Yoshio Mabuchi

Folliculo‐stellate cells and intercellular communication within the rat anterior pituitary gland

Folliculo‐stellate (FS) cell are agranular and arranged around a follicle. They contain the S‐100 protein and β‐adrenergic receptors. It has been suggested that they can act as stem cells, since they show mitotic figures, and could transform into granular or chromophilic cells according to the concept of a “cell renewal system.” Cell‐to‐cell interactions among pituitary cells have been described, and recent progress with freeze‐fracture electron microscopy has provided novel observations of the cell surface and gap junctions within the rat or teleost fish pituitary gland, or in cultured rat pituitary cells. In adult rats, the anterior pituitary was composed of lobules incompletely separated by a basement membrane. Follicles consisted exclusively of FS cells. Gap junctions were observed only between adjacent FS cells, in rare cases on the tips of their cytoplasmic processes. Thus, the FS cells, connected by gap junctions, made up a dense cellular network throughout the pituitary. Gap and tight junctions were absent on granular cells. Elongated follicles with columnar FS cells were observed in 10‐day‐old rats and were separated into smaller units. The number of gap junctions rapidly increased with age until 40–45 days of age. Few S‐100 protein positive cells were observed on day 10, along the marginal cell layer and near the so‐called postero‐lateral wing. The frequency of positive cells increased with age and by day 40; numerous cells were observed throughout the anterior lobe.

Ultrastructural changes at the myotendinous junction induced by exercise

BackgroundAlthough exercise is believed to reduce the risk of rupture of the myotendinous junction, exercise-induced structural changes in this region have not been studied. We examined exercise-induced ultrastructural changes in the myotendinous junction of the lower legs in rats.MethodsTen adult male LETO rats were used. Five rats were randomly placed in the Exercise group; the remaining five were used as controls and placed in the non-Exercise group. Running exercise was performed every day for 4 weeks. The tibialis anterior and gastrocnemius muscles were then removed from both legs from each animal in the two groups. The specimens were subsequently examined by transmission electron microscopy (TEM). Numerous finger-like processes were observed at the myotendinous junction. The changes in frequency of branching of the finger-like process (the number of times one finger-like process branched) and the direction of the processes (the angle of the major axis of a finger-like process to the longitudinal direction of the muscle fiber) were studied. To evaluate the two indicators above, each 10 fingerlike process was randomly and separately selected from the tibialis anterior and gastrocnemius muscles of rats, providing 50 finger-like processes of both muscles for evaluation per group.ResultsIn terms of the frequency of branching of the fingerlike processes, the mean values obtained in the non-Exercise group were 0.04 and 0.18 times, respectively, in the tibialis anterior and gastrocnemius muscles and were 0.38 and 1.16 times, respectively, in these two muscles in the Exercise group. Regarding the direction of the finger-like processes, the values were 4.1° and 3.6°, respectively in the non-Exercise group and 10.4° and 14.5°, respectively in the Exercise group. The differences between the two animal groups were significant.ConclusionsMorphological changes in the myotendinous junction occurred as an adaptation to tension increased by exercise.

In vivo uptake of lecithin-coated polystyrene beads by rat hepatocytes and sinusoidal endothelial cells

While phagocytosis by Kupffer cells (stellate perisinusoidal macrophages) is well known and that by endothelial cells also is thought to occur under certain conditions, the uptake of large particles by hepatocytes has not been well studied. We reported previously the selective phagocytic uptake of material by hepatocytes using egg lecithin‐coated silicon particles. In the present work, we describe more precisely this process following the injection of lecithin‐coated polystyrene beads. Additionally, we consider the possible significance of the transcytotic action by endothelial cells.

Angiotensin II‐induced modulation of endothelium‐dependent relaxation in rabbit mesenteric resistance arteries

The role of local endogenous angiotensin II (Ang II) in endothelial function in resistance arteries was investigated using rabbit mesenteric resistance arteries. First, the presence of immunoreactive Ang II together with Ang II type‐1 receptor (AT1R) and angiotensin converting enzyme (ACE) was confirmed in these arteries. In endothelium‐intact strips, the AT1R‐blocker olmesartan (1 μm) and the ACE‐inhibitor temocaprilat (1 μm) each enhanced the ACh (0.03 μm)‐induced relaxation during the contraction induced by noradrenaline (NA, 10 μm). Similar effects were obtained using CV‐11974 (another AT1R blocker) and enalaprilat (another ACE inhibitor). The nitric‐oxide‐synthase inhibitor NG‐nitro‐l‐arginine (l‐NNA) abolished the above effect of olmesartan. In endothelium‐denuded strips, olmesartan enhanced the relaxation induced by the NO donor NOC‐7 (10 nm). Olmesartan had no effect on cGMP production (1) in endothelium‐intact strips (in the absence or presence of ACh) or (2) in endothelium‐denuded strips (in the absence or presence of NOC‐7). In β‐escin‐skinned strips, 8‐bromoguanosine 3′,5′ cyclic monophosphate (8‐Br‐cGMP, 0.01–1 μm) concentration dependently inhibited the contractions induced (a) by 0.3 μm Ca2+ in the presence of NA+GTP and (b) by 0.2 μm Ca2++GTPγS. Olmesartan significantly enhanced, while Ang II (0.1 nm) significantly inhibited, the 8‐Br‐cGMP‐induced relaxation. We propose the novel hypothesis that in these arteries, Ang II localized within smooth muscle cells activates AT1Rs and inhibits ACh‐induced, endothelium‐dependent relaxation at least partly by inhibiting the action of cGMP on these cells.

Distinct effects of CGRP on typical and atypical smooth muscle cells involved in generating spontaneous contractions in the mouse renal pelvis

Background and purpose:  We investigated the cellular mechanisms underlying spontaneous contractions in the mouse renal pelvis, regulated by calcitonin gene‐related peptide (CGRP).

Intercellular Communication Within the Rat Anterior Pituitary: XIV Electron Microscopic and Immunohistochemical Study on the Relationship Between the Agranular Cells and GnRH Neurons in the Dorsal Pars Tuberalis of the Pituitary Gland

Although numerous investigators in 1970s to 1980s have reported the distribution of LH‐RH nerve fibers in the median eminence, a few LH‐RH fibers have been shown to be present in the pars tuberalis. The significance of the finding remains to be elucidated, and there are few studies on the distribution of LH‐RH neurons in the pars tuberalis, especially in the dorsal pars tuberalis (DPT). Adult male Wistar‐Imamichi rats were separated into two groups: one for electron microscopy and the other for immunohistochemistry to observe LH‐RH and neurofilaments. Pituitary glands attached to the brain were fixed by perfusion, and the sections were prepared parallel to the sagittal plane. The typical glandular structure of the pars tuberalis was evident beneath the bottom floor of the third ventricle, and the thick glandular structure was present in the foremost region. Closer to the anterior lobe, the glandular structure changed to be a thin layer, and it was again observed at the posterior portion. Then the pituitary stalk was surrounded with the dorsal, lateral, and ventral pars tuberalis. LH‐RH and neurofilaments fibers were noted in the bottom floor, and some of them vertically descended to the gland. Adjacent to the glandular folliculostellate cells in the pars tuberalis, Herring bodies with numerous dense granules invading into the gland were present between the pituitary stalk and DPT. It was postulated that the “message” carried by LH‐RH might have been transmitted to the cells in the DPT to aid in the modulation of LH release. Anat Rec, 290:1388–1398, 2007.

Calcium phosphate stones produced by Madin-Darby canine kidney (MDCK) cells inoculated in nude mice.

Abstract The canine renal distal tubular cell line Madin-Darby canine kidney (MDCK) forms calcium phosphate microliths during a long-term culture in vitro. We identified osteopontin (OPN) and calprotectin (CPT) from a urinary stone matrix. We recently also detected the expression of OPN and CPT in MDCK cells. The relationship between the mechanism of the stone formation and these stone matrix proteins is not yet known. Here, MDCK cells were cultured and inoculated in the subcutis of nude mice. After 4, 8 and 12 weeks, the inoculated tissues were resected, fixed and immunostained with polyclonal anti-human OPN and polyclonal anti-human CPT antibodies. Some serial specimens were stained with von Kossas procedure. MDCK cells formed some follicular formations in the subcutis of nude mice at least at 12 weeks after transplantation. At 8 weeks after the inoculation, we detected small calcium phosphate stones with MDCK cells trapped in the follicles. The cells forming the stones also expressed both OPN and CPT. The CPT expression sites coincided with the stone formation sites. We confirmed that MDCK cells inoculated in nude mice had stone-forming potential, and we speculate that OPN and CPT play important roles in stone formation by MDCK cells.

Sealing of the follicular lumen of the anterior pituitary gland of the male rat

It is commonly accepted that follicular lumina of the adult rat anterior pituitary gland are tightly sealed by junctional complexes, especially tight junctions. In this report, we describe the presence of follicular lumina that are unsealed. Peroxidase (HRP) was used to study such structures and when injected through the femoral vein, was observed in association with a few follicular lumina, on their microvilli and around the cilia of folliculo-stellate cells. The existence of peroxidase-positive follicles clearly shows that follicles of the hypophysis are not always firmly sealed by tight junctions. The folliculo-stellate cells which faced the peroxidase-positive follicles displayed HRP deposits which were membrane bound within their cytoplasm. These findings suggest an absorptive function for the folliculo-stellate cells.

Electron Microscopical Observations on the Curative Effect of Ursodesoxycholic Acid in Alloxan-Induced Pancreatic Islet Cell Injury

The preventive effect of ursodesoxycholic acid on pancreatic injury by alloxan (alloxan diabetes) has been reported by Watari, et al. (1976). In the following experiment, to pursue the findings further, ursodesoxycholic acid was used curatively for alloxan diabetes. A first group of animals (5 mice) were injected with alloxan (4 mg) twice at the fifth and tenth day. The second group (5 mice) was injected with ursodesoxycholic acid (0.2 mg each) for 14 days during the experiment in addition to the same alloxan dosage/frequency as the first group. A third group of animals (5 mice) served as the control. The animals were sacrificed on the 15th day and the blood sugar levels were examined, using commercial test paper. The pancreatic tissues were fixed in a mixture of 2.5% glutaraldehyde and 2% osmid acid solution, which was adjusted at pH 7.4 with a veronal acetate buffer; the osmotic pressure was also regulated by adding sucrose of 0.045 g/ml. Following dehydration using a series of alcohol concentrations, the tissues were embedded in Epon 812. Thin sections were cut with a Porter-Blum MT-2B ultramicrotome, stained with both uranyl acetate and a lead mixture, and then observed by electron microscopy. The results were as follows: The pancreatic islet cells, especially of B-cells in the first group of animals injected with alloxan only, were seriously damaged and contained myelinated mitochondria. Golgi apparatus, and an increasing number of autophagic vacuoles. Some B-cells revealed hydropic degeneration. Some B-granules changed into vacuoles after diacrine secretion. Pancreatic A-cells were increased in number and showed no cell injuries. On the other hand, the pancreatic B-cells of mice treated with both alloxan and ursodesoxycholic acid maintained almost normal fine structures. In summary, ursodesoxycholic acid has a curative effect on alloxan-induced pancreatic B-cell injury.