富含CpG的脱氧核苷酸片段对破骨细胞形成的调节作用

目的 :研究富含CpG的脱氧核苷酸片段 (ODN)调节破骨细胞 (OC)的形成 .方法 :分离小白鼠骨髓单核细胞 ,单纯骨髓单核细胞或骨髓单核细胞分别与 30 μg/L CSF ;30 μg/L CSF ,2 0 μg/LRANKL ;30 μg/LM CSF以及不同浓度ODN共同培养 1～ 4d .结果 :ODN可以诱导骨髓单核细胞分化为TRAP阳性的破骨细胞 ,这些细胞表达降钙素受体 .Northernblot显示 :随着ODN浓度由 5nmol/L增加至 1 0 0nmol/L ,细胞TNF αmRNA表达量也随之增加 ;骨髓单核细胞与 30 μg/LM CSF ,2 0nmol/LODN 1 82 6共同培养 1 ,2 ,3和 4d ,同样显示随着时间延长 ,细胞TNF α表达也逐渐增加 .结论 :ODN可以刺激破骨细胞的转化 ,为临床骨疾病的治疗开辟了一个新途径 .

鸟类习鸣脑空间的神经发生

鸟类的前脑内与发声、学习有关的神经核团在体内激素水平的影响下,成年脑可有新神经元产生,并替换原有的细胞,参与神经回路。这种神经发生在调节发声运动和听觉反馈间的整合过程,加强学习与记忆神经网络,促进脑的自我修复等方面具有重要意义。

Modulation of Lianbizi Injection (Andrographolide ) on Some Immune Functions

Objective: To study the effects of andrographolide on immune functions and the immune mechanism in clinical therapy.Methods: The amounts of IFN-α,IFN-γ, TNF-α, IL-8 in the peripheral blood mononuclear cells (PBMC) culture supernatants dealt with by different concentrations of LianBiZhi (LBZ) injection, the effective component of which is andrographolide, were detected by biological activity test or ELISA in vitro. The effects of LBZ injection on macrophage phagocytotic function and natural killer cells cytotoxicity were examined by means of macrophage to phagocytize cock erythrocyte and measurement of lactate dehydrogenase (LDH) activity released from the damaged cells, respectively.Results: The LBZ injection could not only enhance the phagocytosis activity of peritoneal macrophage from guinea pig to phagocytosis cock erythrocytes, but also augment the cytotoxicity mediated by natural killer cells from PBMCs.Conclusion: Andrographolide is an immunostimulant agent which can modulate both antigen specific and nonspecific immune function by means of its natural killer cells, macrophage and cytokines.

Cytological Mechanism of Cytoplasmic Inheritance in Pinus tabulaeformis: Ⅰ. Ultrastructural Aspects and Nucleoids of the Male Gametes

The cytological mechanism of plastid and mitochondrion inheritance in Pinus is an interesting research topic with only a limited number of published articles. The results indicate that the sperms of Pinus tabulaeformis Carr. contain abundant plastids, mitochondria and organelle DNA. These data provide reliable cytological evidence of paternal plastid and mitochondrion inheritance in Pinus . The results are in line with the confirmed general rule of paternal plastid inheritance in Pinaceae. But whether mitochondria in sperm cells can be transmitted into the embryos is an issue needs further developmental studies. Another important finding is that contrary to earlier classification of the male gamete of Pinaceae into the male nuclei type, the results reveal that male gametes in Pinus tabulaeformis are actually cells. However, the sperm cells are only surrounded by plasma membranes without cell walls. The larger leading sperm cell in a pollen tube section is long in shape, with a large amount of cytoplasm; while the second sperm cell is smaller, round in shape and contains less cytoplasm. Whether this feature of the male gamete type could be considered as a representative characteristic of the family is discussed and further conclusions await more experimental evidences from studies on plants from different species.

Effect of HSV-2 Infected Monocytes on the Production of TNF-a and IL-6

Objective: In order to detect the role of monocytes inHSV-2 infection, we studied the effect of herpes sim-plex Virus-2 infection on the production of tumor ne-crosis factor (TNF-σ), interleukin-6 (IL-6) secretedby monocytes. Methods: Monocytes were infected by HSV-2 (333Strain). Culture supernatants were collected at 1, 3,5, 7 days post-infection. The levels of TNF-α, IL-6were measured by enzyme-linked immunosorbent as-say (ELISA). Results: The levels of TNF-α secretion by mono-cytes significantly decreased on first day post-infection. The levels of IL-6 significantly decreasedon first and third days post-infection, and then gradu-ally increased to the control on seventh day post-infection. Conclusions: TNF-α and IL-6 production by mono-cytes was inhibited during HSV-2 infection. The pro-duction of cytokines may play an important role inherpes simplex viurs-2 pathogenicity and immunity.

Arsenic trioxide inhibites transgenic tumor necrosis factor-α promoter activity in vascular smooth muscle cells and THP-1 monocytes in vitro

Aim This study was to evaluate the effect of arsenic trioxide （As2O3） on the transgenic TNF-α promoter activity in cultured vascular smooth muscle cells （VSMCs） and THP-1 monocytes. Methods Human TNF-α promoter was constructed by reporter gene system and was transiently transfected into VSMCs and THP-1 in vitro. The promoter activity was tested by luciferase activity with or without LPS and Ang Ⅱ stimulation, before and after different dosage of As2O3 treatment. Results 1. TNF-α promoter effectively expressed in VSMCs and THP-1 compared with CMV promoter （58.3% and 80.9%, respectively）. Both LPS and Ang Ⅱ significantly up-regulated TNF-α promoter activity （P〈0.05）. 2. As2O3 significantly inhibited, both intact and LPS/Ang Ⅱ stimulated promoter activity, in a dose dependent manner （P〈0.05）, and in both cell type. Conclusion These results manifested that, the inhibition effect of As2O3 on the activity of human TNF-α promoter indicated its potential inhibition on pro-inflammatory cytokine genes expression at transcriptional level and its potential anti-inflammatory property in the cardiovascular system.

Neurogenesis by Activation of Inherent Neural Stem Cells in the Rat Hippocampus after Cerebral Infarction

Objective To investigate the changes of neural stem cells (NSCs) in the rat hippocampus after cerebral infarction (CI) and to evaluate the neurogenesis caused by the activation of NSCs. Methods CI models of rats were made and rats were assigned to 6 groups: sham-operated, 1 day, 3 days, 7 days, 14 days, and 28 days after CI. The dynamic expression of bromodeoxyuridine (BrdU), polysialylated neural cell adhesion molecule (PSA-NCAM), glial fibrillary acidic protein (GFAP), and neuronal nuclear antigen (NeuN) were determined by immunohistochemistry and immunofluorescence staining. BrdU was used to mark the proliferated NSCs. PSA-NCAM was used to mark the plasticity of activated NSCs. GFAP and NeuN were used to mark the differentiated NSCs. Results Compared with the controls, the number of BrdU+ cells in the hippocampus increased significantly at 1 day after CI (P<0.05), reached peak at 7 days after CI (P<0.05), decreased but still elevated compared with the controls at 14 days after CI (P<0.05), and nearly unchanged at 28 days after CI. The number of BrdU+/PSA-NCAM+ cells increased significantly at 7 days after CI (P<0.05), reached peak at 14 days after CI (P<0.05), and decreased but still elevated compared with the controls at 28 days after CI (P<0.05). The number of BrdU+/PSA-NCAM+ cells was equal to 60% of the number of BrdU+ cells in all the same period. The number of BrdU+/NeuN+ cells in the hippocampus increased significantly at 14 days after CI (P<0.05) and reached peak at 28 day after CI (P<0.05). The number of BrdU+/GFAP+cells in the hippocampus nearly unchanged after CI. Conclusion CI can stimulate the proliferation of inherent NSCs, and most proliferated NSCs may differentiate into neurons and represent neural plasticity.

Review:Actin-based dynamics during spermatogenesis and its significance

Actin can be found in all kinds of eukaryotic cells, maintaining their shapes and motilities, while its dynamics in sperm cells is understood less than their nonmuscle somatic cell counterparts. Spermatogenesis is a complicated process, resulting in the production of mature sperm from primordial germ cell. Significant structural and biochemical changes take place in the seminif-erous epithelium of the adult testis during spermatogenesis. It was proved that all mammalian sperm contain actin, and that F-actin may play an important role during spermatogenesis, especially in nuclear shaping. Recently a new model for sperm head elonga-tion based on the acrosome-acroplaxome-manchette complex has been proposed. In Drosophila, F-actin assembly is supposed to be very crucial during individualization. In this mini-review, we provide an overview of the structure, function, and regulation characteristics of actin cytoskeleton, and a summary of the current status of research of actin-based structure and movement is also provided, with emphasis on the role of actins in sperm head shaping during spermiogenesis and the cell junction dynamics in the testis. Research of the Sertoli ectoplasmic specialization is in the spotlight, which is a testis-specific actin-based junction very important for the movement of germ cells across the epithelium. Study of the molecular architecture and the regulating mechanism of the Sertoli ectoplasmic specialization has become an intriguing field. All this may lead to a new strategy for male infertility and, at the same time, a novel idea may result in devising much safer contraception with high efficiency. It is hoped that the advances listed in this review would give developmental and morphological researchers a favorable investigating outline and could help to enlarge the view of new strategies and models for actin dynamics during spermatogenesis.

Nucleocytoplasmic shuttling of Smad proteins

Nuclear accumulation of active Smad complexes is crucial for transduction of transforming growth factor β （TGF-β）- superfamily signals from transmembrane receptors into the nucleus. It is now clear that the nucleocytoplasmic distributions of Smads, in both the absence and the presence of a TGF-β-superfamily signal, are not static, but instead the Smads are continuously shuttling between the nucleus and the cytoplasm in both conditions. This article presents the evidence for continuous nucleocytoplasmic shuttling of Smads. It then reviews different mechanisms that have been proposed to mediate Smad nuclear import and export, and discusses how the Smad steady-state distributions in the absence and the presence of a TGF-β-superfamily signal are established. Finally, the biological relevance of continuous nucleocytoplasmic shuttling for signaling by TGF-β superfamily members is discussed.

Multiple biomarkers of colorectal tumor in a differential diagnosis model:A quantitative study

AIM:To evaluate the multiple biomarkers of colorectal tumor and their potential usage in early diagnosis of colorectal cancers. METHODS:Multiple biomarkers (DNA contents,AgNOR, PCNA,p53,c-erbB-2) in 10 normal colorectal mucosae,37 colorectal adenomas and 55 colorectal cancers were analyzed quantitatively in the computed processing imaging system. Discrimination patterns were employed to evaluate the significance of single and multiple indices in diagnosis of colorectal cancers. RESULTS:The mean values of the analyzed parameters increased in order of the normal mucosa,adenoma and adenocarcinoma,and this tendency reflected the progression of colorectal malignancy.The parameters including DNA index,positive rates,densities of AgNOR,c-erbB-2,and p53, shape and density of nucleus were relatively valuable for diagnoses.Then a diagnostic discrimination model was established.The samples were confirmed with the model, the sensitivity rates in cancer group and adenoma group were 96.36% and 89.19%,respectively.The value of proliferating cell nuclear antigen (PCNA) in early diagnosis of colorectal cancers was uncertain. CONCLUSION:The quantitative evaluation of some parameters for colorectal tumor can provide reproducible data for differential diagnosis.The established diagnostic discrimination model may be of clinicopathological value, and can make the early diagnosis of colorectal cancer possible.

Ellagic acid induces apoptosis through inhibition of nuclear factor κB in pancreatic cancer cells

AIM： To determine the effect of ellagic acid on apoptosis and proliferation in pancreatic cancer cells and to determine the mechanism of the pro-survival effects of ellagic acid.METHODS： The effect of ellagic acid on apoptosis was assessed by measuring Phosphatidylserine externalization, caspase activity, mitochondrial membrane potential and DNA fragmentation; and proliferation by measuring DNA thymidine incorporation. Mitochondrial membrane potential was measured in permeabilized cells, and in isolated mitochondria. Nuclear factor κB （NF-κB） activity was measured by electromobility shift assay （EMSA）. RESULTS： We show that ellagic acid, a polyphenolic compound in fruits and berries, at concentrations 10 to 50 mmol/L stimulates apoptosis in human pancreatic adenocarcinoma cells. Further, ellagic acid decreases proliferation by up to 20-fold at 50 mmol/L. Ellagic acid stimulates the mitochondrial pathway of apoptosis associated with mitochondrial depolarization, cytochrome C release, and the downstream caspase activation. Ellagic acid does not directly affect mitochondria. Ellagic acid dose-dependently decreased NF-κB binding activity. Furthermore, inhibition of NF-κB activity using IkB wild type plasmid prevented the effect of ellagic acid on apoptosis. CONCLUSION： Our data indicate that ellagic acid stimulates apoptosis through inhibition of the prosurvival transcription factor NF-κB.

Facilitating effects of berberine on rat pancreatic islets through modulating hepatic nuclear factor 4 alpha expression and glucokinase activity

AIM： To observe the effect of berberine on insulin secretion in rat pancreatic islets and to explore its possible molecular mechanism. METHODS： Pdmary rat islets were isolated from male Sprague-Dawley rats by collagenase digestion and treated with different concentrations （1, 3, 10 and 30 μmol/L） of berberine or 1 μmol/L Glibenclamide （GB） for 24 h. Glucose-stimulated insulin secretion （GSIS） assay was conducted and insulin was determined by radioimmunoassay. 3-（4,5-Dimethylthiazol-2-yl）- 2,5-diphenyltetrazolium bromide （MTT） assay was performed to evaluate cytotoxicity. The mRNA level of hepatic nuclear factor 4 alpha （HAIF4α） was determined by reverse transcription polymerase chain reaction （RT-PCR）. Indirect immunofluorescence staining and Western blot analysis were employed to detect protein expression of HNF4α in the islets. Glucokinase （GK） activity was measured by spectrophotometric method. RESULTS： Berberine enhanced GSIS rather than basal insulin secretion dose-dependently in rat islets and showed no significant cytotoxicity on islet cells at the concentration of 10 μmol/L. Both mRNA and protein expressions of HNF4α were up-regulated by berberine in a dose-dependent manner, and GK activity was also increased accordingly. However, GB demonstrated no regulatory effects on HNF4α expression or GK activity. CONCLUSION： Berberine can enhance GSIS in rat islets, and probably exerts the insulinotropic effect via a pathway involving HNF4α and GK, which is distinct from sulphonylureas （SUs）.

The ultrastructure of the spermatozoon of Octopus ocellatus Gray, 1849 (Cephalopoda: Octopoda)

Morphology of the spermatozoon of Octopus ocellatus was studied by light, scanning electron, and transmission electron microscopes. Sperm are 600-700 um long, with a large number of granules in diameter about 130 um. Each spermatozoon is composed of a head, neck, and tail. The head is made up of an acrosomal complex anterior to the nucleus. The spiral acrosomal complex consists of an electron-lucent vesicle, lacuna, and an electron-dense acrosomal vesicle. Additionally, the spiral acrosomal vesicle has numerous equidistant striations, and is surrounded by many small granules （20 nm diameter）. A long straight nucleus, which is electron-densed, has a deep posterior concavity, the nuclear vacuole. At the terminal end of the nucleus is a sleeve-like structure with a concave posterior nuclear fossa （PNF）. The neck is short connecting the PNF. The basal body is located in the PNF and gives rise to the axoneme. This structure connects the head, neck, and tail. The tail is divided into a middle piece and a principal piece. The middle piece, having a 9＋9＋2 arrangement, is surrounded by a mitochondrial sheath and terminates by an electron-dense fibrous sheath. The principal piece is the longest part of the sperm with coarse fibers tapering posteriorly. The results of this study shall provide some useful information for artificial breeding of this species.

Expression of intestinal trefoil factor, proliferating cell nuclear antigen and histological changes in intestine of rats after intrauterine asphyxia

AIM:To study the expressions of intestinal trefoil factor (ITF) and proliferating cell nuclear antigen (PCNA) and histologic changes in intestine, to investigate the relationship between ITF and intestinal damage and repair after intrauterine hypoxia so as to understand the mechanism of intestinal injury and to find a new way to prevent and treat gastrointestinal diseases. METHODS: Wistar rats, pregnant for 21 d, were used to establish animal models of intrauterine asphyxia by clamping one side of vessels supplying blood to uterus for 20 min, another side was regarded as sham operation group. Intestinal tissues were taken away at 0, 24, 48 and 72 h after birth and stored in different styles. ITF mRNA was detected by RT-PCR. PCNA expression was measured by immunohistochemistry. Intestinal tissues were studied histologically by HE staining in order to observe the areas and degree of injury and to value the intestinal mucosa injury index (IMDI). RESULTS: ITF mRNA appeared in full-term rats and increased with age. After ischemia, ITF mRNA was decreased to the minimum (0.59?.032) 24 h after birth, then began to increase higher after 72 h than it was in the control group (P<0.01). PCNA positive staining located in goblet cell nuclei. The PCNA level had a remarkable decline (53.29±1.97) 48 h after ischemia. Structure changes were obvious in 48-h group, IMDI (3.40±0.16) was significantly increased. Correlation analyses showed that IMDI had a negative correlation with ITF mRNA and PCNA (r= -0.543, P<0.05; r= -0.794, P<0.01, respectively). CONCLUSION: Intrauterine ischemia can result in an early decrease of ITF mRNA expression. ITF and PCNA may play an important role in the damage and repair of intestinal mucosa.

Visualization of bHLH transcription factor interactions in living mammalian cell nuclei and developing chicken neural tube by FRET

Members of the basic helix-loop-helix （bHLH） gene family play important roles in vertebrate neurogenesis. In this study, confocal microscopy-based fluorescence resonance energy transfer （FRET） is used to monitor bHLH protein-protein interactions under various physiological conditions. Tissue-specific bHLH activators, NeuroD 1, Mash 1, Neurogenin 1 （Ngn 1）, Neurogenin2 （Ngn2）, and ubiquitous expressed E47 protein are tagged with enhanced yellow fluorescence protein （EYFP） and enhanced cyan fluorescence protein （ECFP）, respectively. The subcellular localization and mobility ofbHLH fusion proteins are examined in HEK293 cells. By transient transfection and in ovo electroporation, four pairs of tissue-specific bHLH activators and E47 protein are over-expressed in HEK293 cells and developing chick embryo neural tube. With the acceptor photobleaching method, FRET could be detected between these bHLH protein pairs in the nuclei of transfected cells and developing neural tubes. Mashl/E47 and Ngn2/E47 FRET pairs show higher FRET efficiencies in the medial and the lateral half of chick embryo neural tube, respectively. It suggests that these bHLH protein pairs formed functional DNA-protein complexes with regulatory elements of their downstream target genes in the specific regions. This work will help one understand the behaviours of bHLH factors in vivo.

NOD2 and ATG16L1 polymorphisms affect monocyte responses in Crohn's disease

AIM： TO assess whether polymorphisms in NOD2 and ATG16L1 affect cytokine responses and mycobacterium avium subspecies paratuberculosis （MAP） survival in monocytes from Crohn＇s disease （CD） patients METHODS： Monocytes were isolated from peripheral blood of CD patients of known genotype for common single nucleotide polymorphisms of NOD2 and ATG16L1, Monocytes were challenged with MAP and bacterial per- sistence assessed at subsequent time-points. Cytokine responses were assayed using a Milliplex multi-analyte profiling assay for 13 cytokines. RESULTS： Monooltes heterozygous for a NOD2 polymorphism （R702W, P268S, or 1007fs） were more permissive for growth of MAP （P = 0.045） than those without. There was no effect of NOD2 genotype on subsequent cytokine expression. The T300A polymorphism of ATG16L1 did not affect growth of MAP in our model （P = 0.175）, but did increase expression of cytokines interleukin （IL）-10 （P = 0.047） and IL-6 （P = 0.019）. CONCLUSION： CD-associated polymorphisms affected the eliminaUon of MAP from ex v/vo monooltes （NOD2）, or expression of certain oltokines （ATG16LI）, implying independent but contributory roles in the pathogenesis of CD.

Immunization of mice with concentrated liquor from male zooid of Antheraea pernyi

AIM： To study the effects of concentrated liquor from male zooid of Antheraea pernyion immunological mice.METHODS： For each experiment, 40 mice were randomly divided into normal saline group （control group） and three tested groups that were administered different dosages of concentrated liquor from male zooid of A. pernyiand food for 15 d. The typical FSR and HC50 value, monocybe-phagocytic exponent Kand emendated monocyte-phagocytic exponent were determined and calculated respectively.RESULTS： Alter 24 and 48 h, the FSR values of the three tested groups improved significantly in comparison to the conlyol group by variance analysis. The HCso values showed a significant difference between the high dosage group and the control group, as well as between the high dosage group and other two bested groups. The monocyte-phagocytic exponent Kand emendated exponent α showed rising tendencies, but no significant differences were found by variance analysis.CONCLUSION： The concentrated liquor from male zooid of A. pemyican significantly enhance cellular and humoral immune function in mice, but has no distinct influence on the monocyte-phagocytic system in mice.

Fractalkine and TGF-β1 levels reflect the severity of chronic pancreatitis in humans

MM： To clarify whether serum chemokine and cytokine levels can become useful biological and functional markers to assess the severity of chronic pancreatitis （CP）. This study aimed at clarifying whether serum chemokine and cytokine levels can become useful biological and functional markers to assess the severity of CR METHODS： Serum monocyte chemoattractant protein-1 （MCP-1）, transforming growth factor beta-1 （TGF-βI）, and soluble type fractalkine （s-fractalkine） concentrations were examined in patients with CP （n = 109） and healthy controls （n = 116）. Severity of disease was classified in patients with CP by a staging system. Relationships between stage-specific various clinical factors and serum MCP-1, TGF-β1, and s-fractalkine levels were investigated. Furthermore, 57 patients with non-alcoholic CP were similarly evaluated in order to exclude influence of alcohol intake. RESULTS： Patients with CP showed significant higher levels of serum TGF-β1 and s-fractalkine, but not MCP-1, compared to the controls. Serum TGF-β1 in the severe stage and s-fractalkine in the mild and thesevere stage of CP significantly increased compared to those of controls. However, it was observed that both TGF-β1 and s-fractalkine levels were affected by alcohol intake. In patients with non-alcoholic CP, serum TGF-β1 showed significant increase in the moderate stage of CP, and serum s-fractalkine revealed significant increase in the early stage of CP. CONCLUSION： It is suggested that the measurement of serum F-fractalkine is useful to diagnose early- stage CP. Moreover, the combined determination of both, s-fractalkine and TGF-β1, in human sera may be helpful in evaluating the severity status of CP.

Aberrant activation of nuclear factor of activated T cell 2 in lamina propria mononuclear cells in ulcerative colitis

AIM:To investigate the role of nuclear factor of activated T cell 2(NFAT2),the major NFAT protein in peripheral T cells,in sustained T cell activation and intractable inflammation in human ulcerative colitis(UC). METHODS:We used two-dimensional gel-electrophoresis, immunohistochemistry,double immunohistochemical staining,and confocal microscopy to inspect the expression of NFAT2 in 107,15,48 and 5 cases of UC, Crohn's disease(CD),non-specific colitis,and 5 healthy individuals,respectively. RESULTS:Up-regulation with profound nucleo- translocation/activation of NFAT2 of lamina propria mononuclear cells(LPMC)of colonic mucosa was found specifically in the affected colonic mucosa from patients with UC,as compared to CD or NC(P<0.001,Kruskal- Wallis test).Nucleo-translocation/activation of NFAT2 primarily occurred in CD8+T,but was less prominent in CD4+T cells or CD20+B cells.It was strongly associated with the disease activity,including endoscopic stage (τ=0.2145,P=0.0281)and histologic grade(τ=0.4167, P<0.001). CONCLUSION:We disclose for the first time the nucleo-translocation/activatin of NFAT2 in lamina propria mononuclear cells in ulcerative colitis.Activation of NFAT2 was specific for ulcerative colitis and highly associated with disease activity.Since activation of NFAT2is implicated in an auto-regulatory positive feedback loop of sustained T-cell activation and NFAT proteins play key roles in the calcium/calcineurin signaling pathways,our results not only provide new insights into the mechanism for sustained intractable inflammation,but also suggest the calcium-calcineurin/NFAT pathway as a new therapeutic target for ulcerative colitis.

CONSTRUCTING ADENO-ASSOCIATED VIRUS-TGFβ_3 AND COMPARING ITS BIOLOGICAL EFFECT ON PROTEOGLYCAN SYNTHESIS IN DEDIFFERENTIATED NUCLEUS PULPOUS CELLS WITH ADENOVIRUS-TGFβ_1

Objective To construct adeno-associated virus (AAV) expression system for transforming growth factor β3 (TGFβ3) and detect its biological effect on proteoglycan synthesis of the earlier and later dedifferentiated rabbit lumbar disc nucleus pulpous (NP) cells, which was compared with that of adenovirus (AV) expression system for TGFβ1. Methods TGFβ3 gene was obtained using PCR. Its upstream contained restriction enzyme site Kpn Ⅰ, and its downstream contained restriction enzyme site SalⅠ. Using the restriction enzyme sites of PCR product of TGFβ3 and the corresponding multiple cloning site (MCS) in plasmid AAV, TGFβ3 was subcloned into AAV. The recombinant plasmid AAV-TGFβ3 was transfected into H293 cells with LipofectamineTM 2000, and the expression of TGFβ3 gene was detected using immunofluorescent analysis. After AAV-TGFβ3 virus particle with infectious activity was packaged, TGFβ3 expression in NP cells was detected by immunoblotting, and its biological effect on proteoglycan synthesis was detected by antonopulos method and compared with that of AV-TGFβ1 in the earlier and later dedifferentiated NP cells. Results For the earlier dedifferentiated NP cells, AAV-TGFβ3 slowly and stably enhanced proteoglycan synthesis, but AV-TGFβ1 rapidly and transiently enhanced its synthesis. For the later dedifferentiated NP cells, AAV-TGFβ3 stably enhanced proteoglycan synthesis, but AV-TGFβ1 inhibited its synthesis. Conclusion AAV expression system can mediate TGFβ3 gene to be expressed stably, and AAV-TGFβ3 can enhance proteoglycan synthesis of the earlier and later dedifferentiated NP cells.