Naonori Sugai

Human follicular dendritic cells in vitro and follicular dendritic-cell-like cells.

Abstract.Human follicular dendritic cell (FDC)-like cells (FLC) have been utilized for the in vitro analysis of germinal center reactions. However, there is no consensus whether FLC represent FDC in vitro. The purpose of the present study has therefore been to determine distinguishing features of FDC and FLC in vitro. The expression of CD40, CD54, CD49d, cytokine (γ-IFN and IL-4)-dependent MHC-class II, and CD106 was observed to be specific for the determination of FDC in long-term culture. The cytokine-dependent emperipolesis of germinal center B cells was establised as another discriminating property for FDC in vitro. In 2 out of 72 long-term cultures of FDC, we encountered dividing cells among the non-dividing population of FDC. The dividing cells expressed accessory molecules similar to those of FDC but showed emperipolesis only for the initial few days of their growth. FDC did not enhance the CD40-dependent proliferation of germinal center B cells; in contrast, FLC augumented it. Both types of cells produced a significant amount of cytokine-dependent IL-6. Further studies are needed to determine whether FLC originate from FDC in vitro.

Follicular dendritic cells in vitro are not susceptible to infection by HIV-1.

Objectives To investigate whether follicular dendritic cells (FDC) are target cells for HIV-1 infection. Design Based on the principle that if FDC are true target cells, HIV-1 particles will bind to the surface of FDC and then invade their cytoplasm and nuclei. Methods Freshly isolated tonsilar FDC were exposed to two strains (HE and JR-FL) of HIV-1 and cultured. They were then examined for HIV-1 replication, using p24 antigen-capture enzyme-linked immunosorbent assay, immunolabelling, and polymerase chain reaction (PCR) amplification. We used FDC clusters as the FDC source, since the culture system for FDC clusters has various advantages over other methods for the successful long-term culture of FDC. Results After 2 h incubation, particles of both HIV-1 strains bound to the surfaces of FDC, as well as to CD4+ T cells, although FDC do not have CD4 receptors. The FDC gradually released the particles into the culture supernatant. More HIV-1 particles were bound to fresh FDC than to dedifferentiated FDC or to control fibroblasts. However, HIV-1 particles bound to the FDC did not seem to enter the cells. We found no evidence of HIV-1 proviral DNA synthesis in FDC. Conclusions Our results suggest that FDC are not readily infected with HIV-1 in situ, although we found that FDC in vitro were not infected by HIV-1.

Localization of Carbonic Anhydrase in Guinea Pig Bowman's Glands

We examined the histochemical localization of carbonic anhydrase (CA) in Bowmans glands by light and electron microscopy. Neither CAI nor CAII was detected immunohistochemically in the duct cells. However, by enzyme histochemistry the duct cells revealed electron-dense precipitates demonstrative of CA in the microvilli and intercellular digitations. The reaction product was also noted in small vesicles in the cytoplasm of duct cells. In cells of the acini, the well-developed short microvilli, basolateral cell membrane, and mitochondria along the basolateral membrane showed strong deposits indicating CA activity. Dense reaction product of CA was also detected in a small core within the electron-lucent granules of the secretory cells, although CAI and CAII were not detected by immunostaining in the secretory granules. Although the functional significance of CA in Bowmans glands is obscure, the enzyme may play a role in regulation of pH and ion balance in the mucous layer covering the olfactory epithelium. The presence of CA activity in the ducts suggests that these structures are not simple tubes serving as a conduit for secretory substances but participate in modifying the luminal content by secreting CA.

Histochemical localization of carbonic anhydrase in the trachea of the guinea pig

Tissue specimens from guinea pigs were examined using an enzyme-histochemical reaction to explore the presence of carbonic anhydrase (CA) activity in the trachea. CA activity was detected in a group of morphologically distinct epithelial cells, in goblet cells, and in glands of the tracheal mucosa. The epithelial cells showing CA activity were distributed singly and sparsely throughout the entire trachea. These cells showed a wide morphological variability and were clearly different from those forming the pseudostratified ciliated epithelium. Their number was higher in sections closer to the tracheal bifurcation than in those near the larynx. Although the nature of these cells is unknown, based on their morphological and histochemical characteristics and their distribution, they may represent a specialized chemoreceptor. To our knowledge, this is the first report of CA localized in tracheal epithelial cells.

Innervation of supporting cells in the guinea pig cochlea detected in bloc-surface preparations

We immunohistochemically examined the distribution of nerve fibers among supporting cells of the cochlea by using the bloc-surface preparation. The existence of these nerve fibers was not very clear in the standard avidin–biotin complex (ABC) method. However, the standard ABC method complemented with silver intensification procedure provided very fine details of the nerve fibers. The nerves started to appear at low density about 55% of the distance from the apex, and their density gradually increased toward the upper turn. In each portion, the nerve fibers increased in thickness and length as well as the number of synapses made with the nuclei. Moreover, the distribution of these nerves in the fetal cochlea was similar to that in the adult. However, the functional significance and importance of these nerves remains to be determined. Our study also indicates that the silver intensification procedure combined with the standard ABC method is useful for the detailed observation of stereoscopic innervation in thick tissue preparations like such as the cochlea.

Adhesion and costimulatory molecules on human FDC in vitro.

In the previous study1, we reported a unique accessory function of follicular dendritic cel1s( FDC ), emperipolesis of germinal center lymphoid cells, which was maintained in FDC after long-term culture. The first intercellular phenomenon for emperipolesis is considered the adhesion between lymphoid cells and FDC. This suggests that some specific adhesion molecules are expressed on the surface of FDC in vivo and in vitro2. In this study, we attempted to demonstrate the adhesion and costimulatory factors in FDC after long-term culture using FACS analysis to evaluating their properties of emperipolesis.