Plasticity of button-like junctions in the endothelium of airway lymphatics in development and inflammation.

Endothelial cells of initial lymphatics have discontinuous button-like junctions (buttons), unlike continuous zipper-like junctions (zippers) of collecting lymphatics and blood vessels. Buttons are thought to act as primary valves for fluid and cell entry into lymphatics. To learn when and how buttons form during development and whether they change in disease, we examined the appearance of buttons in mouse embryos and their plasticity in sustained inflammation. We found that endothelial cells of lymph sacs at embryonic day (E)12.5 and tracheal lymphatics at E16.5 were joined by zippers, not buttons. However, zippers in initial lymphatics decreased rapidly just before birth, as buttons appeared. The proportion of buttons increased from only 6% at E17.5 and 12% at E18.5 to 35% at birth, 50% at postnatal day (P)7, 90% at P28, and 100% at P70. In inflammation, zippers replaced buttons in airway lymphatics at 14 and 28 days after Mycoplasma pulmonis infection of the respiratory tract. The change in lymphatic junctions was reversed by dexamethasone but not by inhibition of vascular endothelial growth factor receptor-3 signaling by antibody mF4-31C1. Dexamethasone also promoted button formation during early postnatal development through a direct effect involving glucocorticoid receptor phosphorylation in lymphatic endothelial cells. These findings demonstrate the plasticity of intercellular junctions in lymphatics during development and inflammation and show that button formation can be promoted by glucocorticoid receptor signaling in lymphatic endothelial cells.

Absence of lymphatic vessels in the dog dental pulp: an immunohistochemical study.

In spite of numerous investigations it has not been precisely determined whether lymphatic vessels are present in the dental pulp of dogs. Therefore, this study attempted a specific immunohistochemical detection of lymphatic endothelium. The canine teeth of 19 healthy beagle dogs were dissected into three segments (apical, intermediate and occlusal). After decalcification, specimens were embedded in paraffin wax and histologic cross-sections were stained immunohistochemically using a reliable antibody (anti-Prox-1) against the homeobox transcription factor Prox-1, which is located within the nucleus of lymphatic endothelium. Anti-Prox-1 reacted positively with canine control tissues (lymph nodes, gingiva, nasal mucosa), but showed no staining in tissue sections of the dental pulp. The dog dental pulp contained no vascular structures lined with lymphatic endothelium. This suggests that drainage of interstitial fluid makes use of other routes, i.e. extravascular pathways.

Absence of lymphatic vessels in human dental pulp: a morphological study.

Few and controversial data are available in the literature regarding the presence of lymphatic vessels in the human dental pulp. The present study was designed to examine morphologically the existence of a lymph drainage system in human dental pulp. Human dental pulp and skin sections were immunohistochemically stained with specific antibodies for lymphatic endothelium (D2-40, LYVE-1, VEGFR-3 [vascular endothelial growth factor receptor-3], and Prox-1), with the pan-endothelial markers CD31 and von Willebrand factor (vWF), and with the blood-specific marker CD34. Several blood vessels were identified in human pulps and skin. Lymphatic vessels were found in all human skin samples but in none of the pulps examined. Western blotting performed on human dermis and on pulps treated with collagenase (to remove odontoblasts) confirmed these results. Transmission electron microscopy indicated that vessels which, by light microscopy, appeared to be initial lymphatic vessels had no anchoring filaments or discontinuous basement membrane, both of which are typical ultrastructural characteristics of lymphatic vessels. These results suggest that under normal conditions human dental pulp does not contain true lymphatic vessels. The various theories about dental pulp interstitial fluid circulation should be revised accordingly.

Identification of lymphatics in the ciliary body of the human eye: a novel "uveolymphatic" outflow pathway.

Impaired aqueous humor flow from the eye may lead to elevated intraocular pressure and glaucoma. Drainage of aqueous fluid from the eye occurs through established routes that include conventional outflow via the trabecular meshwork, and an unconventional or uveoscleral outflow pathway involving the ciliary body. Based on the assumption that the eye lacks a lymphatic circulation, the possible role of lymphatics in the less well defined uveoscleral pathway has been largely ignored. Advances in lymphatic research have identified specific lymphatic markers such as podoplanin, a transmembrane mucin-type glycoprotein, and lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1). Lymphatic channels were identified in the human ciliary body using immunofluorescence with D2-40 antibody for podoplanin, and LYVE-1 antibody. In keeping with the criteria for lymphatic vessels in conjunctiva used as positive control, D2-40 and LYVE-1-positive lymphatic channels in the ciliary body had a distinct lumen, were negative for blood vessel endothelial cell marker CD34, and were surrounded by either discontinuous or no collagen IV-positive basement membrane. Cryo-immunogold electron microscopy confirmed the presence D2-40-immunoreactivity in lymphatic endothelium in the human ciliary body. Fluorescent nanospheres injected into the anterior chamber of the sheep eye were detected in LYVE-1-positive channels of the ciliary body 15, 30, and 45 min following injection. Four hours following intracameral injection, Iodine-125 radio-labeled human serum albumin injected into the sheep eye (n = 5) was drained preferentially into cervical, retropharyngeal, submandibular and preauricular lymph nodes in the head and neck region compared to reference popliteal lymph nodes (P < 0.05). These findings collectively indicate the presence of distinct lymphatic channels in the human ciliary body, and that fluid and solutes flow at least partially through this system. The discovery of a uveolymphatic pathway in the eye is novel and highly relevant to studies of glaucoma and other eye diseases.

Orbital lymphatics: do they exist?

INTRODUCTION: Although the lymphatic system was first described almost 400 years ago, it is only in very recent years that researchers have been able to identify lymphatic channels with reasonable accuracy. Through advances in molecular biology and the development of endothelial cell markers the long held view that the human orbit is devoid of lymphatics has now been challenged. DISCUSSION: This review discusses the current evidence on this topic, which confirms the presence of orbital lymphatics in lachrymal gland and optic nerve sheath.

Role of CXC chemokine ligand 13, CC chemokine ligand (CCL) 19, and CCL21 in the organization and function of nasal-associated lymphoid tissue.

Nasal-associated lymphoid tissue (NALT) orchestrates immune responses to Ags in the upper respiratory tract. Unlike other lymphoid organs, NALT develops independently of lymphotoxin-alpha (LTalpha). However, the structure and function of NALT are impaired in Ltalpha(-/-) mice, suggesting a link between LTalpha and chemokine expression. In this study we show that the expression of CXCL13, CCL19, CCL21, and CCL20 is impaired in the NALT of Ltalpha(-/-) mice. We also show that the NALT of Cxcl13(-/-) and plt/plt mice exhibits some, but not all, of the structural and functional defects observed in the NALT of Ltalpha(-/-) mice. Like the NALT of Ltalpha(-/-) mice, the NALT in Cxcl13(-/-) mice lacks follicular dendritic cells, BP3(+) stromal cells, and ERTR7(+) lymphoreticular cells. However, unlike the NALT of Ltalpha(-/-) mice, the NALT of Cxcl13(-/-) mice has peripheral node addressin(+) high endothelial venules (HEVs). In contrast, the NALT of plt/plt mice is nearly normal, with follicular dendritic cells, BP3(+) stromal cells, ERTR7(+) lymphoreticular cells, and peripheral node addressin(+) HEVs. Functionally, germinal center formation and switching to IgA are defective in the NALT of Ltalpha(-/-) and Cxcl13(-/-) mice. In contrast, CD8 T cell responses to influenza are impaired in Ltalpha(-/-) mice and plt/plt mice. Finally, the B and T cell defects in the NALT of Ltalpha(-/-) mice lead to delayed clearance of influenza from the nasal mucosa. Thus, the B and T cell defects in the NALT of Ltalpha(-/-) mice can be attributed to the impaired expression of CXCL13 and CCL19/CCL21, respectively, whereas impaired HEV development is directly due to the loss of LTalpha.

[Functional significance of "blind" outgrowths of the lymphatic capillary network]. / Funktsional'noe znachenie "slepykh" vyrostov seti limfaticheskikh kapilliarov.

The work was aimed at the study of the structural peculiarities of "blind" outgrowths of the lymphatic capillaries and to define their role in the pathogenesis of some diseases. In experiments performed in rabbits by studying the resorption of colloidal solutions, suspensions and hen's blood the subsidiary functional significance of the portion of "blind" outgrowths of the lymphatic capillary network of tendinous center of diaphragm was established. The clavate outgrowths with the narrow mouth may accumulate and store the exogenous material in their lumen. It is concluded that they may accumulate the microorganisms that provoke the relapses of diseases.

[Structure of terminal lymphangion of the thoracic duct]. / Konstruktsiia terminal'nogo limfangiona grudnogo protoka.

Peculiarities of terminal lymphangion wall in humans and dogs were studied in its different parts using various morphological methods. Main results were obtained with the use of a total preparation method after A.V. Borisov. Heterogenous distribution of myocytes in different regions of lymphangion (muscular cuff, valve sinus wall) was noted. In the terminal lymphangion myocytes from three anatomically and topographically different muscles: cuff muscle, tensor muscle of the distant valve, tensor muscle of the proximal valve (thoracic duct orifice valve). Some features of terminal lymphangion construction similar to those seen in other lymphangions as well as certain specific characteristics and species peculiarities, are described.

Tissue-specific expression of Le(Y) antigen in high endothelial venules of human lymphoid tissues.

In this study, we demonstrated that the anti-Le(Y) antibody (BM-1) especially reacted with high endothelial venules (HEVs) in peripheral lymph nodes of blood group O individuals. The Le(Y) expression on HEVs showed a unique tissue-specific pattern, i.e., a large amount of the Le(Y) expression in peripheral lymph nodes and no or small amounts in mesenteric lymph node. Statistical analysis showed that there was the significant difference between the percentage of Le(Y)-positive HEVs in peripheral lymph nodes and mesenteric lymph nodes. No expression of Le(Y) was observed in vessels of Payer's patch, thymus, spleen and other non-lymphoid organs. In blood group A or B individuals, the reactivity between HEVs and anti-Le(Y) antibody increased after enzyme digestion with alpha-N-acetylgalactosaminidase or alpha-galactosidase. These findings show that the expression of difucosylated blood group ABH antigens are especially expressed on HEVs in peripheral lymph nodes. Furthermore, the tissue-specific pattern suggests that these antigens may be related to intercellular adhesion between lymphocytes and HEVs.

Lymphatic vessels of the human heart: precollectors and collecting vessels. A morpho-structural study.

Only topographic and distributional data are available on the lymphatic outflow vessels of the human heart. Here we describe their structural and ultrastructural features. Fragments of the atria, ventricles and fat surrounding the major coronary branches were obtained from hearts of dilated cardiomyopathy patients. Serial semithin sections were observed under light microscopy and used for tridimensional reconstructions. Ultrathin sections were observed by transmission electron microscopy. Precollectors, the initial lymphatic outflow routes of the heart, are small valved vessels with irregular, discontinuous musculature. They originate in the subepicardial region from a network of epicardial, and from scattered myocardial absorbing lymphatic vessels and drain into the collecting vessels accompanying the major coronary branches. Collecting vessels are larger but structurally similar to precollectors. Wall musculature is independent of the size of the vessel. Their ultrastructure is the same as that of precollectors. Endothelial cells have many Weibel-Palade bodies, cytoplasmic filaments and focal adhesions. The basement membrane is discontinuous and anchoring filaments are frequent and conspicuous. The subendothelial layer contains much elastin. Human heart collecting vessels and precollectors may only be distinguished by their size. The scarcity of musculature suggests that lymph progression in this district is mainly ensured by cardiac revolutions. Their ultrastructural features are determined by adaptation to dynamic forces. The architecture of these vessels (random, disorderly, discontinuous, lacking any exact plan) and their large variations in caliber are in line with the ontogenetic hypothesis that peripheral lymphatic vessels originate from the coalescence of mesenchymal lacunae.

[The theory of the design of the lymphangion]. / Teoriia konstruktsii limfangiona.

The results of research in the construction of the lymphangion as a structural and functional unit of human and mammalian lymph vessel are presented. The total preparation method developed by the author was used. Muscle of the lymphatic valve discovered by the author is described for the first time. Lymphangion is proved to have two muscles--the one of the cuff and of the lymphatic valve. Lymph vessels contractile activity is considered as the main factor of the lymph flow. Peculiarities of the innervation of lymphangion different functional portions (cuff, wall of the valvular sinus, valvular torus--muscle of the lymph valve) are demonstrated. Results of the author's longstanding investigations (lymphedema diagnosis and treatment as well as endolymphatic therapy) are introduced into clinical practice.

[The valvular apparatus and tissue organization of the endothelium of the thoracic duct]. / Klapannyi apparat i tkanevaia organizatsiia éndoteliia grudnogo protoka.

The structure of ostial valves and valves located along the thoracic duct and of its branches ostial valves and right lymphatic duct ostial valves were studied in 30 experimental outbred dogs and 46 cats. Cryodestruction of thoracic duct was performed in 28 outbred cats. 1, 3, 7 and 14 days later perfusive fixation with intercellular borders impregnation was carried out with simultaneous examination of intact regions of intravalvular segments, cisterna chyli and area of thoracic duct trunks connection with valvular surfaces. Tissue organization in ageing was studied using the intervalvular segment of old animals. Specimens were studied by means of scanning electron microscopy and film preparations of endothelium. Valves, located along the thoracic duct length are bicuspid formations, while ostial ones are falciform and cuneiform respectively in 80 and 20%. Endotheliocytes of cuspids are characterized with high content of microfilaments bundles in the cytoplasm and low content of microvesicles. Cells of the valvular free margin cross the cuspid edge and have adaptive changes preventing their desquamation: fusiform shape, long basal processes and bundles of microfilaments in the cytoplasm. Peculiar "pericyte-like" cells alike with myofibroblasts lie deep in the cuspid thickness close to the sinusal venous side. Fascicles of the duct smooth myocytes reach the base of the valve. Besides, in the ostial valve stroma there is elastic membrane, better displayed along the cuspid venous side. Increased polymorphism and changes of the endotheliocytes metric characteristics were demonstrated in the zones of turbulent lymph flow. Analysis of the newly formed endothelium tissue mosaics allows to reveal mechanisms of monolayer repair: spreading and migration of endotheliocytes on the first day, their proliferation within three days, desquamation of newly formed endotheliocytes and spreading of adjacent cells on later stages.

[Morphology and histochemistry of lymphatic vessels of the upper aerodigestive tract: a clinically oriented study]. / Morphologie und Histochemie von Lymphgefässen der oberen Luft- und Speisewege: Eine klinisch orientierte Untersuchung.

BACKGROUND: Exact knowledge of lymphatic morphology and histochemistry is required to understand the clinical importance of the lymphatic system of the upper aerodigestive tract. Publications in this field are rare. METHODS: Light and electronmicroscopic examinations were carried out on lymphatics of the upper aerodigestive tract. Enzyme, immune and histochemical investigations were also performed on 764 tissue specimens from this region. RESULTS: The discontinuous basement membrane surrounding the initial lymphatic contains laminin, type IV collagen, and fibronectin and is usually continued by a fibrillar elastic apparatus that is less developed than dermal lymphatics. 5'-nucleotidase, adenylatcyclase, and guanylatcyclase activity in lymphatics is not as high as it is in blood capillaries. The endothelium of collecting lymphatics contains high concentrations of factor VIII associated antigen, while it is found in the endothelium of initial lymphatics only in small amounts or not at all. Histochemical studies of the endothelium of lymphatics show a positive lectine binding reaction for UEA I, PNA, DBA, GS I, MPA, and RCA I. CONCLUSIONS: The histochemical results obtained allow easy differentiation of lymphatics and blood capillaries. The vessel type differentiation is relevant for the examination of organ-related lymphatic systems, to determine whether tumor cell nests are located in lymphatics, blood capillaries, or artificial tissue gaps.

Lymphatic vessels of the human dental pulp in different conditions.

The characteristics of the lymphatic vessel endothelial wall have been investigated in human normal and inflamed dental pulps. In normal pulps the endothelial wall is characterized by the presence of micropinocytotic vesicles and intraparietal channels. In the inflamed pulpal tissue, where an increase in interstitial fluid pressure occurs, the distended endothelial wall presents open junctions between endothelial cells and the openings of the intraparietal channels. Moreover the micropinocytotic vesicles disappear. The cytoplasm of the endothelial cells is characterized by the presence of numerous Weibel-Palade bodies, which increase in number in the dilated vessels. In the fibrillar apparatus surrounding the lymphatic vessel wall collagen fibrils are the prevalent component, while elastic fibers are not present. The different morphological properties of the lymphatic vessels are compared and discussed with regard to the variation of the functional conditions of the tissue.

Enzyme-histochemical study on the fine distribution of the intramural lymphatics at the ileocecal junction of the monkey intestine.

The fine distribution of the intramural lymphatics at the ileocecal junction of the monkey intestine, especially in the lamina propria of the ileocecal valve, was examined by light and electron microscopy using enzyme-histochemical staining. The distinction between the lymphatics and the blood vessels was made by light microscopy on cold glycol methacrylate resin (JB-4) sections using 5'-nucleotidase (5'-Nase)-alkaline phosphatase (ALPase) double staining. The lymphatics were found to show strong 5'-Nase activity and to comprise irregularly shaped vessels or spaces. The central lymphatic vessels (central lacteals) in low villi were seen to lie deep within the ALPase-positive subepithelial capillary network. In the ileum side of the ileocecal junction, the 5'-Nase-positive lymphatics were seen both in the superficial layer and the deep layer of the lamina propria. On the contrary, in the cecum side the mucosal lymphatics were less numerous in the superficial layer and were distributed mainly in the deep layer near the lamina muscularis mucosae. These lymphatics ran through the lamina muscularis and merged into the lymphatic network in the submucosa. The submucosal lymphatics communicated with each other at the ileocecal junction and formed a well-developed network. Collecting lymphatics with valves were also seen near the tunica muscularis (sphincter muscle) in the deep submucosa. These lymphatics traversed the muscle layer and drained into the subserosal lymphatics.

Demonstration of lymphatics in human synovial tissue.

Using a cocktail of monoclonal antibodies PAL-E and DE-U-10 (anti-desmin), combined in double labelling techniques with the lectin Ulex europaeus agglutinin I (UEAI), vessels consistent with lymphatics were demonstrated in normal human synovial tissue. These vessels were negative for the monoclonal cocktail and positive for UEAI, were thin-walled and were located close to deep arterioles and venules as expected. Elastin was not found to assist identification of lymphatics in synovium. In rheumatoid arthritic synovium no vessels staining in the manner of normal lymphatics were found. This may indicate absence or change of phenotype of this type of endothelium in disease.

[Open lymph vessels of the ligamentum falciforme hepatis in the human]. / Offene Lymphgefässe im Ligamentum falciforme hepatis des Menschen.

The falciform ligament and the ligamentum teres of the liver are the so-called ventral mesohepaticum. These ligaments are covered on both sides by one layer of mesothelial cells and consist of a dense network of lymphatic capillaries. Preparations of semithin sections were examined light microscopically, ultrathin sections by means of transmission electron microscopy and gold sputtered surface preparations by scanning electron microscopy. It could be shown for the first time that in the mesothelial cell surface there are so-called "stomata", holes which are considered to be connections between the peritoneal cavity and lymphatic capillaries. This network of lymphatics is able to rapidly absorb fluids and particles like blood cells. The knowledge of localisation, structure and function of the submesothelial lymphatic system is of high importance for surgical processing, because in most infections and tumor diseases it is also affected.