Characterization of the dental lymphatic system and identification of cells immunopositive to specific lymphatic markers

The lymphatic system is important for immune barrier function and for tissue fluid balance. During inflammation, lymphangiogenesis takes place to enhance the transport of filtered fluid, proteins, and immune cells. Dental tissue is frequently exposed to inflammatory insults, but the lymphatic system and its responses to injury have not been investigated in detail using specific lymphatic markers. We aimed to study this system and to establish whether lymphangiogenesis takes place during wound healing. Immunostaining of the lymphatic endothelial hyaluronan receptor-1 (LYVE-1) and vascular endothelial growth factor receptor-3 (VEGFR-3) demonstrated initial lymphatics in the coronal molar pulp, whereas in incisors the initial lymphatics were found only in the apical part. In molars, lymphatic vessels exit the pulp through the apex and lateral canals. In interdental bone, transverse lymphatics were found, raising the possibility that an infection can be spread from the periodontal ligament to a neighbouring tooth. LYVE-1⁺ and VEGFR-3⁺ immune cells were found in both molar and incisor pulps, and phenotyping of the cells showed that they are of a monocytic lineage. In inflamed pulp these cells were not observed. Macrophages are suggested to contribute directly to the formation of lymphatic vessels after pulp exposure.     

Distribution and organization of lymphatic vessels in the mouse gingiva: An immunohistochemical study with LYVE-1

OBJECTIVE: This study introduced the usefulness of LYVE-1 immunoreactivity for identification of lymphatic vessels in decalcified tissues, and demonstrated the fine distribution and organization of these vessels in mouse gingiva. DESIGN: After confirming the specificity of anti-mouse LYVE-1, frozen sections of mouse decalcified gingiva were immunostained with the antibody. RESULTS: The LYVE-1-positive lymphatic vessels were clearly found in the connective tissue under the gingival epithelium; these vessels appeared to pass through the lamina propria of the gingiva toward the alveolar crest and run along the external surface of the alveolar bone. The lymphatic vessels were sparse and apart from the oral gingival and sulcular epithelia, while they were dense adjacent to the junctional epithelium. CONCLUSIONS: The dense network of the lymphatic vessels adjacent to the junctional epithelium, which is apparently exposed to foreign antigens, may act as an efficient drainage pathway of the excessive interstitial fluid and immune cells, and play an active role in the immune defense of the gingiva. The present study also revealed the absence of lymphatic connection between gingiva and periodontal ligament.     

Lymphatic growth factors are expressed in human gingiva and upregulated in gingival fibroblasts after stimulation

1 Background

The lymphatic growth factors vascular endothelial growth factor (VEGF)‐C and ‐D are important for maintenance and growth of lymphatic vessels (lymphangiogenesis), but their localization in human gingiva is unknown. This study investigated the expression of VEGF‐C and ‐D in human gingiva and isolated human gingival fibroblasts (HGFs). In addition, the localization of their main receptor VEGFR‐3 was explored.

2 Methods

Non‐inflamed gingiva from six donors was used for immunohistochemistry or isolation of HGFs. HGFs were stimulated with either E.coli lipopolysaccharide (LPS) or IL‐6/soluble IL‐6 receptor (sIL‐6R) complex for 1, 6, and 24 hours. Quantitative real‐time polymerase chain reaction (qRT‐PCR) was used to quantify the relative changes in gene expression of VEGF‐A, ‐C, and ‐D and enzyme‐linked immunosorbent assay (ELISA) for quantification of protein levels.

3 Results

VEGF‐C, ‐D and VEGFR‐3 were seen in keratinocytes, blood vessels and in scattered single cells in gingiva. VEGFR‐3 was also found in lymphatic vessels and VEGF‐C in cells with fibroblastic appearance. Gene analysis showed no expression of VEGF‐D in the HGFs, but showed constitutive expression of VEGF‐C and ‐A. Stimulation of HGFs with LPS or IL‐6/sIL‐6R complex was followed by gene upregulation of VEGF‐C and ‐A and increased protein levels in cell culture supernatant (P ≤0.05).

4 Conclusions

The localization of VEGF‐C, ‐D, and VEGFR‐3 expression imply that signaling via VEGFR‐3 is linked to vascular homeostasis and keratinocyte function under normal conditions in gingiva. Inflammatory stimulation of HGFs upregulates VEGF‐C and ‐A expression and may contribute to angiogenesis and lymphangiogenesis.     

大鼠牙周组织淋巴管的酶组织化学法研究

目的 :本研究旨在应用酶组织化学双重染色法 ,对大鼠牙周组织中的淋巴管 ,在光镜和电镜下进行观察。方法 :将牙周膜伸展标本固定、超薄切片作电镜观察 ;另将含牙和牙槽骨的标本经EDTA脱矿制作组织冰冻切片 ,进行 5′ Nase染色 ,5′ Nase&ALPase(5′ 核苷酸酶和碱性磷酸酶 )双重染色。结果 :在光镜下观察该染色法的组织切片 ,可证明牙周组织中的淋巴管的存在 ,并将淋巴管与血管相区别 ,即呈现褐色的 5′ Nase阳性的淋巴管 ,呈现兰色的ALPase阳性的血管。用透射电镜可见 5′Nase反应产生的致密颗粒状沉淀物 ,存在于淋巴管内皮细胞的管腔面和基底侧 ,而血管中无此现象。结论 :5′ Nase&ALPase双重染色是研究牙周组织淋巴管的好方法。     

Vascular changes during the development of the rat gingiva

Permeability of the dento-gingival vessels was studied during the development of the gingiva in rats. Groups of animals, aged 19 to 150 days, were injected intravenously with colloidal carbon, and the gingival vessels were observed both macroscopically and microscopically. As the tooth cusps emerged into the oral cavity, vessels labelled with carbon were observed in the connective tissue overlying the tooth and, two to four days later, tracer particles were found in the vessels adjacent to the junctional epithelium. The leaking vessels had a linear pattern beneath the dento-gingival junction in young animals, while in adult rats these vessels formed loops. In the interdental papilla, labelled vessels were observed initially only in the gingiva adjacent to the distal of the first molars, and two days later also in the gingiva facing the mesial of the second molars. There was a progressive increase in gingival blood vessels labelled with carbon until the rats were 150 days old. The results indicate that the inflammatory process and the increased vascular permeability in the gingiva are initialed during tooth eruption and show quantitative and morphological modifications during the organization of the sulcular and junctional epithelia.     

Vascular endothelial growth factors and progression of periodontal diseases.

BACKGROUND: Tissues become hemorrhagic and edematous coincident to periodontal diseases; however, there is little information concerning the biologic mechanisms which may produce these changes. Vascular endothelial growth factor (VEGF) is a macromolecule which enhances blood vessel growth and permeability. However, there is no information concerning gingival VEGF concentrations within normal or diseased gingiva. The purpose of this study was to assess changes in gingival concentrations of VEGF during initiation and progression of periodontal diseases and compare them to changes in the number of blood vessel profiles and concentration of recognized markers of periodontal disease severity (interleukin-6[IL-6]). METHODS: Normal (non-hemorrhagic gingiva adjacent to a < or =3 mm gingival sulcus) and inflamed gingiva (hemorrhagic gingiva adjacent to a < or =3 mm, 4 to 6 mm, or >6 mm periodontal pocket) were studied. VEGF and IL-6 concentrations were assessed by ELISA and the number of blood vessels determined by histomorphometric techniques. Data were placed into one of the following groups: < or =3 mm, normal; < or =3 mm, diseased; 4 to 6 mm, diseased; and >6 mm, diseased. These groups were compared by factorial ANOVA and Scheffe comparisons. In addition, groups were compared by simple and multiple regression and regression ANOVA to determine possible correlations between them. RESULTS: VEGF and IL-6 concentrations were significantly lower within normal than within diseased gingiva. The number of blood vessel profiles and mean IL-6 concentrations were highest in diseased tissues adjacent to >6 mm sulci and were significantly correlated with sulcular depth (P <0.001). In contrast, VEGF concentrations were highest within diseased gingiva adjacent to 4 to 6 mm periodontal pockets (P <0.001) and were not correlated with sulcular depth. CONCLUSIONS: VEGF may be a factor in initiation and progression of gingivitis to periodontitis, possibly by promoting expansion of the vascular network coincident to progression of the inflammation.     

Vascular endothelial growth factors signalling in normal human dental pulp: a study of gene and protein expression

In the well‐vascularized dental pulp vascular endothelial growth factor A (VEGF‐A) is expressed. Vascular endothelial growth factor A is a member of the VEGF family, which includes VEGFs‐B, ‐C, and ‐D. The latter three have not been investigated in the pulp. Vascular endothelial growth factors C and D are the only ligands for vascular endothelial growth factor receptor (VEGFR)‐3, which is usually expressed in lymphatic endothelium. They can also activate VEGFR‐2, the main angiogenic receptor. We aimed to study VEGFs signalling in human dental pulp at the gene level and to identify the cellular source for protein expression using immunolabelling. All VEGFs (‐A, ‐B, ‐C, and ‐D) were expressed in the pulp and may exert both autocrine and paracrine effects in blood vessels and immune cells found to be equipped with VEGFRs‐2 and ‐3. Lymphatic vessel endothelial hyaluronan receptor‐positive macrophages, known to be involved in angiogenesis, were found in the pulp, whereas lymphatic vessels were not detected. Twenty‐six of 84 VEGF signalling genes, including VEGFR‐3, were expressed at a significantly higher level in the pulp than in the control periodontal ligament. In conclusion, the normal human pulp represents a tissue with relatively high VEGF signalling involving both immune responses and vascular activity.     

Leptin within healthy and diseased human gingiva

BACKGROUND: Plasma leptin concentrations are reported to be elevated in patients with inflammatory diseases. There is no consensus concerning the biological mechanism for this phenomenon. To date, tissue leptin concentrations have not been assessed within normal or inflamed gingiva. The purpose of this study was to assess concentrations of human leptin within healthy and diseased gingiva to define its possible role in periodontal disease progression. METHODS: Healthy (non-hemorrhagic gingiva adjacent to a < or =3 mm gingival sulcus) and inflamed gingiva (hemorrhagic gingiva adjacent to a > or =3 mm periodontal pocket) were studied. Leptin, vascular endothelial growth factor (VEGF; to assess potential vascular expansion), and interleukin-6 (IL-6; to assess periodontal disease activity and severity) concentrations were assessed within solubilized gingival biopsies by enzyme-linked immunosorbent assay. Data were grouped by sulcular depth and compared by factorial analysis of variance, regression analysis, and Scheffé comparisons. RESULTS: Leptin concentrations were highest within gingiva adjacent to a < or =3 mm sulcus and progressively declined within gingiva adjacent to a > or =3 mm sulcus. VEGF concentrations were highest within gingiva adjacent to 4 to 6 mm pockets and nearly equivalent in healthy (< or =3 mm sulcus) and severely diseased gingiva (>6 mm sulcus). IL-6 was positively correlated and leptin negatively correlated with adjacent probing depth; IL-6 concentration was significantly higher and leptin significantly lower in gingiva adjacent to >6 mm pockets compared to sites adjacent to <6 mm pockets (P<0.001). CONCLUSIONS: Human leptin is present within healthy and marginally inflamed gingiva and decreases in concentration as the adjacent probing depth increases. When leptin concentrations decreased (> or =3 mm sulcus), VEGF concentrations increased, suggesting that leptin could be released from gingiva coincident to vascular expansion. Thus, gingiva, in addition to adipose tissue, could be a source of circulating leptin in patients with periodontal disease. This possibility requires further investigation.     

人淋巴管内皮细胞的分离和鉴定

目的:分离培养人淋巴管内皮细胞(LECs),为研究淋巴管新生在淋巴管瘤复发及肿瘤淋巴结转移中的作用奠定基础。方法:HE染色和免疫组织化学法了解人淋巴管瘤的组织病理特征;胶原酶消化法从淋巴管瘤组织中分离淋巴管内皮细胞;光镜和透射电镜观察淋巴管内皮细胞的形态和超微结构;MTT法检测细胞体外增殖。结果:HE染色显示舌淋巴管瘤中具有大量扩张的管腔,免疫组织化学染色显示管腔内皮细胞表达FLT-4、和LYVE-1;体外培养的淋巴管内皮细胞呈"铺路石"样外观;在透射电子显微镜下,观察到淋巴管内皮细胞特征性细胞器如weibel-palade小体、胞质小突和质膜小泡;MTT结果显示细胞于第3天开始进入增殖活跃期,第5天进入高峰期,第6、7天进入平台期;在Ⅰ型胶原凝胶中细胞形成管腔样结构。结论:应用胶原酶消化法可以有效的从淋巴管瘤组织中分离得到淋巴管内皮细胞。     

Vascular response to modified Widman flap surgery in monkeys

Revascularization of the periodontal tissues was studied after flap surgery. Modified Widman flap procedures were performed in four young Rhesus monkeys. All premolars and molars were involved. Experimental periods covered from 1 day to 90 days. The animals were sacrificed by exsanguination and perfused through the carotid arteries with a combined solution of equal parts of India ink and 10% buffered formalin solution. After fixation and decalcification, part of the blocks were processed to obtain cleared specimens following the Spalteholtz method. The remaining blocks were processed for routine histologic examination. The findings, mainly from the cleared specimens, showed that 1 day after surgery the supraperiosteal vessels in the flap demonstrated an overt hyperemic response. Patency of the vessels in the periodontal membrane and cancellous bone was evident; however, vessels were not depicted in the exposed cortical bone, nor in the clot. Vascular continuity apical to the surgical detachment was observed. In 3 to 4 days vascular proliferation within the organizing clot was observed and anastomosis between the gingiva and cancellous bone or periodontal membrane started to develop. At 7 days anastomoses were evident and vasodilation was present throughout the tissues. At 15 days and thereafter, vascular organization started to decrease, and the only hyperemic foci remaining were associated with areas of delayed bone remodeling, cementum remodeling or persistent marginal inflammation.     

Microvascular volumes in healthy and inflamed gingiva in humans

It has been suggested that the vascular alterations found in inflamed gingiva may be of significance in the enhanced extension of the pathological process into the periodontium. The purpose of this investigation was to measured the changes in blood vascular volume occurring in gingiva with the onset of gingivitis and its resolution. Twenty-six individuals participated in this study. Gingival biopsies were taken following a 21-day experimental gingivitis, following resolution of a 21-d experimental gingivitis and during a 6-month experimental gingivitis and a 6-month period of optimal oral hygiene. A total of 126 biopsies was obtained, from which 378 sections were cut at 2 microns for stereological analysis. At low magnification, reference volumes were estimated using point counting procedures and expressed as mm3 of gingiva per mm length of vestibular gingiva, in a vestibulo-lingual plane. At higher magnification the ratio between the volume of vessels and connective tissue was calculated. The final results were expressed as mm3 of vessels per mm length of vestibular gingiva, in a vestibulo-lingual plane. The mean vessel volume expressed per unit length of vestibular gingiva ranged from 0.010 to 0.024 mm3/mm. No statistically significant differences in vascular volumes were found between inflamed and non-inflamed gingiva. It was concluded that the changes in vascular architecture during early gingivitis described in the literature had either taken place in the subjects prior to the time of experimentation or that any vascular changes (cytologic or functional) which had taken place may be compensatory for the changes in architecture described in the literature.     

Lymphangiogenesis in human dental pulp

AIM: To investigate the impact of inflammation on lymphangiogenesis in human dental pulp. METHODOLOGY: Eleven samples of dental pulp without inflammation and 11 dental pulps with moderate to intense mononuclear cell inflammatory infiltrate associated with dentine caries were selected. The streptavidin-biotin complex stain was used to detect CD31, vascular endothelial growth factor receptor-3 (VEGFR-3) and alpha-smooth muscle actin. The number of lymphatic vessels was obtained by counting the number of vessels positive for CD31 and VEGFR-3 and negative for alpha-smooth muscle actin. RESULTS: The results demonstrated that the mean number (+/-SD) of vessels positive for CD31 and VEGFR-3 (lymphatic vessels) in the group with inflammation (6.09 +/- 1.81) was statistically higher (P = 0.0123) than the mean number in the group without inflammation (3.73 +/- 2.20). CONCLUSION: Increased co-immunostaining of CD31 and VEGF-3 in vessels associated with human dental pulp inflammation occurred, which suggests lymphangiogenesis.     

Computer‐assisted morphometric analysis of lymphatic vessel changes in hamster tongue carcinogenesis

J Oral Pathol Med (2010) 39 : 518–524 Background: To characterize lymphangiogenesis in early‐stage hamster tongue carcinoma development, morphological features and spatial relationships of lymphatic vessels. Methods: Lymphatic vessels were examined histochemically, using 5′‐Nase‐ALPase enzyme and combined light and electron microscopy to measure lymphatic vessel area (LVA) and lymphatic vessel density (LVD). Results: In atypical hyperplastic tissues, LVA was found to be 1429.97 and LVD was found to be 39, in carcinoma in situ LVA was 2538.33 and LVD was 48, and in micro‐invasive carcinoma LVA was 5733.74 and LVD was 59. Increased lymphangiogenesis was seen in pre‐neoplastic states and in early‐stage oral squamous cell carcinoma (OSCC). Small regular lymphatic vessels predominated in atypical hyperplasia, and large, irregular lymphatic vessels in early‐stage OSCC. Lymphatic endothelial vessels were stretched and porous over large areas. Conclusions: Newly formed lymphatics and patulous intercellular junctions may be optimally suited for tumor cell metastasis through lymphatic channels in early‐ and middle‐phase carcinogenesis. Lymphatic capillary LVA and LVD became enlarged, and positively correlated, with malignancy, but show no correlation with 7,12‐dimethylbenz[a]anthracene‐induced time.     

Microscopic evaluation of the healing and revascularization of free gingival grafts

The objective of this study was to observe the microscopic appearance of the healing and revascularization of free gingival grafts when placed over periosteum. Seven rhesus monkeys were used to provide observation periods of 0, 2, 4, 5, 7, 8, 11, 14, 17, 21, 28, and 42 days following surgery. After dissection and excision of a split flap in the anterior region to prepare the graft bed, a mucoperiosteal graft of attached gingiva from the buccal of the premolar area was sutured in place. The animals were injected with carbon serum just before sacrifice and after preparation the histologic and cleared specimens were studied under the microscope.
The results indicate that the healing and revascularization of free gingival graft and skin grafts are similar. Cellular nutrition was maintained for the first few days by a "plasmatic" circulation. Revascularization occured initially by anastomoses between vessels of the graft bed and pre-existing vessels in the graft and later by capillary budding which penetrated the graft. Carbon serum was present in the vessels of the graft at four days. There was increasing vascular density up to seven days followed by a gradual reduction and return to a normal vascular pattern after 14 days. The graft was initially separated from the bed by a thin layer of fibrinous exudate. Fibroblasts proliferated into the area and a firm fibrous union to the periosteum was apparent between 7–11 days. Initial disintegration of most of the graft epithelium ccured. Re-epithelialization resulted from migration of new epithelium from adjacent gingiva and from surviving basal cells of the graft epithelium. The epithelium had a relatively normal appearance by 14 days although new keratinization was not apparent for 28 days. The connective tissue disintegrated in portion of some of the graft and was replaced by granulation tissue which delayed healing.     

Chronic glycine treatment inhibits ligature-induced periodontal disease in Wistar rats

OBJECTIVE: Dysregulation of immune and stress responses plays a significant role for the development and progression of inflammatory diseases, including periodontal disease. The non-essential amino acid glycine modulates immune and central nervous system (CNS) responses, and has been shown to beneficially affect tissue destructive inflammatory conditions. The purpose of this study was to test the ability of orally administered glycine to influence periodontal disease progression, as well as immune and hypothalamic-pituitary-adrenal (HPA) responses following lipopolysaccharide stimulation. METHODS: Glycine was supplied in the drinking water during the whole experiment to male Wistar rats, starting 3 days before the induction of experimental ligature-induced periodontal disease. Control rats were given tap water only. The periodontal breakdown was assessed after the ligatures had been in place for 34 days. Following intraperitonal lipopolysaccharide stimulation, concentrations of the proximal cytokines tumour necrosis factor-alpha (TNF-alpha) and interleukin-10, as well as the HPA axis-derived hormone corticosterone, were measured in blood serum. RESULTS: Orally administered glycine significantly reduced periodontal bone loss as measured by digital X-rays (p = 0.007). Bone loss was negatively correlated with increased serum glycine, whereas no significant relationship was found with TNF-alpha, interleukin-10, or corticosterone. CONCLUSION: Chronic ingestion of glycine supplied in the drinking water significantly reduced periodontal bone loss. No effect of glycine on immune and HPA-axis responses was revealed. Further studies are needed to clarify the mechanisms of action.     

Effect of inflammation on collateral circulation of the gingiva

The effect of inflammation on collateral circulation of the gingiva and the vasculature of the periodontal ligament was studied in 12 squirrel monkeys. Gingival inflammation was induced by tying silk ligature at the cervical area of mandibular bicuspids and molars to encourage plaque accumulation. At 2, 10 and 26 weeks after ligature placement the animal's vasculature was perfused with India ink and calf serum. Nonligatured bicuspids and molars from the same animals served as controls. At all time points, on all experimental tooth surfaces examined, the vascularity (% grid intersections falling over vessel lumen) of the cervical portion of the periodontal ligament was significantly higher than the controls. The number of vessels connecting vessels of the periodontal ligament with supracrestal vessels was also significantly higher. There was a tendency for the number of vessels perforating either the alveolar bone proper or the alveolar crest to increase when inflammation was induced. The results of this study demonstrate the development of collateral circulation from the periodontal ligament to the gingiva in response to inflammation and help to explain the extension of inflammation into the deeper structures of the periodontium, including the periodontal ligament.     

HGF的表达与口腔鳞癌VEGF-C表达及淋巴管生成关系的研究

目的：探讨口腔鳞状细胞癌（OSCC）中肝细胞生长因子（HGF）的表达与血管内皮生长因子-C（VEGF-C）的表达及淋巴管生成的关系。方法：纳入经病理证实的70例口腔鳞癌患者的手术切除标本,采用SP免疫组化技术检测HGF及VECF-C的表达情况并分析其与淋巴管密度（LVD）的关系,并结合临床病理参数作相关统计学分析。结果：淋巴结转移阳性病例的VEGF—C表达明显升高（P=0．001）,VEGF—C的表达与其他病理参数无相关,另外,HGF的表达与临床病理参数无相关。VEGF-C高表达组的瘤周淋巴管密度（PLD）明显高于VEGF—C低表达组（P=0．001）,HGF表达及VEGF—C低表达组与瘤内及瘤周淋巴管的密度无相关性。另外,HGF的表达与VEGF—C的表达间无相关。结论：VEGF—C的表达与口腔鳞癌淋巴结转移密切相关,可能主要通过参与诱导口腔鳞癌淋巴管生成促进淋巴结转移。     

Involvement of vascular endothelial growth factor, CD44 and CD133 in periodontal disease and diabetes: an immunohistochemical study

Aim: The aim of this study was to investigate the relationship between expression of angiogenic and regeneration markers and periodontal disease in subjects with/without diabetes mellitus.
Material and Methods: Immunohistochemical detection of vascular endothelial growth factor (VEGF), CD44 and CD133 was performed in 16 samples each of (1) healthy gingiva from non-diabetic subjects (controls), (2) gingiva from non-diabetic subjects with periodontitis, (3) gingiva from subjects with type 1 diabetes and periodontitis, (4) gingiva from subjects with type 2 diabetes and periodontitis.
Results: Diseased gingivae from patients with diabetes and periodontitis had greater clinical measures of periodontal disease than those with periodontitis only. VEGF expression was significantly enhanced in epithelial and endothelial cells from patients with periodontitis compared with controls ( p <0.05). Epithelial CD44 expression was strong in all groups, while CD44 was significantly enhanced ( p <0.05) in connective tissue cells from both diabetic groups. Epithelial and endothelial CD133 expression was comparable in all patients except those with type 2 diabetes and periodontitis, where it was not detected. Stromal CD133 expression was significantly lower in patients with type 2 diabetes and periodontitis and was increased in periodontitis patients ( p <0.05).
Conclusions: The involvement and high expression of VEGF, CD44 and CD133 in periodontal disease may predict a greater regeneration capacity of gingival tissue.     

Changes in microvascular patterns following the periodontal flap surgery

Serial changes in the microvascular pattern beneath the inner epithelium and in the periodontal connective tissue attachment on the upper incisors in adult mongrel dogs, were studied by scanning electron microscopy to elucidate the process of reconstructing the vascular architecture following mucoperiosteal flap surgery. In the early stage of wound healing, capillaries beneath the epithelium in the marginal gingiva were transformed into glomeruli, in the course of the predominantly elongation type of angiogenesis. At the junctions, newly formed sinusoidal vessels were formed in the course of predominantly sprouting type of angiogenesis, and anastomosed with vasculatures between the flap and the periodontal membrane. In these newly formed vessels, vascular glomeruli in the marginal gingiva and newly formed sinusoidal vessels at the coronal side of the junctions were transformed into a capillary network beneath the inner epithelium accompanying the epithelization. At the apical side of the junctions, newly formed sinusoidal vessels showed a series of changes to cope with alterations of the tissue environment accompanying connective tissue attachment, but the vasculatures established after surgery showed a dense and irregular arrangement compared with the control side. These results show that the attachment mechanism obtained by wound healing differ from the original one.     

舌癌细胞淋巴道转移能力与淋巴管生成的关系

目的：诱导建立小鼠淋巴管良性肿瘤模型以及异种移植瘤模型,观察不同淋巴道转移能力的舌癌细胞对淋巴管生成的影响。方法：C57BL/6小鼠腹腔注射弗氏不完全佐剂,对诱导出的小鼠腹腔淋巴管瘤进行病理组织学鉴定。诱导的淋巴管瘤在纤维蛋白凝胶内应用Tca8113和LNMTca8113细胞条件培养基培养,倒置显微镜下观察淋巴管分支形成。此外,利用这2种细胞进行体外成瘤,免疫组化染色观察淋巴管生成情况。实验结果应用SPSS11.5软件包对数据进行分组t检验,Mann-Whitney U检验。结果：实验小鼠中膈肌腹腔面、肝脏表面可见边界清楚的散在分布白色肿瘤样组织,组织学病理证实为良性淋巴管瘤。与Tca8113细胞比较,在LNMTca8113细胞条件培养基作用下,从淋巴管瘤块长入纤维蛋白凝胶内的微淋巴管分支数目增加;在LNMTca8113肿瘤中,淋巴管密度显著提高。结论：小鼠腹腔注射弗氏不完全佐剂可稳定诱导小鼠腹腔淋巴管瘤,高淋巴道转移能力的舌鳞状细胞癌细胞对淋巴管生成具有更强的促进作用,适应淋巴道转移的需要。