我们在以往研究中，引人选择性增强体液免疫效应的新型分子佐剂C3d。成功构建了重组避孕疫苗hCGlβ-C3d3，通过免疫Th2型优势的BALB/c小鼠和，Thl型优势的C57BL/6小鼠，显示分子佐剂C3d在不同品系小鼠均使免疫效应从Thl型细胞免疫向Th2型体液免疫偏倚[1-3]。

目的：探讨环孢素A（cyclosporin A, CsA）作用于人滋养层细胞差异表达的功能基因。方法：应用抑制性消减杂交筛选110μmol·L-1 CsA作用于JAR细胞系前后差异表达的功能基因，经RT2PCR及蛋白质印迹进一步验证CsA作用前后原代分离的人早孕期滋养层细胞及JAR细胞株titin 表达的变化。结果：CsA作用于人JAR细胞系前后出现6个具有差异表达的基因，其中1 个为已知基因titin，2个为功能未知的mRNA，3个为位于16号染色体上的EST。RT2PCR显示，CsA作用于人滋养层细胞24h后出现titin mRNA的表达，72h达高峰，并呈现明显的剂量依赖性，110μmol·L-1 CsA作用最明显。蛋白质印迹显示，CsA可诱导titin的表达。结论：CsA可能通过诱导滋养层细胞titin的高表达，改变其生物学行为,从而有利于胚胎着床及早期发育。

目的探讨脱氢表雄酮（DHEA）对成骨细胞（oste0blasts，OB）及CD4+T细胞表达协同刺激分子的调控作用。方法颅骨酶解法培养鼠oB，体外模拟雌激素撤退；免疫磁珠细胞分选（rnag-c cell 9ort，MAC. S）法分离CD4+T细胞；将oB或CD4 T细胞分为对照组、E2及DHEA处理组，并以LPS刺激；以流式细胞术分析OB表面CDS0、CD86以及CD4+T细胞表面CD28、CⅡA4的表达。结果经E2及DHEA处理后，OB表达CDS0、CD86显著增加（P<0.05，P<0.01）；CsA可降低对照组及DHEA处理组OB CD80、CD86的表达（P<0.01）。除DHEA组CD28 T细胞百分比增加外（P<0.05），其余各组CD4 T细胞CD28和CIIA4的表达无显著改变（P>0.05）。结论DHEA可上调鼠OB协同刺激分子CDS0、O386表达，该作用可被CsA阻滞；DHEA还上调CD4 T细胞cD28的表达，提示可改善骨。免疫调节网络。

Objective: To express the hCGβ-C3d3 fusion protein in a CHO cell continual expression system to investigate further the adjuvant effects of C3d on contraceptive vaccination. Method: We constructed a plasmid pcDNA3-hCGβ-C3d3 which contains three copies of murine C3d cDNA and the hCGβ gene by cloning the chimerical hCGβ-C3d3 cDNA into the eukaryotic vector pcDNA3 downstream of the CMV promoter. The plasmid was transfected into a COS-7 cell transient expression system and a CHO cell continual expression system. RIA was used to detect hCGβ in the culture supernatant. Western blot and Raji cell immunohistochemical assays were performed to evaluate the expressed protein. Then, 6-/8-week-old female BALB/c mice were inoculated intramuscularly with pcDNA3-hCGβ and pcDNA3-hCGβ-C3d3, and ELISA was used to assess anti-hCGβ IgG antibody in serum. Results: In 72 h after COS-7 cells were transfected with the plasmid pcDNA3-hCGβ-C3d3, 1.0/105 cells could secrete 152 ng of the recombinant protein (calculated by hCGβ contained). The transfected CHO cells, which were then screened by G418, could continuously secrete the fusion protein at 660 ng/106 cells/48 h. The hCGβ-C3d3 protein was purified by anti-hCGβ immunoaffinity chromatography. Raji cell immunohistochemical assay demonstrated that both the hCGβ and C3d3 were successfully fused. After DNA immunization intramuscularly, the anti-hCGβ IgG antibody titer in the pcDNA3-hCGβ-C3d3 immunized group was 243-fold higher than that of the pcDNA3-hCGβ immunized group. Conclusion: We have expressed the hCGβ-C3d3 protein successfully, both in a transient expression system (COS-7 cells) and in a stable expression system (CHO cells). The C3d3 molecular adjuvant can enhance significantly the immunogenecity of hCGβ antigen in DNA immunization.

Chemokines and chemokine receptors have been implicated as pivotal players in many physiological and pathological situations, but little is known about the expression and function of chemokines and chemokine receptors at the materno-fetal interface. In this study, we first analyzed the transcription of 18 chemokine receptors in first-trimester human trophoblast cells. Among these receptors, CXCR4 was found highly transcribed. We demonstrated afterward that both CXCR4 and CXCL12 (stromal cell-derived factor-1; SDF-1) were expressed in trophoblast cells. Primary cultured trophoblast cells were also found secreting CXCL12 spontaneously. To identify the functional role of CXCR4/CXCL12 in these cells, we treated trophoblast cells with recombinant human (rh) SDF-1a and analyzed the cell viability and signaling pathway. The results showed that rhSDF-1a increased the viability of trophoblast cells and the activation of extracellular signal-regulated kinases signaling pathway in vitro. Our findings suggest that first-trimester trophoblast cells express functional CXCR4/CXCL12, which may play an important role in early pregnancy such as stimulating trophoblast cell proliferation or differentiation in an autocrine manner.