Lymphatic Stomata Research Articles

On the Mouse Lymphatic System and the Lymphatic Stomata

On the Mouse Lymphatic System and the Lymphatic Stomata

Lymphatic Endothelial Cell in Endemic Bancroftian Filariasis: A Focus on the Lymphatics of the Tunica Vaginalis Testis.

Background In endemic areas, lymphangiectasia is the fundamental alteration to live Wuchereria bancrofti adult worms which, in adult males, are usually found in the lymphatic vessels of the spermatic cord; accordingly, hydrocele/filaricele is the most common clinical manifestation of bancroftian filariasis. The pathogenic role of the lymphatic endothelial cells (LECs) and the status of mesothelial cells (MCs) samples of the parietal layer (PL) of the tunica vaginalis testis were examined. Methods The PL of thirty-two patients, excised for different reasons, was examined by histology and immunohistochemistry using the D2-40 monoclonal antibody for identification of LECs and CK-7 antibody for recognition of mesothelial cells (MCs). Results The most important findings were (a) marked lymphangiectasia, especially in hydroceles with minor evolution time; (b) the first report of lymphatic stomata and submesothelial lacunae in filarial acute hydrocele; (c) the likely participation of LECs in filarial granuloma; (d) the potential phenotypic transition of LECs into myofibroblasts in severe chylocele; and (e) mesothelial reactive hyperplasia, a hallmark of filaricele, varying in intensity from mild to severe, sometimes mimicking a mesothelial neoplasia. Conclusion The data suggest that LECs have an active role in the pathogenesis of bancroftian hydrocele and, possibly, in other clinical forms of lymphatic filariasis.

Effect of the intra-abdominal infection on peritoneal lymphatic stomata and regulatory mechanism of PD123319 on peritoneal lymphatic stomata

Objective To investigate the effect of the peritoneal lymphatic stomata on intra-abdominal infection and the regulatory mechanism of angiotensin Ⅱ receptor specific inhibitor PD123319 on peritoneal lymphatic stomata. Methods The experimental study was adopted. Forty rats were divided into the control group, sham operation group, intra-abdominal infection group and intra-abdominal infection drug intervention group by the random number table, every group had 10 rats. The classic appendix perforation (CLP) intra-abdominal infection model was established in the abdominal infection group. After establishing the model of abdominal infection, PD123319 solution was injected intraperitoneally immediately (0.2 g/kg) in the abdominal infection drug intervention group. Abdominal cavity of the rats in the sham operation group was opened, and then was shut after flipping the intestine. The rats in the control group, sham operation group and intra-abdominal infection group were treated with intraperitoneal injection of 1ml stroke-physiological saline solution. After 2 hours, the rats were sacrificed, and peritoneal tissue was taken for the following tests. (1) The aperture size and distribution density of peritoneal lymphatic stomata were observed by scanning electron microscope (SEM). (2) The nitric oxide (NO) concentration in the peritoneal tissues was detected using nitric oxide nitric acid reduction method. (3) The expressions of endothelial nitric oxide synthase(eNOS)and Phospho-eNOS (P-eNOS) were detected by the Western blot. (4) The intracellular Ca2+ concentration were detect by flow cytometry. Measurement data with normal distribution were presented as ±s. The comparison among groups was analyzed using the ANOVA and pairwise comparison was analyzed by the LSD test. Results (1) The aperture size and distribution density of the peritoneal lymphatic stomata in the control group, sham operation group, intra-abdominal infection group and intra-abdominal infection drug intervention group were respectively (2.3±0.4)μm, (2.5±0.5)μm, (4.7±0.5)μm, (3.8±0.5)μm and (2.0±0.8)×108/m2, (2.1±0.7)×108/m2, (6.2±1.3)×108/m2, (4.6±1.4)×108/m2, with statistically significant differences among the 4 groups (F=98.130, 56.780, P 0.05). (2) The concentrations of NO in the peritoneal tissues in the control group, sham operation group, intra-abdominal infection group and intra-abdominal infection drug intervention group were respectively (0.380±0.024)μmol/gprot, (0.450±0.020)μmol/gprot, (1.253±0.033)μmol/gprot and (0.579±0.035)μmol/gprot, with a statistically significant difference among the 4 groups (F=52.725, P 0.05). (3) The results of Western blot showed that the expressions of eNOS and P-eNOS in the control group, sham operation group, intra-abdominal infection group and intra-abdominal infection drug intervention group were respectively (0.591±0.028)U/mg, (0.603±0.007)U/mg, (0.615±0.027)U/mg, (0.626±0.026)U/mg and (0.578±0.003)U/mg, (0.603±0.071)U/mg, (0.773±0.033)U/mg, (0.710±0.012)U/mg, with no statistically significant difference in the expression of eNOS among the 4 groups (F=0.902, P>0.05) and with a statistically significant difference in the expression of P-eNOS among the 4 groups (F=205.062, P 0.05). Conclusion The intra-abdominal infection could increase aperture size and distribution density of peritoneal lymphatic stomata, and PD123319 may be through inhibiting the activation of NO synthase to decrease the concentration of NO, enhance the concentration of Ca2+ in peritoneal mesothelial cells and reduce the opening of peritoneal lymphatic stomata. Key words: Abdominal cavity infection; Peritoneal lymphatic stomata; Nitric oxide; Angiotensin Ⅱ receptor specific inhibitor

The discovery of the synovial lymphatic stomata and lymphatic reabsorption in knee effusion.

To illustrate the mechanism of lymphatic reabsorption in knee joint effusion. The current investigation employed transmission electron microscopy (TEM) and scanning electron microscopy (SEM) techniques to reveal the ultrastructure of the knee synovial membrane in New Zealand rabbits and human. Ultrastructural changes of the synovial lymphatic stomata were observed by using trypan blue absorption and sodium hydroxide (NaOH) digestion methods, and the animal models of synovitis. New Zealand rabbits and human synovial membranes were composed of two types of synovial cells: type A and type B. No lymphatic stomata were found among type A synovial cells, whereas lymphatic stomata with the diameters ranging 0.74-3.26 µm were found in type B synovial cells, and some stomata were closed. After the NaOH digestion, a number of sieve pores, similar to lymphatic stomata in size and shape, were observed in the dense fibrous connective tissue underneath the type B synovial cells. After injecting trypan blue into the rabbit knee joint cavity, absorption of trypan blue through the lymphatic stomata was observed, suggesting the absorption function of the synovial lymphatic stomata. In the rabbit knee joint synovitis models, the synovial lymphatic stomata diameter enlarged. Some macrophages migrated from the lymphatic stomata, indicating that the synovial lymphatic stomata were involved in the joint effusion absorption and inflammatory response. Our study is the first to report the existence of synovial lymphatic stomata in the New Zealand rabbits and human knee joints. Lymphatic stomata may have an important role in the reabsorption of joint effusion.

Physiology of the pleural space.

The pleural cavity is created between the 4(th) and 7(th) week of embryologic development. These embryonic components of visceral and parietal pleurae develop different anatomic characteristics with regard to vascular, lymphatic, and nervous supply. There are two layers: a superficial mesothelial cell layer facing the pleural space and an underlying connective tissue layer. The pleura might present inflammatory response and maintenance of the pleural fluid is observed. The latter function is especially important in the mechanical coupling of the lung and chest wall. Fluid is filtered into the pleural space according to the net hydrostatic oncotic pressure gradient. It flows downward along a vertical pressure gradient, presumably determined by hydrostatic pressure and resistance to viscous flow. There also may be a net movement of fluid from the costal pleura to the mediastinal and interlobar regions. In these areas, pleural fluid is resorbed primarily through lymphatic stomata on the parietal pleural surface. In the current review we will present the physiology of the pleural space in a step by step manner.

Effect of regulation of peritoneal lymphatic stomata on endotoxin induced lung injury in rats with early abdominal infection

Effect of regulation of peritoneal lymphatic stomata on endotoxin induced lung injury in rats with early abdominal infection

Lymphatic Stomata in the Adult Human Pulmonary Ligament.

Lymphatic stomata are small lymphatic openings in the serosal membrane that communicate with the serosal cavity. Although these stomata have primarily been studied in experimental mammals, little is known concerning the presence and properties of lymphatic stomata in the adult human pleura. Thus, adult human pleurae were examined for the presence or absence of lymphatic stomata. A total of 26 pulmonary ligaments (13 left and 13 right) were obtained from 15 adult human autopsy cases and examined using electron and light microscopy. The microscopic studies revealed the presence of apertures fringed with D2-40-positive, CD31-positive, and cytokeratin-negative endothelial cells directly communicating with submesothelial lymphatics in all of the pulmonary ligaments. The apertures' sizes and densities varied from case to case according to the serial tissue section. The medians of these aperture sizes ranged from 2.25 to 8.75 μm in the left pulmonary ligaments and from 2.50 to 12.50 μm in the right pulmonary ligaments. The densities of the apertures ranged from 2 to 9 per mm(2) in the left pulmonary ligaments and from 2 to 18 per mm(2) in the right pulmonary ligaments. However, no significant differences were found regarding the aperture size (p=0.359) and density (p=0.438) between the left and the right pulmonary ligaments. Our study revealed that apertures exhibit structural adequacy as lymphatic stomata on the surface of the pulmonary ligament, thereby providing evidence that lymphatic stomata are present in the adult human pleura.

Ultrastructure of Lymphatic Stomata in the Tunica Vaginalis of Humans

Lymphatic stomata are small openings of lymphatic capillaries on the surface of the mesothelium that lines the serous cavity and have the function of active absorption. They play an important role in physiological and pathological conditions. The cavity of the tunica vaginalis is a typical serous cavity of the testis, but the lymphatic stomata of the tunica vaginalis of humans have never been reported. Here, we studied their ultrastructure by scanning and transmission electron microscopy. The submesothelial connective tissue with foramina was investigated after the mesothelial cells were digested using NaOH solution. We found the lymphatic stomata in cuboidal mesothelial cell regions of the parietal layer of the tunica vaginalis of humans with a diameter of about 1-2 μm. Sometimes, closed lymphatic stomata could be observed. Our study is the first to report the existence of lymphatic stomata of the tunica vaginalis of humans. We found that the tunica vaginalis cavity is connected with the lymphatic system through the stomata, which might provide a morphological basis for the drainage of hydrocele and tumor metastasis of the tunica vaginalis of humans.

The discovery of lymphatic stomata and its ultrastructure in mouse tunica vaginalis

Background/PurposeThe communicating hydrocele is caused by the failure of the processus vaginalis closure. After the patent processus vaginalis is closed, the hydrocele can subside spontaneously, but the mechanism responsible for the drainage of hydrocele has not been expounded. Former studies showed that lymphatic stomata between the mesothelial cells lining the peritoneum and pleura were responsible for the drainage of ascites and pleural effusion. Although the tunica vaginalis is also lined by mesothelial cells, the existence of lymphatic stomata in it has never been reported. Therefore, we investigated the presence and ultrastructure of lymphatic stomata in mouse tunica vaginalis. MethodsWe studied the ultrastructure of mouse tunica vaginalis by scanning and transmission electron microscopy. The submesothelial connective tissue with foramina were investigated after the mesothelial cells were digested by NaOH solution. Typan blue was used as a tracer to show the absorptive function of lymphatic stomata. ResultsThe lymphatic stomata were found between the mesothelial cells. The trypan blue was directly absorbed by the lymphatic stomata. Milky spots which possessed immune function were observed, and the lymphatic stomata were found within the milky spots. ConclusionOur study is the first to report the presence and ultrastructure of lymphatic stomata in mouse tunica vaginalis. They might be related to the lymphatic drainage in the tunica vaginalis cavity.

Recent Advances in the Research of Lymphatic Stomata

Lymphatic stomata are small openings of lymphatic capillaries on the free surface of the mesothelium. The peritoneal cavity, pleural cavity, and pericardial cavity are connected with lymphatic system via these small openings, which have the function of active absorption. The ultrastructure of the lymphatic stomata and their absorption from the body cavities are important clinically, such as ascites elimination, neoplasm metastasis, and inflammatory reaction. The lymphatic stomata play an important role in the physiological and pathological conditions. Our previous study indicated for the first time that nitric oxide (NO) could regulate the opening and absorption of the lymphatic stomata. It could decrease the level of free intracellular calcium [Ca(2+)] through increasing the cyclic guanosine monophosphate (cGMP) level in the rat peritoneal mesothelial cells, thus regulating the lymphatic stomata. This process is related with the NO-cGMP-[Ca(2+)] signal pathway. In this review, we summarize the recent advances in understanding the development and the function of the lymphatic stomata. The ultrastructure and regulations of the lymphatic stomata are also discussed in this review.

Cell Signal Transduction Mechanism for Nitric Oxide Regulating Lymphatic Stomata and its Draining Capability

We have previously demonstrated that nitric oxide (NO) is involved in the regulation of the lymphatic stomata. However, the related mechanisms are still unknown. The present study was designed to test the hypothesis that NO-cyclic guanosine monophosphate (cGMP) -mediated cytosolic Ca(2+) concentration ([Ca(2+)]i) signaling may contribute to the regulation of the lymphatic stomata and lymph drainage. Using trypan blue as a tracer, the effects of NO-cGMP-Ca(2+) signal cascade on the lymphatic stomata and lymph absorption were examined by means of scanning electron microscopy. Then, the role of NO in cGMP and [Ca(2+)]i of rat peritoneal mesothelial cells (RPMCs) was measured by radioimmunoassay and a confocal laser scanning microscope. Our results showed that NO-donor spermine/nitric oxide complex (Sper/NO) could broaden the opening area of the lymphatic stomata and enhance lymph absorption in a dose-dependent manner. These NO-mediated changes could be blocked by 1H-[1,2,4] oxadiazolo [4,3-a] quinoxalin-1-one (ODQ), a specific inhibitor of soluble guanylyl cyclase, and mimicked by calcium channel blocker nifedipine. Furthermore, Sper/NO enhanced the cGMP level and lessened [Ca(2+)](i) in RPMCs, which was completely abrogated at the presence of ODQ. Nifedipine induced an immediate and marked decrease of [Ca(2+)](i) in the RPMCs, which was not attenuated by addition of Sper/NO, indicating that the Sper/NO-cGMP signaling system induced [Ca(2+)](i) change was related to the L-type voltage-gated calcium channel in the RPMCs. Our results suggest that NO enlarges the opening area of the lymphatic stomata to strengthen the lymph drainage of tracer by means of NO-cGMP-[Ca(2+)]i signal transduction pathway in the RPMCs.

Ultrastructure and Estrogen Regulation of the Lymphatic Stomata of Ovarian Bursa in Mice

The ovarian bursa is a key player in maintaining adaptive ovarian microenvironment for ovulation. The lymphatic stomata are believed to be a major contributor to execute the function of the ovarian bursa, whereas little is known about their ultrastructure and regulation. Here, we examined the ultrastructure of lymphatic stomata in mouse ovarian bursa by scanning electron microscopy and transmission electron microscopy and investigated its regulation by estrogen. We found that the mesothelium on the visceral layer of mouse ovarian bursa was composed of the cuboidal and flattened cells. The lymphatic stomata with round and oval shapes were mainly among the cuboidal cells. The particles, cells, and fluid passed through the stomata and entered into the lymphatic drainage unit composed of connective tissue and lymphatic endothelial cells beneath the stomata. We also used trypan blue as a tracer and found that the absorption of trypan blue through the lymphatic stomata was increased by estrogen that enlarged the average opening area of lymphatic stomata. Furthermore, we detected that there existed estrogen receptors in the nuclei of the mesothelial cells on the visceral ovarian bursa by using immunoelectron microscopy. Taken together, these data suggest that both the absorption and opening area of the lymphatic stomata in mouse ovarian bursa may be influenced by estrogen.

Role of human visceral pleura in pleural fluid turnover: Need for morphological evidence of lymphatic stomata

To the Editor: We read with great interest the article by Gao and his colleagues1 proposing a new schema and hypothesis regarding pleural fluid turnover. In the proposed hypothesis, the visceral pleura possesses a more active role in humans than what was believed. The authors support the notion that in humans the pulmonary lymphatic vessels originate directly from the mesothelium of the visceral pleura and that it has an effect on both infiltration and reabsorption of pleural fluid in human beings. However, there are some points that are worth commenting. Should the lymphatic vessels originate from the visceral mesothelium, they would form stoma-like projections on the pleura cavity, as implied by the authors. According to the study of Leak and Rahil2 and the study of Nakatani et al,3 the morphology of a typical orifice of a lymphatic stoma under the scanning electron microscopy (SEM) consists of two margins: an outer microvillous margin formed by a perimeter of cuboidal mesothelial cells and an inner, relatively microvillous-free margin, formed by lymphatic endothelial cells. A study on the existence of lymphatic stomata in humans failed to demonstrate any stomas in specimens of 30 patients.4 Peng et al5 found stomas only in 2 of 9 human specimens. Additionally, Li6 demonstrated stomas only in human diaphragmatic parietal pleura but none in both the thoracic parietal pleura and in the visceral pleura in this study. Another study on the sheep pleura, which is thick and morphologically resembles the human pleura,7 revealed that the pleura mesothelium was composed of two types of mesothelial cells: flattened cells with well developed tight junctions and cuboidal cells with lesser tight junctions. The visceral pleura consists mainly of flat cells, indicating a tighter mesothelium. The parietal pleura consists of both flattened and cuboidal mesothelial cells as mentioned above around the orifice of a stoma. The lower permeability of the visceral sheep pleura compared to the parietal was also verified by our research group by means of electrophysiology.8 The absence of many cuboidal cells in the visceral pleura and the finding of a tighter mesothelium other than the parietal pleura, do not favor the existence of visceral lymphatic stomata. It will be of great interest if the authors could provide any figures of SEM. Should these figures depict stomas, the theory proposed by Gao and colleagues will be strengthened. Nevertheless, further experimental evaluation of this interesting new theory is imperative. If it is proven to be valid, research into the pleural fluid turnover will be redirected.

Apparent successful mesothelial cell transplantation hampered by peritoneal activation

Mesothelial cell transplantation has been suggested to improve mesothelial repair after surgery, recurrent peritonitis and peritoneal dialysis. In this study we evaluated mesothelial cell transplantation during the resolution phase of experimentally thioglycollate-induced peritonitis in rats. To this end 4 x 10(6) DiO-labeled autologous mesothelial cells were transplanted 1 week after peritonitis induction. Peritoneal inflammation and permeability characteristics were evaluated after another week. Mesothelial cell transplantation after peritonitis resulted in incorporation of these cells in the parietal mesothelial lining, leading to an acute transient submesothelial thickening which was not seen in transplanted animals without prior peritonitis induction. Long-term functioning of these repopulated mesothelial cells leaded to peritoneal activation as evidenced by a approximately twofold increase in peritoneal lymphocytes (P < 0.01) and omental mast cell counts (P < 0.05), accompanied by the induction of inflammation markers monocyte chemoattractant protein-1 (MCP-1) (P < 0.01) and hyaluronan (P < 0.01) in the transplanted peritonitis group, but not in rats with peritonitis without mesothelial cell transplantation or in control rats without mesothelial cell transplantation (all four parameters P < 0.01). In addition, trapping of transplanted mesothelial cells in the milky spots of omental tissue and lymphatic stomata of the diaphragm both in control and thioglycollate rats seems to increase microvascular permeability, reflected by apparent increased diffusion rates of small solutes and proteins. Altogether, our data underscore the importance of controlling peritoneal (patho)physiology and function in mesothelial transplantation protocols.

Morphological analysis of transportation pathways of microspheres after their introduction into the uterine horn cavity in cyclic pigs.

Countercurrent transfer is thought to be one of the most important mechanisms involved in the transfer of substances between the uterus and oviduct. The present study was aimed at recognizing other putative transportation pathways from the uterine cavity through the oviduct onto the surface of and into internal ovarian structures. Microspheres (latex beads, 0.8 microm in diameter) were introduced into the uterine horn cavity of pigs, for 30 min, at various days of the estrous cycle. The transportation pathways of the beads were then analyzed by light and electron microscopy. The transport of microspheres through the oviduct canal into ovarian tissues took place on each day of the estrous cycle. The largest numbers of microspheres passed through the tunica albuginea to the corpora lutea. Some of microspheres also reached the surface of the uterine ligament through the oviduct canal, where they attained the lumen of blood and lymphatic vessels, mainly of the vascular subovarian (VSP) and paraovarian lymphatic plexus (PLP), via the lymphatic stomata pathway. Transport of microspheres also took place simultaneously through the uterine and oviduct walls and from particular organs through blood and lymphatic vessels. Although the present results do not exclude the participation of countercurrent transfer between venous, lymphatic, and arterial vessels, they provide morphological evidence for the presence of direct transportation pathways of substances, released into the uterine lumen, into ovarian tissues through the oviduct canal.

Pleural mechanics and fluid exchange.

The pleural space separating the lung and chest wall of mammals contains a small amount of liquid that lubricates the pleural surfaces during breathing. Recent studies have pointed to a conceptual understanding of the pleural space that is different from the one advocated some 30 years ago in this journal. The fundamental concept is that pleural surface pressure, the result of the opposing recoils of the lung and chest wall, is the major determinant of the pressure in the pleural liquid. Pleural liquid is not in hydrostatic equilibrium because the vertical gradient in pleural liquid pressure, determined by the vertical gradient in pleural surface pressure, does not equal the hydrostatic gradient. As a result, a viscous flow of pleural liquid occurs in the pleural space. Ventilatory and cardiogenic motions serve to redistribute pleural liquid and minimize contact between the pleural surfaces. Pleural liquid is a microvascular filtrate from parietal pleural capillaries in the chest wall. Homeostasis in pleural liquid volume is achieved by an adjustment of the pleural liquid thickness to the filtration rate that is matched by an outflow via lymphatic stomata.

Ultrastructural study of pleural lymphatic drainage unit and effect of nitric oxide on the drainage capacity of pleural lymphatic stomata in the rat

The objective of this study was twofold: first to investigate the ultrastructure of the lymphatic drainage unit on the costal pleura of rats by electron microscopy, and secondly to examine the effect of nitric oxide on the pleural lymphatic stomata and fluid absorption from the pleural cavity. The lymphatic drainage unit of the rat costal pleura is composed of three special components: the lymphatic stomata between the mesothelial cells, the initial part of the lymphatic vessels and the underlying connective tissue containing many foramina. The unit is the main passage to drainage fluid, particles and cells in the pleural space. To investigate the regulator of the lymph drainage, nitric oxide synthase inhibitor and nitric oxide donor were injected into the peritoneal cavity of the rats, respectively. Trypan blue was used as tracer. The ultrastructural changes of pleural lymphatic stomata were observed under scanning electron microscope and analyzed by a computer image processing system. It turned out that the area and density of the pleural lymphatic stomata were positively correlated with the nitric oxide quantity (p < 0.05). After the tracer was injected into the pleural cavity, the nitric oxide donor group exhibited a higher trypan blue concentration than the control group (p < 0.05). The ability of the pleura to absorb trypan blue was enhanced because of the larger opening of the lymphatic stomata (p < 0.05). It is suggested that nitric oxide can increase lymphatic absorption of the pleura by opening pleural lymphatic stomata.

Development of the peritoneal lymphatic stomata and lymphatic vessels of the diaphragm in mice

The generation and development of the peritoneal lymphatic stomata (PLS) and lymphatic vessels of the diaphragm were studied in mice at gestational ages from the embryonic to the postnatal period with TEM, SEM and enzyme histochemistry and the PLS data were quantitatively analyzed with computer-assisted image processing technology (Elescope image analysis software). The results showed that the diaphragmatic mesothelium was covered only by flattened mesothelial cells (FMC) at the 13th embryonic day (ED 13). At ED 15, some cuboidal mesothelial cells (CMC) and immature lymphatic stomata (NLS) were found scattered on the diaphragmatic mesothelium. The sub-peritoneal lymphatic capillaries did not appear until ED 18. However, no absorptive function was observed in NLS when trypan blue granules were injected into the peritoneal cavity. At postnatal day 1 (PND 1), the endothelial cytoplasm processes of the diaphragm lymphatic capillaries span the connective tissue fibers and the basal membrane of CMC to form the subperitoneal channels. These channels were connected with NLS and serve as the absorptive route between the peritoneal cavity and the sub-peritoneal lymphatic vessels. The trypan blue absorption test demonstrated that postnatal PLS possessed an absorptive function and had transformed to mature lymphatic stomata (MLS) by PND 1. Thus, NLS were renamed of MLS. At PND 5, the cuboidal mesothelial cell ridge (CMCR) appeared with increased CMC areas. At PND 10, CMCR were fused to form the band-like CMC area with much more MLS distributed in the muscular portion of the diaphragm. With distribution area and density of PLS increasing and growth of lymphatic vessels, an increased absorptive function from the peritoneal cavity was observed in the experiment.

Ultrastructure and three-dimensional study of the lymphatic stomata in the costal pleura of the rabbit.

The aim of this report was to investigate the ultrastructure and three-dimensional organization of the pleural lymphatic stomata in the adult rabbit costal pleura by electron microscopy. A computer image processing system attached to the scanning electron microscopy was used to get statistical evaluation of the dimensions of pleural lymphatic stomata. Mesothelial cells were digested by 2 mol/L NaOH solution in order to expose the submesothelial connective tissue with macula cribriformis. Two kinds of mesothelial cells were observed on the costal pleura: the flattened cells and the cuboidal ones. Both had microvilli on their surface. Pleural lymphatic stomata were located only in the regions of cuboidal mesothelial cells. The average area of a stoma was 7.20 +/- 3.69 microm2 and their average density 121 +/- 72/mm2. Pleural cavity is connected with the lympo-vascular system by lymphatic stomata and the interstitial layer with a dense network of lamina cribriformis. The macula cribriformis (7-60 microm in diameter) were found in subpleural connective tissue below the cuboidal mesothelial cells. Consequently on cross-section, the pathway represents a channel consisting of the stoma, the connective tissue space, and the gap between endothelial cells of the lymphatics. Closed lymphatic stomata and the milky spots composed of macrophages could be observed on the costal pleura. Our results suggest that the pleural cavity is connected with the lymphatic capillaries through the lymphatic stomata and the subpleural channel. This is the only "highway" from the pleural cavity to the vessels. This pathway may be importantly involved in the material exchange and the immunity of the pleura cavity.

The distribution and morphology of lymphatic vessels on the peritoneal surface of the adult human diaphragm, as revealed by an ink-absorption method.

Application of india ink to the peritoneal and pleural surfaces of the adult human diaphragm allowed visualization of the distribution and morphology of the lymphatic vessels by light microscopy and scanning electron microscopy. The diaphragms examined had been fixed and stored in 10% formalin. Numerous lymphatic vessels were stained black with india ink, presenting reticular, radial-meshwork, ladder-like and lacy patterns. They were distributed throughout the entire sternocostal part. Analysis by light and scanning electron microscopy of the areas indicated by india ink revealed the presence of primary lymphatic vessels that formed lymphatic lacunae and stomatal openings to the peritoneal cavity. A layer of secondary collecting lymphatic vessels was located cranially with respect to the layer of primary lymphatic vessels. Thus, the peritoneum had at least two layers of lymphatic vessels. These lymphatic vessels were not tubular vessels but resembled flat cisternae, as has been suggested in the case of the mouse diaphragm. The pleura lacked lymphatic stomata and had no such double-layered lymphatic organization. This is the first report that showed distribution and morphology of the lymphatic vessels in the diaphragmatic peritoneum of the formalin-fixed, adult human diaphragm. The method and results in the present study may contribute to morphological analysis of the lymphatic system in the wall of the human body cavity.