Ultrastructure and immunocytochemistry of the neuroepithelial bodies in the lung of the tiger salamander,Ambystoma tigrinum (Urodela,amphibia)

Light and electron microscopy of the lungs of Ambystoma tigrinum (Urodela) revealed a relatively complex pattern of the neuroendocrine (NE) cells. In the apical parts of smaller septa single NE cells not associated with nerve fibres were covered and surrounded by pneumocytes. The larger septa possessed small areas of ciliated epithelium, in which the NE cells were grouped in a form of neuroepithelial bodies (NEB) consisting of 3–5 cells and covered by goblet cells. NE cells possessed a large nucleus with patches of condensed chromatin, clear cytoplasm, and membrane-bound vesicles of variable morphology and size, of these dense core vesicles (DCV) ranged from 70–140 nm, while rarely the larger ones exhibited a diameter of 300–600 nm. In some NEB a second type of NE cells was observed for the first time in an amphibian species: these cells communicated with the air space and exhibited on their surface microvilli and a single modified cilium with a 8+1 microtubule arrangement. Their cytoplasm contained two types of DCV: dense core granules with a diameter of 140–260 nm and vesicles 320–700 nm in diameter with a moderately electron dense interior. The NEB were associated with intracorpuscular, sensory nerve terminals morphologically afferent and efferent. By immunocytochemistry, the NE cells revealed the presence of serotonin, met-en-kephalin, and leu-enkephalin. A paracrine and chemoreceptor role is proposed for NEB of Ambystoma tigrinum.© Willey-Liss, Inc.     

猕猴输精管内注射高分子聚合物HFMC后对睾丸组织结构的影响

本实验选用具有生育力成年雄性猕猴７只，在直视下行双侧ＨＦＭＣ输精管内注射，每侧剂量分别为３０ｍｇ１只，６０ｍｇ和１００ｍｇ各３只；于注射后２．５年和３．５年分别处死动物，取睾丸组织进行光镜和电镜观察．结果发现：猕猴注射ＨＦＭＣ２．５年后，睾丸光镜大部分曲细精管生精上皮结构完整，排列整齐。仅见局部少数管腔生精上皮层数减少，上皮细胞轻度水样变性等病理改变。电镜下曲细精管内除支持细胞内脂褐素增多，轻度基底膜增厚和精母细胞内质网扩张外，各级生精细胞，支持细胞及细胞间连接复合体等超微结构未见明显异常。注射ＨＦＭＣ３．５年后猕猴的光镜、电镜结果与注射后２．５年结果相似，但局部改变较２．５年组轻。上述结果表明：猕猴输精管内注射一定剂量ＨＦＭＣ节育不会引起睾丸组织的严重病理改变。但是，由于注射ＨＦＭＣ后，ＨＦＭＣ释放Ｈ＋及其对输精管的暂时阻塞，改变了精子生存的内环境，使睾丸出现局部轻度病理改变，随着ＨＦＭＣ逐渐溶解排出，睾丸功能相继恢复正常，配对产仔。为ＨＦＭＣ应用提供了安全性依据。     

The osmotic behaviour of toad skin epithelium (Bufo viridis)

The effect of saline adaptation on the intracellular Na, K, Cl, P concentrations and dry weight content of the toad skin epithelium (Bufo viridis) was studied using the technique of electron microprobe analysis. The measurements were performed on isolated abdominal skins either directly after dissection or after additional incubation in Ussing-type chambers.Adaptations of the toads to increasing NaCl concentrations for 7 days resulted in increased blood plasma osmolarity and a parallel increase in the cellular electrolyte, P and dry weight concentrations of the epithelium, the K increase representing the most significant fraction of the intracellular osmolarity increase. No evidence was obtained to show that the nucleus and cytoplasm reacted differently from each other and all living epithelial cell types basically showed the same response.Incubation of the isolated skins under control conditions showed a drastic inhibition of the transepithelial Na transport after adaptation to high salinities. In spite of the large variations in the transport rate almost identical intracellular electrolyte concentrations were observed. In tap water adapted toads the average cellular concentrations were 8.8 mmole/kg wet weight for Na, 109.6 for K, 41.5 for Cl, and 135.3 for P, respectively. Incubation of the skin with Ringer's solution of different osmolarities demonstrated that the epithelial cells are in osmotic equilibrium with the inner bathing solution. The results are consistent with the view that the osmotic adaptation is mainly accomplished by the movement of water.This work was supported by grants from the Deutsche Forschungsgemeinschaft and the Stiftung Volkswagenwerk     

Expression of luminal and basal cytokeratins in human breast carcinoma

We have examined basal and luminal cell cytokeratin expression in 1944 cases of invasive breast carcinoma, using tissue microarray (TMA) technology, to determine the frequency of expression of each cytokeratin subtype, their relationships and prognostic relevance, if any. Expression was determined by immunocytochemistry staining using antibodies to the luminal cytokeratins (CKs) 7/8, 18 and 19 and the basal markers CK 5/6 and CK 14. Additionally, assessment of alpha-smooth muscle actin (SMA) and oestrogen receptor status (ER) was performed. The vast majority of the cases showed positivity for CK 7/8, 18 and 19 indicating a differentiated glandular phenotype, a finding associated with good prognosis, ER positivity and older patient age. In contrast, basal marker expression was significantly related to poor prognosis, ER negativity and younger patient age. Multivariate analysis showed that CK 5/6 was an independent indicator for relapse free interval. We were able to subgroup the cases into four distinct phenotype categories (pure luminal, mixed luminal/basal, pure basal and null), which had significant differences in relation to the biological features and the clinical course of the disease. Tumours classified as expressing a basal phenotype (the combined luminal plus basal and the pure basal) were in a poor prognostic subgroup, typically ER negative in most cases. These findings provide further evidence that breast cancer has distinct differentiation subclasses that have both biological and clinical relevance.     

Glomerular expression of cell-cycle-regulatory proteins in human crescentic glomerulonephritis

To elucidate the mechanism underlying crescentic formation, we assessed the phenotypic characterization and cell-cycle protein expression in human crescentic glomerulonephritis (CRGN). Kidney tissue specimens taken from CRGN patients (10 patients with pauci-immune type rapidly progressive glomerulonephritis (RPGN), 2 patients with Henoch-Schönlein purpura nephritis, and 1 patient with IgA nephropathy) were examined immunohistochemically. Most of the cellular components of the crescents expressed cytokeratin, whereas few cells expressed PHM-5. CD68-positive cells were minor components of cellular crescents, indicating that the major principal cellular component of the crescents is made up of cells with the parietal glomerular epithelial cell (PEC) phenotype. Additionally, serial section analysis revealed that Ki-67-positive cells in the crescents were frequently cyclin-A positive and Bcl-2 positive, but seldom cyclin-B₁ positive. Moreover, the expression of cyclin-dependent kinase inhibitor p27^Kip1 was low in the cellular crescents, despite being exclusively positive in podocytes within the same section. We concluded that the major component of the cellular crescents is made up of PECs and that apparent expression of cyclins and Bcl-2 and restrained expression of p27^Kip1 may be synergistically associated with the development of cellular crescents in human CRGN.     

Analysis of thymic epithelial cell proliferation in vitro by combining bromodeoxyuridine and keratin labeling in an immunofluorescence assay

A simple method of analyzing thymic epithelial cell (TEC) proliferation has been developed by combining bromodeoxyuridine (BrDU) and keratin labeling in an immunofluorescence assay. The first reagent specifically visualizes the cells entering the S phase of the cell cycle, whereas the second immunostaining reveals which of the proliferating BrDU-positive cells actually belong to the epithelial lineage. This method, besides being rapid and free of radioactivity, appears to be reliable in view of the minor variations in the percentages of BrDU+ TEC observed in several distinct experiments. Thus, BrDU/keratin immunolabeling appears to represent a useful tool for the analysis of in vitro TEC proliferation.     

Nasal mucosa in natural colds: effects of allergic rhinitis and susceptibility to recurrent sinusitis

The mechanisms of virus-induced airway hyperresponsiveness in asthma and allergy and the failure of host defence in patients suffering from secondary airway infections are still largely unknown. The aim of this study was to examine whether the presence of allergic rhinitis or susceptibility to recurrent sinusitis affects the structural and cellular changes in nasal mucosa during natural colds and convalescence. We compared the mucosal changes in biopsy samples during acute natural colds (days 2-4 of illness) and convalescence (3 weeks later) in patients with allergic rhinitis (n = 9), patients with susceptibility to sinusitis (n = 19) and healthy controls (n = 20). We saw similarly increased numbers of mucosal T and B lymphocytes and mast cells and increased vascular density during the acute colds compared to convalescence in all the three groups. The allergic subjects had elevated levels of eosinophils in the acute phase (P = 0.03), and the allergic and sinusitis-prone subjects had elevated levels of epithelial T cells (P = 0.04) and low levels of mast cells (P = 0.005) in convalescence compared to the control group. The sinusitis-prone subjects lacked intraepithelial cytotoxic cells in convalescence. In the allergic subjects, the reticular basement membrane was thicker in the acute phase compared to the convalescence (P = 0.05). These results suggest that various cells of the airways, including inflammatory and structural cells, are involved during viral respiratory infections in subjects with allergic rhinitis. The small numbers of mast cells and cytotoxic lymphocytes in the sinusitis-prone subjects may be related to their susceptibility to bacterial complications.     

Na+/H+ exchange regulates intracellular pH of rat gastric surface cells in vivo

Intracellular pH (pH_i) and viability of gastric surface cells of the rat stomach in response to luminal acidification, and the role of Na⁺/H⁺ exchange in maintaining pH_i homeostasis were studied in vivo using a fluorescent microscopic technique. pH_i was measured during superfusion with buffers of pH 1.2–7.4. When the pH of the superfusate was 7.4, baseline pH_i was unchanged. Superfusion with pH 3 buffer rapidly decreased pH_i to 6.7, with subsequent recovery to baseline pH_i within 15 min despite continuing acid exposure. Superfusion with buffers of pH 1.7 and 1.2 decreased pH_i continuously to below 6.2 with no recovery observed. Despite the relentless decline in pH_i during superfusion with pH-1.2 and –1.7 solutions, over 75% of the surface cells were still viable, as measured by exclusion of the vital dye propidium iodide. We then examined the role of Na⁺/H⁺ exchange in the regulation of pH_i. Superfusion with amiloride did not affect recovery of pH_i from intracellular acidification induced by a NH₄Cl prepulse. Exposure to the potent, lipophilic Na⁺/H⁺ exchange inhibitor 5-(N,N-hexaniethylene)-amiloride (HMA), either in the superfusate or by close arterial perfusion, decreased baseline pH_i from 7.1 to 6.8. Close arterial perfusion of HMA additionally attenuated the recovery of pH_i to baseline during superfusion with pH 3 buffer. We conclude that luminal protons permeate into the cytoplasm of gastric surface cells, where they are eliminated by an Na⁺/H⁺ exchanger, most probably localized to the basolateral membrane.