This paper reviews recent findings of the synovial membrane, in particular the morphology, function and development of synovial lining cells, in the temporomandibular joint (TMJ). Electron microscopic studies have confirmed the synovial membrane in TMJ consists of macrophage-like type A cells and fibroblast-like type B cells identical to those in other systematic joints. The macrophage-like type A cells react with anti-macrophage and macrophage-derived substances including the major histocompatibility class II molecule, and show a drastic increase in their number in the inflamed synovial membrane. In addition, they have the ability to produce substances involved in the progression of TMJ inflammation such as nitric oxide and inducible nitric oxide synthase. Observation of osteopetrotic mice revealed that macrophage-like type A cells in TMJ are derived from monocyte lineage. Immunocytochemistry for 25kDa heat shock protein was able to depict the entire shape of fibro-blast-like type B cells including their unique processes. The expression of an estrogen receptor α-immunoreaction in the fibroblast-like type B cells may explain the etiology of temporomandibular disorders at a higher frequency in females than in males, suggesting that TMJ is a target tissue for estrogen. Furthermore, fibroblast-like type B cells are equipped with a basement membrane to serve as an adhesion molecule for the fibroblast-like type B cells to keep their epithelial arrangement. A clear understanding of the morphology of the intact synovial membrane will serve to clarify the etiology and development of temporomandibular disorders.

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Aquaporin-5 (AQP5), an isoform of membrane water channel aquaporins, is expressed in the salivary and lacrimal glands. We surveyed the expression and immunohistochemical localization of AQP5 in the rat digestive system. RT-PCR analysis revealed that AQP5 is expressed in the submandibular gland, tongue, gastric corpus, pyloric region, duodenum, and liver. Immunofluorescence microscopy using AQP5-specific antibodies showed that AQP5 protein is present in the minor salivary glands of the tongue, the pyloric glands, and duodenal glands. To distinguish apical and basolateral domains of the plasma membrane of epithelial cells, double-immunofluorescence staining for AQP5 and tight junction protein occludin was performed. In the minor salivary gland, AQP5 was present in both the serous and mixed secretory end portions. AQP5 was found in the apical membrane of the secretory cells including intercellular secretory canaliculi demarcated with occludin. At higher magnifications, omega-shaped indentations of AQP5 labeling were seen along the apical membrane, suggesting a dynamic process for the apical membrane in exocytosis. Only weak labeling for AQP5 was detected in the basolateral domain. In the stomach, AQP5 was detected in the apical membrane of the pyloric gland secretory cells. In the duodenum, AQP5 was restricted to duodenal glands, where it was localized to the apical membrane. AQP5 was not detected in the intestinal glands or cells in the villi. These observations show that AQP5 is localized mainly in the apical membrane, including intercellular secretory canaliculi of secretory cells in the minor salivary glands, pyloric glands, and duodenal glands. AQP5 appears to play an important role in water transfer in these glands.

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The liver, which produces a large volume of lymph, has a lymphatic system which can be classified into three categories: portal, sublobular, and superficial lymphatic vessels. As little is known about the origin and pathways of sublobular lymph, this study demonstrates pathways of interstitial fluid flowing into sublobular lymphatic vessels. Livers from cats whose thoracic ducts were either ligated or non-ligated were examined by light-, transmission electron- and scanning electron-microscopy (SEM). Complete ligation of the thoracic duct caused significant dilation of the hepatic sinusoids, the space of Disse, and channels passing through the limiting plate. Sublobular interstitial space and sublobular lymphatic vessels were also expanded. The channels between hepatocytes forming the limiting plate contained collagen fibers, and connected the space of Disse with a sublobular interstitial space. The alkali-water maceration/SEM confirmed that collagen fibers traversing the layer of the limiting plate independently of blood vessels connected collagen fibers in the space of Disse with those in the sublobular space. Complete ligation of the thoracic duct also showed an accumulation of mast cells and plasma cells in the sublobular interstitial space. Our data suggest that fluid in the space of Disse flows along collagen fibers in channels traversing the limiting plate as well as those along the sinusoids and central veins that drain into sublobular veins, and enters the sublobular interstitial space to finally lead into sublobular lymphatic vessels. Our study has also shown that hepatic lymphostasis causes the accumulation of mast cells and plasma cells in the sublobular interstitial space, which may be involved in lymphangiogenesis and fibrogenesis.

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The present study investigated the development of plasma cells in the early rat yolk sac (days 10-16 of gestation) by light microscopy, transmission electron microscopy, immunoelectron microscopy, and indirect immunofluoresce techniques. Cells delineating the morphology of plasma cells in the yolk sac were observed as early as 12 days of embryonic life. As for positive immune staining for the intra-cyto-plasmic immunoglobulin (Ig) production (IgA, IgM and IgG), the intensity of the immune staining was very weak on days 10 and 11 of gestation, while it turned very dense on day 12 of gestation. At 14 days of gestation, the number of positive cells was markedly reduced. Immunoelectron microscopy visualized products of the immune reaction in cisterns of the rough endoplasmic reticulum. Conventional electron microscopic examination of 12, 13, and 16-day yolk sacs confirmed the development and differentiation of plasma cells with their well-known ultrastructural features, making this the first study to demonstrate these in the early rat yolk sac. The development of plasma cells in the early yolk sac implies the ability of the yolk sac to effect a humoral immune response at this stage of fetal life. The probable role of plasma cells in the yolk sac is also discussed.

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Gastrin-releasing peptide (GRP) has been proposed as a novel regulatory peptide in the reproductive tract. We previously demonstrated that GRP immunoreactivities are found predominantly in the uterine gland epithelial cells of nonpregnant and pregnant cows. The present study focused on the distribution of GRP immunoreactivity and the expression of GRP mRNA in the bovine endometrium during the estrous cycle. Tissues were collected from 21 uterine horns and bodies during the estrous cycle. RT-PCR showed the expected GRP mRNA fragments (284 bp) in the tissues from all stages of the cycle. In situ hybridization results ascertained the expression of the GRP mRNA in the uterine gland epithelial cells and superficial epithelial cells of the endometrium. Positive staining of GRP immunoreactivity in the uterine gland epithelial cells was detected in both the uterine horn and body from all stages of the cycle. In metestrus and diestrus stages, GRP was also detected in the superficial epithelial cells of horn, but not in the body. The degrees of GRP mRNA expression and intensities of GRP immunoreactivity in the endometrium increased from proestrus to diestrus stages. These findings suggest that GRP may be important both in the endometrial remodeling during the estrous cycle and in the implantation and development of blastocysts.

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To determine the progenitor nature of centroacinar cells (CACs), we attempted to compare the expression pattern of endocrine cell markers and PDX-1 (pancreatic duodenal homeobox gene 1) in CACs of both the quiescent and the regenerating rat pancreas. In the normal pancreas, most CACs were relatively small cells with sparse cytoplasm and oval or elongated nuclei. In addition, we noticed a distinct population of a small number of large cells with round nuclei in the centroacinar region. By immunohistochemistry, 0.21% and 0.3% of CACs in normal rat pancreas were respectively found positive for glucagon and insulin, being large CACs and designated as GL-CAC and IL-CAC. They also exhibited the mRNA of each hormone by in situ hybridization (ISH). The ISH signal for glucagon but not insulin was also detected in a subset of small CACs (designated GS-CAC). The expression of PDX-1 was also observed in subsets of small and large CACs (PS-CAC and PL-CAC, respectively). After a 90% pancreatectomy, the relative frequency for GS-CACs, but not those for other CACs, was significantly reduced in two days after surgery. On day 7 after surgery, the number of GS-CACs recovered to preoperative levels, whereas GL-CACs, IL-CACs, PS-CAC, and PL-CAC gradually increased to about double in number. From these results, a portion of CACs was suggested to differentiated into endocrine cells. A possible cell lineage is discussed for endocrine neogenesis during pancreatic regeneration.

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Previous mapping studies examinig the distribution and pattern of staining for connexin-43 expression (the major ventricular gap junction protein) across the ventricular wall have yielded variable findings. The aim of this study was to determine if variations in the distribution of connexin-43 were due to histochemical detection problems, i.e. cross-linking of antigenic sites as a consequence of aldehyde fixation and/or due to low levels of protein expression within the epicardial or endocardial regions of the heart. Immunoperoxidase staining of connexin-43 using the ABC method was carried out in crosssections of rabbit hearts at the level of the papillary muscle. The following treatments were examined: the antibody (Ab) only, Ab with 1/2 Tyramide Signal Amplification (TSA) or full TSA; antibody with microwave antigen retrieval (AR); Ab + 1/2 TSA + AR and finally Ab + TSA + AR. Under light microscopy and using computerized image analysis the percentages of ventricular cross-sectional transmural staining for the different treatment groups were calculated: Ab amounted to only 55%; Ab + 1/2 TSA 63%; Ab + TSA 78%; Ab + AR 72%; Ab + AR + 1/2 TSA 72% and Ab + AR + TSA 88%. The percentages of transumural connexin-43 staining in both TSA + Ab and Ab + TSA + AR groups when compared to Ab only were significantly greater p <0.01. The antigenic cross-linking due to aldehyde fixation and low levels expression of connexin-43 are contributing factors that influence the immunohistochemical detection of connexin-43 in the mammalian heart. Methodological enhancement for the detection of connexin-43 in this study was derived primarily from amplification of low background levels of connexin-43 being expressed using the TSA protocol. This is supported by the significant differences encountered when TSA was utilized in the protocol and compared with antibody treatment only.

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A 35 kD protein was isolated and purified from conditioned media of Bcl-2 cDNA-transfected PC12 cells and its cDNA cloned. A database analysis showed that the 35 kD protein is a rat homologue of the human FLRG protein. The biochemical as well as morphological properties of the rat FLRG protein in PC12 cells were examined and its distribution in rat tissues determined. The levels of FLRG mRNA expressed were low during the fetal period, compared with those of follistatin mRNA. The distribution of FLRG and follistatin mRNAs differed from each other after birth; the expression levels of FLRG mRNA were abundant in the adrenal gland and testis, whereas those of follistatin mRNA and activin A were markedly high in the ovary. The presence of FLRG mRNA and/or protein was confirmed in spermatocytes at various differentiating stages and in endocrine cells of both the adrenal cortex and medulla. When overexpressed in PC12 cells, the FLRG protein was found to be stored in secretory granules of the cells and largely secreted by a regulated pathway, while activin A enhanced the constitutive secretion of the FLRG protein from wild-type PC12 cells, indicating that the FLRG protein possesses dual properties in secretory pathways. The different distribution between FLRG and follistatin mRNA suggests that, like follistatin in the ovary, the FLRG protein may be involved in the maintenance of spermatogenesis in the testis and the growth and function of adrenal tissue cells, probably by regulating the functions of its binding partners such as the TGF-β superfamily members.

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