A hypothesis on the role of primitive macrophages in initial embryonic lymphatic development

Recent evidences have shown that macrophages are tightly related to pathological lymphangiogenesis. However, the effects which primitive macrophages take in embryonic lymphatic development remains unclear. Here, we postulate that the primitive macrophages may play an important role in initial embryonic lymphatic development. The possible mechanism: primitive macrophages induce BECs to transdifferentiate into LECs and initiate the budding, moreover, the lymph sacs are not only formed by LECs but also some scattered cells with macrophage characteristics preferentially located in the loose mesenchyme.     

Proteomics analysis of two mice hepatocarcinoma ascites syngeneic cell lines with high and low lymph node metastasis rates provide potential protein markers for tumor malignancy attributes to lymphatic metastasis

Lymph node metastasis (LNM) is recognized as an important factor involved in the tumor malignancy progression. Our previous study has indicated that the hepatocarcinoma cell line with 75% of LNM (Hca‐F)‐cell‐induced neoplasia and the hepatocarcinoma cell line with 25% of LNM‐induced neoplasia are accompanied with high (75%) and low (25%) incidences of LNM. In the current study, 62 and 54 protein spots were observed up‐regulated and down‐regulated in Hca‐F cell relative to the hepatocarcinoma cell line with 25% of LNM by 2‐D DIGE. Totally, 113 unique proteins were identified by HPLC‐nano ESI‐MS/MS analysis. The expression levels of Annexin A7, Ulch3, and ER protein 29 were validated by Western blotting analyses. The abnormally regulated proteins were categorized and annotated by protein analysis through evolutionary relationships analysis with the aid of the database for annotation, visualization and integrated discovery tool. Seventeen gene candidates concordantly expressed both at mRNA and protein levels. By making a challenge, we detected expression levels of Annexin A7 in primary gastric cancer (GC) and primary GC cancer tissues with LNMs by immunohistochemisty. Higher ratio of positive and strong expressions Annexin A7 in GC might correlate with the tumor progression. The repression of Annexin A7 inhibits the mobility and invasion abilities of Hca‐F cell, increases the apoptosis rate of Hca‐F cell. Current study narrows and provides certain specific protein candidates potentially playing important roles in LNM‐associated cancers.     

粘附分子在组织内淋巴管内皮和体外培养淋巴管内皮细胞的表达

目的研究粘附分子PECAM-1、ICAM-3、VCAM-1和CD44在组织淋巴管和培养淋巴管内皮细胞(LECs)的表达.方法取狗的空肠和腹前壁皮肤,作冰冻切片,用免疫组织化学染色法标记粘附分子在组织淋巴管内皮的表达.分离和培养狗胸导管的LECs,用免疫荧光染色法标记粘附分子在培养LECs的表达,在共聚焦激光扫描显微镜下观察.用图像分析系统分析粘附分子表达的光密度值(OD值),并与肿瘤坏死因子α(TNF-α)或脂多糖(LPS)刺激细胞的表达进行比较.结果组织内淋巴管内皮的PECAM-1和ICAM-3呈免疫反应阳性.培养LECs表达PECAM-1、ICAM-3和CD44,而VCAM-1的表达不明显.用TNF-α刺激细胞后,PECAM-1和ICAM-3表达的OD值与正常对照组无明显差别,CD44的OD值低于正常对照组.用LPS刺激细胞后,PECAM-1、ICAM-3和CD44表达的OD值均无显著性变化.用TNF-α或LPS刺激LECs后,VCAM-1呈弱表达.结论粘附分子PECAM-1、ICAM-3、VCAM-1和CD44的表达在培养LECs和组织内淋巴管内皮细胞存在着差异.用TNF-α或LPS刺激后,LECs的VCAM-1表达增强.这可能与淋巴管内皮细胞的功能状态有关.     

Role of Angiopoietins and Tie receptor tyrosine kinases in angiogenesis and lymphangiogenesis

This review focuses on the signaling system involving the Angiopoietin1/Tie2 receptor, which appears to be involved in the secondary stages of blood vessel formation. Although this system is crucial for blood and lymphatic vessel formation, identifying its precise role in embryonic and adult vascular biology has been a major challenge. The evidence for the key role of the Angiopoietin/Tie system is discussed, and some of the other members of the system (Ang2, Tie1) are mentioned. A comparison is made with the VEGF signaling system, which seems to provide a complementary, and somewhat more tractable, signaling system. Some of the basic unanswered questions concerning Tie/Angiopoietin biology and the secondary stages of blood vessel formation are also highlighted.     

Mechanics of interstitial-lymphatic fluid transport: theoretical foundation and experimental validation

Interstitial fluid movement is intrinsically linked to lymphatic drainage. However, their relationship is poorly understood, and associated pathologies are mostly untreatable. In this work we test the hypothesis that bulk tissue fluid movement can be evaluated in situ and described by a linear biphasic theory which integrates the regulatory function of the lymphatics with the mechanical stresses of the tissue. To accomplish this, we develop a novel experimental and theoretical model using the skin of the mouse tail. We then use the model to demonstrate how interstitial–lymphatic fluid movement depends on a balance between the elasticity, hydraulic conductivity, and lymphatic conductance as well as to demonstrate how chronic swelling (edema) alters the equipoise between tissue fluid balance parameters. Specifically, tissue fluid equilibrium is perturbed with a continuous interstitial infusion of saline into the tip of the tail. The resulting gradients in tissue stress are measured in terms of interstitial fluid pressure using a servo-null system. These measurements are then fit to the theory to provide in vivo estimates of the tissue hydraulic conductivity, elastic modulus, and overall resistance to lymphatic drainage. Additional experiments are performed on edematous tails to show that although chronic swelling causes an increase in the hydraulic conductivity, its greatly increased distensibility (due to matrix remodeling) dampens the driving forces for fluid movement and leads to fluid stagnation. This model is useful for examining potential treatments for edema and lymphatic disorders as well as substances which may alter tissue fluid balance and/or lymphatic drainage.     

The passage of macrophages and lymphocytes from the interstitium across the lymphatic endothelium of rat lacteals

Summary The passage of cells across the lymphatic endothelium of rat lacteals in both normal and non-pathological experimental conditions (fasting, lymphatic stasis) was studied by means of serial thin sections and three-dimensional models. Two different pathways of transendothelial migration were observed: (1) macrophages enter the lymphatic lumen via the cytoplasm of endothelial cells, without involvement of intercellular junctions, whereas (2) lymphocytes migrate through intraendothelial channels, dynamic structures organized by the lymphatic endothelium under physiological conditions.     

Abnormal recruitment of periendothelial cells to lymphatic capillaries in digestive organs of angiopoietin-2-deficient mice

The fine structure of lymphatic capillaries in the digestive organs of angiopoietin-2 (Ang2) knockout mice was studied by using both immunohistochemistry and electron microscopy. The genetic deletion of Ang2 yielded hypoplasia and disorganization of the lymphatic capillaries, with their shapes being irregular, and an aberrant recruitment of vascular periendothelial cells immunopositive for smooth muscle actin to the lymphatic capillaries. The abnormal lymphatic periendothelial cells were considered to be a type of pericyte for the lymphatic capillaries after the deletion of Ang2, because they were ultrastructurally characterized by abundant thin myofilaments in their cytoplasm and long cytoplasmic extensions similar to those shown by blood vascular pericytes. The genetic replacement of Ang2 with Ang1 rescued the defects, viz., the disorganization and disordered structure of the lymphatic capillaries. The present findings suggest that Ang2 serves the morphogenesis of lymphatic capillaries as an agonist for the receptor, Tie2, and that Ang1 can replace Ang2 in guiding lymphatic formation and development. H. Shimoda and M. Witte were supported by an Exchange Visitor Program Grant from the Ministry of Education, Culture, Sports, Science and Technology, Japan, and by Contract 6011 from the Arizona Disease Control Research Commission, respectively.     

Maldevelopment of dermal lymphatics in Wnt5a-knockout-mice

Maintenance of tissue homeostasis and immune surveillance are important functions of the lymphatic vascular system. Lymphatic vessels are lined by lymphatic endothelial cells (LECs). By gene micro-array expression studies we recently compared human lymphangioma-derived LECs with umbilical vein endothelial cells (HUVECs). Here, we followed up on these studies. Besides well-known LEC markers, we observed regulation of molecules involved in immune regulation, acetylcholine degradation and platelet regulation. Moreover we identified differentially expressed WNT pathway components, which play important roles in the morphogenesis of various organs, including the blood vascular system. WNT signaling has not yet been addressed in lymphangiogenesis. We found high expression of FZD3, FZD5 and DKK2 mRNA in HUVECs, and WNT5A in LECs. The latter was verified in normal skin-derived LECs. With immunohistological methods we detected WNT5A in LECs, as well as ROR1, ROR2 and RYK in both LECs and HUVECs. In the human, mutations of WNT5A or its receptor ROR2 cause the Robinow syndrome. These patients show multiple developmental defects including the cardio-vascular system. We studied Wnt5a-knockout (ko) mouse embryos at day 18.5. We show that the number of dermal lymphatic capillaries is significantly lower in Wnt5a-null-mice. However, the mean size of individual lymphatics and the LEC number per vessel are greater. In sum, the total area covered by lymphatics and the total number of LECs are not significantly altered. The reduced number of lymphatic capillaries indicates a sprouting defect rather than a proliferation defect in the dermis of Wnt5a-ko-mice, and identifies Wnt5a as a regulator of lymphangiogenesis.