Structural and functional abnormalities in the spleen of an mFtz-F1 gene-disrupted mouse.

The spleen has two main functions. The first is to provide a proper microenvironment to lymphoid and myeloid cells, whereas the second involves clearance of abnormal erythrocytes. Ad4BP/SF-1, a product of the mammalian FTZ-F1 gene (mFTZ-F1), was originally identified as a steroidogenic, tissue-specific transcription factor. Immunohistochemical examination of the mammalian spleens confirmed the expression of Ad4BP/SF-1 in endothelial cells of the splenic venous sinuses and pulp vein. In mFtz-F1 gene-disrupted (KO) mice, several structural abnormalities were detected in the spleen, including underdevelopment and nonuniform distribution of erythrocytes. Examination of the spleen of KO fetuses showed failure of development of certain tubular structures during embryogenesis. These structures are normally assembled by Ad4BP/SF-1 immunoreactive cells, and most likely form the vascular system during later stages of development. Other structural abnormalities in the spleen of the KO mice included defects in the tissue distribution of type-IV collagen, laminin, c-kit, and vimentin. These morphologic defects in the vascular system were associated with a decrease in the proportion of hematopoietic cells, although differentiation of these cells was not affected significantly. A high number of abnormal red blood cells containing Howell-Jolly bodies were noted in the KO mice, indicating impaired clearance by the splenic vascular system. We also detected the presence of an mRNA-encoding cholesterol side-chain cleavage P450 in the spleen, resembling the findings in steroidogenic tissues such as the gonads and adrenal cortex. The mRNA transcript was not involved in splenic structural defects as it was detected in the spleens of both normal and KO mice, indicating that the regulatory mechanism of the P450 gene in the spleen is different from that in steroidogenic tissues. Our results indicate that a lack of the mFtz-F1 gene in mice is associated with structural and functional abnormalities of the splenic vascular system.

High density culture of immortalized liver endothelial cells in the radial-flow bioreactor in the development of an artificial liver.

Liver endothelial cells are important components of the tissue along the hepatic sinusoid. They are responsible for microcirculation in the liver and scavenger functions. It would therefore be important to include these cells in any hybrid type of artificial liver in addition to hepatocytes. However, it is difficult to culture these cells in vitro. The development of a liver endothelial cell line, which maintains the characteristics of the primary culture, would thus be of great benefit in the development of an artificial liver. In the present study we established immortalized liver endothelial cells from the liver of an H-2Kb-tsA58 transgenic mouse, which harbors the SV40 TAg gene. Hepatic sinusoidal cells isolated from H-2Kb-tsA58 mouse proliferated in the presence of gamma-interferon at 33 degrees C. Four clones were established, out of which clone M1 had the highest amounts of PGI2 production, as well as plasminogen activator activity and internalized acetylated low density lipoprotein. On culture dishes the M1 cells grew individually and spread. Sieve plates on the cell surface were not readily visible, but small pores were detected under electron microscopic observation. These results suggest that M1 clone cells originated from liver endothelial cells. Moreover it was possible to culture the immortalized liver endothelial cells in a radial-flow bioreactor for 5 days, with a maximum 6-keto prostaglandin F1alpha production of 25 microg per day. This suggests that immortalized liver endothelial cells and a radial-flow bioreactor can prove useful tools in the development an artificial liver.

Phenotypic characteristics and significance of reticular meshwork surrounding splenic white pulp of mice.

This report describes the phenotypic characteristics of the reticular meshwork (RM) localized between the marginal zone and white pulp of mice. Considered from an anatomical point of view, RM contributes to the migration of lymphocytes into splenic white pulp. We divided the RM into two parts: one surrounding the periarterial lymphoid sheaths (PALS) and the other surrounding the follicles. Throughout both locations of RM, alkaline phosphatase activity was noted on the plasma membrane of the reticular cells and tenascin was expressed on the collagen fibrils in the matrix of the reticular fibers. In contrast, the expression of MAdCAM-1, one of the homing receptors of lymphocytes, was solely confined to the plasma membranes of the fiber-forming reticular cells and processes of the perifollicular RM. In aly mice with defective perifollicular RM and loss of MAdCAM-1 expression, lymphocyte homing to the white pulp was remarkably inhibited, and the compartmentalization of T and B cells did not occur. This fact supports the notion that perifollicular RM plays an essential role in lymphocyte homing and compartmentalization.

The splenic marginal zone is absent in alymphoplastic aly mutant mice.

aly is a unique spontaneous autosomal recessive mutation in mice that causes a deficiency in the systemic lymph nodes (LN) and Peyer's patches (PP). aly also induces abnormal histological findings in the spleen and a deficiency in humoral and cell-mediated immune functions, although lymphocytes of the aly/aly mouse show virtually normal immune responses in vitro. We studied the structure of the spleen of aly mice in detail by immunohistochemistry and electron microscopy. We found that the spleen of the aly/aly mouse was deficient in the expression of specific antigens for marginal metallophils (MM), marginal zone macrophages (MZM) and fibroblastic reticular cells (FRC), which are components of the marginal zone (MZ) of the spleen. Morphological analysis indicated that the aly/aly spleen is deficient in the structure of MZ, which may, in part, account for the severe immunodeficiency in the aly/aly mouse. We then performed reciprocal bone marrow transplantation experiments (BMT) between normal and aly mice and found a clear correlation between the formation of LN and the development of the splenic MZ in these mice; i.e. successful development of LN was invariably associated with the appearance of the MZ structure, whereas the failure of LN development was always associated with the absence of the development of MZ. These BMT results suggest that a common factor may regulate the generation of both LN and MZ of the spleen.

A case of congenital onychoheterotopia on both fifth fingers.

The present report illustrates the extremely rare clinical features of congenital onychoheterotopia (ectopic nail). The amino-acid composition of congenital ectopic nail was estimated, and the surface of nail clippings was compared to the patient's normal nail using scanning electron microscopy.

Fine architecture of the splenic terminal vascular bed as revealed by arterial and venous pressure-loading perfusion fixation.

The three-dimensional fine architecture of the red pulp of human and animal spleens, which as fixed by a modified version of the arterial and venous pressure-loading perfusion fixation (AVPL perfusion fixation) method, is demonstrated by scanning and transmission electron microscopy. In the human spleen, changes in splenomegalias associated with hereditary spherocytosis and chronic portal hypertension are also introduced in addition to the normal architecture of the red pulp of spleens removed from patients with stomach cancer. The AVPL perfusion fixation of these spleens clearly visualized complicated three-dimensional fine architecture of the red pulp and provided much important information on in situ morphology and dynamic change of the terminal vascular bed, including venous pressure-dependent size change of the stomata and three-dimensional shapes of the capillary terminal, with positive proof of their opening into the cordal reticular tissue. In studies of the spleen associated with portal hypertension, the AVPL perfusion fixation is considered a necessary technique for analysis of the structural deviation closely relating to a very high venous pressure.

Scanning electron microscopic study of the red pulp of ferret spleen.

Scanning electron microscopy of the red pulp of ferret spleen showed areas of relatively wide splenic cord and rather poorly developed splenic sinuses of two different types. The inner aspect of most sinuses was covered with rod-shaped cells running in the longitudinal direction of the sinus. Stomata of these sinuses were recognized between the rod-like cells in a relatively regular lattice-like arrangement. The lining of other sinuses showed characteristics of the poorly differentiated sinuses, having a remarkable irregularity in form, size, and distribution of the lining cells and the stomata. The outer surface of the splenic sinuses was covered with attenuated reticular cell processes supporting the sinus. Sheathed capillaries which have a well-developed sheath structure were frequently found in the cordal area. Cordal capillaries were exclusively open into labyrinthine reticular tissue space of the cord with a funnel-shaped or a perforated saccular or ampullar terminal.

Human case of dracunculiasis in Japan.

Dracunculus medinensis was recovered from a 42-year-old man residing in Shizuoka, Japan. Part of an adult female worm was removed from an abscess in the subcutaneous tissue on his right gastrocnemial muscle. The worm's morphology and that of the embryos found in its uterus revealed that the parasite was Dracunculus medinensis. This is the first human case of dracunculiasis acquired in Japan.

Stereoscopic scanning electron microscopy of the red pulp of dog spleen with special reference to the terminal structure of the cordal capillaries.

In order to obtain direct morphological information about the three-dimensional fine structure of the splenic terminal vascular bed, especially the terminating mode of the cordal capillaries, stereoscopic scanning electron microscopy of perfusion-fixed and freeze-fractured red pulp of a normal dog spleen was undertaken. An improved method of perfusion-fixation was utilized in which the hydrostatic pressures of the splenic artery and vein were maintained at approximately the same levels as those in the living state. Stereoscopic observations of scanning electron micrographs clearly demonstrated the three-dimensional fine architecture of the splenic sinuses, the spongy cordal reticular tissue and the intracordal vasculature. The cordal capillaries terminate in the labyrinthine cordal space according to a certain mode in which the walls of the terminals are transformed into a meshwork structure continuous with the cordal reticular tissue owing to an increase in number and size of fenestrations. No evidence could be detected to prove or suggest any direct continuity of the capillary end with the splenic sinus. These results strongly support the concept of an "open circulation", at least in the red pulp of the dog spleen, with the possibility of a "functionally closed circulation" under some physiological conditions.