including a control group, a model group, a Sin(1.5 mg·kg~(-1)) group, a Sin-SLN(1.5 mg·kg~(-1)) group, and a dexamethasone(positive drug, 1.0 mg·kg~(-1), ig) group. The control group and the model group only received puncture treatment without drug injection. After drug administration, the local skin temperature and knee joint diameter were monitored every day. The knee joint diameter and the local skin temperature were lower in the drug administration groups than in the model group(P<0.05, P<0.01). FELCM recorded the morphological alterations of the cartilage of knee joint. The Sin-SLN group showed compact tissue structure and smooth surface of the cartilage. Enzyme-linked immunosorbent assay(ELISA) was employed to determine the serum le-vels of interleukin-1(IL-1) and tumor necrosis factor-α(TNF-α). The findings revealed that the Sin-SLN group had lower IL-1 and TNF-α levels than the model group(P<0.05, P<0.01). Hematoxylin-eosin(HE) staining was employed to reveal the pathological changes of the synovial tissue, which were significantly mitigated in the Sin-SLN group. The prepared Sin-SLN had uniform particle size and high stability. Through joint injection administration, a drug reservoir was formed. Sin-SLN effectively alleviate joint swelling and cartilage damage of rabbit, down-regulated the expression of inflammatory cytokines, and inhibited the epithelial proliferation and inflammatory cell infiltration of the synovial tissue, demonstrating the efficacy in treating RA.

Application of microneedle-assisted percutaneous drug delivery system in treatment of rheumatoid arthritis:a review / 中国中药杂志

Rheumatoid arthritis(RA) is a chronic degenerative joint disease characterized by inflammation. Due to the complex causes, no specific therapy is available. Non-steroidal anti-inflammatory agents and corticosteroids are often used(long-term, oral/injection) to interfere with related pathways for reducing inflammatory response and delaying the progression of RA, which, however, induce many side effects. Microneedle, an emerging transdermal drug delivery system, is painless and less invasive and improves drug permeability. Thus, it is widely used in the treatment of RA and is expected to be a new strategy in clinical treatment. This paper summarized the application of microneedles in the treatment of RA, providing a reference for the development of new microneedles and the expansion of its clinical application.

Comparative study on establishing an immune tolerance model for allergic conjunctivitis in mice with different methods / 国际眼科杂志(Guoji Yanke Zazhi)

AIM:To establish an immune tolerance model for allergic conjunctivitis in newborn mice with different methods and observe the impact of environmental factors on allergic conjunctivitis in early life.METHOD: A total of 50 Balb/c newborn mice were randomly divided into blank control group, ovalbumin(OVA)+subcutaneous injection group, OVA+nebulized inhalation group, OVA+gastric group, ragweed pollen(RW)+subcutaneous injection group, RW+nebulized inhalation group, RW+gastric group, house dust mite(HDM)+subcutaneous injection group, HDM+nebulized inhalation group, HDM+intragastric group(n=5 animals/group). Except for the blank control group, mice in each group were individually exposed to the corresponding antigens to induce immune tolerance early in life and stimulated with the corresponding antigens in adulthood. The ocular surface was visualized by anterior segment photography. The relative expression level of conjunctival RANTES and IL-17 mRNA was measured by RT-qPCR and serum IL-17 concentration was measured by ELISA.RESULTS: Compared with the blank control group, the relative expression level of conjunctiva IL-17 mRNA in RW+gastric group was the highest, and it was the lowest in RW+subcutaneous group(all P&#x003C;0.05). The relative expression level of conjunctiva RANTES mRNA was the highest in RW+gastric group(P&#x003C;0.001). Compared with the blank control group, the serum concentration of IL-17 was increased in all treatment groups except OVA+nebulizer group and RW+subcutaneous group(P&#x003C;0.05).CONCLUSION: The immune tolerance of allergic conjunctivitis induced by subcutaneous injection of antigen was the most suitable method in the early life of mice.

Primary Study on Chronic Myeloid Leukemia in NCG Mice from Qinba Mushroom / 中国实验血液学杂志

OBJECTIVE@#The present study was to evaluate the anti-tumor effects of acidic RNA protein complex (FA-2-b-β) extracted from the wild edible Qinba mushroom in inducing of apoptosis and immunoregulation of tumor cell.@*METHODS@#Cell proliferation inducing rate of FA-2-b-β to K562 cell was measured using CCK-8. Apoptosis rate was detected by using flow cytometry. Chronic myeloid leukemia model was developed by tail vein injection/subcutaneous inoculation of K562 cells in NCG mice. The tumor burden of mice was observed. The general condition of the mice was monitored twice daily. The peripherivcal full blood counts of mice was tested daily. RT-qPCR and Western blot was FA-2-b-β performed to determine involvement of apoptotic-related gene and protenin, Immunofluorescence and immunohistochemistry was used to detected the expression of CD3, CD4 and CD8.@*RESULTS@#The proliferation and apoptosis of K562 cell could be inhibitied and induced by FA-2-b-β, there was 100% successful in the tumor formation in vivo, after treated by drug for 21 days there were significantly increased peripheral leucocytes, but decreased hemoglobin of mice treated by FA-2-b-β as compared with those in control group. The CD3, CD4 and CD8 showed positive in mice, and the propotation was imbalance, but it showed reserved after treated by FA-2-b-β.@*CONCLUSION@#FA-2-b-β is strong anti-leukemia effect in vitro and in vivo, suggesting the traditional Chinese medicine maybe contribute to the anti-cancer and immunoregulation research.

Research progress on circular RNA in oral squamous cell carcinoma / 华西口腔医学杂志

Circular RNA, a non-coding RNA that forms a covalently closed continuous loop, exists widely in eukaryotic cells. The biogenesis and biological function of this type of RNA indicate that it can play a crucial role in diseases such as tumors, neural system diseases, and cardiovascular diseases; moreover, this RNA may have great potential use as a biomarker in these diseases. Oral squamous cell carcinoma (OSCC) is a common malignancy in oral surgery that is difficult to cure, metastasizes easily, and has poor prognosis. In this review, we summarize the loop-forming mechanisms and functions of circular RNA and describe the progress of current research in the development of oral cancer.

Inhibitory Effects of Agaricus Blazei Murrill (FA-2-b-β) on Proliferation of Chronic Myeloid Leukemia Cells in vivo and Its Mechanism / 中国实验血液学杂志

OBJECTIVE@#To investigated the anti-tumor in vivo effect and mechanism of the acid RNA protein complex (FA-2-b-β) of Agaricus blazei Murrill extract.@*METHODS@#CCK-8 method was used to detected the inhibitory effect of FA-2-b-β on proliferation of primary CML cells from newly diagnosed CML patients, the CML mouse model was established by trail-venous injection of primary CML cells, and the survival time, blood cell count and body weight were observed, the immunoflouresence and immunehistochemistry analysis, RT-qPCR, Western bolt were used to detemine the expression of caspase-3 signal pathway-related apoptosis genes and proteins.@*RESULTS@#The experiments in vitro showed that the proliferative inhibitory rate in drug-treated group increased with concentration- and time-dependent manner (r@*CONCLUSION@#The FA-2-b-β can induce apoptosis of primary CML cells and prolong the survival time of CML model mouse, which may be related with the caspase-3 signal pathway related genes and proteins.

Cross-neutralizing Anti-hemagglutinin Antibodies Isolated from Patients Infected with Avian Influenza A (H5N1) Virus / 生物医学与环境科学（英文）

Objective@#To recover broad-neutralizing monoclonal antibodies (BnAbs) from avian influenza A (H5N1) virus infection cases and investigate their genetic and functional features.@*Methods@#We screened the Abs repertoires of expanded B cells circulating in the peripheral blood of H5N1 patients. The genetic basis, biological functions, and epitopes of the obtained BnAbs were assessed and modeled.@*Results@#Two BnAbs, 2-12D5, and 3-37G7.1, were respectively obtained from two human H5N1 cases on days 12 and 21 after disease onset. Both Abs demonstrated cross-neutralizing and Ab-dependent cellular cytotoxicity (ADCC) activity. Albeit derived from distinct Ab lineages, , V 1-69-D2-15-J 4 (2-12D5) and V 1-2-D3-9-J 5 (3-32G7.1), the BnAbs were directed toward CR6261-like epitopes in the HA stem, and HA I45 in the hydrophobic pocket was the critical residue for their binding. Signature motifs for binding with the HA stem, namely, IFY in V 1-69-encoded Abs and LXYFXW in D3-9-encoded Abs, were also observed in 2-12D5 and 3-32G7.1, respectively.@*Conclusions@#Cross-reactive B cells of different germline origins could be activated and re-circulated by avian influenza virus. The HA stem epitopes targeted by the BnAbs, and the two Ab-encoding genes usage implied the VH1-69 and D3-9 are the ideal candidates triggered by influenza virus for vaccine development.

Characterization and analysis of VH1-2-encoded heterosubtypic antibodies isolated from an avian H5N1 patient / 中华实验和临床病毒学杂志

Objective@#To isolate the cross-reactive antibodies against hemagglutinin of influenza virus and identify its biological function.@*Methods@#The antibodies gene reservoir of cross-reactive and H5N1 pseudotype particles neutralizing B cell circulating in peripheral blood of a human H5N1 case was recovered by in vitro B cell culture, screening, RT-PCR and expression vector cloning techniques. The Ab gene pairing was screened by transient transfection of human kidney 293T cells and detected using ELISA and neutralization test. The heterosubtypic antibodies were prepared and characterized.@*Results@#We discovered the VH1-2-based heterosubtypic antibodies from two B cell lineages could neutralize GX-H5N1 pseudotype particles and have broader binding with Group 1 (including H1, H5, H6 and H9) and H7 subtype.@*Conclusions@#Cross-reactive antibodies can be induced by H5N1 infection.

Protective effects of zingiber officinalis extract on the lens of diabetic rats / 眼科新进展

Objective To observe the protective mechanisms of zingiber officinalis extract on the lens of diabetic rats induced by streptozotocin (STZ).Methods Totally 60 SD rats were randomly divided into A group (normal control rats),B group (diabetic rats),C1 group (DM +50 mg · kg-1 ginger gavage),C2 group (DM + 100 mg ·kg-1 ginger gavage) and C3 group (DM + 300 mg · kg-1 ginger gavage).Each group had 12 rats.And rats in A group received intraperitoneal injection of the same volume of normal saline,while the other groups were intraperitoneally injected with 65 mg · kg-1streptozotocin (STZ).When the animal model was successfully established,both A group and B group were administered orally with normal saline,and the C1,C2 and C3 groups were administered orally with ginger rhizome extract.The changes in blood glucose and lens were observed every week.The rats were sacrificed in succession at 4 week,8 week,12 week,and the lens was removed immediately.The content of aldose reductase (AR) was detected by ELISA,and the expression of glycosylation end products (AGEs) was detected by Western blot.The superoxide dismutase (SOS) activity and malondialdehyde (MDA) content were also detected.Fluorescent Tunel staining was used to detect the apoptosis of the lens epithelial cells.And finally,scanning electron microscopy was used to observe the ultrastructural changes in the lens.Results The activity of SOD in the lens of diabetic rats showed a decreasing trend with statistical significance (all P ＜ 0.05).Changes in the content of MDA in the lens of rats in B,C1,C2 group were statistically significant in 4,8 and 12 weeks after successful modeling (all P ＜ 0.05),but no significant difference in A group and C3 group (both P ＞ 0.05).The persistent increase of AR in group B (P =0.003).The content of AR of the lens in C1,C2,C3 group showed a decreasing trend,and the differences were statistically significant (all P ＜ 0.05).There were significant differences in the expression of AGEs in C1,C2 and C3 group (all P ＜ 0.05).The degree of cortical fiber destruction in group B was progressively aggravated,but the degree of cortical destruction in C1,C2 and C3 group decreased with the increase of ginger gavage concentration through the scanning electron microscopy.It was observed that there was no significant difference in the apoptotic rate of LECs in A group (P =0.191),but the apoptosis of LECs in the rest group showed a rising trend,and the differences were statistically significant (all P ＜ 0.05).Conclusion The effects of ginger gavage extract can delay the opacification of lens and slow down the diabetic development in rats with a dose-independence manner.Ginger gavage extract may play a protective role on the lens of diabetic rats by inhibiting the activity of AR,oxidative stress and the production of AGEs as well as suppress the apoptosis of lens epithelial cells.

Isolation and identification of exosomes derived from bone marrow mesenchymal stem cells / 基础医学与临床

Objective To develop a method of isolation and identification of exosomes of bone marrow mesenchymal stem cells.Methods The mesenchymal stem cells were cultured by whole bone marrow adherence method.A new reagent -exosomes extraction kit was used to isolate and collecte exosomes.The exosomes were identified by elec-tron microscopy,particle size detection,flow cytometry and Western blot.Results The expression of CD45 on the surface of the third generation bone marrow mesenchymal stem cells was negative,and CD73 and CD105 were posi-tive;exosomes derived from bone marrow mesenchymal stem cells were round or oval,the size is non-uniform,the diameter is 30~100 nm,have a complete membrane structure,and containing low-density substances;Particle size detection particle diameter of the main peak was 61.25 nm, in which the diameter of particles was about 20-200 nm accounted for 72.4%;exosome expressed CD63 and CD81;The expression of CD9 and CD63 from cell cul-ture supernatants was positive.Conclusions The exosomes can be collected in the medium of mesenchymal stem cells.The exosomes derived from bone marrow mesenchymal stem cells can be identified by electron microscopy, particle size detection,flow cytometry and Western blot.

Phylogenetic and Molecular Analysis of an H7N7 Avian Influenza Virus Isolated in East Dongting Lake in 2012 / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>In March 2012, an H7N7 subtype avian influenza virus (AIV) named A/wild goose/Dongting/PC0360/2012 (H7N7) (DT/PC0360) was recovered from a wild goose in East Dongting Lake. We performed whole-genome sequencing of the isolate, and analyzed the phylogenetic and molecular characterization.</p><p><b>METHODS</b>RNA was extracted from environment samples (including fecal samples from wild bird or domestic ducks, and water samples) for detecting the presence of Influenza A Virus targeting Matrix gene, using realtime RT-PCR assay. The positive samples were performed virus isolation with embryonated eggs. The subtype of the isolates were identified by RT-PCR assay with the H1-H16 and N1-N9 primer set. The whole-genome sequencing of isolates were performed. Phylogenetic and molecular characterizations of the eight genes of the isolates were analyzed.</p><p><b>RESULTS</b>Our results suggested that all the eight gene segments of DT/PC0360 belonged to the Eurasian gene pool, and the HA gene were belonged to distinct sublineage with H7N9 AIV which caused outbreaks in Mainland China in 2013. The hemagglutinin cleavage site of HA of DT/PC0360 showed characterization of low pathogenic avian influenza virus.</p><p><b>CONCLUSION</b>Strengthening the surveillance of AIVs of wild waterfowl and poultry in this region is vital for our knowledge of the ecology and mechanism of transmission to prevent an influenza pandemic.</p>

A novel reassortant H2N3 influenza virus isolated from China / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To analyze the genetic composition of a novel H2N3 virus isolate identified from a duck cage swab in a live poultry market (LPM) in 2009 in Guangdong province of China.</p><p><b>METHODS</b>PCR-positive specimens were inoculated into embryonated chicken eggs and subtyped by conventional RT-PCR. All segments of the virus A/environment/Guangdong/2/2009 were sequenced, and phylogenetic trees were constructed and analyzed.</p><p><b>RESULTS</b>The genes of this virus belong to Eurasian-lineage avian viruses. The virus is a reassortant with the HA gene from an H2N2 virus and the NA gene from an H5N3 virus. The PB1, PB2, and NP genes were from an H4N6 virus, the PA was from an H3N8 virus, the M gene was from an H1N3 virus, and the NS gene was from an H10N6 virus.</p><p><b>CONCLUSION</b>A novel avian-origin reassortant H2N3 influenza virus was detected in a live poultry market. Its potential impacts and evolution should be closely monitored.</p>

Age-related differences in the biological parameters of vertebral cancellous bone from Chinese women / 中华医学杂志(英文版)

<p><b>BACKGROUND</b>With aging, the human fracture risks gradually increase. This is mainly due to the corresponding changes of the biomechanical parameters of human bone presents with aging. We measured the microstructural parameters of lumbar bone from women in several age groups by micro-computed tomography and scanning electron microscopy. We observed changes in lumbar cancellous bone mineral density and in biomechanical parameters with aging to elucidate the relationship between age and risk of fracture. We provide theoretical support for human pathology, fracture risk increased with age and the individualized of each age group.</p><p><b>METHODS</b>Thirty-two fresh L3 vertebral bodies were donated from 32 women, aged 20-59 years and were divided into four age groups: 20 to 29 years (group A); 30 to 39 years (group B); 40 to 49 years (group C); and 50 to 59 years (group D). Conventional lumbar separation was performed by removing soft tissue and subsidiary structures, leaving only the vertebral body. The vertebral body was cut into halves along the median sagittal plane, maintaining the upper and lower end-plates of each half, and used for biomechanical, morphological, and density measurements.</p><p><b>RESULTS</b>Comparing group A to B, the rod-like trabecular thickness (Tb.Th) decreased; the trabecular spacing (Tb.Sp) increased; the plate-like Tb.Th decreased; bone mineral density, tissue mineral density, bone volume fraction, and bone surface fraction decreased, and the elastic modulus and the ultimate stress decreased (all changes P < 0.05). Similar significant (P < 0.05) trends were obtained when comparing group C to D. With aging, the collagen cross-linking capacity declined, the thickness of the collagen fibrils was variable (ranging from almost the same to loose, sparse, or disordered), and the finer collagen fibrils between the thick filaments were disorganized.</p><p><b>CONCLUSIONS</b>In women aged 20 to 59 years, the rod-like and plate-like Tb.Th of the vertebral body decreased, while Tb.Sp increased. Additionally, the density, elastic modulus, and ultimate stress of the cancellous bone decreased with age. These associated changes in bone microstructure, density, and biomechanics with age may lead to an increasing risk of osteoporosis and fracture.</p>

A novel reassortant H3N8 influenza virus isolated from drinking water for duck in a domestic duck farm in Poyang Lake area / 生物医学与环境科学（英文）

<p><b>OBJECTIVE</b>To conduct a full genome sequence analysis for genetic characterization of an H3N8 influenza virus isolated from drinking water of a domestic duck farm in Poyang Lake area in 2011.</p><p><b>METHODS</b>The virus was cultivated by specific pathogen free (SPF) chicken embryo eggs and was subtyped into hemagglutinin (HA) and neuraminidase (NA) by real-time PCR method. Eight gene segments were sequenced and phylogenetic analysis was conducted.</p><p><b>RESULTS</b>The NA gene of this virus belongs to North American lineage; other seven genes belong to Eurasian lineage. Compared with the viruses containing NA gene, the PB2 and PB1 gene came from different clades. And this indicates that the virus was a novel reassortant genotype. The HA receptor binding preference was avian-like and the cleavage site sequence showed a low pathogenic feature. There was no drug resistance mutation of M2 protein. The mutations of Asn30Asp, and Thr215Ala of the M1 protein implied the potential of pathogenicity increase in mice.</p><p><b>CONCLUSION</b>The finding of novel genotype of H3N8 virus in drinking water in this duck farm near Poyang Lake highlighted the importance of strengthening the surveillance of avian influenza in this region, which could contribute to pinpointing the influenza ecological relations among avian, swine, and human.</p>

Expression and purification of avian influenza virus H5N1 NP protein, and screening interaction proteins of host cells in vitro / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To express and purify H5N1 influenza virus (A/Anhui/1/2005) NP in prokaryotic system and to explore the NP-interacting proteins of human bronchial epithelial cells BEAS-2B in vitro.</p><p><b>METHODS</b>The full length H5N1 NP gene fragment was amplified by PCR, inserted into prokaryotic expression vector (pET30a) to generate NP expression plasmid pET30a-NP. After transforming pET30a-NP into E. coli (BL21), the expression of soluble NP protein was induced by IPTG. The expressed NP protein was purified by two steps with metal chelation chromatography and ion exchange chromatography. Then the total proteins of BEAS-2B cells was extracted for screening the components which have protein-protein interaction with purified NP by pull-down and LC-MS/MS methods.</p><p><b>RESULTS</b>The expression of H5N1 NP protein could be induced by IPTG in bacterial system using expression plasmid pET30a-NP. The soluble NP was purified. Twenty proteins were found by pull-down and LC-MS/MS, the further experiments may be needed to prove protein-protein interaction between them.</p><p><b>CONCLUSION</b>The soluble H5N1 NP fusion protein with high purity was obtained and twenty proteins were found which could interact with it by pull-down and LC-MS/MS.</p>

The study of multiple RT-PCR-based reverse dot blot hybridization technique for detecting influenza viruses / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To establish a multiplex RT-PCR-based reverse dot blot hybridization technique to detect influenza viruses.</p><p><b>METHODS</b>Obtain the HA nucleotide sequences of seasonal influenza H1N1, seasonal influenza H3N2, influenza H1N1 and human avian influenza H5N1 from GenBank. Design primers in conservative district and probes t in high variable region respectively, after analyzing the HA nucleotide sequences of influenza virus through the Vector NTI 9.0. Establish and optimize multiple RT-PCR system by comparing amplification efficiency and specificity at different primer concentrations. Establish the reverse dot hybridization system after optimizing the concentration of probes. To compare the sensitivity and specificity of this technique and the general RT-PCR Method through extracting the viral RNA of the mentioned influenza virus which are to be the reference substance.</p><p><b>RESULTS</b>Successfully establish a multiplex RT-PCR-based reverse dot blot hybridization technique for detecting influenza viruses. This technique is 100-1000 times more sensitive than gel electrophoresis method, and it has a good specificity.</p><p><b>CONCLUSION</b>Successfully established multiplex RT-PCR-based reverse dot blot hybridization technique for detecting influenza viruses.</p>

Characterization of pseudotyped viruses coated with hemagglutinin of H5N1 avian influenza / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To construct pseudovirus bearing H5N1 HA based on a lentivirus vector system. Then we study the biological feature of the pseudovirus. With the newly established viral particles, we performed the serological tests.</p><p><b>METHODS</b>H5N1 avian influenza virus that isolated from human case was cloned to construct pLP-HA, then pLP-HA co-transfected with lentivirus vector plasmids pLP1, pLP2 and pEmGFP into 293T cells. The supernatant was harvested 48h post-transfection. Concentrated by super centrifuge, the pseudotyped viruses were analyzed by infection test, HA test and micro-neutralization test. At the same time, optimized HA gene and a Vietnam H5N1 HA gene were used to construct pseudotyped virus for comparison.</p><p><b>RESULTS</b>Pseudotyped virus particles can be observed with electronic microscope. Western-blot revealed that HA glycoprotein can be expressed in the virions. Our neutralization assay by using the pseudoviruses was comparable with the conventional microneutralization assay with wild-type viruses. A high degree of correlation was detected.</p><p><b>CONCLUSION</b>Pseudotyped Viruses coated with HA of H5N1 High Pathogenic Avian Influenza were successfully constructed; it can be used to for the microneutralization assay. The HA gene from different sources affect the efficiency of the packaging of the pseudovirus. But the optimized HA gene can not obviously improve packaging efficiency of the pseudovirus.</p>

Study on immunogenicity of a recombinant adenovirus vaccine containing neuraminidase gene of H5N1 influenza virus (A/Anhui/1/2005) in mice / 中华实验和临床病毒学杂志

<p><b>OBJECTIVE</b>To investigate immunity of a recombinant adenovirus vaccine (rAdV) containing codon-modified neuraminidase (Mod. NA) gene of H5N1 influenza virus in BALB/c mice and to screen for appropriate dose.</p><p><b>METHODS</b>BALB/c mice were immunized with the rAdV-Mod.NA vaccine intramuscularly twice (double injection at 0 and 4th week) in three groups, low dosage (10(5) TCID50 per dose), medium dosage (10(7) TCID50 per dose) and high dosage (10(9) TCID50 per dose). The effect of humoral and cell-mediated immunity were analysed at 5th week.</p><p><b>RESULTS</b>(1) The rAdV-Mod.NA vaccine could elicit both humoral and cell-mediated robust NA specific immunity in mice by neuraminidase inhibitor assay and IFN-gamma ELISpot assay; (2) 10(7) TCID50 per dose was the appropriate dose; (3) Peptide NA(109-124): CRTFFLTQGALLNDKH and peptide NA(182-199): AVAVLKYNGIITDTIKSW were the dominant epitopes for neuraminidase-immunized BALB/c mice, which was screened out from the whole length of neuraminidase of an H5N1 virus, A/Anhui/1l/2005.</p><p><b>CONCLUSION</b>The recombinant adenovirus NA could induce specific humoral and cellular immune responses in BALB/c after immunization, which suggest rAdV-Mod.NA vaccine was a potential vaccine candidate against H5N1 influenza and worthy of further investigation.</p>