Immunohistochemical distribution of serum proteins in living mouse heart with In vivo cryotechnique

In vivo cryotechnique (IVCT), which immediately cryofixes target organs in situ, was used to clarify the morphological features of beating heart tissue of living mice. IVCT was performed for diastolic heart tissue under the condition of monitoring with electrocardiogram (ECG). Other mouse hearts were prepared with conventional perfusion-fixation (PF-DH) or immersion-fixation followed by dehydration (IM-DH), and quick-freezing of resected heart tissues (FQF). Immunolocalizations of albumin, immunoglobulin G1 (IgG1), intravenously injected bovine serum albumin (BSA), and connexin 43 were examined after different intervals of BSA injection. In the case of IVCT, the exact stop time of beating mouse hearts was recorded by ECG, and open blood vessels with flowing erythrocytes were observed with less artificial tissue shrinkage than with conventional preparation methods. Both albumin and BSA were well preserved in intercalated discs and t-tubules of cardiomyocytes in addition to blood vessels and interstitial matrices. IgG1 was immunolocalized in interstitial matrices of heart tissues in addition to their blood vessels. At 4 hr after BSA injection, it was immunolocalized in the intercalated discs of cardiomyocytes and lost later at 8 hr. IVCT should prove to be more useful for the morphofunctional examination of dynamically changing heart tissue than conventional preparation methods.     

The pathogenesis of experimentally induced Trypanosoma brucei infection in the dog. I. Tissue and organ damage.

Inoculation of dogs with Trypanosoma brucei produced an acute fetal disease similar to that seen following natural infection. The disease was characterised by high levels of parasitaemia, moderately severe anemia, and marked changes in the lymphoid system. Extravascular invasion by large numbers of trypanosomes was widespread throughout the body and was accompanied by severe tissue damage. Tissue invasion by trypanosomes was associated with marked cellular infiltration involving lymphoid cells and plasma cells followed by macrophages and polymorphonuclear leukocytes. Associated with these reactions, severe cellular degeneration and focal necrosis occurred. While these changes were widespread and were found in the majority of tissues examined, consistently severe lesions were found in the heart, eyes and central nervous system. In many organs, lymphatic vessels were distended with fluid, trypanosomes, and a cell population similar to that in the surrounding tissue; fibrin deposition and thrombus formation was sometimes observed within the lymphatic lumens. Thrombosis was also found in the blood vessels of the pampiniform plexus, the venous plexus of the ovary, and branches of the renal vein. A severe necrotizing vasculitis affecting only the coronary vessels was a prominent feature in some animals.     

睾丸间质细胞及淋巴管的超微结构

目的　探明睾丸间质细胞及淋巴管的超微结构。方法　灌流固定后半薄超薄切片光镜和电镜下观察。结果　睾丸间质细胞有二种亚群 ,各有其特点。淋巴管存在于纵隔内 ,小叶内未见典型淋巴管。结论　睾丸间质细胞分明细胞和暗细胞二种 ,纵隔内有丰富的淋巴管及毛细淋巴管 ,而小叶内则存在广泛的淋巴间隙     

Pathways for movement of fluid and cells from hepatic sinusoids to the portal lymphatic vessels and subcapsular region in rat livers

It has long been a mystery how fluid and migrating cells in the hepatic sinusoids reach lymphatic vessels in the portal tract. Here we describe previously-unknown channels that connect the space of Disse with the portal tract in the rat liver. Transmission electron microscopy was performed on livers injected with either horseradish peroxidase (HRP) or lipopolysaccharide, and scanning electron microscopy was carried out on livers macerated with KOH. Transmission electron microscopy revealed the presence of channels with collagen fibers traversing the limiting plate. A tracer study showed that HRP was in the channels as well as along inlet venules. Dendritic cells in the hepatic sinusoids or between hepatocytes of the limiting plate were also observed extending their pseudopodia through the channels in the limiting plate to the interstitial space of the portal tract. Scanning electron microscopy further showed that many channels (1-3microm in diameter) penetrated through the limiting plate independently of blood vessels and connected the space of Disse with the interstitial space of the portal tract. In addition, the portal tract possessed prelymphatic vessels that were lined with fibroblast-like cells and frequently contained dendritic cells. The initial segment of the portal lymphatic vessels opened to the interstitial tissue space. These results indicate that fluid and dendritic cells in the hepatic sinusoids probably pass through both the space of Disse and the channels traversing the limiting plate, enter the interstitial space of the portal tracts, and finally move from the prelymphatic vessels to the portal lymphatic vessels.     

Lymph pathways associated with three types of follicle structure found in gut-associated lymphoid tissue of horse ileum

In the horse ileum, lacteals in the villi are continuous with prelymphatic intercellular channels and a plexus of lymphatic sinuses in the lamina propria that encircle the domes of the follicle/dome structures and proprial follicles. These sinuses may act as the major entry site for many of the lymphocytes migrating from gutassociated lymphoid tissue via the lymphatic system. Vessels from this plexus penetrate the muscularis mucosae and lymph flows into lymphatic vessels within the interfollicular tissue between the follicles of both follicle/dome structures and lymphoglandular complexes (LGCs). No lymphatic vessels leave the follicles, but intercellular pathways of the follicles are continuous with those in the surrounding interfollicular tissue and follicular sinuses around the base of the follicles. These pathways appear to provide the only available lymphatic route for lymphocytes leaving LGCs to enter the lymphatic system. Lymph from the interfollicular tissue enters deep submucosal lymphatic vessels, containing prominent valves, which drain into other vessels transporting lymph from the surface of the ileum.     

Morphologic characteristics of initial lymphatics of the healthy and diseased human gingiva.

Investigation was performed on healthy and inflamed human gingivae. In the healthy mucosa lymphatic vessels generally appeared as flattened channels with a reduced lumen. Only in very inflamed tissue were some more evident vessels with a distended wall detectable. Ultrastructurally, most of the vessels had the characteristics of capillaries and they were delimited by a thin and irregular endothelial wall with large intercellular spaces. These observations indicate that in the gingival tissues, which are continuously exposed to inflammatory agents and need a really efficient draining system, the pathway of interstitial exudation and cell migration may include both the lymphatic vessel system and the intercellular spaces of the permeable junctional epithelium.     

An Autopsy Case of Chronic Rheumatic Carditis with Myocardial Aschoff Bodies

An autopsy case of chronic rheumatic carditis is described. The patient was a woman aged 32 years and the duration of the disease was more than eight years.
At autopsy, the heart showed typical mitral stenosis with verrucous scarring, calcification and contraction of the valve. Areas of degenerating muscle cells and proliferation of the interstitial connective tissue, partly with perivascular infiltration, were seen throughout the myocardium, and heart muscles showed narrowing and fragmentation. There was fibrinoid degeneration of the wall in small coronary vessels. Numerous Aschoff bodies of various structures were found mostly in the proliferated interstitial connective tissue, between damaged muscle bundles and adjacent to the small coronary vessels of the myocardium. A small number of Aschoff bodies were situated in the subendocardial tissue in which there was no muscle.     

淋巴管阻断对大鼠睾丸影响的自动图像分析

目的 :研究淋巴管阻断后对睾丸的定量组织学影响。方法 :用电灼法将双侧睾丸淋巴管阻断 ,对双侧睾丸淋巴管阻断术后 2天、6天、1 2天、2 4天和 48天的 2 5只大鼠进行研究。另 2 5只大鼠为假手术对照。将睾丸组织切片输入TAS plus型自动图像分析仪进行定量组织学测定。结果 :在睾丸淋巴管阻断术后 6天、1 2天和 2 4天 ,睾丸间质的面积占总面积的比值增加 ,而生精小管的面积占总面积的比值则减少。在睾丸淋巴管阻断术后 2天、6天、1 2天、2 4天和 48天 ,生精上皮的面积占总面积的比值均减少 ,而间质面积与生精上皮面积的比值则均增加 ,生精小管的直径和生精上皮的厚度均变小。结论 :以上结果表明睾丸淋巴管阻断对生精上皮的影响比对间质的影响更为明显。     

Morphologic characteristics of initial lymphatics of the healthy and diseased human gingiva

Investigation was performed on healthy and inflamed human gingivae. In the healthy mucosa lymphatic vessels generally appeared as flattened channels with a reduced lumen. Only in very inflamed tissue were some more evident vessels with a distended wall detectable. Ultrastructurally, most of the vessels had the characteristics of capillaries and they were delimited by a thin and irregular endothelial wall with large intercellular spaces. These observations indicate that in the gingival tissues, which are continuously exposed to inflammatory agents and need a really efficient draining system, the pathway of interstitial exudation and cell migration may include both the lymphatic vessel system and the intercellular spaces of the permeable junctional epithelium. Anat Rec 255:175–179, 1999. © 1999 Wiley‐Liss, Inc.     

小鼠胰腺淋巴管的分布及其与胰岛的关系

目的:探讨小鼠胰腺淋巴管的形态分布及其结构特点。方法:对小鼠胰组织切片进行HE染色,5核苷酸酶(5-Nase)和碱性磷酸酶(ALP)双重染色,光镜、透射电镜、扫描电镜二次电子和背散射电子图像(SEI/BEI)观察。结果:小鼠胰腺的淋巴管结构较典型,在胰腺的小叶间结缔组织内,较大淋巴管与血管和导管相互伴行;毛细淋巴管起自胰腺腺泡周围,并且均匀地分布于整个小叶内;小叶内有单独走行的集合淋巴管,亦存在与血管并行情况;在胰岛内部未发现毛细淋巴管,但胰岛周围可见丰富的毛细淋巴管。结论:小鼠胰腺小叶间和小叶内结缔组织中,均有淋巴管分布;胰岛内部虽无淋巴管,但胰岛与周围毛细淋巴管的关系较密切。     

Intrinsic interrelation of lymphatic endothelia with nerve elements in the monkey urinary bladder

Histochemical staining techniques for 5'-nucleotidase (5'-Nase) and acetylcholinesterase (AChE) were undertaken to localize the lymphatic network and nerve plexus in the monkey urinary bladder. Abundant 5'-Nase-positive lymphatic networks were characterized by increased number of valve-like structures and decreased calibre of blind-ends from the subepithelium to the subserosa. AChE-positive nerve fibers were visible throughout the vesical walls as fine plexuses, the densest being the neuromuscular plexus among the detrusor muscle cells or in each muscle bundle. AChE-positive nerve fibers or terminals were more frequently discernible around blood vessels than around lymphatics, and showed more intimate association with the lymphatics in the muscularis than those in the subepithelium. The nerve terminals in the subepithelium were frequently separated from attenuated lymphatic endothelium by the long processes of fibroblasts or some connective tissue cells. An ultrastructural observation revealed that unmyelinated nerve fibers with numerous neurofilaments and neurotubules run in close apposition to the lymphatic endothelium. Noteworthily, fewer terminal varicosities containing numerous small agranular vesicles (30-50 nm) and mitochondria, partially or completely bare of their Schwann cell covering in the vicinity of the lymphatic endothelium, were found in the subendothelium of initial lymphatics than in collecting ones. These terminals were occasionally identified at a distance of 120-350 nm from the subendothelial aspect of valve-originating roots, although no direct innervation of the vascular muscle cells could be found. A loose fibro-elastic connective tissue was usually interlaced between glial cell covering and lymphatic endothelium. The intrinsic interrelation of the lymphatic wall with the nerve plexus implies that the twisted subendothelial nerve terminals might be involved in intramural lymph drainage of the bladder.     

家兔回肠淋巴管三维结构的扫描电镜观察

目的　探讨回肠淋巴管的三维结构及流注关系。方法　将Mercox注入家兔回肠壁内制成淋巴管铸型 ,在扫描电镜下观察回肠各层淋巴管的三维结构。结果　回肠的粘膜层、粘膜下层、肌层和浆膜层都有毛细淋巴管网及淋巴管。在小肠绒毛内存有中央乳糜管 ,铸型样品上可见其立体结构 ,可见中央乳糜管注入粘膜层和粘膜下层的毛细淋巴管网 ,该网汇合而成的淋巴管穿过肌层进入浆膜层 ,之后以淋巴集合管的形式入肠系膜淋巴管而离开回肠。铸型标本可见淋巴管呈串珠状外观 ,管壁表面有双凹切迹 ,相当于淋巴管瓣膜的部位。铸型表面还可见淋巴管内皮细胞的椭圆形压迹 ,光镜下可见粘膜下与肌层之间存有三角形间隔 ,其内动脉与两条淋巴管并行。结论　中央乳糜管注入粘膜下毛细淋巴管网 ,该网与粘膜下淋巴管网相连 ,此淋巴管再穿肌层入浆膜层后离开回肠 ,并见淋巴通道存在于中央糜管周围。     

整片组织免疫染色显示淋巴管构筑

目的探讨整片组织内淋巴管免疫染色,并与间接注射法和组织切片免疫染色法比较。方法用普鲁士蓝间接注射新生儿肝被膜内淋巴管,观察肝浅淋巴管的构筑。通过火棉胶切片分析肝浅淋巴管的流向。取人大腿皮肤,作石蜡切片和冷冻切片,免疫染色后观察淋巴管的分布。取大鼠耳和背部的皮肤、膈和小肠,将整片组织作免疫染色,观察组织内淋巴管构筑。结果间接注射普鲁士蓝可显示淋巴管构筑和流向,但淋巴管染色缺乏特异性。组织切片免疫染色可显示淋巴管分布,然而不能准确地分析淋巴管密度。整片组织的免疫染色很理想,毛细淋巴管盲端、毛细淋巴管网和淋巴管丛清晰可见。结论整片组织免疫染色能够很好地显示组织内完整的淋巴管构筑。     

Lymph Circulation in the Liver

The liver produces a large amount of lymph, which is estimated to be 25 to 50 % of lymph flowing through the thoracic duct. The hepatic lymphatic system falls into three categories depending on their locations: portal, sublobular, and superficial lymphatic vessels. It is suggested that 80 % or more of hepatic lymph drains into portal lymphatic vessels, while the remainder drains through sublobular and capsular lymphatic vessels. The hepatic lymph primarily comes from the hepatic sinusoids. Our tracer studies, together with electron microscopy, show many channels with collagen fibers traversing through the limiting plate and connecting the space of Disse with the interstitial space either in the portal tracts, or around the sublobular veins. Fluid filtered out of the sinusoids into the space of Disse flows through the channels traversing the limiting plate either independently of blood vessels or along blood vessels and enters the interstitial space of either portal tract or sublobular veins. Fluid in the space of Disse also flows through similar channels traversing the hepatocytes intervening between the space of Disse and the hepatic capsule and drains into the interstitial space of the capsule. Fluid and migrating cells in the interstitial space pass through prelymphatic vessels to finally enter the lymphatic vessels. The area of the portal lymphatic vessels increases in liver fibrosis and cirrhosis and in idiopathic portal hypertension. Lymphatic vessels are abundant in the immediate vicinity of the hepatocellular carcinoma (HCC) and liver metastasis. HCCs expressing vascular endothelial growth factor‐C are more liable to metastasize, indicating that lymphangiogenesis is associated with their enhanced metastasis. Anat Rec, 291:643–652, 2008. © 2008 Wiley‐Liss, Inc.     

Blood vessels of the Peyer's patch in the mouse: I. Topographic studies

The topographic distribution of blood vessels in Peyer's patches of mice was studied by light and scanning electron microscopy with whole mounts of flattened gut segments and vascular corrosion casts. Peyer's patches are imbedded in the intestinal wall and share its blood supply. Two to four mural trunks may contribute to the area of the patch. In and around the lymphoid nodules the microcirculation is highly specialized. The nodule is permeated by a meshwork of fine capillaries that is supplied by arterioles entering on the serosal and lateral surfaces. Blood flow to the lymphoid nodule appears tobe monitored by arterial sphincters; the dense lymphatic tissue can also beby passed by arteriovenous communications. An extensive venous network encircles the nodule. Most of these venules are lined by high endothelium which is penetrated by lymphocytes. The geometry of these vessels suggests a slow and turbulent flow in these vascular segments that may aid margination of lymphocytes. A planar capillary plexus lies subjacent to the mucosal epithelium in the dome area.     

Intraperitoneal dye injection method for visualizing the functioning lymphatic vascular system in zebrafish and medaka

A hierarchically organized lymphatic vascular system extends throughout the vertebrate body for tissue fluid homeostasis, immune trafficking, and the absorption of dietary fats. Intralymphatic dye injection and serial sectioning have been the main tools for visualizing lymphatic vessels. Specific markers for identifying the lymphatic vasculature in zebrafish and medaka have appeared as new tools that enable the study of lymphangiogenesis using genetic and experimental manipulation. Transgenic fishes have become excellent organisms for visualizing the lymphatic vasculature in living embryos, but this method has limited usefulness, especially in later developmental stages. The functional lymphatic endothelium predominantly takes up foreign particles in zebrafish and medaka. We utilized this physiological activity and lymph flow to label lymphatic vessels. Intraperitoneal injection of trypan blue is useful for temporal observations of the lymphatic ducts, which are essential for large-scale genetic screening, while cinnabar (HgS) injection allows identification of the lymphatic endothelium under electron microscopy, avoiding artefactual damage. This injection method, which is not high in cost and does not require high skill or special devices, is applicable to any live fish with functioning lymphatic vessels, even mutants, with high reproducibility for visualizing the entire lymphatic vascular system.     

Structural organization and fine distribution of lymphatic vessels in periodontal tissues of the rat and monkey: a histochemical study

The structural organization and fine distribution of lymphatic vessels in the periodontal tissues (gingiva, periodontium and alveolar process) were examined by light and electron microscopy using an enzyme-histochemical method. Whole mount preparations of periodontal membranes peeled from the teeth and cryostat sections of normal or decalcified tissues treated with EDTA were double-stained using 5'-nucleotidase (5'-Nase)-alkaline phosphatase (ALPase) and examined by light microscopy. This staining procedure allowed the lymphatic vessels in the periodontal tissue to be differentiated from blood vessels. Well-developed 5'-Nase-positive lymphatics were observed in the gingiva and periodontium. The histochemical aspects of 5'-Nase activity in lymphatic vessels are discussed in detail, with special reference to the supply of Mg++ ions. A network of 5'-Nase-positive lymphatics was observed in whole mount preparations of the periodontal membrane for the first time. This network was also observed in the tissue sections. More 5'-Nase-positive lymphatics were seen in the root area of the periodontium than in the cervical area. 5'-Nase-positive lymphatics in residual tissue blocks remaining after cryostat sectioning and in whole mount preparations were highlighted with good contrast and resolution on backscattered electron images produced by scanning electron microscopy. Dense granular precipitations resulting from the 5'-Nase reaction were observed on the luminal surface of the lymphatic endothelial cells as well as on the basal side but were absent in the blood vessels.     

Lymphatic amyloidosis, a previously unrecognized form of amyloid deposition in generalized amyloidosis

Although amyloid deposition in relation to blood vessels is a well-recognized feature of generalized amyloidosis, lymphatic vessel amyloidosis is not mentioned in the literature. Systematic investigation of tissue removed at autopsy from patients with generalized amyloidosis and biopsy specimens from cases of localized amyloidosis and familial Mediterranean fever showed that amyloid deposition around lymphatics is by no means uncommon. The material investigated was mainly large and small bowel, lung, heart and kidney. Amyloid was identified by green birefringence with the Congo red stain on cross-polarization and lymphatics by their lack of immunostaining for CD34. Involvement of lymphatics was noted in 20 of the 42 organs from which specimens were examined, and was always accompanied by involvement of blood vessels and/or the interstitium. In the intestine, lymphatic amyloidosis was found mainly in the submucosa and subserosa, and was also demonstrated by electronmicroscopy in one case. Although lymphatic amyloidosis was equally common in the heart, lung and kidney, it was usually less prominent here than in the intestine. No lymphatic involvement was seen in localized amyloidosis. As the lymphatics play a central role in the resorption of interstitial proteins, they are probably also involved in the resorption of amyloid proteins. Amyloid deposition in the vicinity of lymphatics is probably the result of decompensation of this process.     

Observation of lymphatic vessels in orbital fat of patients with inflammatory conditions: a form fruste of lymphangiogenesis?

In the absence of antibodies specific for lymphatic vessels, analysis of lymphatic vessels within different tissues has been widely performed with light microscopic and, most importantly, electron microscopic techniques. In regard to lymphatic vessels in the ocular globe and the periocular structures, controversy remains about the specific distribution of lymphatic channels. It is postulated that bulbar and retrobulbar tissues are devoid of lymphatic vessels, but lymphatic vessels have been demonstrated in lacrimal gland and epibulbar conjunctiva. In this study, we analyzed orbital fat for the presence of lymphatic tissue using D2-40, a monoclonal antibody, specific for lymphatic vessels. We found lymphatic vessels present within bulbar conjunctiva extending to the level of the ciliary apparatus. No lymphatics were identified in healthy anterior orbital adipose tissue. In two cases of orbital mucor-mycosis and one case of panendophthalmitis, significant lymphovascular proliferation was present within granulation tissue associated with the acute inflammation. We conclude that lymph vessel proliferation may be induced in inflammatory conditions in tissues which are normally devoid of lymph channels.