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.     

豚鼠卵巢囊淋巴孔的发现及卵巢囊超微结构研究

In order to explore its structure and speciality of species, the ovary bursa of guinea pig was studied by using dissecting microscopy and scanning electron microscopy. In this paper, the lymphatic stomata on both internal and external layer of the ovary bursa was frist reported in guinea pig. The results suggested that the stomata in bursa not only provided a pathway to connect the bursal cavity with the lymphatic vessels of bursa and the peritoneal cavity, but also may be involved in the reproduction and local immunity of the ovary. The stomata may play an important role in physiological function of the ovary. At the same time, the structural differences were identified in ovary bursa between guinea pig and golden hamster.     

豚鼠卵巢囊淋巴孔的发现及卵巢囊超微结构研究

本实验通过扫描电镜等方法研究豚鼠卵巢囊及卵巢囊淋巴孔的结构,并探讨了卵巢囊及其淋巴孔的种属差异.实验结果首次报道豚鼠卵巢囊内、外层上皮均存在淋巴孔;豚鼠卵巢囊结构在输卵管走行、囊闭合程度及囊表面超微结构等方面与金仓鼠存在差异.结果提示豚鼠卵巢囊内、外层淋巴孔是沟通卵巢囊腔、卵巢囊淋巴系及腹膜腔的形态基础,可能是三者间物质转运的重要途径.卵巢囊淋巴孔可能影响物种的繁殖并参与囊腔内局部免疫反应.卵巢囊结构和发育程度与对应的输卵管伞端等结构及其他生殖特点相适应,研究结果丰富了生殖形态学和比较解剖学资料.     

Electronmicroscopic observations on the visceral and parietal rat's pleura after contralateral pneumonectomy

The mechanism of compensatory growth and healing of the pleura remains unresolved. Contralateral visceral and parietal (diaphragmatic and costal) pleura were investigated by transmission electron microscopy, following an experimental pneumonectomy (EP). Fifteen young-adult Wistar rats were divided into three groups and with survival times of 1, 5 and 8 days respectively after EP. Three sham-operated (thoracic cavity opened and closed) and three unoperated rats served as controls. One day following EP the superficial mesothelial cells have more microvilli and microvesicles, but a lower number of specialized contacts. Multiplication of extravasal cells leads to an increase of the thickness of the layer over the basal lamina and of the submesothelial layer. Five days after EP the superficial cells show a stratified arrangement in longer sectors of both pleural sheets. Along with typical mesothelial cells there are three new populations of cells: (1) with an abundant granular endoplasmic reticulum and secretory granules, (2) with fibroblast-like characteristics and (3) with a more extensive lysosomal system. The submesothelial layer is thickened due to newly formed blood vessels and collagen bundles. Eight days after EP the mesothelial cells build multi-row arrangement sectors and surround intercellular dilatations covered with microvilli. 'Activated' high mesothelial cells characterize the monolayer sectors. The submesothelial layer remains thicker due to larger collagen bundles and elastic fibers. The changes in the mesothelium and in the connective tissue layer suggest the existence of two periods. The first one is characterized by different mesothelial cell populations, new vasculogenesis and starting of fibrillogenesis. In the second period there are 'activated' mesothelial cells, pleural villi, groups of lymphatic lacunae and significant fibrillogenesis.     

妊娠和激素干预对豚鼠卵巢囊淋巴孔的影响

In order to study the influence of pregnancy and hormone on the lymphatic stomata of guinea pig's ovary bursa, the lymphatic stomata of ovary bursa were observed during pregnancy and by using exogenous hormone, including serum gonadotrophin (PMSG) combined with human chorionic gonadotropin (HCG) to promote ovulation and androgen to inhibit the ovary. Then the lymphatic stomata in the inner layer of ovary bursa and their absorption functions were observed with scanning electron microscope (SEM) and trypan blue as a tracer respectively. The lymphatic stomata were also counted by the Elescope computer image processing system. We found that the quantity of tracer absorbed by the lymphatic stomata of ovary bursa in pregnant group was more than that in ovulation-promoted group, and least in the androgen-treated group, which had statistical significance between each couples of them (P < 0.05). The opening area of the lymphatic stomata in pregnant, ovulation-promoted and androgen-treated groups were 189.9 +/- 48.7 micron 2/1000 micron 2, 104.4 +/- 31.2 micron 2/1000 micron 2 and 40.5 +/- 18.7 micron 2/1000 micron 2 respectively, which had remarkable statistical difference between each couples (P < 0.01). Under SEM observation, the secretory granules in ciliated columnar epithelium were less in pregnant group than the other two, while both the secretory granules and the cilium were most active in ovulation-promoted group. The results indicated that both the openings and absorption functions of the lymphatic stomata in guinea pig's ovary bursa could be affected by pregnancy and exogenous homone. There existed relationship between ovary function and the regulation of the lymphatic stomata in ovary bursa.     

Functional arrangement of rat diaphragmatic initial lymphatic network

Fluid and solute flux between the pleural and peritoneal cavities, although never documented under physiological conditions, might play a relevant role in pathological conditions associated with the development of ascitis and pleural effusion and/or in the processes of tumor dissemination. To verify whether a pleuroperitoneal flux might take place through the diaphragmatic lymphatic network, the transdiaphragmatic pressure gradient (Delta P(TD)) was measured in five spontaneously breathing anesthetized rats. Delta P(TD) was -1.93 cmH2O (SD 0.59) and -3.1 cmH2O (SD 0.82) at end expiration and at end inspiration, respectively, indicating the existence of a pressure gradient directed from the abdominal to the pleural cavity. Morphometrical analysis of the diaphragmatic lymphatic network was performed in the excised diaphragm of three additional rats euthanized with an anesthesia overdose. Optical and electron microscopy revealed that lymphatic submesothelial lacunae and lymphatic capillaries among the skeletal muscles fibers show the ultrastructural features of the so-called initial lymphatic vessels, namely, a discontinuous basal lamina and anchoring filaments linking the outer surface of the endothelial cells to connective tissue or to muscle fibers. Primary unidirectional valves in the wall of the initial lymphatics allow entrance of serosal fluid into the lymphatic network preventing fluid backflow, while unidirectional intraluminar valves in the transverse vessels convey lymph centripetally toward central collecting ducts. The complexity and anatomical arrangement of the two valves system suggests that, despite the existence of a favorable Delta P(TD), in the physiological condition no fluid bulk flow takes place between the pleural and peritoneal cavity through the diaphragmatic lymphatic network.     

Compartmentalized mast cell degranulations in the ovarian hilum, fat pad, bursa and blood vessel regions of the cyclic hamster: relationships to ovarian histamine and blood flow.

Ovaries from hamsters on each day of the oestrous cycle at 09.00 h were observed for the number of mast cells, the pattern of mast cell degranulation, histamine concentration and blood flow. On day 4 (pro-oestrus), ovaries were also observed at 9.00, 15.00 and 21.00 h. Mast cell degranulation was evaluated by 3 criteria: (1) no degranulation = less than 5 granules dispersed from the cell; (2) moderate degranulation = 5 or more granules dispersed but less than 15, and (3) extensive degranulation = 15 or more granules released. Blood flow was determined using radio-active microspheres in anaesthetized animals. Mast cells were observed in fat pad (beyond 2 mm of the bursal mesothelium), bursa (within 2 mm of the bursal mesothelium), hilum and near ovarian blood vessels (these 4 regions are collectively called the ovarian complex). The distribution of ovarian mast cells was not uniform. Most mast cells were near ovarian blood vessels (42.2%) and in the fat pad (37.2%). A moderate number of cells were in the bursal wall (20%) and only a few cells were observed in the hilum (0.64%). Mast cell number remained unchanged on days 1-4 of the cycle in each ovarian compartment. However, summation of the number of mast cells in the entire ovarian complex revealed a significant decline in number at 15.00 h on pro-oestrus. Alterations in mast cell degranulation were primarily restricted to 2 periods of the cycle (pro-oestrus and di-oestrus). An increase in moderate but not extensive degranulation was observed in only the fat pad and bursa on day 2 when compared with day 1 values. In most ovarian compartments on pro-oestrus, degranulation was higher than on any other day of the cycle. At 15.00 h on pro-oestrus, extensive degranulation in bursa, fat pad and blood vessel regions (but not hilum) coincided with an increase in ovarian histamine and decline in number of mast cells; ovarian blood flow also increased at the time but remained unchanged the remainder of the cycle.(ABSTRACT TRUNCATED AT 250 WORDS)     

Proliferating cell nuclear antigen immunoreactivity in the ovarian surface epithelium of mice of varying ages and total lifetime ovulation number following ovulation

Everytime an oocyte is released at ovulation, the ovarian surface epithelium (OSE) is ruptured and must be restored by epithelial cell proliferation. Ovulation site closure was studied in mice of various ages along with total lifetime ovulation number to investigate the known association of these factors with the risk of epithelial ovarian cancer. Ovaries from Swiss Webster mice were collected at various time points postovulation from 3-mo virgin animals (estimated median total lifetime ovulation number, 92; n = 40 mice), 8-mo virgin animals subject to incessant ovulation (estimated median total lifetime ovulation number, 652; n = 15 mice), and 12-mo breeders (estimated median total lifetime ovulation number, 208; n = 35 mice). Diameters of ovulation sites were estimated by scanning electron microscopy. No differences were found in the rate of ovulation site closure between the groups. Sections of ovaries were analyzed using immunohistochemistry for proliferating cell nuclear antigen (PCNA). The highest density of immunoreactive cells was observed in all animal groups in the cuboidal cells around the rupture site the day after ovulation. Despite the similarity in ovulation site closure rates between groups, the total number of OSE cells that were positive for PCNA in both the 8- and 12-mo animals was significantly reduced, so the number of stained cells appeared to be insufficient to cover the ovulation site. These data suggest that other mechanisms, such as proliferation of the extraovarian mesothelium, may play a role in the re-epithelialization of the ovary.     

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。     

Absorption from the peritoneal cavity: SEM study of the mesothelium covering the peritoneal surface of the muscular portion of the diaphragm.

Colored tracers, injected intraperitoneally in mice, are taken up by diaphragmatic lymphatics, outlining their large, terminal cisterns, the so-called lacunae. The lacunae occur exclusively on the muscular portion of the diaphragm. The mesothelium covering non-lacunar and lacunar areas of the muscular portion was examined with the SEM. Mesothelial cells overlying non-lacunar areas are extremely flat, and their boundaries are indistinct. Mesothelial cells overlying lacunae protrude towards the lumen of the peritoneal cavity and have distinct outlines. There are openings or stomata, 4-12 micron in diameter, between them. Some of the stomata overlie a deep pit; others overlie a shallower pit in which the surface of another cell can be seen beneath the opening. It seems likely that the bulk of the fluid draining from the peritoneal cavity passes through these stomata into underlying lymphatic lacunae.     

Curschmann's spirals in pleural and peritoneal effusions

Curschmann's spirals were found in smears and cell block preparations of five spontaneously occurring pleural and peritoneal fluids. The spirals were similar to those seen in the respiratory tract, although generally much smaller. In three of the five cases, the fluids also contained mucus-secreting adenocarcinoma cells; it is postulated that the spirals formed from mucus secreted by these cells. In the other two cases, there was evidence of serosal inflammation; it is suggested that the spirals in these cases developed from submesothelial connective tissue mucosubstances that entered the serosal cavity through a mesothelium of increased permeability due to the inflammation. No simple explanation can be accepted as to the exact mode of spiral formation, which is presumed to be a complex physical and biochemical phenomenon.     

Penetration of an ascitic reticulum cell sarcoma of the golden hampster into the body wall and through the diaphragm. A scanning electron microscopic (SEM) study.

SEM studies on infiltration of the ascitic form of the hamster reticulum cell sarcoma HaTu 25 into the ventral body wall and through the diaphragm were performed during 6 consecutive days after intraperitoneal transplantation. The findings allow an interpretation of the course of events based on 3 main stages: 1) Contraction of mesothelial cells with partial exposure of the submesothelial stratum. 2) Preferential attachment of tumor cells to these denuded areas. 3) Advance of tumor cells within defects gradually extening from the submesothelial stratum of the musculature. These stages were more pronounced and took a more rapid course at the peritoneal side of the diaphragm than at the body wall. At the pleural side of the diaphragm the appearance of single tumor cells within widened intercellular spaces of the mesothelium was recorded prior to the onset of penetration at the peritoneal surface. The rapid migration of tumor cells through the diaphragm as well as the particularly intensive tumor infiltration into this organ is thought to be connected with the mechanism of intravasation of tumor cells into the lymphatic plexus of the diaphragm. During the whole sequence of events, HaTu 25 cells were found to have maintained their spherical configuration and characteristic surface architecture. Apparently, growth pressure is of minor or no importance in this spacial mode of tumor penetration, rather the action of proteolytic enzymes elaborated by the tumor cells has to be taken into consideration.     

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.     

Reactive growth of the mesothelium of the parietal leaf of the peritoneum]

In the process of reparative regeneration of the mesothelium caused by suturing of a portion of the jugular vein into the peritoneum defect the inflammatory mesothelial growings are formed. They have different configuration, consist only of the mesotheliumor the connective tissue covered with the mesothelium and are observed from the 4th day till 3 months after operation, being always faced to the abdominal cavity. The capacity of mesothelial cells to high mitotic activity and wedging out from the layer underlies the formation of posttraumatic structures of the mesothelium.     

Microscopic and ultrastructural study of the lymphatic system in the human parotid gland

The localization and fine structure of the lymphatic system vessels are examined in the human parotid gland. A network of lymphatic capillaries extends in the intralobular connective septa around the striated ducts. These lymphatics converge in collectors frequently bordering the excretory ducts. On the contrary, no lymphatics are present next to the intercalated ducts and adhenomers. Ultrastructurally, the lymphatic capillaries are characterized by a very thin endothelial wall and by slightly complicated intercellular adhesions. Open junctions are also present. The presence of numerous lymphatic capillaries bordering the striated ducts and their blood microvasculature is discussed in relation to the functional activities of the striated ducts in the modification of the saliva.     

Embryonic and post-embryonic development of the parietal and visceral peritoneum in white mice]

Light and electron microscopy were used in order to investigate histogenesis of the parietal and visceral peritoneum of white mice in embryonic and postembryonic periods of development. Four periods were distinguished, during which gradual differentiation of the primordium material into tissue structures (mesothelium and the connective tissue) of the peritoneum were observed. Asynchronous differentiation of the mesothelium as well as certain correlation in the degree of differentiation of mesothelial and mesenchymal cells took place at all stages of the embryonic and postembryonic development. More differentiated cells of prolonged shape were predominant in the mesenchyma even at early stages (11 days) in those portions where the lining of the secondary cavity of the body resembled mesothelim in its structure.     

Brugia pahangi: infections and their effect on the lymphatic system of Mongolian jirds (Meriones unguiculatus)

Brugia pahangi has been found to be primarily a lymphatic-dwelling parasite in jirds when infections are induced by the subcutaneous injection of infective larvae or by allowing infected Aedes aegypti to feed.Migration to the regional lymphatics occurred as early as 1–4 days. Although some injected larvae remained in the skin for as long as 30 days and some became localized in the heart, lungs, pleural cavity, or peritoneal cavity, about three-fourths of the recovered filariae were found in the regional lymphatics. In contrast, when larvae were injected peritoneally they remained largely in the peritoneal cavity for at least 30 days.The relevant lymphatics and their drainage patterns in jirds have been described.The major pathological changes noted in jirds involved the regional lymphatic vessels and nodes, which were severely affected when they contained dead worms. Pulmonary granulomas due to dead microfilariae and occasionally to dead larvae or adult worms were noted.Observations are included on the susceptibility and course of B. pahangi infections in jirds.     

Inhibition of active lymph pump by simulated microgravity in rats

During spaceflight the normal head-to-foot hydrostatic pressure gradients are eliminated and body fluids shift toward the head, resulting in a diminished fluid volume in the legs and an increased fluid volume in the head, neck, and upper extremities. Lymphatic function is important in the maintenance of normal tissue fluid volume, but it is not clear how microgravity influences lymphatic pumping. We performed a detailed evaluation of the influence of simulated microgravity on lymphatic diameter, wall thickness, elastance, tone, and other measures of phasic contractility in isolated lymphatics. Head-down tail suspension (HDT) rats were used to simulate the effects of microgravity. Animals were exposed to HDT for 2 wk, after which data were collected and compared with the control non-HDT group. Lymphatics from four regional lymphatic beds (thoracic duct, cervical, mesenteric, and femoral lymphatics) were isolated, cannulated, and pressurized. Input and output pressures were adjusted to apply a range of transmural pressures and flows to the lymphatics. Simulated microgravity caused a potent inhibition of pressure/stretch-stimulated pumping in all four groups of lymphatics. The greatest inhibition was found in cervical lymphatics. These findings presumably are correlated to the cephalic fluid shifts that occur in HDT rats as well as those observed during spaceflight. Flow-dependent pump inhibition was increased after HDT, especially in the thoracic duct. Mesenteric lymphatics were less strongly influenced by HDT, which may support the idea that lymph hydrodynamic conditions in the mesenteric lymphatic during HDT are not dramatically altered.     

Ontogeny of oestrogen-sensitive mesenchymal cells in the bursa of Fabricius of the chick embryo. An immunohistochemical study on progesterone receptor

The expression of progesterone receptor (PR) and its induction by oestradiol during the embryogenesis of the chick bursa of Fabricius (BF) were studied by immunohistochemistry using three different polyclonal antibodies to the chicken oviduct PR. Mesenchymal cells of the cloacal area surrounding the bursa primordium in controls (without exogenous oestrogen) express the PR between 9 and 11 days of incubation. In the same cells, PR was induced experimentally by oestradiol at 9 days. Mesenchymal cells in the bursa did not express PR after oestradiol treatment before the age of 11 days. The PR was not inducible in the bursal epithelium or in haemopoietic cells. None of the bursal cells expressed the PR to a detectable level during embryonic life without exogenous treatment. Some haemopoietic cells showed strong artefactual staining in their nuclei. It is concluded that (1) the embryonic bursa of Fabricius is a sex-steroid-sensitive organ, (2) exogenous oestradiol is able to induce progesterone receptor in the mesenchymal cells, but (3) the PR is not expressed without exogenous oestrogen. This indicates that the PR becomes oestrogen inducible well before it is naturally expressed during sexual maturation and that the level of endogenous oestrogen during embryonic life is not high enough to affect the bursa significantly.     

Effect of environmental antigens on the Ig diversification and the selection of productive V-J joints in the bursa

In chickens, a single set of unique functional segments of both Ig H and L chain genes is rearranged during early embryogenesis to generate a pool of B cell progenitors that will be diversified in the bursa by gene conversion, forming the preimmune repertoire. After hatching, bursal cells are exposed to environmental Ags in the bursal lumen. We prepared B cells from each single bursal follicle and used PCR-directed Ig L chain gene analysis to study the differentiation of B cells and the effect of antigenic stimulation from the bursal lumen on the neonatal chicken B cell repertoire formation. Selective amplification of B cell clones with a productive V-J joint was observed during the late embryonic stage, possibly by the interaction with ligands expressed on the bursal stroma and further accelerated in the neonatal chicken. Administration of the artificial Ags into the bursal lumen before the isolation of bursa by bursal duct ligation in the embryo caused a significant increase in lymphocytes with a productive V-J joint in the neonatal chicken bursa compared with the isolated bursa. Intra- and interclonal diversity of a complementarity-determining region measured by an evolutionary distance increased during bursal development. Clonal diversification did not require stimulation by artificial Ags from the bursal lumen. Thus, the preimmune repertoire in the bursa is generated by gene conversion during Ag-independent B cell proliferation, and antigenic stimulation from the bursal epithelium to bursal B cells plays roles in the selection of clones with a productive V-J joint.