Multiple injections of indocyanine green for improving the outcomes of lymphatic-venous anastomosis reconstruction of lymphatic return under microscopy: A case report

Abstract

This study explores effective treatment methods for chronic secondary lymphedema after radical cervical cancer surgery combined with pelvic lymphadenectomy. In cases where conservative treatment was ineffective, we investigated whether multiple injections of indocyanine green can effectively improve the outcomes of lymphatic-venous anastomosis under microscopy. Preoperative lymphatic imaging was used to localize functional vessels, guiding distal left lower limb lymphatic reconstruction. Intraoperatively, supermicroscopy (40×) and real-time indocyanine green imaging enabled eight end-to-end anastomoses between 0.5- and 1.3-mm lymphatic capillaries and subcutaneous veins. Each anastomosis took ≤30 min, with intraoperative lymphangiography confirming patency. Immediate postoperative lymphatic diversion to veins was observed. By day 3, reconstructed pathway efficiency reached 83% of the healthy side, with visual analog scale pain scores decreasing from 5 to 2. At 3–6 months, mid-thigh and calf circumferences reduced progressively; ultrasound confirmed edema resolution and complete cessation of exudation. High-precision multipoint supermicrosurgical anastomosis achieved anatomical and functional reconstruction of chronic secondary lymphedema, overcoming traditional size and functional limitations. Innovations included multipoint design to prevent reobstruction and dynamic imaging-guided precision. This case establishes a radical treatment strategy for chronic secondary lymphedema.

Introduction

Chronic secondary lymphedema (CSL) is a progressive inflammatory-fibrotic disorder caused by lymphatic drainage dysfunction, primarily secondary to tumor treatments (e.g. lymph node dissection and radiotherapy), infections, or trauma. Clinically, it manifests as progressive limb swelling, cutaneous lymphatic follicular hyperplasia, tension-type pain, and persistent exudation. The condition may advance to severe lymphedema, such as “rubber legs” or “rubber hands,” significantly impairing patients’ daily life and work. In microsurgical treatment for CSL, lymphatic-venous anastomosis (LVA) is considered the gold standard procedure, aiming to reconstruct lymphatic drainage pathways and alleviate symptoms. However, its efficacy critically depends on precise intraoperative identification of functional lymphatic vessels. ¹ Traditional intraoperative lymphatic visualization relies on anatomical experience or single-point injections of indocyanine green (ICG) for fluorescence imaging. In chronic pathological regions, lymphatic vessels are often obscured due to fibrosis, narrow lumens, or sparse distribution, complicating visualization. ² Multisite injection can cover the main lymphatic drainage area of the lower limbs, significantly improve the detection rate of lymphatic vessels, and avoid the imaging obstacles caused by the anatomical differences of lymphatic vessels. In addition, multisite injection can locate the unblocked “compensatory lymphatic vessels” by increasing the number of injection points, providing more options for anastomosis. Recent advancements in ICG multipoint injection techniques involve subcutaneous injections of contrast agents across multiple regions, combined with near-infrared fluorescence imaging to dynamically track multisegment lymphatic networks. This approach significantly improves lymphatic visualization coverage and targeting efficiency. The technique differentiates patent lymphatics with drainage potential from occluded pathways, aiding surgeons in selecting optimal anastomotic targets while minimizing tissue damage. Furthermore, real-time multipoint imaging with dynamic assessment enables postoperative evaluation of lymphodynamic improvements, providing a foundation for surgical refinement and promoting precision individualized LVA protocols.

Case report

A 76-year-old woman was admitted to the Affiliated People’s Hospital of Ningbo University in January 2025 due to left lower limb edema for 8 years and worsening for 2 years. The medical history and personal history were unremarkable. The patient developed left lower limb edema and mild pain >8 years ago after cervical cancer surgery, without numbness in the lower limbs, fever, chills, abdominal pain, diarrhea, nausea, vomiting, lower limb movement disorders, headache, dizziness, chest tightness, shortness of breath, hematemesis, melena, chest pain, or palpitations. The patient did not pay attention to these symptoms and did not visit the hospital. Over the past 2 years, the swelling in the left lower limb has worsened, accompanied by significant pain. The patient’s visual analog scale (VAS) score for pain was 5. Preoperative magnetic resonance imaging (MRI) enhancement (3.0T) of the left lower limb suggested a difference in circumference between both sides (Figure 1(a)). Physical examination also indicated a left thigh circumference of 47.5 cm, right thigh circumference of 40.5 cm, left calf circumference of 33.2 cm, and right calf circumference of 27.3 cm (Figure 1(b)). The skin of the left lower limb showed sclerodermatous changes, and the skin temperature was slightly lower. Preoperative venography of the left lower limb showed poor visualization of the left common iliac vein, revealing a compression shadow with significant collateral circulation formation. The deep veins of the left lower limb, including the anterior tibial vein, posterior tibial vein, popliteal vein, and femoral vein, were clearly visualized and uniformly filled (Figure 2(a) to (c)). Furthermore, left lower extremity ultrasound revealed subcutaneous soft tissue edema and thickening with a cobblestone-like pattern, accompanied by subcutaneous lymphatic vessel dilation. The widest segment was observed at the medial malleolus, measuring approximately 1.3 mm in diameter. Based on the patient’s condition, LVA was the recommended primary treatment for lower limb lymphedema. When diameter congruence is achieved between lymphatic and venous vessels, LVA employs an end-to-end anastomosis. Conversely, an end-to-side configuration is utilized when dimensional mismatch persists, optimizing patency and hemodynamic compatibility. LVA has good efficacy for early- to mid-stage (stage 0 to II) lymphedema, but the treatment effect for late-stage (stage III) lymphedema remains controversial. In the early stages, LVA can significantly reduce lymphedema. However, over time, the pressure in the lymphatic vessels gradually decreases until the pressure difference between the lymphatic vessels and veins returns to normal. This may lead to complications such as lymph fluid reflux and thrombosis, thereby affecting the long-term efficacy of LVA. Therefore, multiple injections of ICG are used to achieve precise anastomosis between veins and lymphatic vessels and reduce postoperative complications. The patient provided informed consent for both treatment and publication of the case report, with study approval granted by the Institutional Ethics Committee. We have deidentified all patient details.

Figure 1.

(a) Preoperative contrast-enhanced magnetic resonance imaging (3.0T) demonstrated marked edema in the left lower limb compared with that in the right lower limb. (b) The patient’s left lower limb showed full and raised skin, illustrating the clinical presentation of preoperative left lower limb edema and (c) the patient presented with postoperative skin folds, showing postoperative improvement, with reduced edema in the left lower limb relative to preoperative findings.

Figure 2.

(a) Preoperative left lower extremity venography revealed suboptimal visualization of the left common iliac vein with an indentation and prominent collateral circulation, further corroborating chronic secondary lymphedema postradical hysterectomy with pelvic lymphadenectomy. In contrast, it demonstrated normal opacification of the anterior tibial, posterior tibial, popliteal, and femoral veins, with uniform filling and no evidence of venous thrombosis or arteriovenous fistula (b, c).

Perioperative management

The circumference of the left limb 10 cm above and below the knee was measured weekly before and after surgery, and the average of multiple measurements was taken. ICG was used for lymphatic vessel imaging to clarify the patency of lymphatic vessels before and after surgery, and incision markings were made.

Surgical procedure

After successful general anesthesia, the patient was placed in the supine position. Preoperatively, intradermal injections of ICG (0.05 mL, 2.5 mg/mL) were administered at the first, second, third, and fourth interdigital spaces as well as the medial and lateral aspects of the plantar arch. Following injection, localized cutaneous massage was performed for 30 min. Near-infrared fluorescence imaging was performed, the ICG fluorescence excitation and reception equipment was set up, and ICG lymphangiography was performed (Figure 3(a)). Based on the imaging results, the clearly displayed lymphatic vessels were marked for future use. At the upper thigh root on the left side, a tourniquet was applied, and the left lower limb was routinely disinfected and draped. Under a microscope with 40×–60× magnification, a 3-cm incision was made on the surface of the marked lymphatic vessel at the left medial malleolus. The skin was incised, and the lymphatic vessel was located and identified within the superficial fascia layer. An appropriate small vein was selected nearby (Figure 3(b)), and the distal end of the lymphatic vessel and the proximal end of the small vein were anastomosed end-to-end with 10-0, 11-0, and 12-0 sutures. The direction of the needle was at an angle of 30° to the long axis of the blood vessel. At the same time, the proximal end of the lymphatic vessel and the distal end of the vein were ligated, and the anastomosis was rinsed with heparin (Figure 3(c)). After the anastomosis was completed, another ICG lymphangiography was performed and the distal lymphatic vessels were massaged, revealing the lymphatic fluid entering the venous lumen to confirm the patency of the anastomosis. Two lymphatic vessels were anastomosed to the vein at the medial malleolus (Figure 3(d)). Using the same method, two lymphatic vessels were anastomosed to the veins at the lateral malleolus, one lymphatic vessel to a vein at the mid-calf, one lymphatic vessel to a vein below the knee, and two lymphatic vessels to the veins above the knee. After the surgery, the left lower limb was wrapped with an elastic bandage for compression. The subcutaneous tissue of the wound was intermittently sutured with 4-0 absorbable suture, and the skin was continuously sutured with lock stitch. After the operation, the left lower limb was bandaged with elastic bandage under pressure; the affected limb was elevated and immobilized; the incision was kept clean and dry; and the bleeding, skin temperature, and limb swelling were closely observed. Ankle pump exercises were gradually performed, along with high-protein and low-sodium diet. Given the patient's long-term bed rest after surgery and to prevent deep venous thrombosis, enoxaparin (1 mg/kg, q12h) was used for anticoagulation.

Figure 3.

(a) Preoperative near-infrared fluorescence imaging with indocyanine green (ICG) injections at multiple sites near the ankle of the left lower limb demonstrated lymphatic vessels. (b) Following this, the skin was incised, and the lymphatic and venous structures within the superficial veins were dissected. (c) A functionally patent lymphatic vessel with anatomic compatibility in diameter to the adjacent vein was selected for anastomosis and (d) intraoperative ICG angiography was subsequently performed to confirm successful lymphatic-venous anastomosis and patency of the reconstructed pathway.

Results and prognosis

Postoperatively, the patient experienced mild pain in the left lower limb, with a VAS score of 2. The patient was alert and exhibited normal mood, with no abnormalities in the cardiopulmonary system. There was slight swelling on the dorsum of the foot, and the left lower limb was wrapped with an elastic bandage for compression. The lower limb incision was dry with no exudate or hematoma, and the dorsalis pedis artery pulse was normal (Figure 1(c)). The lymphoscintigraphy results within 3 days postsurgery are summarized in Table 1. Additionally, the patient showed no signs of chills and fever, no lower limb numbness, no lymphatic leakage, and no skin necrosis, and the left lower limb had good mobility. Laboratory tests indicated that D-dimer and coagulation function were both normal. Additionally, the patient required anticoagulant therapy postoperatively to prevent deep vein thrombosis. At the same time, attention should be paid to the monitoring of the swelling and peripheral circulation of the left lower limb. The swelling characteristics of the left lower limb 3–6 months postoperatively are shown in Table 2. A comparison of the circumference of the patient's lower limbs before and after surgery is detailed in Table 3. The lower limb circumference presented in Table 3 indicates the average of the diameters above and below the knee of a single limb. The reporting of this study conforms to the Case Report (CARE) guidelines. ³

Table 1.

Preoperative versus postoperative lymphodynamic comparison (dynamic indocyanine green fluorescence imaging).

	Contralateral side (right lower limb)	Preoperative affected side (left lower limb)	Postoperative affected side (left lower limb)	Improvement rate (%)
Lymphatic vessel density (vessels/cm2)	8.2 ± 1.3	2.1 ± 0.7	6.9 ± 1.1	229
Tracer flow velocity (mm/min)	12.5 ± 2.1	3.2 ± 1.4	10.4 ± 1.8	225
Drainage efficiency (%)	100	23 ± 5	83 ± 7	261
Lymphatic visualization rate (%)	95	18	78	333

Table 2.

Preoperative versus postoperative circumference comparison of the left lower limb (above and below the knee).

	Above the knee		Below the knee
	Mean circumference (cm)	Percentage reduction in circumference (%)	Mean circumference (cm)	Percentage reduction in circumference (%)
Preoperative	47.50	0	33.20	0
Postoperative 3 months	43.10	9.26	30.30	8.73
Postoperative 6 months	40.30	15.16	27.50	17.17

Table 3.

Comparison of circumference of both lower limbs between preoperative and postoperative patients.

	Lower limb circumference		Swelling rate (%)
	Left lower limb (cm)	Right lower limb (cm)
Preoperative	40.35	33.90	19.03
Postoperative 3 months	36.70	33.90	8.26
Postoperative 6 months	33.90	33.90	0

Discussion

In microsurgical LVA, the multipoint injection technique of ICG enhances the visualization of the lymphatic system, allowing the clear identification of functional lymphatic vessels and veins. This technique involves multiple subcutaneous or intradermal injections (usually 3–8 sites per limb), combined with a near-infrared fluorescence imaging system, which can dynamically display the anatomical structure and drainage dynamics of lymphatic vessels, providing important guidance for the intraoperative selection of functional lymphatic vessels. The role of this technology mainly manifests in several aspects. The ICG multipoint injection technique can achieve precise anastomosis of functional lymphatic vessels and veins, increasing the success rate of anastomosis and reducing postoperative complications. Additionally, the ICG multipoint injection technique can shorten the surgery time and improve surgical efficiency. This case further demonstrates that by combining advanced fluorescence imaging technology with meticulous microsurgical techniques, key challenges in lymphatic vessel repair can be effectively addressed, including insufficient visualization of diseased lymphatic vessels and the maintenance of patency at the anastomosis site. Therefore, the combination of multipoint ICG injection and LVA shows significant advantages in the treatment of chronic lymphedema. Traditional single-point ICG injection can only show local drainage pathways, while multipoint injection can comprehensively assess the compensatory status of the lymphatic network. In a comparative study of 25 patients with upper limb lymphedema, the multipoint injection group achieved a lymphatic vessel detection rate of 92.3% (23/25 cases), significantly higher than that (68%; 17/25 cases) observed in the single-point injection group. ⁴ Especially in cases of chronic transplant limb lymphedema, multipoint injection can identify secondary lymphatic vessels with diameters of 0.3–0.8 mm (an average of 4.2 ± 1.1 vessels per case), of which 78.6% have sustained drainage function (flow rate ≥0.1 cm/s). ⁵ Animal experiments further confirmed that using the second-generation near-infrared imaging system (NIR-II), multipoint injections can achieve a resolution of 0.2 mm for lymphatic imaging in pig models, showing a 42% improvement over the traditional NIR-I system. ⁶ This is crucial for identifying small anastomotic targets. The integration of ICG multipoint injection with near-infrared real-time imaging demonstrates a high signal-to-noise ratio (SNR = 4.8), enabling effective penetration through fibrotic tissue. Dynamic fluorescence monitoring can screen functional lymphatic vessels with an ICG clearance rate of >50% at 10 min for anastomosis, achieving an immediate patency rate of 100%, whereas traditional end-to-side anastomosis has a patency rate of 78%–84% at 6 months. Three months postsurgery, ICG lymphangiography showed that the directed blood flow velocity recovered to 0.8 cm/s (preoperative 0.2 cm/s), and the subcutaneous fluid volume decreased by 82%, which is better than the 33% recurrence rate of simple compression therapy. This technique can also avoid 38% of sclerosed lymphatic vessels (lumen occlusion >50%), shorten the surgery time by 40 min, and maintain a safe ICG dosage (≤10 mg). ⁷ The key to the anastomosis technique lies in precisely connecting the lymphatic vessels to the venous vessels to ensure that lymph fluid can effectively drain from the obstructed area. The multipoint localization strategy guided by real-time ICG imaging allows the surgeon to select the best anastomosis site based on the hemodynamic characteristics of the lymphatic vessels. In a cohort of 17 patients undergoing overlapping lock-stitch anastomosis, multipoint injection-assisted lymphatic mapping enabled an average of 5.2 ± 1.8 anastomoses per case, with a postoperative 1-week limb circumference reduction rate of 24.3% ± 5.6%. For cases with diameter mismatch (lymphatic vessel:vein = 1:1.5–2.0), ICG visualization revealed dynamic relationships between lymphatic dilation rates and venous compliance, allowing overlapping end-to-end anastomosis to achieve a 100% success rate with an average operative time of 5.3 ± 1.2 min. ⁸ In addition, in the treatment of recurrent lymphatic fistulas, multipoint injection localization maintained 14 anastomoses in 8 patients patent for 1 year postoperatively, achieving a 100% closure rate for lymphatic fistulas. ⁹ In addition, during the anastomosis technique, the biocompatibility and stability of the anastomosis site should be considered to ensure long-term drainage effectiveness. In recent years, the end-to-end anastomosis technique for lymphatic-venous vessels has also been continuously advancing. Synthetic hydrogels, by optimizing their physical and biochemical properties, can effectively promote the sprouting of collecting lymphatic vessels and the reconstruction of lymphatic networks. ¹⁰ This method not only provides lymphatic vascular graft materials to restore lymphatic continuity at the damaged site but also avoids secondary damage at the donor site. Although the end-to-end anastomosis technique of lymphatic-venous vessels has potential in reconstructing lymphatic return, this field still faces numerous challenges. For example, the precision required for anastomosis techniques is high, the surgical difficulty is substantial, and the postoperative outcomes are influenced by various factors. Therefore, we need to improve the success rate of surgeries as well as explore new biomaterials and treatment methods to enhance the therapeutic effects and quality of life for lymphedema patients. For example, using bioengineered materials as carriers for growth factors can more precisely activate lymphangiogenesis at the injury site. Correspondingly, one method mentioned in the literature is the use of hydrophobic silicone tubes to create artificial lymphatic drainage channels, which provides a new approach for the treatment of lymphedema. ¹¹ Additionally, studies have reported the stimulation of lymphangiogenesis through gene delivery technology as well as the use of prevascularized lymphoid tissue implants to promote the formation of new lymphatic capillary networks at the injury site. ¹² The future development of LVA technology can shift from purely mechanical channel reconstruction to a new model that combines functional repair with bioengineering. Through interdisciplinary innovation, it may be possible in the future to overcome the anatomical limitations of traditional surgery and achieve physiological regulation of lymphatic reflux. Driven by precision medicine and regenerative medicine, this field is evolving toward a three-pronged approach of developing intelligent devices, dynamic functional assessments, and targeted molecular regulation, providing more comprehensive solutions for lymphatic system diseases.

In microsurgical techniques, the use of hybrid operating rooms makes it possible to combine open surgery and endovascular techniques for comprehensive treatment. The setup of this hybrid operating room is equally enlightening for lymphatic reconstruction surgeries, as it allows for both open surgery and endovascular procedures to be performed simultaneously in the same operation. Additionally, for stubborn lymphatic complications that are unresponsive to traditional treatments, multiple ICG injections have shown unique advantages. In a case of refractory ascites secondary to postradiation abdominal lymphatic injury, eight intradermal ICG injection sites were established in the lower limbs to localize functional lymphatics, successfully creating 14 cross-regional LVAs. Postoperative 3.5-year follow-up confirmed complete resolution of ascites. ¹³ In cases of chronic transplant limb rejection accompanied by lymphatic congestion, quantitative analysis of multiple injection sites showed that the lymphatic drainage speed in the transplant segment reduced by 63% compared with that in the normal limb (0.12 vs. 0.32 cm/s). Based on this, three proximal high-flow (>0.2 cm/s) lymphatic vessels were selected for anastomosis, resulting in a 58% reduction in edema volume 6 months postsurgery. ¹⁴ Although ICG multipoint injection has significant advantages, its effectiveness is affected by the degree of tissue fibrosis. In inflammatory stage lymphedema (ISL), especially stage III edema patients, subcutaneous fat fibrosis reduced the success rate of ICG imaging to 67% (six of the nine cases), ¹⁵ at which point magnetic resonance (MR) lymphangiography needs to be combined for three-dimensional reconstruction. Photoacoustic imaging (PAI)-based postoperative evaluation revealed an 86.7% anastomotic patency rate (26/30) in the multipoint injection group, although 13.3% of anastomoses exhibited intermittent visualization due to lymphatic cyclic contraction, suggesting the necessity of dynamic observation protocols. ¹⁶ Additionally, studies have demonstrated that increasing ICG injection concentration from 0.1% to 0.3% enhances lymphatic visualization rates in severely fibrotic regions from 42% to 68%. ¹⁷ However, the analysis of existing clinical data shows that the multipoint ICG injection technique, by displaying the structure and function of the lymphatic system in multiple dimensions, has transformed LVA surgery from an experience-based approach to a precision medicine model. In the future, by combining NIR-II imaging and dynamic flow analysis, it is expected to further optimize injection site selection and anastomosis strategies. In summary, microsurgical techniques have broad application prospects in lymphatic reconstruction. Through continuous innovation and technological improvements, these techniques are expected to provide more effective treatment options for patients with lymphedema and other lymphatic drainage disorders. This is a case study with certain limitations. Further efforts can be made to increase the sample size and conduct multicenter research in the future.

Conclusion

This study successfully achieved the three-dimensional structural repair and physiological drainage function restoration of the lymphatic system in patients with CSL through an innovative multiregion anastomosis strategy and real-time imaging navigation technology. Unlike traditional surgical methods that rely on a single large lymphatic vessel, this technical system effectively avoids the risk of secondary obstruction caused by local tissue fibrosis through submillimeter ultra-microsurgical operations and multichannel drainage path design. During surgical procedures, dynamic fluorescence tracing technology enables precise identification of functional microlymphatic networks, while the integration of cross-regional gradient pressure balance principles ensures that targeted anastomotic sites achieve sustained lymphatic drainage. This innovative technique provides a reproducible solution for addressing chronic lower limb edema and tissue remodeling associated with advanced lymphatic circulatory dysfunction.