Ramin Shayan

Sox18 induces development of the lymphatic vasculature in mice

The lymphatic system plays a key role in tissue fluid regulation and tumour metastasis, and lymphatic defects underlie many pathological states including lymphoedema, lymphangiectasia, lymphangioma and lymphatic dysplasia. However, the origins of the lymphatic system in the embryo, and the mechanisms that direct growth of the network of lymphatic vessels, remain unclear. Lymphatic vessels are thought to arise from endothelial precursor cells budding from the cardinal vein under the influence of the lymphatic hallmark gene Prox1 (prospero homeobox 1; ref. 4). Defects in the transcription factor gene SOX18 (SRY (sex determining region Y) box 18) cause lymphatic dysfunction in the human syndrome hypotrichosis-lymphoedema-telangiectasia, suggesting that Sox18 may also play a role in lymphatic development or function. Here we use molecular, cellular and genetic assays in mice to show that Sox18 acts as a molecular switch to induce differentiation of lymphatic endothelial cells. Sox18 is expressed in a subset of cardinal vein cells that later co-express Prox1 and migrate to form lymphatic vessels. Sox18 directly activates Prox1 transcription by binding to its proximal promoter. Overexpression of Sox18 in blood vascular endothelial cells induces them to express Prox1 and other lymphatic endothelial markers, while Sox18-null embryos show a complete blockade of lymphatic endothelial cell differentiation from the cardinal vein. Our findings demonstrate a critical role for Sox18 in developmental lymphangiogenesis, and suggest new avenues to investigate for therapeutic management of human lymphangiopathies.

Lymphangiogenesis and lymphatic vessel remodelling in cancer

The generation of new lymphatic vessels through lymphangiogenesis and the remodelling of existing lymphatics are thought to be important steps in cancer metastasis. The past decade has been exciting in terms of research into the molecular and cellular biology of lymphatic vessels in cancer, and it has been shown that the molecular control of tumour lymphangiogenesis has similarities to that of tumour angiogenesis. Nevertheless, there are significant mechanistic differences between these biological processes. We are now developing a greater understanding of the specific roles of distinct lymphatic vessel subtypes in cancer, and this provides opportunities to improve diagnostic and therapeutic approaches that aim to restrict the progression of cancer.

VEGF-D Promotes Tumor Metastasis by Regulating Prostaglandins Produced by the Collecting Lymphatic Endothelium

Lymphatic metastasis is facilitated by lymphangiogenic growth factors VEGF-C and VEGF-D that are secreted by some primary tumors. We identified regulation of PGDH, the key enzyme in prostaglandin catabolism, in endothelial cells of collecting lymphatics, as a key molecular change during VEGF-D-driven tumor spread. The VEGF-D-dependent regulation of the prostaglandin pathway was supported by the finding that collecting lymphatic vessel dilation and subsequent metastasis were affected by nonsteroidal anti-inflammatory drugs (NSAIDs), known inhibitors of prostaglandin synthesis. Our data suggest a control point for cancer metastasis within the collecting lymphatic endothelium, which links VEGF-D/VEGFR-2/VEGFR-3 and the prostaglandin pathways. Collecting lymphatics therefore play an active and important role in metastasis and may provide a therapeutic target to restrict tumor spread.

A role for bone morphogenetic protein-4 in lymph node vascular remodeling and primary tumor growth

Lymph node metastasis, an early and prognostically important event in the progression of many human cancers, is associated with expression of VEGF-D. Changes to lymph node vasculature that occur during malignant progression may create a metastatic niche capable of attracting and supporting tumor cells. In this study, we sought to characterize molecules expressed in lymph node endothelium that could represent therapeutic or prognostic targets. Differential mRNA expression profiling of endothelial cells from lymph nodes that drained metastatic or nonmetastatic primary tumors revealed genes associated with tumor progression, in particular bone morphogenetic protein-4 (BMP-4). Metastasis driven by VEGF-D was associated with reduced BMP-4 expression in high endothelial venules, where BMP-4 loss could remodel the typical high-walled phenotype to thin-walled vessels. VEGF-D expression was sufficient to suppress proliferation of the more typical BMP-4-expressing high endothelial venules in favor of remodeled vessels, and mechanistic studies indicated that VEGF receptor-2 contributed to high endothelial venule proliferation and remodeling. BMP-4 could regulate high endothelial venule phenotype and cellular function, thereby determining morphology and proliferation responses. Notably, therapeutic administration of BMP-4 suppressed primary tumor growth, acting both at the level of tumor cells and tumor stromal cells. Together, our results show that VEGF-D-driven metastasis induces vascular remodeling in lymph nodes. Furthermore, they implicate BMP-4 as a negative regulator of this process, suggesting its potential utility as a prognostic marker or antitumor agent.

A system for quantifying the patterning of the lymphatic vasculature

The lymphatic vasculature is critical for immunity and interstitial fluid homeostasis, playing important roles in diseases such as lymphedema and metastatic cancer. Animal models have been generated to explore the role of lymphatics and lymphangiogenic growth factors in such diseases, and to study lymphatic development. However, analysis of lymphatic vessels has primary been restricted to counting lymphatics in two-dimensional tissue slices, due to a lack of more sophisticated methodologies. In order to accurately examine lymphatic dysfunction in these models, and analyse the effects of lymphangiogenic growth factors on the lymphatic vasculature, it is essential to quantify the morphology and patterning of the distinct lymphatic vessels types in three-dimensional tissues. Here, we describe a method for performing such analyses, integrating user-operated image-analysis software with an approach that considers important morphological, anatomical and patterning features of the distinct lymphatic vessel subtypes. This efficient, reproducible technique is validated by analysing healthy and pathological tissues.

Lymphatic vessel density in primary melanomas predicts sentinel lymph node status and risk of metastasis

Shayan R, Karnezis T, Murali R, Wilmott J S, Ashton M W, Taylor G I, Thompson J F, Hersey P, Achen M G, Scolyer R A & Stacker S A 
(2012) Histopathology 61, 702–710

Adipose-derived stem cells in radiotherapy injury: a new frontier.

Radiotherapy is increasingly used to treat numerous human malignancies. In addition to the beneficial anti-cancer effects, there are a series of undesirable effects on normal host tissues surrounding the target tumor. While the early effects of radiotherapy (desquamation, erythema, and hair loss) typically resolve, the chronic effects persist as unpredictable and often troublesome sequelae of cancer treatment, long after oncological treatment has been completed. Plastic surgeons are often called upon to treat the problems subsequently arising in irradiated tissues, such as recurrent infection, impaired healing, fibrosis, contracture, and/or lymphedema. Recently, it was anecdotally noted – then validated in more robust animal and human studies – that fat grafting can ameliorate some of these chronic tissue effects. Despite the widespread usage of fat grafting, the mechanism of its action remains poorly understood. This review provides an overview of the current understanding of: (i) mechanisms of chronic radiation injury and its clinical manifestations; (ii) biological properties of fat grafts and their key constituent, adipose-derived stem cells (ADSCs); and (iii) the role of ADSCs in radiotherapy-induced soft-tissue injury.

Autologous fat grafting: current state of the art and critical review.

To the Editor: We read with interest the article entitled ‘‘Autologous fat grafting: current state of the art and critical review.’’1 We would like to congratulate Dr Richard J. Ross, Dr Mark W. Ashton, and colleagues for their publication, reviewing scientific databases to critically appraise the current body of literature in fat grafting, providing a framework to guide application and comparison. We believe that a systematic elaboration analyzing donor site, effect of infiltration solution, harvest method, effect of centrifugation, reinjection method, supplementation, role of adipose-derived stem cells, and scaffolding encourages a scientific debate. Considering all currently available reinjection methods listed in the article, we would like to add our personal experience using an 18-gauge angiographic sharp needle (Cordis Corp, Bridgewater, NJ) as published first time in 2008. Our studies gave evidence of better results in treatment of burn scars, surgical scars, hard-to-heal wounds, and radiodystrophic tissues. These conditions are all characterized by hard fibrous tissue. Performing fat grafting using the sharp angiographic cannula, compared with the blunt ones, we achieved better results. Because of needle cannula, we can overcome tissue resistance, thus obtaining better control in fat depositioning and ‘‘creating space’’ for fat transplant. Our technique has 2 key points, namely, multiple skin accesses breaking scar tissue and retrograde fat injection into the scar tissue. By multiple skin accesses, we obtain scar release, stimulating new collagen deposition and remodeling scar fibrous tissue as demonstrated in our studies. Histologic examination of scar tissue punch biopsies showed patterns of new collagen deposition, local hypervascularity, and dermal hyperplasia in the context of new tissue. Treated tissue magnetic resonance imaging showed optimal fat survival too. By retrograde fat replacement, we avoid intravascular fat injection, leading us to have low complications incidence, and we obtain lower cysts formation, increasing surface between graft and recipient bed. In conclusion, we believe that our technique can be considered in a systematic review about state of the art of fat grafting, our clinical results support autologous fat transplant using an 18-gauge angiographic sharp needle in selected cases as we demonstrated.

Vascular Endothelial Growth Factor-d Modulates Caliber and Function of Initial Lymphatics in the Dermis

The lymphatic vasculature is important for skin biology as it maintains dermal fluid homeostasis. However, the molecular determinants of the form and function of the lymphatic vasculature in skin are poorly understood. Here, we explore the role of vascular endothelial growth factor-d (Vegf-d), a lymphangiogenic glycoprotein, in determining the form and function of the dermal lymphatic network, using Vegf-d-deficient mice. Initial lymphatic vessels in adult Vegf-d-deficient mice were significantly smaller than wild-type but collecting lymphatics were unaltered. The uptake/transport of dextran in initial lymphatics of Vegf-d-deficient mice was far less efficient, indicating compromised function of these vessels. The role of Vegf-d in modulating initial lymphatics was further supported by delivery of Vegf-d in skin of wild-type mice, which promoted enlargement of these vessels. Vegf-d-deficient mice were subjected to cutaneous wounding to challenge lymphatic function: the resulting wound epithelium was highly edematous and thicker, reflecting inadequate lymphatic drainage. Unexpectedly, myofibroblasts were more abundant in Vegf-d-deficient wounds leading to faster wound closure, but resorption of granulation tissue was compromised suggesting poorer-quality healing. Our findings demonstrate that Vegf-d deficiency alters the caliber of initial lymphatics in the dermis leading to reduced functional capacity.

Preoperative radiation and free flap outcomes for head and neck reconstruction: a systematic review and meta-analysis.

There is a general consensus among reconstructive surgeons that preoperative radiotherapy is associated with a higher risk of flap failure and complications in head and neck surgery. Opinion is also divided regarding the effects of radiation dose on free flap outcomes and timing of preoperative radiation to minimize adverse outcomes. Our meta‐analysis will attempt to address these issues.