Expression Level And Localization Of CD80 And CEA In Fetal Thymus

BackgroundThe thymus is optimal microenvironment for T cell development. The T cell development and maturation process strictly positive and negative selections in the thymus. The negative selection not only requires the 1^st TCR signal on the thymocyte, but also the combined 2^nd costimulatory signal (MHC-self peptide), on both thymocyte and APC (antigen presending cell). The costimulatory molecule plays a critical role in the negative selection, and may promote or terminate the immune response, ie, may regulate the immune response. It has been showed that the B7-1(CD80) and CD28 are the primary costimulators, and the dendritic cell (DC), macrophage and some epithelioreticlar cells (ERC) can express B7-1 molecule.According to the location and function, the ERC in broad sense can be classified into 6 patterns. The ERC in broad sense, mainly including ERC and DC (interdigitating cells) in narrow sense, can express CD80. The DC is capable to present antigen potently, its antigen presenting capacity is 100-1000 folds stronger than that of the macrophage.Materials and Methods1. The 12 artificial abortion specimens of human fetal thymus from pregnant volunteers, including 8 cases aged 5-8 month (later aged group) and 4 cases aged 3-4 month (earlier aged group).2. Each specimen was divided two aliquots. One aliquot was prepared into frozen sections, stained with HE, ANAE histochemisyty, CD80 and CEA immunohistochemistry. The signals were observed under oil-emersion.3. The protein was extracted with Trizol reagent from another aliquot. The CD80 Western blotting and CEA ELISA were performed. The immunoblotting of CD80 andβ-actin was scanned with a Shimadu scanner (Japan).The ratio of CD80/β-actin was calculated and the OD value was examined with an enzyme labeling detector (Austria) in each group.4. The ANOV was detected with SPSS 10.0 for statistical analysis,andα=0.05 was considered as significant level.Results1 .HE staining The development of the earlier aged group was poor. The cortex was thinner and the reticular space in the medulla was smaller. The later aged group developed progressively with thicker cortex and larger reticular space. The classicalHassall's corpuscle could be found in the medulla.2.CD80 immunostaining CD80-IR (immunoreactivity, IR) appeared as fine brownishgranules, expressed in the ERC with 6 patterns. The CD80 expression was not distinct in the earlier aged group. The expression of CD80, especially in the medulla, was enhanced in the later aged group. Type I ERC in flatten shape located at the inner surface of trabecula or capsule. Type II ERC in small star shape distributed in the cortex. Type III and type IV ERC located at the division between the cortex and the medulla. Type V ERC in large satellite shape distributed in the medulla and the type VI ERC located at the Hassall's corpuscle.3.ANAE staining The T cell and its subpopulations can be shown with ANAE staining, such as the dot pattern with 1-3 distinct red-colored dots in the CD4⁺, the diffuse granular pattern with scattered red-colored granules in the CD8⁺. T subpopulation cells in respective cluster form were located in the medulla of later aged group. The Hassall's corpuscle and the macrophage appeared as strong ANAE staining with the flame pattern.4.CEA immunostaining In the earlier aged group the CEA-IR was located in themedullary ERC and the Hassall's corpuscle; in the later aged group the strongerCEA-IR was mostly concentrated in the Hassall's corpuscle.5.Western blotting The ratio of CD80/β-actin was 0.66±0.09 in the earlier aged group;while that was 0.89±0.11 in the later aged group,p<0.05.6.CEA-ELISA The OD value was 0.181±0.03 in the earlier aged group; while thatwas 0.302±0.04 in the later aged group,p< 0.05.Conclusions1 .The CD80 immunoblotting and the OD value of CEA ELISA in the later aged group were stronger than those in the earlier aged group, indicating that the expression level of both CD80 and CEA in the later aged group were higher than those in the earlier aged group.2.The development of the earlier aged group was poor. In the later aged group the ERC with c CD-80-IR ould be divided into 6 types. The type I ERC and type III /TV ERC possessed separation function as the barrier. The type II and type V ERC were DC in narrow sense. The type VI ERC was located at the Hassall's corpuscle. The results suggest that the thymus microenvironmrnt may be further divided into some sub-microenvironmrnts.3.In the earlier aged group the CEA-IR was located in the medullary ERC and the Hassall's corpuscle; in the later aged group the stronger CEA-IR was mostly concentrated in the Hassall's corpuscle. The corpuscle with CEA-IR combined with CD80-IR and strong ANAE staining suggest that the corpuscle may have biological significance at certain extent.4.In the medulla of later aged group the T subpopulation cells exhibited respective cluster forms, suggesting that each T subpopulation may be located in separate reticular space sub-microenvironmrnt.