III – Pathophysiology
Guest lecture
Chairperson: M. Malouf
Why do varicose veins develop?
N. Labropoulos
N. Labropoulos gave an overview of current knowledge and theories on the pathophysiology of varicose veins. He reminded the audience of the high prevalence of this disease, particularly in developed countries, and its specificity to humans, which renders investigations on animal models impossible. The talk focused on primary venous disease, hence excluding postthrombotic syndrome, in which obstruction and valve destruction occurs. Venous hypertension plays a major role in the progression of the disease. It provokes vein dilatation and inflammation. Vein dilatation leads to distortion, leakage, and altered shear stress, which is responsible for inflammation. The inflammation may cause vein valve and wall changes responsible for the reflux, which further increases the venous hypertension. Several theories have been developed to explain initiation of the disease: 1) Primary valve failure leading to its incompetence, with retrograde progression through the venous system; 2) Existence of multiple arteriovenous fistulae; 3) Turbulences above the valve leading to secretion of noradrenaline, hypoxia, secretion of free radicals, and inflammation.
Current theory focuses on weakening and dysfunction of the venous wall due to changes in collagen and elastin structures. A deficiency in type 3 collagen is observed that could be due to its destruction by overexpressed matrix metalloproteases. Contrary to what was previously thought, reflux does not always extend in a retrograde fashion from a primarily damaged vein. In a longitudinal study of 31 patients with GSV reflux, follow-up examination revealed either local extension of the reflux in 17 patients (7 retrograde, 7 anterograde, and 3 twodirectional extensions) or appearance of reflux in a new vein segment in 14.
The role of genetics remains unclear, even if a family predisposition is often reported. Whether venous disease is a local or systemic disease is also debated. Vein wall pathology may have not only local effects. On the other hand, normal venous segments adjacent to varicose veins share the same biochemical properties, suggesting that wall changes could precede biological abnormalities. The increased hydrostatic pressure could provoke increased vein wall tension and endothelial cell injuries, leading to inflammation and modification of matrix metalloprotease expression. This could eventually lead to vein dilatation and valve dysfunction through an altered collagen structure.
Much work still needs to be done to better understand the pathophysiology of venous disease. Data from epidemiological studies need to be combined with clinical studies, genetics, cellular and wall matrix investigations, and studies of flow dynamics and wall mechanics.
When asked by the audience if these new concepts should have an impact on patients’ clinical management, N. Labropoulos came back to the debate on whether or not patients with chronic venous disease should be treated as early as possible to stop or at least delay the progression of the disease. It also appears that some venoactive drugs may have a favorable modulating action on metalloproteases.
Satellite workshop
Tissular repair in phlebology
P. Sansilvestri-Morel presented her investigations on venous disease. She focused on vein wall distensibility, elasticity, and contractility. In patients with varicose veins, disorganization of the vessel media extracellular matrix is observed. Collagen plays a major role in the structure of vein wall. She compared collagen characteristics in cellular cultures obtained from varicose veins (vein stripping specimens) and from control veins (healthy veins removed for arterial bypass). The main result was a significant quantitative increase of collagen 1 in smooth muscle cells, and a decrease in collagen 3. An overexpression of collagen 1 was observed by RNA analysis, but no difference was seen in the expression of collagen 3 gene. Overexpression of collagen 1 could be due to collagen 3 deficiency. In fact, adding collagen 3 to the cell culture provoked a decrease in collagen 1 expression. Since expression of collagen 3 is normal, the hypothesis of an enhanced destruction of collagen 3 was raised, supported by the fact that adding matrix metalloproteases (responsible for collagen destruction) increased the levels of collagen 3. Similar results were obtained when comparing skin biopsies of patients with venous ulcer with biopsies obtained in healthy subjects.
Discussion with the audience raised the question of a possible systemic disease since the same pattern is seen in the skin and in the vein wall. Note that skin biopsy specimens were taken from the groin, which could partly account for the results. It would be interesting to compared data from skin biopsies taken from other parts of the body, the perivenous derma, and from arteries close to the diseased veins.
J-C. Kerihuel reported on his research on healing, which comprises four phases: coagulation, inflammation, proliferation, and remodeling. The healing process is altered in patients with chronic wounds. As compared with acute wounds, fewer mitoses are seen in the derma, fibroblasts tend to be in apoptosis, metalloproteinases are increased, the inflammation process is not controlled, and the extracellular matrix is damaged. Metalloproteinases play a role in the different phases of the healing process. They have proteolytic and angiogenic properties, interact with immunoglobins, cytokines and chemokines. They could become a major target for intervention in patients with vein ulcers. The micronized flavonoid fraction might exhibit a regulatory action on metalloproteinases, which could at least partly account for its efficacy in the treatment of venous ulcers.
Venous pathophysiology
Chairperson: V. Blazek
Moderator: F. Boccardo
Unmyelinated C fibers and inflammatory cells are present in the wall of human varicose veins. A clinicopathological study
A. Vital
The work by A. Vital is based on current hypotheses that pain in chronic venous disease has a local inflammatory origin. Over the last five years, indicators suggesting an inflammatory reaction in varicose veins have accumulated dramatically. However, little is known about the nociceptors, likely to be C fibers, their innervation and their relationship with inflammatory cells in human blood vessels, and particularly in veins. C fibers have yet to be found in human varicose vein samples.
In addition, the precise mechanisms governing the interaction between the mediators of inflammation and venous nociceptors, which may account for the variability of pain in venous disease, remain to be clarified.
The aim of the study presented was to localize C fibers and define their relationship with inflammatory cells in varicose vein wall. The material examined consisted of segments of the great saphenous vein harvested during saphenectomy from patients with documented chronic venous disease and venous pain scored > 4 on a visual analogue scale.
Five segments per patient were immunostained with anti-S100 protein and anti- CD45 to identify nerve fibers and inflammatory cells, respectively. Light microscopy was completed by electron microscopy.
Immunostaining indicated that most sections had a low density of nerve fibers. CD45-staining showed a low density of inflammatory cells. At microscopy, most nerve fibers appeared scattered in the media and some were identified in the vicinity of the vasa vasorum, close to the adventitia, while mast cells were mainly located in the media, often close to the adventitia, but not in close contact with C fibers. Inflammatory cells were mainly macrophages, rather than neutrophils and lymphocytes, which were not observed.
The presence of unmyelinated C fibers in the wall of varicose veins suggests the existence of a neurological component able to diffuse pain signals from the vein into the spinal chord and finally to the brain. Inflammatory cells and particularly mast cells that could be responsible for activation of those C fibers were also found in the pathological vein wall.
C fibers and mast cells may form a functional unit that could contribute significantly to mechanisms of venous pain arising at earlier and/or later stages of chronic venous disease.
Proteomic comparison of varicose veins in humans and in those that develop in a new porcine model
A. Van Rij
A. Van Rij and his team have developed and characterized a pig-based model of superficial varicose veins by means of femoral arteriovenous fistulae fashioned in the right thigh. Animals were examined at postoperative times up to 15 weeks to determine the development of varicose veins and the associated protein expression. Protein expression associated with varicose vein disease was examined both in humans and the porcine model.
Gel electrophoresis and mass spectrometry were used to identify proteins expressed in the varicose porcine model and in control animals. The same process was used to compare human primary varicose and control superficial thigh vein samples. In humans and the pig model, cytoskeletal and contractility-related proteins were upregulated (actin, tropomyosin, desmin, and vimentin), as well as some heat shock proteins (in particular HSBP1). These profiles suggest a similar process of mild inflammation and tissue remodeling in both human and porcine varicose veins. Superficial varicose vein formation in this porcine model, which mimics human venous disease, may represent a considerable advance in the development and assessment of phlebotropic pharmacological agents.
Venous ulcer: the final stage in chronic venous disease – Managing it in 2009 (Pierre Fabre Symposium)
Chairperson: P. Carpentier
In this symposium, the speakers summarized the latest information regarding to pathophysiology, clinical evaluation, and therapeutic management of chronic venous disease and venous ulcers.
As an introduction P. Carpentier emphasized that painful and disabling venous ulcers, which occur at high incidence and generate huge costs, remains a major public health issue. Understanding the pathogenetic mystery of end-stage chronic venous disease is crucial since its management is based mainly on this knowledge. A. Nicolaides gave us a useful summary of what we know about pathophysiology and about the levels of clinical evaluation in CVI. Changes in the superficial and/or deep veins are the starting events at the level of macrovasculature. Primary varicose veins occur in the absence of previous DVT; secondary ones are the consequence of DVT or superficial thrombophlebitis. Recanalization may give rise to relative obstruction and incompetence of deep, superficial, and perforating veins. Relative obstruction means that the affected vein becomes rigid, losing the ability to dilate in the case of effort. In 30% of patients with deep venous reflux, the etiology is primary valvular incompetence. Spontaneous lysis occurs in 50-70% following DVT and early thrombus resolution is associated with a higher incidence of valve competence. Postthrombotic syndrome (PTS) is the result of venous hypertension arising from deep valve incompetence and/or outflow obstruction after a previous episode of DVT. Venous hypertension leads to skin capillary damage, lipodermatosclerosis, and ultimately ulceration. The prevalence of PTS is variable and depends on the extent and location of thrombosis and on the results of treatment. The risk of PTS is lower in patients who have adequate and early anticoagulation therapy, and early mobilization with continuous compression. Patients with both chronic obstruction and reflux have the highest incidence of skin changes or ulceration. The risk of PTS is higher in patients with recurrent thrombosis and in the presence of thrombophilia. Incompetent perforating veins (IPVs) usually occur in the presence of superficial and/or deep venous reflux. The number and diameter of IPVs, and the volume and velocity through them increase linearly with clinical severity of CVI. Superficial and perforating vein incompetence with normal deep veins is found in 40% of patients with skin changes and ulceration. In the pathogenesis of primary varicose veins the leukocyte- endothelium interaction plays a key role inducing chronic inflammatory process and remodeling in the venous wall and venous valves, generating reflux and venous hypertension. Leukocytes also play an important role in edema formation. The speaker emphasized that there is no single test that can provide all information needed to make clinical decisions and plan a management strategy. Understanding the pathophysiology is the key to selecting the appropriate investigations. For reflux and obstruction several questions need to be answered: are they present or absent, where are they (anatomic extent) and how, many are there (quantitation)? For the demonstration of reflux, the CW hand-held Doppler is frequently sufficient. However, to determine the anatomic extent we need duplex ultrasound or descending venography. Air plethysmography is best for quantitative measurements of reflux . There are three levels of investigations regarding the CEAP class of the disease. In class C0/1, level I investigations (history, clinical examination, which may include hand-held Doppler or duplex) are sufficient. In class 2, level II (duplex scanning with or without plethysmography) should be used in the majority of patients and is mandatory in those being considered for intervention. In class 3 or more, level II investigations are utilized to determine whether or not reflux or obstruction in the deep veins is responsible for edema or skin changes. If obstruction is demonstrated or suspected as a result of a noninvasive test, level III studies (invasive investigations or complex imaging studies) to investigate the deep venous system should be considered.
Later on P. Blanchemaison briefly presented the therapeutic options in the management of venous ulceration. Compression (four-layer bandage, short stretch bandages), dressings (wet dressings and hydrocolloids), physiotherapy (structured exercise improving calf muscle pump function, aquatic physiotherapy), venotonic (MPFF at a dose of 500 mg, Cyclo 3) and other drugs (pentoxifylline, stanozol), sclerotherapy, surgery (stripping, laser, radiofrequency, deep venous surgery, skin grafts) vacuumassisted closure therapy, hemodilution, autohemotherapy, hyperbaric oxygen, larval therapy (application of maggots –Lucilla sericata – to a necrotic lesion) all have a more or less evidence-based role in the management of venous ulcer. The main risk factors for chronic ulceration in patients with varicose veins are: heredity, age, obesity, diabetes, history of DVT, fixed ankle joint and impairment of the calf muscle pump. Regarding the role of superficial venous surgery and compression therapy in the management of venous ulcers, a systematic review by Howard el al (Eur J Vasc Endovasc Surg, 2008) and the ESCHAR randomized controlled trial conducted by Gohel (BMJ 2007) concluded that the surgical correction of superficial venous reflux in addition to compression therapy does not improve ulcer healing, but reduces the recurrence of ulcers at 4 years. Deep venous surgery (venoplasty, stenting, by-pass procedures, valvuloplasty, vein transposition, vein transplantation, neovalve formation, external cuff) might be effective and can be clinically beneficial, but there is a lack of evidence. The speaker concluded that conservative treatment should precede surgery. Compression therapy should consist of 4-layer bandages, class 3 stockings, or an Unna boot. The nutritional status of the patient, elevation of the leg and mobilization are also important factors. Venotonic drugs are always indicated and microcirculatory and hemodynamic follow-up is recommended
