Study on the ultrastructure of the peritoneal stomata in humans.

In 16 human specimens the topography and organization of stomata and mesothelial cells of the diaphragmatic, pelvic wall and anterior abdominal wall peritoneum were studied by transmission electron microscopy, scanning electron microscopy and the image processing technique. The mesothelial cells were organized into two discrete populations, cuboidal cells and flattened cells. The stomata were found only among cuboidal cells, either on the muscular portion or on the tendinous portion of the diaphragm. The size and shape of stomata, which were arranged in a cluster or a strip, were often irregular. The average area of a stoma on the muscular portion was 10.43 +/- 1.61 microns2, on the tendinous portion 7.93 +/- 1.67 microns2. Stomata opened to submesothelial connective tissue, under which numerous lymphatics were observed. Stomata were not discovered in the pelvic and anterior abdominal wall peritoneum. In animal experiments intraperitoneally injected trypan blue particles were rapidly removed from the peritoneal cavity through stomata of the diaphragmatic peritoneum in rabbits. It is suggested that stomata may be the main pathway for draining matter from the peritoneal cavity and that the diaphragmatic peritoneum shows the strongest absorption in all parts of the peritoneum.     

Light and electron microscope observations of the lymphatic drainage units of the peritoneal cavity of rodents

Fluid, particles, and cells are taken up from the peritoneal cavity by lymphatic drainage units, which, in the mouse and rat, are located along the peritoneal surface of the muscular portion of the diaphragm. The drainage units are composed of three specifically differentiated components: a lymphatic lacuna, a covering of lacunar mesothelium, and intervening submesothelial connective tissue. The units are drained by connecting lymphatic vessels that cross the diaphragm to empty into collecting lymphatic vessels running along the pleural surface of the diaphragm. The collecting lymphatics empty into parasternal lymphatic trunks. In this report, we briefly review critical features of the drainage apparatus and describe new observations, summarized below, about their structure. Around the rim of stomata, the mesothelial openings that lead into the lymphatic lacunae, plasma membranes of lacunar mesothelial cells and of lacunar endothelial cells abut but are not linked to one another by recognizable junctional specializations. Lacunar endothelial cells often extend valve-like processes that bridge the distal end of the channel beneath the stoma. The configuration of the endothelial processes may be complex. Occasionally, processes from fibroblasts in the submesothelial connective tissue adjacent to stomata make contact with the interstitial surface of lacunar endothelial cells. A discontinuous elastic layer in the submesothelial connective tissue spans the roof of each lacuna. Connecting and collecting lymphatics, which drain lymphatic lacunae, possess endothelial valves. Possible functions for each of these newly described structural features are discussed.     

Distribution of diaphragmatic lymphatic lacunae.

The morphology of the submesothelial lymphatic lacunae on the pleural and peritoneal surface over the tendinous and muscular portion of the diaphragm was studied in 10 anesthetized rabbits. The lymphatic network was evidenced by injecting 1 ml of colloidal carbon solution in the pleural (n = 5) or the peritoneal (n = 5) space. After 1 h of spontaneous breathing, the animal was killed and the diaphragm was fixed in situ by injection of approximately 5 ml of fixative in pleural and peritoneal spaces. Then both cavities were opened and the diaphragm was excised and pinned to a support. According to which cavity had received the injection, the peritoneal or the pleural side of the diaphragm was scanned by sequential imaging of the whole surface by use of a video camera connected to a stereomicroscope and to a video monitor. The anatomic design appeared as a network of lacunae running either parallel or perpendicular to the major axis of the tendinous or muscular fibers. The lacunae were more densely distributed on the tendinous peritoneal area than on the pleural one. Scanty lacunae were seen on the muscular regions of both diaphragmatic sides, characterized by large areas without lacunae. The average density of lacunae on tendinous and muscular regions was 6 and 1.7/cm2 for the pleural side and 25 and 3.4/cm2 for the peritoneal side, respectively. The average width of lacunae was 137.9 +/- 1.6 and 108.8 +/- 1.7 microns on the tendinous pleural and the peritoneal side, respectively, and 163 +/- 1.8 microns on the muscular portion of the pleural and peritoneal surfaces.     

Mesothelial stomata overlying omental milky spots: Scanning electron microscopic study

Summary The presence in the rat omentum of intercellular pores (the classical stomata of von Recklinghausen) between the mesothelial cells overlying aggregates of lymphoreticular cells (the classical milky spots of Ranvier) and the apparent migration of lymphocytes through these stomata were recorded for the first time by scanning electron microscopy. Previous studies on passage of cells across the peritoneum and omentum used experimentally administered cells, while in the present study no cells were administered to the rats and their own lymphocytes were observed in situ. The possible role of lymphocytes in the peritoneal cavity is also briefly discussed.     

小鼠膈腹膜淋巴孔和膈淋巴管的发育

应用透射电镜、扫描电镜和酶组织化学方法,研究胚胎期和出生后不同时期小鼠膈腹膜淋巴孔（PLS）和膈淋巴管的发生和发育,并用Elescope计算机图像处理技术对PLS进行定量分析。结果发现：胚胎13天时,膈腹膜仅由扁平形间皮细胞（FMC）组成;胚胎15天时,FMC间出现立方形间皮细胞（CMC）和早期腹膜淋巴孔（NLS）;胚胎18天时,膈毛细淋巴管出现,台盼蓝吸收试验显示NLS无物质吸收功能;出生后1天（PND1）,膈毛细淋巴管内皮细胞向PLS伸出胞质突起,并横跨CMC下结缔组织纤维和基底膜,形成腹膜下小管。后者与PLS沟通,建立了腹膜腔内物质转归通路。台盼蓝吸收试验表明,出生后PLS具有物质吸收功能,即为成熟腹膜淋巴孔（MLS）。PND5,立方细胞嵴（CMCR）发生,膈毛细淋巴管数量增多。PND10,大量立方细胞嵴融合,形成条带状分布的立方细胞区域,其上分布有大量MLS。随着发育进展,MLS平均面积和平均分布密度逐渐增大,且随着膈淋巴管的发育,吸收功能逐渐增强。     

Recombinant GPI-anchored TIMP-1 stimulates growth and migration of peritoneal mesothelial cells

BACKGROUND: Mesothelial cells are critical in the pathogenesis of post-surgical intraabdominal adhesions as well as in the deterioration of the peritoneal membrane associated with long-term peritoneal dialysis. Mesothelial denudation is a pathophysiolocigally important finding in these processes. Matrix metalloproteinase (MMP) biology underlies aspects of mesothelial homeostasis as well as wound repair. The endogenous tissue inhibitors of metalloproteinases (TIMPs) moderate MMP activity. METHODS AND FINDING: By modifying human TIMP-1 through the addition of a glycosylphosphatidylinositol (GPI) anchor, a recombinant protein was generated that efficiently focuses TIMP-1 on the cell surface. Treatment of primary mesothelial cells with TIMP-1-GPI facilitates their mobilization and migration leading to a dramatic increase in the rate of wound experimental closure. Mesothelial cells treated with TIMP-1-GPI showed a dose dependent increase in cell proliferation, reduced secretion of MMP-2, MMP-9, TNF-α and urokinase-type plasminogen activator (uPA), but increased tissue plasminogen activator (t-PA). Treatment resulted in reduced expression and processing of latent TGF-β1. CONCLUSIONS: TIMP-1-GPI stimulated rapid and efficient in vitro wound closure. The agent enhanced mesothelial cell proliferation and migration and was bioactive in the nanogram range. The application of TIMP-1-GPI may represent a new approach for limiting or repairing damaged mesothelium.     

The effect of the polyethylene intrauterine device on peritoneal fluid cell distribution of adult female mice

At Hahnemann Medical College and Hospital in Philadelphia, researchers attempted to determine whether uterine cavity IUD-induced inflammation extends into the abdominal cavity. Adult female mice were used. In each experimental animal (14), a piece of polyethylene intramedic tubing (1.3 cm long) was fixed in place by suture within each uterine horn. In the sham-operated controls (15), a silk suture was sutured into each horn. 3 weeks later peritoneal fluid specimens were obtained from the 2 groups and compared. The IUD resulted in significant increases in histiocytes (pl.001) and polymorphonuclear leukocytes (pl.001). Mesothelial cells were significantly decreased (pl.001), but mesothelial cells were somehwat higher in the controls than is usually found. No significant changes in lymphocytes or monocytes were observed. Only experimental animals showed daisy cells. Thus, the IUD resulted in a definite abdominal inflammatory effect. Judging by the spontaneous activity measured on an Electronic Activity Monitor, the IUD produced no animal discomfort. 25% of the IUDs were expelled.     

Functional Stomata in a Variegated Leaf Chimera of Pelargonium zonale L. without Guard Cell Chloroplasts

Fluorescence microscopy indicated that chlorophyll was absentfrom epidermal and guard cells overlying all white areas andgreen areas (of certain leaves) in variegated leaves of Pelargoniumzonale, cv. Chelsea Gem. Stomata with chlorophyll-free guardcells, in general, responded normally to light and CO₂ as gaugedby direct measurements of stomatal aperture and by transpirationalwater loss studies, although stomata from white regions of variegatedleaves were more reluctant to open than stomata from green regionsof the leaves. Thus, functional stomata without guard cell chloroplastshave been discovered in another genus, namely Pelargonium, besidesthat originally discovered in Paphiopedilum. When stomata withchlorophyll-free guard cells opened, K⁺ accumulated in the guardcells. This indicates that chloroplasts are not essential forthe normal functioning of stomata and that the energy sourcefor driving stomatal movements can come from sources other thanphotophosphorylation. Key words: Guard cell chloroplasts, Leaf chimera, Pelargonium, Stomata     

Structure and development of stomata on the primary root of Ceratonia siliqua L

Stomata of various sizes are produced on the primary root of Ceratonia siliqua L. Most are generated during embryogenesis, prior to seed desiccation. They can be detected on the dry embryo in a wide zone just above the root tip. Initially, large stomata are formed. These have the ability to induce divisions of their neighbouring cells, creating particular cell patterns around them. Later, small perigenous stomata are generated. As the root grows following seed germination, the stomatal zone overlaps with that of the root hairs. Although root stomata of C. siliqua undergo a structural differentiation that seems almost identical to that of the elliptical stomata formed on leaves, they are unable to move and remain permanently open. Polarizing microscopy of fully differentiated stomata and young stomata at the stage of stomatal pore formation revealed deposition of radial cellulose microfibril systems on their periclinal walls. However, these systems were less developed than those on leaf stomata, a feature that might be responsible for their inactivity. Besides, plastids of the root guard cells (GCs) do not differentiate into chloroplasts but function solely as amyloplasts. Root stomata have a short life span. During rapid and intense root growth, GCs cannot keep pace with the elongation of their neighbouring rhizodermal cells. They therefore split in their mid-region, transversely to the stoma axis. The two parts of the transversely torn stoma are dragged apart and a large opening is formed on the root surface, just above the substomatal cavity. The root stomata, together with these openings, may facilitate increased gaseous exchange during respiration and/or an increased transfer of some nutrients and water in the rapidly growing primary root.     

Immunohistochemical localisation of ET-1 and eNOS in lymphatic stomata of the porcine broad ligament of the uterus

Immunohistochemical localization and distribution of endothelin (ET-1) and nitric oxide synthase (eNOS) were investigated in lymphatic stomata in areas of their special accumulation in the porcine broad ligament during the estrous cycle. The study was performed using polyclonal antibody for ET-1 and monoclonal antibody for eNOS. ET-1 and eNOS immunoreactivities were demonstrated in some thin endothelial lymphatic lacuna walls throughout the estrous cycle. In the mesothelial cell layer, ET-1 and eNOS were detected only in stomata-related cuboidal mesothelial cells, however, the intensity of the immunostaining and distribution of the positive cells varied during the cycle. These results suggest that ET-1 and eNOS can play a role in mechanisms regulating the tone of lymphatic stomata during the absorption and passage of fluids, particles and cells from the peritoneal cavity to lymphatic vessels in the porcine broad ligament.     

Specialized structures in the leaf epidermis of basal angiosperms: morphology, distribution, and homology

The morphology of specialized structures in the leaf epidermis of 32 species of basal (ANITA: Amborella, Nymphaeales, Illiciales, Trimeniaceae, and Austrobaileyaceae) angiosperms, representing all seven families and 11 of 14 genera, was investigated using light and scanning electron microscopy. Distribution, density, and size of structures were also measured, and character evolution was analyzed. Hydropotes are a synapomorphy of Nymphaeales and ethereal oil cells are a synapomorphy of Austrobaileyales, but uniseriate nonglandular trichomes appear to have arisen independently several times. Specialized structures are frequently characterized by adjacent epidermal cells that have striking similarities in their form and arrangement (i.e., architecture) to subsidiary cells of certain types of stomatal complexes. Additionally, forms intermediate to oil cells and stomata, to trichomes and stomata, and to hydropotes and oil cells are present in some taxa. Thus, all of these specialized structures and their adjacent epidermal cells form complexes that may be homologous with, and evolutionarily derived from stomatal complexes, and the specialized structure, or portion thereof, may be homologous to the stoma or guard mother cell. Improved knowledge of the morphology and evolution of these structures in the earliest branching extant angiosperm lineages has a bearing on many diverse areas of botany.     

A scanning electron microscope study of mouse peritoneal mesothelium.

As seen in the scanning electron microscope, peritoneal mesothelial cells of the mouse diaphragm, anterior abdominal wall and intestinal serosa carry numerous microvilli. These microvilli are absent over certain areas of the cell surface and are sometimes, interlocked in meshwork patterns or coronal formation. The apical cell membranes of the mesothelium at the base of the microvilli, are invaginated by many plasmalemmal vesicles and vacuoles and carry a number of protruding spherical structures. Deep circular craters, giving the impression of stomata, are also visible.     

Significance of pericellular lacunae in cell blocks of effusions.

A retrospective study was conducted to assess the usefulness of pericellular lacunae in cell block sections of serous effusions in diagnosis. From January to December 1988, 286 cell blocks were prepared in our laboratory from pleural, pericardial and peritoneal fluids; 62 of them were excluded from this study because of inadequate cellularity, diagnostic uncertainty or lack of a proteinaceous background. The remaining consisted of 148 benign effusions from 128 patients and 76 malignant effusions from 56 patients. A single specimen from each patient was selected and reviewed to assess the presence and number of pericellular lacunae and to determine the relationship of this feature to cell arrangement (single cells versus cell clusters). Pericellular lacunae were found in 42 (75%) of the malignant effusions as compared to 41 (32%) of benign specimens. In the majority of malignant cases with lacunae, this feature was associated with greater than two-thirds of the cells, whereas in benign cases, when present, it was seen in less than one-third of the cells. Neoplasms characterized by large cell clusters more frequently had lacunae than did those with small groups or single cells. Lacunae were not evident in cases of malignant melanoma and lymphoma. We conclude that although pericellular lacunae are more often associated with malignant cells, their presence in itself cannot be used as a reliable indicator of malignancy in body cavity fluids.     

A quantitative analysis of the interspecific plasmodesmata in the non-division walls of the plant chimera Laburnocytisus adamii (Poit.) Schneid.

Non-division walls in petals of the chimera Laburnocytisus adamii (Poit.) Schneid, were screened for the occurrence and distribution of symplasmic connections. The secondary plasmodesmata (PD) between epidermal cells of Cytisus purpureus Scop, and subepidermal cells of Laburnum anagyroides Medik. were compared with the PD of corresponding cell walls in petals of the two parental species. The non-division walls in the petals of L. adamii were traversed mainly by continuous PD and a few half-PD, both being grouped in pit fields. The secondary PD were characterized by a high percentage of branching (82%), with more than 40% consisting of a single strand at the Cytisus cell side interconnected by a median cavity with two strands of the Laburnum subepidermal cell. In addition, more than 30% of all PD showed secondary branching in the subepidermal wall portion. As a consequence, the cross-sectional areas of plasmodesmatal strands on each side of the central cavity differed remarkably in size, representing a “bottleneck” in the epidermal wall portion. In contrast, PD in the petals of the parental species were symmetrically branched. The comparison of cross-sectional areas of PD in the cell wall between the epidermis and subepidermis of petals of L. anagyroides showed a well-tuned system. The occurrence of half-PD in the intraspecific wall indicates a secondary origin. We conclude that, in the chimera, both genotypically different cells take part in the formation of the interspecific PD.     

Surface architecture of the mucosal epithelium of the cat trachea: I. Cartilaginous portion

The mucosal covering of the cartilaginous portion of the cat trachea was studied by correlated light, transmission-electron, and scanning-electron microscopy. While in some areas the ciliated pseudostratified epithelial lining is fairly smooth in contour, in other areas it contains longitudinally oriented, cilia-lined clefts. Ducts from submucosal glands sometimes open into the base of these clefts, or into funnel-shaped stomata that are lined by either ciliated or microvillus-rich cells. Specialized epithelial cells are occasionally associated with the clefts or with other regions of surface epithelium. In single sections, these cells appear to contain a cilium-lined vacuole, but serial sectioning has demonstrated that these apparent vacuoles actually are long intracellular invaginations in enormously elongated cells that extend longitudinally in the plane of the epithelium. The function of these cells is undetermined. Basal cells are attached to the lamina densa by means of hemidesmosomes that consistently lack peripheral densities; in contrast, the tall columnar cells have no hemidesmosomes.     

Phagocytosis of dying chondrocytes by osteoclasts in the mouse growth plate as demonstrated by annexin-V labelling

Endochondral ossification in the epiphyseal growth plate of long bones is associated with programmed cell death (PCD) of a major portion of the chondrocytes. Here we tested the hypothesis that at the ossification front of the epiphyseal growth plate osteoclasts preferentially phagocytose chondrocytes that are undergoing PCD. We injected biotin-labelled annexin-V (anx-V-biotin, an early marker of PCD) intravenously in young adult mice. After 30 min of labelling, long bones were recovered and the tissue distribution examined of anx-V-biotin-labelled cells in the growth plate using ABC-peroxidase histochemistry. Positive staining for anx-V-biotin was detected in hypertrophic chondrocytes still present in closed lacunae at some distance from the ossification front. At the ossification front, chondrocyte lacunae were opened and close contacts were seen between tartrate-resistant acid phosphatase-positive osteoclasts and hypertrophic cartilage cells. Osteoclasts were significantly more frequently in contact with anx-V-biotin-labelled chondrocytes than with unlabelled chondrocytes. Osteoclasts also contained labelled and unlabelled phagocytic fragments within their cytoplasm. We conclude that in the growth plate osteoclasts preferentially phagocytose hypertrophic chondrocytes that are dying, suggesting these dying cells may signal osteoclasts for their removal.     

Distribution of diaphragmatic lymphatic stomata

In seven anesthetized rabbits we measured the size, shape, and density of lymphatic stomata on the peritoneal and pleural sides of the diaphragm. The diaphragm was fixed in situ and processed for scanning electron microscopy. Results are from 2,902 peritoneal and 3,086 pleural fields (each 1,620 microns 2) randomly chosen from the various specimens. Stomata were seen in 9% of the fields examined, and in 30% of the cases they appeared grouped in clusters with 2-14 stomata/field. Stoma density was 250 +/- 242 and 72 +/- 57 (SD) stomata/mm2 on peritoneal and pleural sides, respectively, and it was similar over the muscular and tendinous portion of the two surfaces. The maximum diameter ranged from less than 1 to approximately 30 microns, with an average value of 1.2 +/- 3.1 micron. The ratio of the maximum to the minimum diameter and the surface area averaged 2 +/- 1.4 and 0.7 +/- 2.4 micron 2, respectively. The maximum and minimum diameter and surface area values followed a lognormal frequency distribution, suggesting that stomata geometry is affected by diaphragmatic tension.     

Ultrastructural immunocytochemical study of the massa caudalis of the subcommissural organ-Reissner's fiber complex in lamprey larvae (Geotria australis): Evidence for a terminal vascular route of secretory material

Summary The massa caudalis of the subcommissural organ-Reissner's fiber complex of lamprey larvae (Geotria australis) was studied immunocytochemically at the ultrastructural level by use of the immunoperoxidase-silver methenamine procedure. An antiserum raised against bovine Reissner's fiber was utilized as primary antibody.The caudalmost portion of the central canal and its ampulla caudalis communicate, via wide intercellular spaces in their dorsal wall, with large cavities or lacunae. In addition, distinct openings in the dorsal wall of the ampulla establish an open communication between the latter and the lacunae. The lacunae are lined by slender processes of cells of unknown nature. No junctional complexes can be observed between these cells, which lack a basal lamina. The lacunae communicate with structures resembling blood capillaries, however, they are devoid of a basal lamina. These peculiar vessels, in turn, are in direct communication with characteristic blood capillaries.Reissner's fiber (RF) and its massa caudalis are strongly immunoreactive with the antiserum used. The wide intercellular spaces in the dorsal wall of the central canal and the ampulla, as well as the lumina of the (i) lacunae, (ii) modified vessels and (iii) blood capillaries are filled with a flocculent, strongly immunoreactive material. No immunoreactive material was found outside these structures. Thus, the blood capillaries appear to represent the only final target of RF-material arriving at the ampulla caudalis.Supported by Grant I 38259 from the Stiftung Volkswagenwerk, Federal Republic of Germany, Grant S-85-39 from the Dirección de Investigaciones, Universidad Austral de Chile, and Grant 6027 from Fondo Nacional de Desarrollo Científico y Tecnológico, Chile. The authors express their gratitude to Mrs. Elizabeth Santibáñez and Mr. Julio Lamilla for providing the lamprey larvae and to Mr. Humberto Molina for preparing the three-dimensional drawing     

The Floral Nectary of Tropaeolum majus L.--The Nature of the Secretory Cells and the Manner of Nectar Secretion

The floral nectary of Tropaeolun majus L. was studied with theaid of a microscope with transmitting and incident light, atransmission electron microscope and a scanning electron microscope.The Gomori method was used for the localization of acid phosphatase.As a result of this investigation the previously accepted viewthat nectar in this plant is secreted only from the hair tipsof the inner epidermis of the calyx spur was found to be inaccurate.The present studies showed that the parenchyma cells locatedbetween the inner epidermis and the region of the vascular bundlesof the lowest third of the spur, are the main nectar-secretingelements of the nectary. These secretory cells release the nectarsolution into intercellular spaces leading to modified stomata,through which it is exuded into the spur cavity. The modifiedstomata occur in the lowest portion of the spur only. At thestage of secretion small droplets of liquid of high viscositywere observed on the epidermal hairs. These droplets presumablycontain polysaccharides and a certain amount of sugar.     

Organization and differentiation of the segmentary plate in common toad embryo Bufo bufo L., following heat shock (author's transl)

A heat shock greatly disturbs the shaping and the growth of embryos with 4 or 5 somites. One part of a segmentary plate with a size including four control somites stays inorganized, whereas posterior segmentation gets on normally. The differentiation of the unsegmented somitic block (fusing myoblasts, myogenesis) is the same as the one of the previous somites; the dermatome and the sclerotome issued from the unsegmented block are not cut up. Ultrastructural observations of treated segmentary plates show that the somitic block undergoes important changes. The non-segmented somitic mesoderm of the control animals obtains a loose aspect; its constitutive cells separated by lacunae are brought into contact by the means of filopods and junctions. 18 hours after the heat shock, the cells of the unsegmented block keep close to each other and are tightened together by many "pseudo-desmosomes", the intercellular spaces are reduced to small meatus. The nuclei of the somitic cells are clear and provided with a restricted amount of chromatine spots; dark granular nuclear like chromatin material is thrown out of the nucleus. In embryos fixed 40 hours after heat shock, several large plurinucleated cells, are formed in the non-segmented somitic block. Inside these plurinucleated cells, there is synthesis of myofilaments and differentiation of myofibrilles. Numerous intercellular lacunae are formed, some yet apparent "pseudo-desmosomes" form "pinchings" between the membranes of muscular cells and therefore confine series of intercellular lacunae.