Evidence for dermal angiogenesis in breast cancer related lymphedema demonstrated using dual-site fluorescence angiography

OBJECTIVE: To determine whether the skin expansion associated with breast cancer related lymphedema (BCRL) reduces microvessel density or causes a compensatory angiogenesis. METHODS: Dermal microvessel density was measured by simultaneous fluorescence video angiography in the ipsilateral and contralateral forearms of 25 patients treated for breast cancer, 12 with BCRL (excess volume 30 +/- 20%) and 13 without. RESULTS: Microvessel density was as high in the swollen arm (27.2 +/- 7.2 mm(-2)) as in the contralateral arm (24.2 +/- 6.2 mm(-2)) despite a 16 +/- 12% increase in local circumference. Total microvessels in a 1-mm annulus of skin was 30% greater in the swollen arm (752 +/- 211) than the contralateral arm (578 +/- 157; p = 0.005, t-test). Microvessel density was similar in each arm in control patients (ipsilateral 24.1 +/- 6.2 mm(-2), contralateral 26.1 +/- 8.8 mm(-2)). CONCLUSIONS: The number of microvessels increased in the expanded skin to maintain microvessel density at a normal level. Therefore, angiogenesis occurs during chronic limb swelling. The control group results indicate that angiogenesis is not caused by the cancer treatment.     

Increased dermal lymphatic density in the human leg compared with the forearm.

During orthostasis, capillary filtration pressure is higher in the leg than in the arm due to the effect of gravity. We investigated the possibility that the lymphatic network in leg skin might be adapted to cope with a greater fluid load. The dermal lymphatics of the forearm and lower leg were studied in white-skinned and brown-skinned men using fluorescence video microscopy. From video print lymphangiograms the following were determined: lymphatic length density at a series of radii from the centre of the lymphangiogram (LDr); maximum lymphatic density (LDmax); total length of lymphatic vessel (LL); maximum spread of lymphatic vessel; number and size of lymphatic rings (continuous circuits of vessel); and vessel diameter. There were no differences between the two racial types, but clear differences between the arm and leg. In the leg, mean (+/- SD) peak LDr (25.13 +/- 5.65 cm-1), LDmax (32.95 +/- 6.89 cm-1), LL (40.17 +/- 27.42 cm), and spread (1.39 +/- 0.08 cm) were all significantly higher than in the arm (18.03 +/- 5.48 cm-1, 23.91 +/- 7.21 cm-1, 11.76 +/- 5.47 cm, and 1.00 +/- 0.05 cm respectively, P 相似文献   

Distal Extremity Swelling with Pitting Oedema in Rheumatoid Arthritis

Distal extremity swelling with pitting oedema due to altered lymphatic drainage has been reported in some patients with rheumatoid arthritis (RA). The resistant-to-therapy oedema usually affected the upper limbs in an asymmetrical pattern. Until now, extensor tenosynovial involvement has not been described in RA patients suffering from distal extremity swelling with pitting oedema. Three patients are described: two of them had predominant extensor tenosynovial involvement in their hands, with impaired lymphatic drainage demonstrated by (MRI) and lymphoscintigraphy, respectively. In both cases the oedema was chronic and not responsive to treatment. One patient had extensor tenosynovial involvement without impaired lymphatic drainage. In this case, the oedema remitted completely after a few days of corticosteroid therapy. None of them showed differences in serum levels of vascular endothelial growth factor (VEGF), whether they were RA patients with no pitting oedema or healthy volunteers. Received: 6 January 2000 / Accepted: 20 September 2000     

Safety study of axillary reverse mapping in the surgical treatment for breast cancer patients

Purpose  

With the purpose of minimizing arm lymphedema after axillary staging surgeries in breast cancer patients, the axillary reverse mapping (ARM) technique has been developed to identify and preserve arm drainage system during axillary surgery. This study aimed to clarify risk factors for metastasis in arm lymphatic drainage system in breast cancer patients with clinically negative axillary nodes.     

Interleukin-8 reduces post-surgical lymphedema formation by promoting lymphatic vessel regeneration

Lymphedema is mainly caused by lymphatic obstruction and manifested as tissue swelling, often in the arms and legs. Lymphedema is one of the most common post-surgical complications in breast cancer patients and presents a painful and disfiguring chronic illness that has few treatment options. Here, we evaluated the therapeutic potential of interleukin (IL)-8 in lymphatic regeneration independent of its pro-inflammatory activity. We found that IL-8 promoted proliferation, tube formation, and migration of lymphatic endothelial cells (LECs) without activating the VEGF signaling. Additionally, IL-8 suppressed the major cell cycle inhibitor CDKN1C/p57^KIP2 by downregulating its positive regulator PROX1, which is known as the master regulator of LEC-differentiation. Animal-based studies such as matrigel plug and cornea micropocket assays demonstrated potent efficacy of IL-8 in activating lymphangiogenesis in vivo. Moreover, we have generated a novel transgenic mouse model (K14-hIL8) that expresses human IL-8 in the skin and then crossed with lymphatic-specific fluorescent (Prox1-GFP) mouse. The resulting double transgenic mice showed that a stable expression of IL-8 could promote embryonic lymphangiogenesis. Moreover, an immunodeficient IL-8-expressing mouse line that was established by crossing K14-hIL8 mice with athymic nude mice displayed an enhanced tumor-associated lymphangiogenesis. Finally, when experimental lymphedema was introduced, K14-hIL8 mice showed an improved amelioration of lymphedema with an increased lymphatic regeneration. Together, we report that IL-8 can activate lymphangiogenesis in vitro and in vivo with a therapeutic efficacy in post-surgical lymphedema.     

Expression of vascular endothelial growth factor-C and the relationship between lymphangiogenesis and lymphatic metastasis in colorectal cancer

AIM: To investigate the expression of vascular endothelial growth factor-C (VEGF-C) and the relationship between VEGF-C and lymphangiogenesis, lymph node metastasis in colorectal cancer. METHODS: Fifty six cases of colorectal cancer were selected randomly. Expression of VEGF-C was detected by immunohistochemistry, and lymphatic vessels were stained by enzyme histochemical method. RESULTS: VEGF-C expression was found in 66.7% (37/56) patients. In VEGF-C positive and negative patients, the lymphatic vessel density was 25.16+/-7.52 and 17.14+/-7.22, respectively (P<0.05). The rate of lymph node metastasis in VEGF-C positive patients (81.1%) was significantly higher than that in the negative group (42.1%). CONCLUSION: VEGF-C expression may induce lymphangiogenesis in colorectal cancer, as a result, tumor cells can entry the lymphatic vessels easily. VEGF-C may serve as a useful prognotic factor in colorectal carcinoma.     

Lymphatic microangiopathy of the skin in systemic sclerosis.

METHODS: The cutaneous capillary lymphatic system in patients with systemic sclerosis was investigated using fluorescence microlymphography. The distal upper limbs of 16 healthy controls (mean age 62.3+/-13.1 yr) and 16 patients with systemic sclerosis (mean age 58.9+/-13.6 yr) were examined and the following parameters were evaluated: (a) single lymphatic capillaries; (b) lymphatic capillary network and cutaneous backflow; (c) extension of the stained lymphatics; (d) diameter of single lymphatic capillaries. RESULTS: At the finger level, lymphatic capillaries were lacking in five patients, while they were present in all controls (P < 0.05). Extension of the stained lymphatics was increased in 11 patients (8.1+/-6.0 mm) compared to the 16 healthy controls (2.0+/-1.2 mm) (P < 0.0001). Cutaneous backflow was observed in three patients (P < 0.05). At the hand level, lymphatic network extension was significantly different between patients (3.8+/-2.4 mm) and controls (1.2+/-0.8 mm) (P < 0.01); however, no significant differences were found at the forearm level. CONCLUSION: Lesional skin in patients with systemic sclerosis exhibits evidence of lymphatic microangiopathy.     

Potential therapeutic strategies for lymphatic metastasis

Physiologically, the lymphatic system regulates fluid volume in the interstitium and provides a conduit for immune cells to travel to lymph nodes, but pathologically, the lymphatic system serves as a primary escape route for cancer cells. Lymphatic capillaries have a thin discontinuous basement membrane, lack pericyte coverage and often contain endothelial cell gaps that can be invaded by immune cells (or tumor cells). In addition, tumor cells and stromal cells in the tumor microenvironment secrete factors that stimulate lymphangiogenesis, the growth of lymphatic endothelial cells and the sprouting of lymphatic capillaries. As a result, many tumors are surrounded by large, hyperplastic, peri-tumoral lymphatic vessels and less frequently are invaded by intra-tumoral lymphatic vessels. Carcinoma cells commonly metastasize through these lymphatic vessels to regional lymph nodes. The presence of metastatic cells in the sentinel lymph node is a prognostic indicator for many types of cancer, and the degree of dissemination determines the therapeutic course of action. Lymphangiogenesis is currently at the frontier of metastasis research. Recent strides in this field have uncovered numerous signaling pathways specific for lymphatic endothelial cells and vascular endothelial cells. This review will provide an overview of tumor lymphangiogenesis and current strategies aimed at inhibiting lymphatic metastasis. Novel therapeutic approaches that target the tumor cells as well as the vascular and lymphatic endothelial compartments are discussed.     

Two cases of distal extremity swelling with pitting oedema in psoriatic arthritis: the different pathological mechanisms

In psoriatic arthritis, swelling and pitting oedema may be caused by different pathogenic mechanisms: on one hand, the involvement of tenosynovial structures; on the other hand, the involvement of lymphatic vessels, which may be rarely implicated by the inflammatory process. This different involvement is responsible for a different response to therapy and a different clinical outcome. In fact, patients with inflammation of the tenosynovial structures and normal lymphatic drainage have a more favourable clinical outcome and response to pharmacologic treatment, whilst patients affected by psoriatic arthritis with chronic lymphatic vascular damage are characterized usually by resistance of oedema to therapy. In this study, we report two cases of psoriatic arthritis with distal extremity swelling and pitting oedema. In the first patient, the swelling and pitting oedema were associated with lymphatic obstruction, as detected by lymphoscintigraphy. In the second, the predominant involvement of the tenosynovial structures, as shown by magnetic resonance, with normal lymphatic flow, may have been the cause of arthritis with oedema. These different pathogenetic mechanisms were associated with different response to therapy. Nevertheless, oedema was resistant to therapy in both patients probably because of other unknown factors, which influence therapy and clinical outcome.     

The importance of volumetry, lymphscintigraphy and computer tomography in the diagnosis of brachial edema after mastectomy

Arm volumes of 360 patients with breast cancer were determined by a new optoelectronic technique. About 42% developed swelling of the arm after modified radical mastectomy which varied from mild (edema-volume 150-400 ml), moderate (400-800 ml), to severe (more than 800 ml) lymphatic edema. Lymphscintigraphy and computer tomography of the arms was also studied in different grades of lymphatic edema. Two years after operation and telecobalt-irradiation, one half of the patients without arm edema showed marked signs of a decreased capacity of lymphatic transport. In patients with severe lymphatic edema, the dynamic as well as the static evaluation of the lymphscintigrams revealed delayed transport, lack of radioisotope accumulation in the axillary region and notable congestion in the upper arm and forearm. Computerized tomography displayed a shift of fluid volume in the epifascial and subfascial tissue compartments and detected progressive structural changes in the soft tissue. Moderate edema not only increased the epifascial but also the subfascial compartments. In severe postmastectomy arm lymphedema, however, expansion of the epifascial space was paradoxically sometimes associated with a decrease in the subfascial compartment.     

Potential therapeutic strategies for lymphatic metastasis

Physiologically, the lymphatic system regulates fluid volume in the interstitium and provides a conduit for immune cells to travel to lymph nodes, but pathologically, the lymphatic system serves as a primary escape route for cancer cells. Lymphatic capillaries have a thin discontinuous basement membrane, lack pericyte coverage and often contain endothelial cell gaps that can be invaded by immune cells (or tumor cells). In addition, tumor cells and stromal cells in the tumor microenvironment secrete factors that stimulate lymphangiogenesis, the growth of lymphatic endothelial cells and the sprouting of lymphatic capillaries. As a result, many tumors are surrounded by large, hyperplastic, peri-tumoral lymphatic vessels and less frequently are invaded by intra-tumoral lymphatic vessels. Carcinoma cells commonly metastasize through these lymphatic vessels to regional lymph nodes. The presence of metastatic cells in the sentinel lymph node is a prognostic indicator for many types of cancer, and the degree of dissemination determines the therapeutic course of action. Lymphangiogenesis is currently at the frontier of metastasis research. Recent strides in this field have uncovered numerous signaling pathways specific for lymphatic endothelial cells and vascular endothelial cells. This review will provide an overview of tumor lymphangiogenesis and current strategies aimed at inhibiting lymphatic metastasis. Novel therapeutic approaches that target the tumor cells as well as the vascular and lymphatic endothelial compartments are discussed.     

Modeling lymphangiogenesis: Pairing in vitro and in vivo metrics

Lymphangiogenesis is the mechanism by which the lymphatic system develops and expands new vessels facilitating fluid drainage and immune cell trafficking. Models to study lymphangiogenesis are necessary for a better understanding of the underlying mechanisms and to identify or test new therapeutic agents that target lymphangiogenesis. Across the lymphatic literature, multiple models have been developed to study lymphangiogenesis in vitro and in vivo. In vitro, lymphangiogenesis can be modeled with varying complexity, from monolayers to hydrogels to explants, with common metrics for characterizing proliferation, migration, and sprouting of lymphatic endothelial cells (LECs) and vessels. In comparison, in vivo models of lymphangiogenesis often use genetically modified zebrafish and mice, with in situ mouse models in the ear, cornea, hind leg, and tail. In vivo metrics, such as activation of LECs, number of new lymphatic vessels, and sprouting, mirror those most used in vitro, with the addition of lymphatic vessel hyperplasia and drainage. The impacts of lymphangiogenesis vary by context of tissue and pathology. Therapeutic targeting of lymphangiogenesis can have paradoxical effects depending on the pathology including lymphedema, cancer, organ transplant, and inflammation. In this review, we describe and compare lymphangiogenic outcomes and metrics between in vitro and in vivo studies, specifically reviewing only those publications in which both testing formats are used. We find that in vitro studies correlate well with in vivo in wound healing and development, but not in the reproductive tract or the complex tumor microenvironment. Considerations for improving in vitro models are to increase complexity with perfusable microfluidic devices, co-cultures with tissue-specific support cells, the inclusion of fluid flow, and pairing in vitro models of differing complexities. We believe that these changes would strengthen the correlation between in vitro and in vivo outcomes, giving more insight into lymphangiogenesis in healthy and pathological states.     

Interstitial flow as a guide for lymphangiogenesis

The lymphatic system is important in tissue fluid balance regulation, immune cell trafficking, edema, and cancer metastasis, yet very little is known about the sequence of events that initiate and coordinate lymphangiogenesis. Here, we characterize the process of lymphatic regeneration by uniquely correlating interstitial fluid flow and lymphatic endothelial cell migration with lymphatic function. A new model of skin regeneration using a collagen implant in a mouse tail has been developed, and it shows that (1) interstitial fluid channels form before lymphatic endothelial cell organization and (2) lymphatic cell migration, vascular endothelial growth factor-C expression, and lymphatic capillary network organization are initiated primarily in the direction of lymph flow. These data suggest that interstitial fluid channeling precedes and may even direct lymphangiogenesis (in contrast to blood angiogenesis, in which fluid flow proceeds only after the vessel develops); thus, a novel and robust model is introduced for correlating molecular events with functionality in lymphangiogenesis.     

VEGF-C induced angiogenesis preferentially occurs at a distance from lymphangiogenesis

AIMS: Vascular endothelial growth factor-C (VEGF-C) has been shown to stimulate both angiogenesis and lymphangiogenesis in some but not all models where VEGF-C is over-expressed. Our aim was to investigate the interaction between lymphangiogenesis and angiogenesis in adult tissues regulated by VEGF-C and identify evidence of polarized growth of lymphatics driven by specialized cells at the tip of the growing sprout. METHODS AND RESULTS: We used an adult model of lymphangiogenesis in the rat mesentery. The angiogenic effect of VEGF-C was markedly attenuated in the presence of a growing lymphatic network. Furthermore, we show that this growth of lymphatic vessels can occur both by recruitment of isolated lymphatic islands to a connected network and by filopodial sprouting. The latter is independent of polarized tip cell differentiation that can be generated all along lymphatic capillaries, independently of the proliferation status of the lymphatic endothelial cells. CONCLUSION: These results both demonstrate a dependence of VEGF-C-mediated angiogenesis on lymphatic vascular networks and indicate that the mechanism of VEGF-C-mediated lymphangiogenesis is different from that of classical angiogenic mechanisms.     

Physiopathology of lymphedema

Edema is defined as an excess of interstitial fluids. Its appearance is always linked to a relative functional insufficiency of the lymphatic drainage, but the use of the word lymphedema is restricted to the conditions where the lymphatic insufficiency is the primary pathogenetic factor of edema formation. The lymphatic insufficiency may result from: 1) an incompetence of the lymphatic capillaries, either due to their aplasia (Milroy's disease) or to their destruction (lipodermatosclerosis of chronic venous insufficiency); 2) an insufficiency of the lymphatic collectors related to their hypoplasia (primary lymphedema), or their destruction by trauma, infection or carcinologic treatments; a drug related functional insufficiency of lymphatic collectors is probable, but more data are needed on this topic; 3) pathologic lymphatic nodes (surgical excision, invasion by hematological disorders or cancer) could interfere with their ability to concentrate lymphatic fluids. Local tissue changes in the lymphedematous limbs are important and should be taken into account in the care management of such patients.     

Severe lymphedema of the arm as a potential cause of shoulder trauma

The aim of this study was to determine whether lymphedema of the arm is associated with traumatic injury to the shoulder and to assess the role of lymphatic physiotherapy in reducing disabling shoulder pain. The study group consisted of 10 women aged 58-81 years (mean 66.9) with arm lymphedema after surgery for breast cancer. The average interval between the operation and the appearance of lymphedema was 9.8 years. All patients complained of shoulder pain. Five patients had a tear in the supraspinatus muscle diagnosed by ultrasound examination, and 5 had chronic bursitis; the nonaffected arm showed no pathology. The mean volume of the affected arm was 568 ml greater. Treatment consisted of manual lymphatic drainage and intermittent sessions of pneumatic compression with the LymphaPress device. This led to an average decrease in arm volume of 170 ml, with improvement of arm mobility and a drastic reduction in shoulder pain. In conclusion, lymphedema of the arm can cause severe shoulder trauma, pain and disability. Proper physiotherapy can reduce these effects. Patients should be referred for early treatment and follow-up to avoid permanent damage to the shoulder muscles.     

诱导型一氧化氮合酶、环氧合酶-2与乳腺癌淋巴管生成的关系及其临床意义

目的探讨诱导型一氧化氮合酶(iNOS)和环氧合酶-2(COX-2)在乳腺癌间质淋巴管生成中的作用机制及其临床意义。方法收集乳腺浸润性导管癌组织90例和乳腺纤维腺瘤组织30例,应用寡核苷酸探针原位杂交法检测组织中iNOS和COX-2的mRNA表达,另以Elivision免疫组化法检测淋巴管内皮细胞透明质酸受体1(LYVE-1)和血管内皮生长因子-C(VEGF-C)蛋白的表达情况,分析iNOS和COX-2与LYVE-1标记的淋巴管密度(LVD)以及VEGF-C的关系。结果在90例乳腺癌中iNOS、COX-2的mRNA阳性率分别为66.7%、68.9%,LVD为(8.03±2.26)个/HPF,VEGF-C表达阳性率为47.8%,与良性对照组比较均有显著差异(P<0.01)。iNOS mRNA表达与乳腺癌肿瘤大小、分期、LVD和VEGF-C表达呈正相关(P<0.01);COX-2 mRNA表达与肿瘤分级、淋巴结转移、分期和LVD呈正相关(P<0.05),与VEGF-C表达无显著相关(P>0.05);iNOS和COX-2的mRNA阳性表达呈正相关(P<0.05)。结论 iNOS和COX-2具有协同促进乳腺癌淋巴管生成及侵袭转移的作用,有望在乳腺癌的临床预后判断及靶向治疗中发挥作用。     

Secondary lymphedema in the mouse tail: Lymphatic hyperplasia, VEGF-C upregulation, and the protective role of MMP-9

Disturbances in the microcirculation can lead to secondary lymphedema, a common pathological condition that, despite its frequency, still lacks a cure. Lymphedema is clinically well described, but while the genetic underpinnings that cause lymphatic malformations and primary lymphedema are being discovered, the pathophysiology and pathobiology of secondary lymphedema remain poorly understood, partly due to the lack of well-described experimental models. Here, we provide a detailed characterization of secondary lymphedema in the mouse tail and correlate the evolution of tissue swelling to changes in tissue architecture, infiltration of immune cells, deposition of lipids, and proliferation and morphology of the lymphatic vessels. We show that sustained swelling leads to lymphatic hyperplasia and upregulation of vascular endothelial growth factor (VEGF)-C, which may exacerbate the edema because the hyperplastic vessels are poorly functional. The onset of lymphatic hyperplasia occurred prior to the onset of lipid accumulation and peak VEGF-C expression. Langerhans dendritic cells were seen in the dermis migrating from the epidermis to the lymphatic capillaries in edematous tissue. Furthermore, these results were consistent between two different normal mouse strains, but swelling was significantly greater in a matrix metalloproteinase (MMP)-9 null strain. Thus, by characterizing this highly reproducible model of secondary lymphedema, we conclude that VEGF-C upregulation and lymphatic hyperplasia resulting from dermal lymphatic ligation and lymphedema leads to decreased drainage function and that MMP-9 may be important in counteracting tissue swelling.     

Secondary lymphedema in the mouse tail: Lymphatic hyperplasia,VEGF-C upregulation,and the protective role of MMP-9

Disturbances in the microcirculation can lead to secondary lymphedema, a common pathological condition that, despite its frequency, still lacks a cure. Lymphedema is clinically well described, but while the genetic underpinnings that cause lymphatic malformations and primary lymphedema are being discovered, the pathophysiology and pathobiology of secondary lymphedema remain poorly understood, partly due to the lack of well-described experimental models. Here, we provide a detailed characterization of secondary lymphedema in the mouse tail and correlate the evolution of tissue swelling to changes in tissue architecture, infiltration of immune cells, deposition of lipids, and proliferation and morphology of the lymphatic vessels. We show that sustained swelling leads to lymphatic hyperplasia and upregulation of vascular endothelial growth factor (VEGF)-C, which may exacerbate the edema because the hyperplastic vessels are poorly functional. The onset of lymphatic hyperplasia occurred prior to the onset of lipid accumulation and peak VEGF-C expression. Langerhans dendritic cells were seen in the dermis migrating from the epidermis to the lymphatic capillaries in edematous tissue. Furthermore, these results were consistent between two different normal mouse strains, but swelling was significantly greater in a matrix metalloproteinase (MMP)-9 null strain. Thus, by characterizing this highly reproducible model of secondary lymphedema, we conclude that VEGF-C upregulation and lymphatic hyperplasia resulting from dermal lymphatic ligation and lymphedema leads to decreased drainage function and that MMP-9 may be important in counteracting tissue swelling.