Lymphaticovenous anastomosis in rabbits: A novel live experimental animal model for supermicrosurgical training.

BACKGROUND: Lymphaticovenous anastomosis is widely used in lymphedema management. Although its effectiveness in reducing edema in patients can be clinically observed, evaluating the long-term outcomes of this technique can be complex. This study established an animal model to assess the outcomes of lymphaticovenous anastomosis technique at 15 and 30-days post-surgery using indocyanine green lymphography, Patent Blue V dye injection, and histopathological examination. METHODS: An experimental model was established in the hindlimbs of 10 rabbits using the popliteal vein and afferent lymphatic vessels in the popliteal area. The subjects were divided into two groups: the first group (n = 5) underwent patency assessment at 0 and 15 days, and the second group (n = 5) at 0 and 30-days, resulting in 20 anastomoses. Patency was verified at 0, 15, and 30-days using indocyanine green lymphography and Patent Blue V injection. Histopathological examinations were performed on the collected anastomosis samples. RESULTS: The patency rate was 90% (19/20) initially, 60% (6/10) at 15 days post-surgery, and 80% (8/10) at 30-days. The average diameter of lymphatic vessels and veins was 1.0 mm and 0.8 mm, respectively. The median number of collateral veins was 3; the median surgical time was 65.8 min. Histopathology revealed minimal endothelial damage and inflammatory responses due to the surgical sutures, with vascular inflammation and thrombosis in a single case. Local vascular neoformations were observed. CONCLUSION: This study highlights the reliability and reproducibility of using rabbits as experimental models for training in lymphaticovenous anastomosis technique owing to the accessibility of the surgical site and dimensions of their popliteal vasculature.

SurLym trial: study protocol for a multicentre pragmatic randomised controlled trial on the added value of reconstructive lymphatic surgery to decongestive lymphatic therapy for the treatment of lymphoedema.

INTRODUCTION: Lymphoedema is a chronic condition caused by lymphatic insufficiency. It leads to swelling of the limb/midline region and an increased risk of infection. Lymphoedema is often associated with mental and physical problems limiting quality of life. The first choice of treatment is a conservative treatment, consisting of exercises, skin care, lymph drainage and compression. Reconstructive lymphatic surgery is also often performed, that is, lymphovenous anastomoses, lymph node transfer or a combination. However, robust evidence on the effectiveness of reconstructive lymphatic surgery is missing. Therefore, the objective of this trial is to investigate the added value of reconstructive lymphatic surgery to the conservative treatment in patients with lymphoedema. METHODS AND ANALYSIS: A multicentre randomised controlled and pragmatic trial was started in March 2022 in three Belgian university hospitals. 90 patients with arm lymphoedema and 90 patients with leg lymphoedema will be included. All patients are randomised between conservative treatment alone (control group) or conservative treatment with reconstructive lymphatic surgery (intervention group). Assessments are performed at baseline and at 1, 3, 6, 12, 18, 24 and 36 months. The primary outcome is lymphoedema-specific quality of life at 18 months. Key secondary outcomes are limb volume and duration of wearing the compression garment at 18 months. The approach of reconstructive lymphatic surgery is based on presurgical investigations including clinical examination, lymphofluoroscopy, lymphoscintigraphy, lymph MRI or CT angiography (if needed). All patients receive conservative treatment during 36 months, which is applied by the patient's own physical therapist and by the patient self. From months 7 to 12, the hours a day of wearing the compression garment are gradually decreased. ETHICS AND DISSEMINATION: The study has been approved by the ethical committees of University Hospitals Leuven, Ghent University Hospital and CHU UCL Namur. Results will be disseminated via peer-reviewed journals and presentations. TRIAL REGISTRATION NUMBER: NCT05064176.

Immediate Partial Breast and Nipple-Areola Complex Reconstruction Using a Superficial Circumflex Iliac Artery Perforator Flap.

The superficial circumflex iliac artery perforator (SCIP) flap is a versatile flap that has been described for various applications, mostly for lower extremity coverage and head and neck reconstructions. However, there are few publications reporting its use for breast reconstruction, mainly because of its low volume availability. In this article, we present the case of a patient who successfully underwent a partial breast and immediate nipple-areola complex (NAC) reconstruction with an SCIP flap. She had been previously reconstructed with an implant after a nipple-sparing mastectomy, but the NAC turned out to be involved with cancer needing further resection. Our goal with this article, is to introduce a novel concept for addressing partial breast and NAC reconstruction and mostly, to illustrate the importance of an adaptable surgical plan based on every individual case emphasizing the versality of microsurgery for breast cancer reconstruction.

Popliteal Vascular Lymph Node Resection in the Rabbit Hindlimb for Secondary Lymphedema Induction.

Lymphedema is a common condition often associated with cancer and its treatment, which leads to damage to the lymphatic system, and current treatments are mostly palliative rather than curative. Its high incidence among oncologic patients indicates the need to study both normal lymphatic function and pathologic dysfunction. To reproduce chronic lymphedema, it is necessary to choose a suitable experimental animal. Attempts to establish animal models are limited by the regenerative capacity of the lymphatic system. Among the potential candidates, the rabbit hindlimb is easy to handle and extrapolate to the human clinical scenario, making it advantageous. In addition, the size of this species allows for better selection of lymphatic vessels for vascularized lymph node resection. In this study, we present a procedure of vascular lymph node resection in the rabbit hindlimb for inducing secondary lymphedema. Anesthetized animals were subjected to circumferential measurement, patent blue V infiltration, and indocyanine green lymphography (ICG-L) using real-time near-infrared fluorescence, a technique that allows the identification of single popliteal nodes and lymphatic channels. Access to the identified structures is achieved by excising the popliteal node and ligating the medial and lateral afferent lymphatics. Special care must be taken to ensure that any lymphatic vessel that joins the femoral lymphatic system within the thigh without entering the popliteal node can be identified and ligated. Postoperative evaluation was performed at 3, 6, and 12 months after induction using circumferential measurements of the hindlimb and ICG-L. As demonstrated during follow-up, the animals developed dermal backflow that was maintained until the 12th month, making this experimental animal useful for novel long-term evaluations in the management of lymphedema. In conclusion, the approach described here is feasible and reproducible. Additionally, during the time window presented, it can be representative of human lymphedema, thus providing a useful research tool.

Delphi survey of intercontinental experts to identify areas of consensus on the use of indocyanine green angiography for tissue perfusion assessment during plastic and reconstructive surgery.

BACKGROUND: In recent years, indocyanine green angiography (ICG-A) has been used increasingly to assist tissue perfusion assessments during plastic and reconstructive surgery procedures, but no guidelines exist regarding its use. We sought to identify areas of consensus and non-consensus among international experts on the use of ICG-A for tissue-perfusion assessments during plastic and reconstructive surgery. METHODS: A two-round, online Delphi survey was conducted of 22 international experts from four continents asking them to vote on 79 statements divided into five modules: module 1 = patient preparation and contraindications (n = 11 statements); module 2 = ICG administration and camera settings (n = 17); module 3 = other factors impacting perfusion assessments (n = 10); module 4 = specific indications, including trauma debridement (n = 9), mastectomy skin flaps (n = 6), and free flap reconstruction (n = 8); and module 5 = general advantages and disadvantages, training, insurance coverage issues, and future directions (n = 18). Consensus was defined as ≥70% inter-voter agreement. RESULTS: Consensus was reached on 73/79 statements, including the overall value, advantages, and limitations of ICG-A in numerous surgical settings; also, on the dose (0.05 mg/kg) and timing of ICG administration (∼20-60 seconds preassessment) and best camera angle (61-90o) and target-to-tissue distance (20-30 cm). However, consensus also was reached that camera angle and distance can vary, depending on the make of camera, and that further research is necessary to technically optimize this imaging tool. The experts also agreed that ambient light, patient body temperature, and vasopressor use impact perfusion assessments. CONCLUSION: ICG-A aids perfusion assessments during plastic and reconstructive surgery and should no longer be considered experimental. It has become an important surgical tool.

RhoA/ROCK2 signalling is enhanced by PDGF-AA in fibro-adipogenic progenitor cells: implications for Duchenne muscular dystrophy.

BACKGROUND: The lack of dystrophin expression in Duchenne muscular dystrophy (DMD) induces muscle fibre and replacement by fibro-adipose tissue. Although the role of some growth factors in the process of fibrogenesis has been studied, pathways activated by PDGF-AA have not been described so far. Our aim was to study the molecular role of PDGF-AA in the fibrotic process of DMD. METHODS: Skeletal muscle fibro-adipogenic progenitor cells (FAPs) from three DMD treated with PDGF-AA at 50 ng/mL were analysed by quantitative mass spectrometry-based proteomics. Western-blot, immunofluorescence, and G-LISA were used to confirm the mass spectrometry results. We evaluated the effects of PDGF-AA on the activation of RhoA pathway using two inhibitors, C3-exoenzyme and fasudil. Cell proliferation and migration were determined by BrdU and migration assay. Actin reorganization and collagen synthesis were measured by phalloidin staining and Sircol assay, respectively. In an in vivo proof of concept study, we treated dba/2J-mdx mice with fasudil for 6 weeks. Muscle strength was assessed with the grip strength. Immunofluorescence and flow cytometry analyses were used to study fibrotic and inflammatory markers in muscle tissue. RESULTS: Mass spectrometry revealed that RhoA pathway proteins were up-regulated in treated compared with non-treated DMD FAPs (n = 3, mean age = 8 ± 1.15 years old). Validation of proteomic data showed that Arhgef2 expression was significantly increased in DMD muscles compared with healthy controls by a 7.7-fold increase (n = 2, mean age = 8 ± 1.14 years old). In vitro studies showed that RhoA/ROCK2 pathway was significantly activated by PDGF-AA (n = 3, 1.88-fold increase, P < 0.01) and both C3-exoenzyme and fasudil blocked that activation (n = 3, P < 0.05 and P < 0.001, respectively). The activation of RhoA pathway by PDGF-AA promoted a significant increase in proliferation and migration of FAPs (n = 3, P < 0.001), while C3-exoenzyme and fasudil inhibited FAPs proliferation at 72 h and migration at 48 and 72 h (n = 3, P < 0.001). In vivo studies showed that fasudil improved muscle function (n = 5 non-treated dba/2J-mdx and n = 6 treated dba/2J-mdx, 1.76-fold increase, P < 0.013), and histological studies demonstrated a 23% reduction of collagen-I expression area (n = 5 non-treated dba/2J-mdx and n = 6 treated dba/2J-mdx, P < 0.01). CONCLUSIONS: Our results suggest that PDGF-AA promotes the activation of RhoA pathway in FAPs from DMD patients. This pathway could be involved in FAPs activation promoting its proliferation, migration, and actin reorganization, which represents the beginning of the fibrotic process. The inhibition of RhoA pathway could be considered as a potential therapeutic target for muscle fibrosis in patients with muscular dystrophies.

Isolation of human fibroadipogenic progenitors and satellite cells from frozen muscle biopsies.

Skeletal muscle contains multiple cell types that work together to maintain tissue homeostasis. Among these, satellite cells (SC) and fibroadipogenic progenitors cells (FAPs) are the two main stem cell pools. Studies of these cells using animal models have shown the importance of interactions between these cells in repair of healthy muscle, and degeneration of dystrophic muscle. Due to the unavailability of fresh patient muscle biopsies, similar analysis of interactions between human FAPs and SCs is limited especially among the muscular dystrophy patients. To address this issue here we describe a method that allows the use of frozen human skeletal muscle biopsies to simultaneously isolate and grow SCs and FAPs from healthy or dystrophic patients. We show that while the purified SCs differentiate into mature myotubes, purified FAPs can differentiate into adipocytes or fibroblasts demonstrating their multipotency. We find that these FAPs can be immortalized and the immortalized FAPs (iFAPs) retain their multipotency. These approaches open the door for carrying out personalized analysis of patient FAPs and interactions with the SCs that lead to muscle loss.

Reverse Lymph Node Mapping Using Indocyanine Green Lymphography: A Step Forward in Minimizing Donor-Site Morbidity in Vascularized Lymph Node Transfer.

BACKGROUND: Reverse lymphatic mapping before harvesting a lymph node flap is crucial to avoid donor-site lymphedema; however, the technique is complex and unavailable in many centers. The authors introduce radioisotope-free reverse lymphatic mapping using indocyanine green and Patent Blue dye. METHODS: The authors conducted a prospective study in patients undergoing free vascularized groin lymph node transfer for postmastectomy upper extremity lymphedema. The day before surgery, 0.2 ml of technetium-99 was injected into the first and second web spaces of the ipsilateral foot. The following day, once the patient was anesthetized, indocyanine green was injected into the same web spaces of the same foot and Patent Blue dye was injected just proximal to the upper margin of the skin paddle of the lymph node flap. The main lymph nodes draining the limb were localized using indocyanine green lymphography and gamma probe. RESULTS: Thirty-nine patients underwent vascularized groin lymph node transfer with or without deep inferior epigastric artery perforator flap breast reconstruction. Navigation of the main lower extremity draining inguinal lymph nodes using the gamma probe and indocyanine green lymphography was identical in all patients. The blue-stained lymphatics in the skin paddle drained to the superficial proximal inguinal lymph node and were targeted for transfer. No donor-site lymphedema was reported, and lymphatic drainage of the lower extremity was preserved in all cases. CONCLUSIONS: Reverse lymphatic mapping using indocyanine green lymphography provides identical results to those using technetium-99 isotope scanning. However, indocyanine green is preferable in terms of safety and reproducibility and also avoids the complexity and hazards of radioisotope mapping. CLINICAL QUESTION/LEVEL OF EVIDENCE: Therapeutic, IV.

Unique Techniques or Approaches in Microvascular and Microlymphatic Surgery.

Several methods can be used for identifying tissues for transfer in donor-site-depleted patients. A fillet flap can be temporarily stored in other parts of the body and transferred back to the site of tissue defect, including covering the amputated stump of the lower extremity. Human arm transplant is rare and has some unique concerns for the surgery and postsurgical treatment. Cosmetics of the narrow neck of transferred second toes can be improved with insertion of a flap. Lymphedema of the breast after cancer treatment can be diagnosed with several currently available imaging techniques and treated surgically with lymphaticovenous anastomosis.

Major Changes in Lymphatic Microsurgery and Microvascular Surgery in Past 20 Years.

This article summarizes the major changes seen in lymphatic microsurgery and microvascular surgery in first 20 years of the 21st century. Lymphatic microsurgery is discussed first, as more advances have been seen in imaging of the lymphatic system, lymphatico-venous anastomosis, and vascularized lymph node transfers. During the past 2 decades, there have been more patient population changes than major technical evolutions in microvascular surgery, although new techniques and modifications emerged and became clinical routines, with the landscape of microvascular surgery evolving in this time period.

Correlation between superficial and deep lymphatic systems using magnetic resonance lymphangiography in breast cancer-related lymphedema: Clinical implications.

BACKGROUND: Magnetic resonance lymphangiography (MRL) has increased our knowledge of lymphatic anatomy and lymphedema pathophysiology and improved the efficacy of microsurgical procedures to manage peripheral lymphedema. The aim of this study is to investigate the ability of MRL to detect communications between superficial and deep lymphatic systems in breast cancer-related lymphedema (BRCL) and to investigate whether these communications could preserve lymphatic drainage of the hand. METHODS: Between 2008 and 2017 we used MRL imaging in 59 women with BCRL. Lymphedema of the arm and hand was detected in 30 patients while the hand was spared in 29. Using axial and coronal MRL reconstruction images we investigated the existence of any communication between the superficial and deep lymphatic systems. RESULTS: Among the 29 patients with spared hand, MRL revealed that 24 had at least one communicating lymphatic perforator at the wrist region (p < 0.001). Lymphatic flow at the axilla was clearly visualized in 16 of the 29 patients (55.2%), no perforating lymphatic vessels were detected in the group with lymphedema in the hand (30 patients). CONCLUSIONS: Communications between the deep and superficial lymphatic systems at the wrist region in BCRL patients without hand lymphedema should be considered when planning microsurgical lymphatic procedures at the wrist and in postoperative compression therapy.

Unique Techniques or Approaches in Microvascular and Microlymphatic Surgery.

Several methods can be used for identifying tissues for transfer in donor-site-depleted patients. A fillet flap can be temporarily stored in other parts of the body and transferred back to the site of tissue defect, including covering the amputated stump of the lower extremity. Human arm transplant is rare and has some unique concerns for the surgery and postsurgical treatment. Cosmetics of the narrow neck of transferred second toes can be improved with insertion of a flap. Lymphedema of the breast after cancer treatment can be diagnosed with several currently available imaging techniques and treated surgically with lymphaticovenous anastomosis.

Intratissue lymphovenous communications in the mechanism of action of vascularized lymph node transfer.

BACKGROUND AND OBJECTIVES: Vascularized lymph node transfer (VLNT) as a surgical treatment for cancer-related lymphedema has shown beneficial effects. The mechanism of action of this procedure involves lymphangiogenesis and lymphovenous communications (LVC) within the lymph nodes. We propose intratissue LVC as an additional element responsible for drainage of lymph to blood in the flap and examine this hypothesis in the current study. METHODS: This prospective study determined the passage of lymph to the venous system via intratissue LVC in 26 free flaps used for breast reconstruction. We evaluated whether fluorescence was positive in the pedicle vein after increasing time intervals following intradermal injection of indocyanine green (ICG) dye into the flaps using near infra-red fluorescence lymphography. RESULTS: We studied 26 free flaps, 22 deep inferior epigastric perforator (DIEP) flaps (84.6%), and 4 superior gluteal artery perforator (SGAP) flaps (15.4%). Fluorescence in the pedicle vein was positive in 22 of the 26 flaps (P = 0.000). The median time for fluorescence evaluation in the pedicle vein was 120 min. CONCLUSIONS: Fluorescence in the pedicle vein after ICG intradermal injection indicates functional intratissue LVC in free flaps. J. Surg. Oncol. 2017;115:27-31. © 2016 Wiley Periodicals, Inc.

Barcelona Lymphedema Algorithm for Surgical Treatment in Breast Cancer-Related Lymphedema.

Background Breast cancer-related lymphedema is a prevalent condition that has a major impact on quality of life. Surgical treatment has become an alternative to help affected patients with good results. However, there is no consensus on surgical procedure and protocol. Methods We analyzed our data in two periods: from June 2007 to December 2011 and from January 2012 to June 2014. Data included the analysis of the limb circumferences and the subjective symptoms felt by patients. Results Of the 200 patients treated in the study, 81 had lymphaticovenous anastomosis, 7 had autologous lymph node transfer, 16 had total breast anatomy restoration, 52 had vibroliposuction, and 44 had combined reconstructive procedures. In the first period, the circumference of the superior limb showed a decrease of 0.9 to 6.1 cm (average 2.75 cm). In the second period, the circumference of the superior limb showed a decrease of 2.9 to 6.1 cm (average 3.85 cm). Clinical results and data from the questionnaires confirmed the improvement and subjective benefits. Conclusion We have obtained considerable improvements in results of limb circumferences and subjective symptoms after incorporating several modifications into our surgical strategy for lymphedema treatment. A detailed preoperative assessment should be performed to determine whether reconstructive surgery or palliative surgery is indicated.

A comprehensive review of the natural lymphaticovenous communications and their role in lymphedema surgery.

Cancer-related lymphedema is a progressive, chronic condition that impairs quality of life. Its pathophysiology and the mechanisms of action of current reconstructive surgical treatments are not fully understood but lymphaticovenous communications may play a key role. We review the available literature on lymphaticovenous communications and their implications in lymphedema surgery, and propose a subclassification of lymphaticovenous communications. J. Surg. Oncol. 2016;113:374-380. © 2016 Wiley Periodicals, Inc.

Combined Surgical Treatment in Breast Cancer-Related Lymphedema.

BACKGROUND: Lymphedema is a well-known sequela of breast but no consensus has been reached about the ideal treatment. Surgical approaches, however, are receiving increased attention. Various microsurgical reconstructive techniques aim to restore anatomy and function of the lymphatic system in upper limb breast cancer-related lymphedema (BCRL). We combined two techniques, lymphaticovenous anastomosis (LVA) and autologous lymph node transplantation (ALNT) after carefully selecting those who may benefit from the surgery. We called this the "combined surgical treatment (CST)" approach. METHODS: From June 2007 to December 2011, we performed CST in 106 patients with upper limb BCRL. Clinical evaluation and diagnostic imaging studies were performed preoperatively in all the patients. CST was offered to patients with stage I/II lymphedema, according to the criteria of the International Society of Lymphology (ISL). RESULTS: Overall 59 of the 106 patients underwent LVA, 7 underwent ALNT, and 40 underwent both the techniques. All 47 lymph node (LN)-flaps survived but 11 (22%) required surgical revision within 3 days. A total of 21 LN-flaps (45%) showed no radiotracer uptake at 1 year. Around 1 to 7 LVAs for each patient (average 3.4) were performed. Preoperative versus postoperative excess circumference decreased between 12 and 86.7% (average 39.72%). Arm circumference decreased between 0.9 and 6.1 cm (average 2.75 cm). The number of episodes of lymphangitis per year decreased from 1.8 to 0.2. CONCLUSION: Preoperative assessment is essential to select patients who can benefit from surgery for lymphedema and to choose the best surgical approach in each case. Our satisfactory results in well-selected cases encourage further research into surgical treatment for BCRL.

Autologous reconstruction of a complex form of Poland syndrome using 2 abdominal perforator free flaps.

Poland syndrome is the most frequent cause of congenital breast aplasia and hypoplasia. Breast and possible chest wall deformities can be treated with several surgical techniques, including implants, and pedicled or free flaps.We describe the case of a young patient with severe Poland syndrome with amastia, athelia, and deformity of the chest wall, and aplasia of 2 ribs. Marked hypoplasia of the ipsilateral latissimus dorsi muscle ruled out a reliable reconstructive option.Two perforator flaps were performed in a single-stage operation. A hemi-deep inferior epigastric perforator flap was harvested to correct the chest deformity, whereas the contralateral superficial inferior epigastric artery flap allowed breast reconstruction.No complications occurred and a subjectively and objectively pleasing cosmetic result was maintained at 3-year follow-up.

A case of donor-site lymphoedema after lymph node-superficial circumflex iliac artery perforator flap transfer.

Vascularised lymph node transfer is a promising technique to treat limb lymphoedema, especially when caused by lymph node dissection. The most common approach is the transfer of superficial inguinal lymph nodes using groin flaps or superficial circumflex iliac artery perforator flaps. Lower-limb lymphatic sequelae are unexpected as these lymph nodes should drain lymph from the lower abdominal wall. Recently, Vignes et al. described two cases out of 26 cases of chronic lymphoedema after superficial inguinal lymph node harvest. From a series of 42 vascularised lymph node transfers performed at our centre, only one patient developed swelling in the donor thigh. The features of this patient who underwent a lymph node-containing superficial circumflex iliac artery perforator flap are reported herein. We recommend maximal accuracy in selecting the appropriate lymph nodes for transfer and provide some tips from our experience.