Lymphangiogenesis and genetics in lymphedemas: a review of the literature

Abstracts

O estudo do genoma humano propiciou recentes descobertas de genes e de complexos mecanismos de controle da linfangiogênese. Neste artigo esses conhecimentos são revistos, com suas implicações na embriogênese e desenvolvimento do sistema linfático e na etiopatogenia de diferentes formas e síndromes de linfedema hereditário. Algumas doenças linfáticas de transmissão genética e síndromes de aneuploidia são descritas nas suas características genotípicas e fenotípicas. Os avanços na compreensão do crescimento e desenvolvimento dos vasos linfáticos devem trazer novas alternativas terapêuticas nas linfangiodisplasias e no controle da disseminação linfática dos tumores.

Linfedema; genética; linfangiogênese

REVIEW ARTICLE

Lymphangiogenesis and genetics in lymphedemas: a review of the literature

Mauro Andrade

Department of Surgery, Faculdade de Medicina, Universidade de São Paulo (USP), São Paulo, SP, Brazil

Correspondence

ABSTRACT

Recent advances in the exploration and manipulation of the human genome provided new insights into the intricate genetics and control of lymphangiogenesis. Implications for embryogenesis and development of the lymphatic system and its role in some familial and aneuploid syndromes are reviewed, along with their genotypic and phenotypic characteristics. Increased understanding of growth and development of the lymphatic vessels may bring new therapeutic options for lymphatic angiodysplasias and control of the lymphatic spread of tumors.

Keywords: Lymphedema, genetics, lymphangiogenesis.

Introduction

Although the first correlation between heredity and lymphedema dates from the 19th century,¹ only in the past years there has been a significant advance in the understanding of the mechanisms of genetic transmission and of the genes involved in lymphangiogenesis.

An attractive hypothesis in the past associated lymphatic malformations with a disruption like "a key, a lock," which means that, if a single effective mechanism or its specific receptor were targeted by mutations, there would be specific disorders in the genesis of lymphatic structures. If this were the case, the identification of this mutation could create a wide field of possibilities in diagnostic and therapeutic management. However, regulation of lymphangiogenesis involves complex and multiform mechanisms, depending on the perfect integration of several signaling proteins and on proper functioning of cell receptors, creating many possibilities of imperfect lymphangiogenesis and phenotypical heterogeneity, related to different altered mechanisms.

Until the present time, the classification of lymphatic disease follows, above all, the clinical characteristics related to one of its most visible manifestations: edema. Over the next years there will certainly be a change in the interpretation of lymphatic insufficiencies in the sense of understanding them based on their pathophysiological and etiopathogenic aspects, resulting from a better understanding of the intimate mechanisms of lymphangiogenesis. This study aims at reviewing the current literature on this issue, showing the relevant data under the perspective of clinical lymphology and its therapeutic applications in future selected cases.

Lymphangiogenesis

The lymphatic system occurs in the sixth or seventh week of embryonic development,² about 4 weeks after the appearance of the first components of blood circulation. In the early 20th century, Sabin's³ and Huntington's⁴ theories were the reason for many discussions in anatomy societies at that time. The development of the lymphatic system in the embryo was studied in live animals through serial sections and injection methods. These studies showed that lymphatic sacs are the origin of the system in birds and mammals (including humans) and that their development occurs in a close association with the venous system. According to its specific location, lymphatic sacs are classified as jugular, subclavian, posterior and retroperitoneal, in addition to cisterna chyli.

Historically, the most widely accepted theory is Sabin's,³ suggesting that early in fetal development, primitive lymphatic sacs are originated by formation of endothelial cells from embryo veins. According to the centrifugal theory, peripheral lymphatic vessels spread throughout tissues and organs from these primitive lymphatic sacs and give origin to lymphatic capillaries.

The centripetal theory by Huntington & McClure,⁴ on the other hand, suggested that primitive lymphatic vessels formed in the mesenchyma from independent lymphatic angioblasts of veins, centripetally growing and later establishing connections with veins.

Although recent reports⁵ support Sabin's centrifuge theory, the existence of primitive lymphatic angioblasts in the mesenchyma has been demonstrated in birds.⁶ Perhaps both theories can exist in combination with lymphatic vessels from the venous endothelium, anastomosing with other lymphatic vessels developed from tissue mesenchymal cells.

Development regulation of the lymphatic and blood systems is performed by a series of cell signalers and receptors. The most important signalers belong to a family of glycoproteins: the vascular endothelial growth factor (VEGF). The VEGF are primary regulators of endothelial proliferation, angiogenesis, vasculogenesis, and vascular permeability.⁷ They are divided into VEGF A, B, C, D, and E, which bond to membrane-specific tyrosine kinase receptors, the vascular endothelial growth factor receptor 1 (VEGFR-1/Flt1), 2 (VEGFR-2/Flk1/KDR) and 3 (VEGFR-3/Flt4) (Figure 1). Activation of the VEGFR-2 by the VEGF is considered as the main path to angiogenesis and mitogenesis of endothelial cells.

Changes in genes that codify VEGF or any of the three receptors (VEGFR-1/Flt1, VEGFR-2/Flk1/KDR or VEGFR-3/Flt4) result in embryonic death due to lack of development of blood vessels.⁹

Experiments have shown that the exaggerated expression of these signaling and receptor paths is translated into hyperplasia of the lymphatic system, whereas their inhibition leads to lymphatic hypoplasia.

Expression of the VEGFR-3 gene starts early during the rat embryonic development, and VEGFR-3 deficient embryos die during pregnancy due to defects in the remodeling of primary blood vascular networks.⁹ In adult tissues, VEGFR-3 expression mainly occurs in the lymphatic endothelium.¹⁰ The VEGFR-3 bonds to two known members of the VEGF-C and VEGF-D signaling family and plays a major role in remodeling of embryo vessels. Later, during development, the VEGFR-3 regulates growth and maintenance of lymphatic vessels. Independent stimulation of the VEGFR-3 stimulates growth and migration of the lymphatic endothelium.

Neuropilins 1 and 2 (NRP-1/2) are other receptors with high affinity for the VEGF, found in the membranes of endothelial and neuronal cells.¹¹ NRP-2 also bonds to the VEGF-C to activate the VEGFR-3 in the lymphatic endothelium.¹²

Angiopoietins (Ang) 1 to 4 also have important activity in vascular remodeling and act through the tyrosine kinase receptor Tie-2. The role of Tie-2 is unknown. While Ang-1 and Ang-4 activate the Tie-2 receptor, the Ang-2 and Ang-3 act inhibiting the action of the Ang-1.²

Genetics and lymphatic angiodysplasias

Milroy described a disease, identified soon after birth, characterized by lower limb edema and not followed by other constitutional symptoms, in a group of 22 patients, from a family of 97 individuals, throughout six generations.¹ Currently the database Online Mendelian Inheritance in Man (OMIN), of Johns Hopkins University, lists 54 syndromes of genetic transmission, many of them with identified genomic change, in which the lymphedema is one of the phenotype aspects.¹³

Gene-related diseases can be divided into two groups: chromosomal aneuploidy and hereditary diseases; their most common representatives are listed in Table 1.¹⁴

Aneuploidies

Among the aneuploidies related to lymphatic system disorders, the most common is Turner syndrome (45X0), present in 1/2,500 and 1/3,000 of live-born fetuses.¹⁵ Most XO embryos (about 99%) die during pregnancy, and findings like fetal hydropsy and voluminous cervical cystic hygromas are very often found,¹⁶ suggesting that the lack of sexual chromosomes causes loss of genes related to lymphangiogenesis, essential for fetus survival. It is believed that the genes involved in Turner syndrome phenotype are homologous present in the missing chromosome, which no longer inhibit their corresponding alleles in the existing chromosome. The haploinsufficiency of the SHOX gene has been proved to account for the low height presented by these patients and some other skeletal anomalies.¹⁷ A gene related to lymphatic dysplasias has not been identified yet. On the other hand, an interesting aspect of the peripheral edema in this syndrome is its frequent regression with the child's growth.

Other aneuploidies that have lymphatic system disorder include trisomies of chromosomes 13, 18, 21 and 22, in addition to some chromosomal deletions and duplications.

Hereditary diseases

The lymphangiogenesis disorder present in some hereditary diseases can be manifested as lymphedema, although structural disorders of the lymphatic system can be much more frequent than presence of edema.¹⁸ The phenotype in most of these diseases has bilateral lymphedema of the lower limbs, restricted to the infragenicular region and observed soon after birth. Other lymphatic system defects may occasionally be associated, such as upper limb, genitalia and face edema, and a large amount of associated malformations (cardiovascular, bone, neurological, etc.).

Milroy edema (type I hereditary lymphedema - OMIM 153100)

Milroy disease is transmitted by dominant autosomal form and is characterized by having hypoplastic lymphatics or lymphatic aplasia in affected areas. It is also important that the fact of patients do not have other associated congenital diseases, although there may be genotypic, phenotypic and lymphoscintigraphic variations in patients with this diagnosis.¹⁹ Analysis of the genome detected a mutation in the q35.3 locus of chromosome 5.^20,21 The gene related to this area was identified as being the Flt-4, a gene responsible for production of VEGFR-3.²²

An experimental model for type I hereditary lymphedema, using rats, was created by mutagenesis, with inactivation of the VEGFR-3 gene. The rats with mutation had hypoplasia of cutaneous lymphatic vessels and leg lymphedema, but without change in visceral lymphatics.²³

There was increase in serum levels of VEGF-D in patients with primary lymphedema.²⁴ The lack of VGEFR-3 stimulation could be responsible for the largest production of VEGF-D, as an attempt to compensate insufficient lymphangiogenesis caused by lack of VGEFR-3.

Meige disease (type II hereditary lymphedema - OMIM 153200) and distichiasis-lymphedema syndrome (OMIN 153400)

Eight years after Milroy's report, type II hereditary lymphedema was described by Meige²⁵ and characterized as an early primary lymphedema with familiar manifestations. It is a disease with dominant autosomal transmission, low penetrance and variable phenotype. The most common clinical presentation is below-the-knee edema, bilateral and symmetrical, with prevalence of the female gender (3:1). Some authors such as Burnand et al.²⁶ and Northrup et al.²⁷ consider Meige disease and distichiasis-lymphedema (DL) syndrome as different entities, with which this author agrees. Others consider that Meige disease represents a specific portion of patients affected by DL, without the associated characteristics of distichiasis, ptosis, cleft palate, yellow nails, and congenital heart disease.²

From the structural perspective, Meige syndrome has distal lymphatic collectors of the lower limbs, hypoplastic or even obstructed,²⁸ while in DL lymphoscintigraphy shows hyperplastic lymphatic collectors, abundant and dilated.²⁹ On the other hand, identification of genetic change related to Meige syndrome is still unknown, possibly due to the interaction of different alleles,²⁶ whereas in DL there was mutation in the FOXC2 gene of chromosome 16q24.3.³⁰

In experimental studies heterozygous rats for FOXC2 showed the phenotypic characteristics of DL, although without presence of lymphedema. Analysis of lymphatics by lymphoscintigraphy and immunohistochemistry showed hyperplasia of vessels and lymph nodes as reflux resulting from valve insufficiency.³¹ An interesting fact is that the FOXC2 is a transcription factor expressed in the kidney and bone mesenchyma and participates in the formation of embryo somites. Its specific function in the development of the lymphatic system has not been identified.³² In addition, FOXC2 mutations cause several different phenotypes,³³ creating distinct hypotheses as to the regulating role of this gene in lymphangiogenesis.

Hypotrichosis-lymphedema-telangiectasia (OMIM 607823)

The rare hypotrichosis-lymphedema-telangiectasia syndrome has been recently described and is manifested with congenital lymphedema.³⁴ Its transmission can be dominant autosomal or recessive autosomal and was correlated with allele q13 mutations of chromosome 20, corresponding to the SOX18 gene,³⁵ which, similarly to the FOXC2, is a transcription-regulating gene.

Cholestasis-lymphedema syndrome or Aagenaes syndrome (OMIM 214900)

The cholestasis-lymphedema syndrome is a recessive autosomal disease, which presents with neonatal cholestasis and lower limb lymphedema,³⁶ in which lymphoscintigraphic studies showed hypoplastic lymphatics. The involved gene was identified in chromosome 15.³⁷.

Perspectives

From the structural, functional and pathogenetic perspective, wide knowledge of the lymphatic system is essential to understand the several clinical manifestations and therapeutic perspectives of its dysfunctions.³⁸

Identification of the genes related to lymphangiogenesis may allow prenatal diagnosis of the potentially affected fetus and genetic counseling in case of history of the disease in relatives. It is important to define whether the activity of these genes occurs only during embryonic development of lymphatics or also in postnatal stage.

Change in genes through genetic engineering or replacement of defective gene by a trophic factor can be clinically useful¹⁸ in the prevention of development of congenital and primary lymphedemas, or also in the reconstitution of normal lymphatic tissue, either promoting normal development of hypoplastic lymphedemas or blocking lymphatic hyperplasia observed in some syndromes.

Perhaps the closest and most essential application of understanding the mechanisms of lymphangiogenesis is prevention of lymphatic metastases, in which the blockade of lymphangiogenesis can play a crucial role in oncological treatment.

References

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Publication Dates

• Publication in this collection
  17 Dec 2008
• Date of issue
  Sept 2008

History

• Received
  08 Jan 2008
• Accepted
  15 May 2008