Applications of Platelet-Rich Plasma in Lymphedema

Introduction

Platelet-rich plasma (PRP) is a part of blood plasma that is rich in growth factors and platelet concentrate. It is essentially an increased concentration of autologous platelets suspended in a small amount of plasma after repeated centrifugation procedures.^1,2 The preparation of PRP began during the 1960s and became a routine through the 1970s. In the 1980s, platelets were found to release wound-healing substances that initiated the repair of injured tissues and vessels in cutaneous ulcers. ³ PRP has been safely used and documented in many different fields, including orthopedics, sports injuries, dental and periodontal surgery, and cosmetic, plastic, cardiovascular, general, and maxillofacial surgery, for many years.^4–7 Recently, PRP has drawn the attention of dermatologists, specifically in the aesthetic field, to promote tissue remodeling as well as wound healing, to rejuvenate the skin, and to treat skin wrinkles.^7–9

Lymphangiogenesis requires the action of a variety of growth factors to ensure funtional lymph vessel restoration. Platelets release main factors for angiogenesis as well as for lymphangiogenesis: platelet-derived growth factor (PDGF), epidermal growth factor (EGF), interleukin 8 (IL-8), transforming growth factor beta (TGF-β), fibroblast growth factor (FGF), vascular endothelial growth factors (VEGFs), insulin-like growth factor 1 and 2 (IGF-1, IGF-2), keratinocyte growth factor (KGF), and connective tissue growth factor.^2,10

Over the past few years, our understanding in molecular regulation of lymphangiogenesis has been expanded. Besides, the relevance of platelets for lymphangiogenesis and tissue repair has been recently described.^11,12 As PRP contains factors for angiogenesis, it might potentially be used to regenerate injured lymphatic vessels and PRP could probably play a crucial role in the lymphedema treatment. We have reviewed the literature regarding the studies that have been conducted to confirm this hypothesis, and to address the clinical application of PRP in lymphedema as a therapeutic option.

Literature Review

We conducted a literature search in the following databases: Medline via PubMed, the Cochrane Library, and EMBASE using the key words for selected indications (“platelet-rich plasma,” “PRP,” “clinical study,” “lymphedema,” and “trial”). Initially, the literature search was conducted and the references were retrieved to be evaluated. In addition to this, reference lists of the selected articles were inspected for additional relevant studies. However, we have been able to identify only two animal studies regarding the use of PRP in lymphedema treatment.

Hadamitzky et al. studied the effects of PRP on autotransplanted avascular lymph nodes and were able to show an improved lymph node ingrowth and regeneration. ¹³ In this study, 44 female Lewis rats were divided into therapy and control groups. The therapy group consisted of 32 animals that were divided into four groups of eight rats each, and it was subjected to a bilateral extirpation of the superficial inguinal lymph nodes. An ipsilateral autologous implantation of whole (group A) and fragmented (groups B, C, and D) lymph nodes followed. In groups A and B, the transplants were left to regenerate without further intervention. In group C, sheep red blood cells were injected intradermally in the lymphatic drainage area of the subcutaneous inguinal pouch 3 weeks after operation. In group D, PRP was injected intradermally 3 weeks after surgery. Transplanted lymph nodes were examined both macro- and microscopically. In the control group, 12 animals were divided into three groups of four rats. No operation was performed. Group E was used as a normal lymph node control. In group F, an intradermal injection of sheep red blood cells was given in the drainage area of the inguinal lymph nodes, and in group G, PRP was injected. Results of this study showed that PRP not only improves the survival rate of transplanted lymph node fragments in group D but also causes a B-cell proliferation in the lymph nodes of control group G. This result is in favor of the positive effect of PRP in regeneration of new lmphatic vessels and a possible prevention of postoperative lymphedema. However, further research is needed to confirm the preliminary results observed with a single PRP injection.

Ackermann et al. investigated the effects of PRP and adipose-derived mesenchymal stem cell (ASC) on lymphedema, wound repair, and vascular architecture in a murine tail lymphedema model. ¹⁴ Thirty male mice were divided into three groups, and depending on the groups, PRP, ASCs, or saline were topically applied to the wounds, which were followed up for 14 days. The results showed that wounds treated by PRP and ASC healed faster and showed a significantly increased epithelialization mainly from the proximal wound margin. The application of PRP induced a significantly increased lymphangiogenesis, whereas the application of ASC did not induce any significant change in this regard. The authors concluded that the PRP has a positive effect on lymphangiogenesis and reduces lymphedema formation in the short term. This study highlights the possible benefits of PRP in lymphedema therapies, but further research is needed with increasing follow-up duration to obtain more reliable results.

Conclusion

The lymphatic system—vessels, nodes, and surrounding tissue—serves to transport lymphatic fluid from interstitial space to the venous system. Blockage of this network leads to atypical accumulation of protein-rich tissue fluid and is called lymphedema. Diseases associated with the lymphatic network range from congenital disorders to vascular disease and/or trauma as well as cancer and cancer's therapies. Lymphedema is characterized by inflammation, chronic accumulation of protein-rich fluid, fibrosis, and adipose tissue hypertrophy. ¹⁵ Fibrosis has been shown to impair tubule formation, lymphatic endothelial cell proliferation and migration, and lymphatic regeneration, and to decrease lymphatic function.^16,17 Existence of fibrosis in lymphedema considers the pathology as a progresive and incurable disease, requiring very long-term treatment. Ghanta et al. indicated that lymphedema-associated macrophages are a major source of VEGF-C and that impaired macrophage responses after lymphatic injury result in decreased lymphatic function. ¹⁸

Current treatment modalities that are divided into conservative (non-operative) and operative methods aim at reducing swelling and associated symptoms, without a final cure. Non-operative treatment of lymphedema is based on manual lymphatic drainage, high-pressure intermittent pneumatic compression, compression garments, therapeutic exercises, low laser therapy, skin hygiene, and medication; whereas operative treatment options include lymphaticovenous shunts, lymphatic bypasses, lymph node transfer, and liposuction.^19–21 Obtaining and preserving the lymphatic vessels even with modern microsurgical techniques still remains difficult. ¹² Any secondary changes, developing as a result of chronic lymphedema, may also lead to surgical reconstruction failure. ²² New methods to prevent lymphedema or to regenerate lymphatic vessels in diseased tissue not only might improve the progression of the pathology but also treat the lymphatic network. This so called “regenerative medicine” should be considered by tissue engineers, which may result in a promising approach for the treatment of lymphatic diseases.

New lymphatic vessels in lymphedema might be regenerated by a platelets's growth factors, since these factors are effective in new vessel regeneration, angiogenesis, and they are also required during embryonic lymphangiogenesis for the separation of the nascent lymphatic vasculature from blood vessels. ²³ One of the common sources of platelets and also these factors is PRP. Although a great number of studies have shown the effect of growth factors and PRP in the regeneration of different tissues for a relatively long period, the relevance of platelets for lymphangiogenesis has been recently described. ¹¹ The International Society of Lymphology indicated that administration of VEGF-C by different methods has not yet been translated into clinical applications inspite of the fact that the basic research and clinical trials related to molecular treatments has increased in recent years. ¹⁹

Vascular endothelial growth factors that stimulate cellular respeonses by binding to their endothelial tyrosine kinase receptors (VEGFR-1, VEGFR-2, and VEGFR-3) are important regulators of vasculogenesis, angiogenesis, and lymphangiogenesis. ²⁴ VEGFR-1 that is expressed on hematopoietic stem cells, monocytes, and vascular endothelial cells is required for the migration of monocytes and macrophages. VEGFR-2 that is expressed on vascular endothelial cells and lymphatic endothelial cells regulates vascular endothelial function, whereas VEGFR-3 that is only expressed on lymphatic endothelial cells is responsible for lymphatic endothelial cell function.^24,25 Activation of both VEGFR-2 and VEGFR-3 might be needed for efficient lymphangiogenesis. ²⁶ VEGF-C improves lymphedema by promoting lymphangiogenesis, stimulating the recanalization of injured lymphatic vessels, restoring the lymph flow, promoting the organization of lymphatic endothelial cells into functional lymphatic vessels, and ameliorating the pump activity of the collecting lymphatics through the activation of VEGFR-2 and VEGFR-3.^27–29 The potential and the specificity of VEGF-C or VEGF-D for stimulating new lymphatic vessels growth have been repesented in the preclinical lymphedema models utilizing recombinant protein application. ³⁰ On the other hand, Cuzzone et al. found that IL-6 is increased as a result of adipose deposition and CD4(+) cell inflammation in lymphedema, and they suggested that IL-6 expression in lymphedema acts to limit adipose accumulation. ³¹ One of the other possible mechanisms of PRP-induced angiogenesis might be due to accumulation of mast cells and their mediators as bFGF after PRP administration.^32,33 Among all these data, using PRP, containing various kinds of growth factors, to prevent or treat lymphedema is a relatively new approach.

Lymphedema can lead to thickening of the dermis and can create chronic skin disorders, skin infections such as cellulitis and lymphangitis, lymphorrhea, and inflammation wounds. ³⁴ Unlike other types of edema, the high protein content in lymphedema represents a very good medium for bacterial growth and increases the risk of developing cellulitis, which probably causes additional lymphatic damage. ³⁵ The skin becomes hard, hyperpigmented, and verrucous as lymphedema progresses. ³⁶ Commonly, skin defects over the edematous area caused by pressure, friction, and shear from wraps or garments occur. Skin defects sometimes turn into chronic wounds due to the fact that an insufficient supply of nutrients and oxygen to the tissues leads to impaired wound healing. ³⁶ The presence of cellulitis and skin lesions have a negative impact on health-related quality of life in primary lymphedema patients. ³⁷ However, prevention of cellulitis and skin lesions is difficult to improve the advanced lymphedema stage.

More recently, various PDGFs, which have been found to stimulate angiogenesis and mitosis, have been used to initiate wound healing.^38,39 Growth factors increase the number of undifferentiated mesenchymal cells at the scar site during tissue regeneration and wound healing. Platelets have an important role in the angiogenesis process that is needed for the activation, proliferation, and migration of endothelial cells to form new blood vessels from pre-existing vessels in the wound-healing response. ⁴⁰ Serra et al. indicated that platelet gel application might be an effective adjuvant treatment to promote wound healing in diabetic dysvascular patients after transmetatarsal amputation. In the literature, it has been shown that topical application of autologous PRP enhanced the process of wound healing after revascularization of lower limbs in patients affected by critical limb ischemia. ³⁸ Considering the importance of angiogenesis in wound healing and the role of platelets in the angiogenesis process, it is tempting to speculate whether PRP can be used to manage skin defects, chronic skin disorders, skin infections such as cellulitis and lymphangitis, lymphorrhea, and inflammation wounds caused by lymphedema.

We have been able to identify two animal studies regarding the clinical application of PRP in lymphedema. These studies had a low level of evidence. Besides, the optimal dosage, frequency of administration, its long-term results, and appropriate stage of lymphedema for the use of PRP have not been investigated. Literature cannot yet give a convincing answer about the value of PRP as a therapeutic strategy. However, it provides positive and promising results for the future application of PRP to improve regeneration of lymphatic vessels, restore disrupted lymphatic circulation, and treat or prevent lymphedema-associated problems such as skin defects, chronic skin disorders, skin infections, and chronic wounds. Consequently, it is clear that clinical trials in humans are needed to yield the strong evidence regarding the clinical application of PRP usage in lymphedema. This promising field of research gives the scientists the ability to expand it extensively.