Update February 2021

Numerous diseases have been reported to affect the lymphatic system. As such, several strategies have been developed to deliver chemotherapeutics to this specific network of tissues and associated organs. Nanotechnology has been exploited as one of the main approaches to improve the lymphatic uptake of drugs. Different nanoparticle approaches utilized for both active and passive targeting of the lymphatic system are discussed here. Specifically, due to the rich abundance of lymphatic capillaries in the dermis, particular attention is given to this route of administration, as intradermal administration could potentially result in higher lymphatic uptake compared to other routes of administration. Recently, progress in microneedle research has attracted particular attention as an alternative for the use of conventional hypodermic injections. The benefits of microneedles, when compared to intradermal injection, are subsequently highlighted. Importantly, microneedles exhibit particular benefit in relation to therapeutic targeting of the lymphatic system, especially when combined with nanoparticles, which are further discussed. However, despite the apparent benefits provided by this combination approach, further comprehensive preclinical and clinical studies are now necessary to realize the potential extent of this dual-delivery platform, further taking into consideration eventual usability and acceptability in the intended patient end-users.

Normal growth and development of lymphatic structures depends on mechanical forces created by accumulating interstitial fluid. However, prolonged exposure to pathologic mechanical stimuli generated by chronically elevated lymph flow results in lymphatic dysfunction. The mechanisms that transduce these mechanical forces are not fully understood. Our objective was to investigate molecular mechanisms that alter the growth and metabolism of isolated lymphatic endothelial cells (LECs) exposed to prolonged pathologically elevated lymph flow in vivo within the anatomic and physiologic context of a large animal model of congenital heart disease with increased pulmonary blood flow using in vitro approaches. To this end, late gestation fetal lambs underwent in utero placement of an aortopulmonary graft (shunt). Four weeks after birth, LECs were isolated and cultured from control and shunt lambs. Redox status and proliferation were quantified, and transcriptional profiling and metabolomic analyses were performed. Shunt LECs exhibited hyperproliferative growth driven by increased levels of Hypoxia Inducible Factor 1alpha (HIF-1alpha), along with upregulated expression of known HIF-1alpha target genes in response to mechanical stimuli and shear stress. Compared to control LECs, shunt LECs exhibited abnormal metabolism including abnormalities of glycolysis, the TCA cycle and aerobic respiration. In conclusion, LECs from lambs exposed in vivo to chronically increased pulmonary lymph flow are hyperproliferative, have enhanced expression of HIF-1alpha and its target genes, and demonstrate altered central carbon metabolism in vitro. Importantly, these findings suggest provocative therapeutic targets for patients with lymphatic abnormalities.

Macrophages are components of the innate immune system with key roles in tissue inflammation and repair. It is now evident that macrophages also support organogenesis, but few studies have characterized their identity, ontogeny and function during heart development. Here, we show that the distribution and prevalence of resident macrophages in the subepicardial compartment of the developing heart coincides with the emergence of new lymphatics and macrophages interact closely with the nascent lymphatic capillaries. Consequently, global macrophage-deficiency led to extensive vessel disruption with mutant hearts exhibiting shortened and mis-patterned lymphatics. The origin of cardiac macrophages was linked to the yolk sac and fetal liver. Moreover, the Cx3cr1 (+) myeloid lineage was found to play essential functions in the remodeling of the lymphatic endothelium. Mechanistically, macrophage hyaluronan was found to be required for lymphatic sprouting by mediating direct macrophage-lymphatic endothelial cell interactions. Together, these findings reveal insight into the role of macrophages as indispensable mediators of lymphatic growth during the development of the mammalian cardiac vasculature.

Fsp1 (a.k.a S100A4 or Metastatin) is an intracellular and secreted protein widely regarded as a fibroblast marker. Recent studies have nonetheless shown that Fsp1 is also expressed by other cell types, including small subsets of endothelial cells. Since no detailed and systematic description of Fsp1 spatio-temporal expression pattern in cardiac vascular cells is available in the literature, we have used a transgenic murine line (Fsp1-GFP) to study Fsp1 expression in the developing and postnatal cardiac vasculature and endocardium. Our work shows that Fsp1 is expressed in the endocardium and mesenchyme of atrioventricular valve primordia, as well as in some coronary venous and lymphatic endothelial cells. Fsp1 expression in cardiac venous and lymphatic endothelium is progressively restricted to the leaflets of cardiac venous and lymphatic valves. Our results suggest that Fsp1 could play a role in the development of atrioventricular valves and participate in the patterning and morphogenesis of cardiac venous and lymphatic vessel valves.

Lymph node metastases in cancer patients are associated with high aggressiveness, poor prognosis, and short survival time. The chemokine receptor 4 (CXCR4)/stroma derived factor 1alpha (CXCL12) biological axis plays a critical role in the spread of cancer cells. Designing effective delivery systems that can successfully deliver CXCR4 antagonists to lymph nodes, which are rich in CXCR4-overexpressing cancer cells, for controlling cancer metastasis remain challenging. In this study, we demonstrated that such a challenge may be alleviated by developing nanometer-sized polyethylene glycol-phosphatidylethanolamine (PEG-PE) micelles for the co-delivery of the CXCR4 antagonistic peptide E5 and doxorubicin (M-E5-Dox). This nanomicelle platform enables the preferential accumulation of cargos into lymph nodes and thus can better inhibit cancer metastasis and enhance antitumor efficacy than either free drugs or single drug-loaded micelles in breast cancer-bearing mouse models. Hence, M-E5-Dox is expected to be a potential therapeutic agent that would improve the clinical benefits of breast cancer therapy and treatment of various CXCR4-overexpressing malignancies.

Pharmacological openers of ATP-sensitive potassium (KATP) channels are effective antihypertensive agents, but off-target effects, including severe peripheral edema, limit their clinical usefulness. It is presumed that the arterial dilation induced by KATP channel openers (KCOs) increases capillary pressure to promote filtration edema. However, KATP channels also are expressed by lymphatic muscle cells (LMCs), raising the possibility that KCOs also attenuate lymph flow to increase interstitial fluid. The present study explored the effect of KCOs on lymphatic contractile function and lymph flow. In isolated rat mesenteric lymph vessels (LVs), the prototypic KATP channel opener cromakalim (0.01–3 micromol/l) progressively inhibited rhythmic contractions and calculated intraluminal flow. Minoxidil sulfate and diazoxide (0.01–100 micromol/l) had similar effects at clinically relevant plasma concentrations. High-speed in vivo imaging of the rat mesenteric lymphatic circulation revealed that superfusion of LVs with cromakalim and minoxidil sulfate (0.01–10 micromol/l) maximally decreased lymph flow in vivo by 38.4% and 27.4%, respectively. Real-time polymerase chain reaction and flow cytometry identified the abundant KATP channel subunits in LMCs as the pore-forming Kir6.1/6.2 and regulatory sulfonylurea receptor 2 subunits. Patch-clamp studies detected cromakalim-elicited unitary K(+) currents in cell-attached patches of LMCs with a single-channel conductance of 46.4 pS, which is a property consistent with Kir6.1/6.2 tetrameric channels. Addition of minoxidil sulfate and diazoxide elicited unitary currents of similar amplitude. Collectively, our findings indicate that KCOs attenuate lymph flow at clinically relevant plasma concentrations as a potential contributing mechanism to peripheral edema. SIGNIFICANCE STATEMENT: ATP-sensitive potassium (KATP) channel openers (KCOs) are potent antihypertensive medications, but off-target effects, including severe peripheral edema, limit their clinical use. Here, we demonstrate that KCOs impair the rhythmic contractions of lymph vessels and attenuate lymph flow, which may promote edema formation. Our finding that the KATP channels in lymphatic muscle cells may be unique from their counterparts in arterial muscle implies that designing arterial-selective KCOs may avoid activation of lymphatic KATP channels and peripheral edema.

The lymphatic vasculature has a pivotal role in regulating body fluid homeostasis, immune surveillance and dietary fat absorption. The increasing number of in vitro and in vivo studies in the last decades has shed light on the processes of lymphatic vascular development and function. Here, we will discuss the current progress in lymphatic vascular biology such as the mechanisms of lymphangiogenesis, lymphatic vascular maturation and maintenance and the emerging mechanisms of lymphatic vascular aging.

Drug delivery to the lymphatic system is gaining increasing attention, particularly in fields such as immunotherapy where drug access to lymphocytes is central to activity. We have previously described a prodrug strategy that facilitates the lymphatic delivery of a model immunomodulator, mycophenolic acid (MPA) via incorporation into intestinal triglyceride transport pathways. The current study explored a series of structurally related glyceride and phospholipid mimetic prodrugs of MPA in an attempt to enhance lymph targeting and to better elucidate the design criteria for lipid mimetic prodrugs. MPA was conjugated to a glyceride or phospholipid backbone at various positions using different spacers employing ester, ether, carbonate and amide bonds. Patterns of prodrug hydrolysis were evaluated in rat digestive fluid, and lymphatic transport and plasma pharmacokinetics were assessed in lymph duct cannulated rats. Prodrugs with different spacers between MPA and the glyceride backbone resulted in up to 70-fold differences in gastrointestinal stability. MPA conjugation at the 2 position of the glyceride backbone and via an ester bond were most effective in promoting lymphatic transport. Phospholipid prodrug derivatives, or glyceride derivatives with MPA attached at the 1 position or when linked via ether, carbonate or amide bonds were poorly incorporated into lymphatic transport pathways.

Metabolism plays a pivotal role in the formation of the lymphatic vasculature. Pyruvate kinase M2 (PKM2) is typically a metabolic marker of proliferating cells and maintains the growth of vascular endothelial cells. In this study, the potential status of PKM2 in lymphatic endothelial cells and the pathogenesis of lymphatic malformations (LMs) was investigated. The glycolysis index, including glucose uptake, ATP, and lactate production, stayed at a relatively high level in human dermal lymphatic endothelial cells (HDLECs) compared with human umbilical vein endothelial cells, whereas the inhibition of PKM2 by shikonin or PKM2 knockdown significantly suppressed glycolysis, migration, tubular formation, and invasion of HDLECs. Moreover, compared with lymphatic vessels in healthy skin, lymphatic vessels of LMs expressed PKM2 highly, and this expression correlated with infection of LMs. Meanwhile, the overexpression of PKM2 in HDLECs strengthened the proliferation, migration, tubular formation, and invasion of HDLECs. The findings from further experiments in a rat LM model support that targeting PKM2 by shikonin significantly impedes the progression of LMs, even in an infected LM rat model. Taken together, these results indicate that PKM2 plays a pivotal role in the activation of LECs and promotes the progression of LMs, whereas the inhibition of PKM2 can effectively suppress the pathogenesis of LM lesions in the rat model.

Orlistat is a pancreatic lipase (PL) inhibitor that inhibits dietary lipid absorption and is used to treat obesity. The oral bioavailability of orlistat is considered zero after administration in standard formulations. This is advantageous in the treatment of obesity. However, if orlistat absorption could be improved it has the potential to treat diseases such as acute and critical illnesses where PL transport to the systemic circulation via gut lymph promotes organ failure. Orlistat is highly lipophilic and may associate with intestinal lipid absorption pathways into lymph. Here we investigate the potential to improve orlistat lymph and systemic uptake through intestinal administration in lipid formulations (LFs). The effect of lipid type, lipid dose, orlistat dose, and infusion time on lymph and systemic availability of orlistat was investigated. After administration in all LFs, orlistat concentrations in lymph were greater than in plasma, suggesting direct transport via lymph. Lymph and plasma orlistat derivative concentrations were approximately 8-fold greater after administration in a long-chain fatty acid (LC-FA) compared to a lipid-free, LC triglyceride (LC-TG) or medium-chain FA (MC-FA) formulation. Overall, administration of orlistat in a LC-FA formulation promotes lymph and systemic uptake which may enable treatment of diseases associated with elevated systemic PL activity.

The lymphatic vasculature has an essential role in maintaining normal fluid balance in tissues and modulating the inflammatory response to injury or pathogens. Disruption of normal development or function of lymphatic vessels can have severe consequences. In the heart, reduced lymphatic function can lead to myocardial oedema and persistent inflammation. Macrophages, which are phagocytic cells of the innate immune system, contribute to cardiac development and to fibrotic repair and regeneration of cardiac tissue after myocardial infarction. In this Review, we discuss the cardiac lymphatic vasculature with a focus on developments over the past 5 years arising from the study of mammalian and zebrafish model organisms. In addition, we examine the interplay between the cardiac lymphatics and macrophages during fibrotic repair and regeneration after myocardial infarction. Finally, we discuss the therapeutic potential of targeting the cardiac lymphatic network to regulate immune cell content and alleviate inflammation in patients with ischaemic heart disease.

Almost 150 papers about brain lymphatics have been published in the last 150 years. Recently, the information in these papers has been synthesized into a picture of central nervous system (CNS) “glymphatics,” but the fine structure of lymphatic elements in the human brain based on imaging specific markers of lymphatic endothelium has not been described. We used LYVE1 and PDPN antibodies to visualize lymphatic marker-positive cells (LMPCs) in postmortem human brain samples, meninges, cavernous sinus (cavum trigeminale), and cranial nerves and bolstered our findings with a VEGFR3 antibody. LMPCs were present in the perivascular space, the walls of small and large arteries and veins, the media of large vessels along smooth muscle cell membranes, and the vascular adventitia. Lymphatic marker staining was detected in the pia mater, in the arachnoid, in venous sinuses, and among the layers of the dura mater. There were many LMPCs in the perineurium and endoneurium of cranial nerves. Soluble waste may move from the brain parenchyma via perivascular and paravascular routes to the closest subarachnoid space and then travel along the dura mater and/or cranial nerves. Particulate waste products travel along the laminae of the dura mater toward the jugular fossa, lamina cribrosa, and perineurium of the cranial nerves to enter the cervical lymphatics. CD3-positive T cells appear to be in close proximity to LMPCs in perivascular/perineural spaces throughout the brain. Both immunostaining and qPCR confirmed the presence of adhesion molecules in the CNS known to be involved in T cell migration.

The present research work was aimed to explore the ability of nanostructured lipid carriers (NLCs) to improve oral bioavailability of Nintedanib esylate (NE) via lymphatic uptake. The NE loaded NLCs (NE-NLCs) were fabricated using high speed homogenization followed by probe sonication method and physiochemically characterized. The NE-NLCs had particle size of 125.7 +/- 5.5 nm, entrapment efficiency of 88.5 +/- 2.5% and zeta potential of -17.3 +/- 3.5 mV. DSC and XRD studies indicated that NE was converted to amorphous form. TEM images showed uniformly distributed spherical shaped particles. In vitro release study of NE-NLCs showed drug release of 6.87 +/- 2.72% in pH 1.2 and 92.72 +/- 3.40% in phosphate buffer pH 6.8 and obeyed higuchi model. Lipolysis study showed higher amount of drug in aqueous layer in NE-NLCs compared to NE-suspension. Tissue distribution study showed deeper penetration of FITC loaded NLCs compared to FITC solution. The cellular uptake across Caco-2 cells exhibited more uptake of FITC loaded NLCs. Cytotoxicity study using A549 cell line revealed higher potential of NE-NLCs in inhibiting tumor cell growth in comparison to that of suspension. The oral bioavailability of NE was ameliorated over 26.31 folds after inclusion into NLCs in contrast to NE-suspension. Intestinal lymphatic uptake of NE-NLCs in cycloheximide treated mice was lower as compared to control without cycloheximide treatment. Thus, the developed NE-NLCs can be an encouraging delivery strategy for increasing oral bioavailability of NE via lymphatic uptake.

Various therapeutic approaches, for example, in case of trauma or cancer require the transplantation of autologous tissue. Depending on the size and the origin of the harvested tissue, these therapies can lead to iatrogenic complications and donor-site morbidities. In future, these side effects could be avoided by transplanting artificially generated tissue consisting of different cell types and matrix components derived from the host body. Tissue that is grown in the patient could be advantageous compared with the more simply structured in vitro-grown alternatives. To overcome the limitations of graft vascularization, the arteriovenous (AV) loop technique has been established for different tissues in the last years and was adapted for lymphatic tissue engineering in the present study. We utilized the AV loop technique to grow human lymphatic vasculature in vivo in the Rowett nude (RNU) rat. A combination of human lymphatic endothelial cells (LECs) and bone marrow-derived mesenchymal stem cells was implanted in a fibrin matrix surrounding the AV loop. After 2 or 4 weeks of implantation, the animals were perfused and the tissue was harvested. It could be demonstrated by immunohistochemistry for human LYVE1, human CD31, and murine podoplanin that the implanted cells formed human lymphatic vasculature in the AV loop chamber. Beside development of murine podoplanin-positive vasculature in the AV loop tissue, vasculature positive for human marker proteins developed in comparable numbers. This suggests that implanted LECs are able to improve the lymphatic vascularization of the newly engineered tissue. Thus, we were able to establish an in vivo tissue engineering method to generate lymphatic vascularized soft tissue. An axially vascularized transplantable lymphatic vessel network was engineered without requiring advanced cell culture equipment, rendering the lymphatic AV loop highly suitable for applied regenerative medicine. Impact statement Various surgical procedures require the transplantation of autologous harvested tissue, for example, the vascularized lymph node transfer for the treatment of lymphedema. Tissue-engineered transplants could be used instead of autologous transplants and thereby help to reduce the side effects of those therapies. However, in vitro tissue engineering of large constructs requires a lot of know-how as well as advanced cell culture equipment, which might not be accessible in every hospital. In vivo tissue engineering approaches like the presented technique for the generation of transplantable networks of lymphatic vasculature could serve as an alternative for in vitro tissue engineering approaches in clinical settings.

The lymphatic system is important for antigen presentation and immune surveillance. The lymphatic system in the brain was originally introduced by Giovanni Mascagni in 1787, while the rediscovery of it by Jonathan Kipnis and Kari Kustaa Alitalo now opens the door for a new interpretation of neurological diseases and therapeutic applications. The glymphatic system for the exchanges of cerebrospinal fluid (CSF) and interstitial fluid (ISF) is associated with the blood-brain barrier (BBB), which is involved in the maintenance of immune privilege and homeostasis in the brain. Recent notions from studies of postmortem brains and clinical studies of neurodegenerative diseases, infection, and cerebral hemorrhage, implied that the breakdown of those barrier systems and infiltration of activated immune cells disrupt the function of both neurons and glia in the parenchyma (e.g., modulation of neurophysiological properties and maturation of myelination), which causes the abnormality in the functional connectivity of the entire brain network. Due to the vulnerability, such dysfunction may occur in developing brains as well as in senile or neurodegenerative diseases and may raise the risk of emergence of psychosis symptoms. Here, we introduce this hypothesis with a series of studies and cellular mechanisms.

The objective of this study is to investigate the effect of hyperthermia-induced improvement of hydraulic conductivity and lymphatic function on both tumoral IFP reduction and nanoparticle delivery to PC3 tumors. We developed a theoretical model for nanoparticle transport in a tumor incorporating Starling's law, Darcy's law, transient convection, and diffusion of chemical species in porous media, and nanoparticle accumulation in tumors. Results have shown that both mechanisms were effective to decrease the IFP at the tumor center from 1600 Pa in the control without heating to 800 Pa in tumors with whole body heating. IFP reductions not only elevate the nanoparticle concentration in the tumor, but also result in a more uniform nanoparticle concentration in the tumor than that in the control without heating. Due to the IFP reductions at the tumor center and/or local blood perfusion increases, the final amount of accumulated nanoparticles in the tumor increased by more than 35–95% when compared to the control without heating. We conclude that increases in the hydraulic conductivity and recovery of lymphatic functions are possible mechanisms that lead to IFP reductions and enhancement in nanoparticle deposition in PC3 tumors observed in our in vivo experimental studies.

Lymphatic vessels provide an anatomical framework for immune surveillance and adaptive immune responses. Although appreciated as the route for Ag and dendritic cell transport, peripheral lymphatic vessels are often not considered active players in immune surveillance. Lymphatic vessels, however, integrate contextual cues that directly regulate transport, including changes in intrinsic pumping and capillary remodeling, and express a dynamic repertoire of inflammatory chemokines and adhesion molecules that facilitates leukocyte egress out of inflamed tissue. These mechanisms together contribute to the course of peripheral tissue immunity. In this review, we focus on context-dependent mechanisms that regulate fluid and cellular transport out of peripheral nonlymphoid tissues to provide a framework for understanding the effects of afferent lymphatic transport on immune surveillance, peripheral tissue inflammation, and adaptive immunity.

The lymphatic system has received increasing scientific and clinical attention because a wide variety of diseases are linked to lymphatic pathologies and because the lymphatic system serves as an ideal conduit for drug delivery. Lymphatic vessels exert heterogeneous roles in different organs and vascular beds, and consequently, their dysfunction leads to distinct organ-specific outcomes. Although studies in animal model systems have led to the identification of crucial lymphatic genes with potential therapeutic benefit, effective lymphatic-targeted therapeutics are currently lacking for human lymphatic pathological conditions. Here, we focus on the therapeutic roles of lymphatic vessels in diseases and summarize the promising therapeutic targets for modulating lymphangiogenesis or lymphatic function in preclinical or clinical settings. We also discuss considerations for drug delivery or targeting of lymphatic vessels for treatment of lymphatic-related diseases. The lymphatic vasculature is rapidly emerging as a critical system for targeted modulation of its function and as a vehicle for innovative drug delivery.

Lymphorrhea is a rare but potentially severe complication that occurs after various surgical procedures. Untreated lymphorrhea may lead to wound dehiscence, infection, and prolonged hospital stay. Currently, there is no standard effective treatment. Early management usually includes leg elevation, drainage, and pressure dressing. However, these methods are associated with prolonged recovery and high recurrence rates. We report a case of lymphorrhea from a calf wound after endoscopic great saphenous vein (GSV) harvesting for coronary artery bypass grafting (CABG). The patient presented with intractable oozing from the postoperative wound on the right calf. Lymphorrhea perGsisted for 6 weeks despite negative-pressure wound therapy with a long-acting somatostatin. We performed unilateral pedal lymphangiography that confirmed wound lymphorrhea, followed by glue embolization. No recurrence was observed after 8 months of follow-up. This case report demonstrates the successful use of lymphangiography with glue embolization in the control of lymphorrhea after GSV harvesting for CABG.

Animal studies implicate meningeal lymphatic dysfunction in the pathogenesis of neurodegenerative diseases such as Alzheimer's disease and Parkinson's disease (PD). However, there is no direct evidence in humans to support this role(1–5). In this study, we used dynamic contrast-enhanced magnetic resonance imaging to assess meningeal lymphatic flow in cognitively normal controls and patients with idiopathic PD (iPD) or atypical Parkinsonian (AP) disorders. We found that patients with iPD exhibited significantly reduced flow through the meningeal lymphatic vessels (mLVs) along the superior sagittal sinus and sigmoid sinus, as well as a notable delay in deep cervical lymph node perfusion, compared to patients with AP. There was no significant difference in the size (cross-sectional area) of mLVs in patients with iPD or AP versus controls. In mice injected with alpha-synuclein (alpha-syn) preformed fibrils, we showed that the emergence of alpha-syn pathology was followed by delayed meningeal lymphatic drainage, loss of tight junctions among meningeal lymphatic endothelial cells and increased inflammation of the meninges. Finally, blocking flow through the mLVs in mice treated with alpha-syn preformed fibrils increased alpha-syn pathology and exacerbated motor and memory deficits. These results suggest that meningeal lymphatic drainage dysfunction aggravates alpha-syn pathology and contributes to the progression of PD.

BACKGROUND: The World Health Organization has targeted lymphatic filariasis (LF) for elimination as a public health problem and recommends, among other measures, post-elimination surveillance of LF. The identification of sensitive and specific surveillance tools is therefore a research priority. The Wuchereria bancrofti-specific antigen Wb123-based enzyme-linked immunosorbent assay (Wb123 ELISA) detects antibodies to the recombinant Wb123 antigen of W. bancrofti and may be useful as a surveillance tool for LF. Six years after stopping mass drug administration to eliminate LF and recording successful results on two post-treatment transmission assessment surveys, a study was conducted in Togo aimed at helping to identify the role of the Wb123 ELISA in post-validation surveillance of LF. METHODS: This was a cross-sectional study in eight previously LF-endemic districts and one non-endemic district in Togo. In each sub-district of these nine districts, two schools were selected and 15 children aged 6 to 9 years old at each school provided finger-stick blood for testing for antibodies to Wb123 using the Filaria Detect IgG4 ELISA kit(R) (InBios, International, Inc., Seattle, WA, USA). RESULTS: A total of 2654 children aged 6 to 9 years old were tested in 134 schools in the nine districts. Overall, 4.7% (126/2654) children tested positive for antibodies to the Wb123 antigen of W. bancrofti. The prevalence of Wb123 antibodies varied across the eight previously endemic LF districts, from 1.56 to 6.62%. The highest prevalence, 6.99%, was found in the non-endemic district, but this was not significantly different from the average of all the LF districts (4.49%, P = 0.062). CONCLUSIONS: The Wb123 ELISA was positive in 4.7% of Togolese school-age children who were almost certainly unexposed to LF. This apparent lack of specificity in the Togo context makes it difficult to establish a seroprevalence threshold that could serve to signal LF resurgence in the country, precluding the use of this test for post-validation surveillance in Togo. There remains a need to develop a useful and reliable test for post-elimination surveillance for LF in humans.

Lymphatic filariasis is the major global cause of non-hereditary lymphoedema. We demonstrate the filarial nematode, Brugia malayi, induces lymphatic remodelling and impaired lymphatic drainage following parasitism of limb lymphatics in a mouse model. Lymphatic insufficiency was associated with elevated circulating lymphangiogenic mediators, including vascular endothelial growth factor C. Lymphatic insufficiency was dependent on type-2 adaptive immunity, interleukin-4 receptor, recruitment of C-C chemokine receptor-2 monocytes and alternatively-activated macrophages with pro-lymphangiogenic phenotype. Oral treatments with second-generation tetracyclines improved lymphatic function, while other classes of antibiotic had no significant effect. Second-generation tetracyclines directly targeted lymphatic endothelial cell proliferation and modified type-2 pro-lymphangiogenic macrophage development. Doxycycline treatment impeded monocyte recruitment, inhibited polarisation of alternatively-activated macrophages and suppressed T cell adaptive immune responses following infection. Our results determine a mechanism-of-action for the anti-morbidity effects of doxycycline in filariasis and supports clinical evaluation of second-generation tetracyclines as affordable, safe therapeutics for lymphoedemas of chronic inflammatory origin.

Clinicians and scientists at St George's University Hospital have collaborated to develop a classification algorithm for primary lymphatic anomalies. Instruction is offered on how to apply the algorithm in clinical practice to refine the diagnosis of primary lymphedema and guide on genetic testing and management. It can also be used to interpret mutation testing results of uncertain significance. The algorithm has evolved as more genes have been discovered, and it remains a “work in progress” as further discoveries are made. This transformational approach has revolutionized the understanding and classification of primary lymphatic anomalies.

Background: The standard examination for diagnosing lymphedema is lymphoscintigraphy, which has a disadvantage in versatility and radiation exposure. We have reported the usefulness of echography in observing the lymphatic degeneration. The purpose of this study was to investigate the usefulness of lymphatic ultrasound in diagnosing lymphedema. Methods and Results: The study included 14 patients (28 lower limbs) who underwent lymphaticovenous anastomosis for lower limb lymphedema. Preoperative echography with a common 18-MHz linear probe was used to detect lymphatic vessels. We evaluated abnormal expansion or sclerosis of lymphatic vessels in the medial legs, which indicated the presence of lymphedema. We proposed the method “D-CUPS” on how to detect and observe the lymphatic vessels. We then performed indocyanine green (ICG) lymphography to diagnose lymphedema. The results of examination were compared. Stage 1 lymphedema was diagnosed in 9 limbs, Stage 2a in 7, Stage 2b in 8, and Stage 3 in 4. Lymphatic vessel detection was possible in all 28 medial thighs and in 27 medial lower legs. The sensitivity and specificity for diagnosis of lymphedema based on echography of the medial leg were 95.0% and 100.0%, respectively. The accuracy rate was 94.6%. We could detect lymphatic vessels with echography in 39 of 54 areas that failed detection using lymphoscintigraphy or ICG lymphography (72.2%). Conclusion: The location and degeneration of lymphatic vessels in lymphedematous limbs can be evaluated with a commonly used ultrasound device. Although exclusion of comorbidities is still necessary, lymphatic ultrasound has potential for use in diagnosis of lymphedema or lymphatic dysfunction.

Secondary lymphedema is a life-long disorder characterized by chronic tissue swelling and inflammation that obstruct interstitial fluid circulation and immune cell trafficking. Regenerating lymphatic vasculatures using various strategies represents a promising treatment for lymphedema. Growth factor injection and gene delivery have been developed to stimulate lymphangiogenesis and augment interstitial fluid resorption. Using bioengineered materials as growth factor delivery vehicles allows for a more precisely targeted lymphangiogenic activation within the injured site. The implantation of prevascularized lymphatic tissue also promotes in situ lymphatic capillary network formation. The engineering of larger scale lymphatic tissues, including lymphatic collecting vessels and lymph nodes constructed by bioengineered scaffolds or decellularized animal tissues, offers alternatives to reconnecting damaged lymphatic vessels and restoring lymph circulation. These approaches provide lymphatic vascular grafting materials to reimpose lymphatic continuity across the site of injury, without creating secondary injuries at donor sites. The present work reviews molecular mechanisms mediating lymphatic system development, approaches to promoting lymphatic network regeneration, and strategies for engineering lymphatic tissues, including lymphatic capillaries, collecting vessels, and nodes. Challenges of advanced translational applications are also discussed.

RASopathies are caused by variants in genes encoding components or modulators of the RAS/MAPK signaling pathway. Noonan syndrome is the most common entity among this group of disorders and is characterized by heart defects, short stature, variable developmental delay, and typical facial features. Heterozygous variants in SOS2, encoding a guanine nucleotide exchange factor for RAS, have recently been identified in patients with Noonan syndrome. The number of published cases with SOS2-related Noonan syndrome is still limited and little is known about genotype-phenotype correlations. We collected previously unpublished clinical and genotype data from 17 individuals carrying a disease-causing SOS2 variant. Most individuals had one of the previously reported dominant pathogenic variants; only four had novel changes at the established hotspots for variants that affect protein function. The overall phenotype of the 17 patients fits well into the spectrum of Noonan syndrome and is most similar to the phenotype observed in patients with SOS1-related Noonan syndrome, with ectodermal anomalies as common features and short stature and learning disabilities as relatively infrequent findings compared to the average Noonan syndrome phenotype. The spectrum of heart defects in SOS2-related Noonan syndrome was consistent with the known spectrum of cardiac anomalies in RASopathies, but no specific heart defect was particularly predominating. Notably, lymphatic anomalies were extraordinarily frequent, affecting more than half of the patients. We therefore conclude that SOS2-related Noonan syndrome is associated with a particularly high risk of lymphatic complications that may have a significant impact on morbidity and quality of life.

Generalized lymphatic dysplasia (GLD), characterized by lymphedema, lymphangiectasias, chylothorax, effusions, represents a recognized cause of fetal hydrops. We describe for the first time recurrent pregnancies showing different ultrasound presentations of lymphatic dysplasia. The first fetus displayed diffuse subcutaneous cysts and septations while the second one presented fetal hydrops. Exome sequencing results at 18 gestational weeks in the second pregnancy showed compound heterozygosity for two novel PIEZO1 variants, afterwards detected also in the first fetus and in the heterozygous parents. Both ultrasound and genetic findings expand the current knowledge of PIEZO1-related GLD. We suggest exome sequencing in hydropic fetuses with normal cytogenetics and in pregnancies with recurrent hydrops/lymphatic dysplasia.

Background: To demonstrate the value of intranodal lymphangiography (INL) and lymphatic embolization (LE) in management of iatrogenic chylous ascites in children who fail conservative management. Methods and Results: Retrospective review of medical records revealed six patients (four males and two females; age range 11–39 months) who underwent eight INLs and three LEs between 2017 and 2019. In one patient, the leak was visualized and embolized. In three patients, the leak was not visualized and no embolization was done, but drain output decreased and INL was not repeated. In two patients, the leak was not visualized and no embolization was done, but drain output did not decrease and INL was repeated. Repeat INL identified a leak in one patient and targeted LE was done. Repeat INL did not identify a leak in the other patient, but empirical LE was performed at the sites suspicious for leakage after multidisciplinary discussion. INL was able to identify the site of lymphatic leak in two patients (33%). In the three patients who underwent LE (two targeted at the site of identified leak and one empirical at sites suspicious for leak), one (33%) was clinically successful and the other two required surgery to address the lymphatic leak. In three patients, chylous ascites resolved after INL alone with no additional interventions. Three patients developed complications after the procedure, but only one appears to be related to the procedure itself. Follow-up for 13.8 months (13–26 months) revealed no long-term complications or mortality. Conclusion: INL with or without LE is a safe treatment for iatrogenic pediatric chylous ascites. Early utilization before more invasive surgical intervention should be considered in light of the response to INL.

Magnetic resonance lymphangiography (MRL) is a noninvasive imaging technique that can be used in the management of lymphatic disorders to delineate the central lymphatic system for treatment planning. This article reviews the MRL technique, its advantages, limitations, indications, and impact on patient management. Level of Evidence 5 Technical Efficacy Stage 3 J. MAGN. RESON. IMAGING 2021;53:374–380.

AIM: To clarify the incidence of lymphatic drainage disorders (LLD) after treatment for cervical cancer (CC) and to establish univariate models for their occurrence. METHODS: A total of 263 eligible patients with CC were identified between 2010 and 2019. We conducted a case-control study and divided the study population into two subsamples of 12 and 251 CC survivors based on the presence/absence of LLD, respectively. The cumulative incidence was evaluated using the Kaplan-Meier method. Univariate models based on Pearson correlation coefficient were used to evaluate associations between explanatory variables and LLD. RESULTS: The cumulative incidence of LLD began at 3.3% after the 7-month follow-up, reaching a plateau of 21.2% between 130 and 250 months of follow-up. We detected correlation between LLD and number of removed para-aortic lymph nodes (r = −0.39), number of pelvic lymphadenopathies (r = 0.16), pelvic lymphadenectomy (PL) (r = 0.16), age at diagnosis of CC (r = −0.1) and primary surgery (r = 0.1). CONCLUSION: We observed a cumulative incidence of LLD of 21.2%, which is in accord with other retrospective studies. Number of removed para-aortic lymph nodes, number of pelvic lymphadenopathies, PL, age at diagnosis of CC and primary surgery were the most influential clinical factors associated with the occurrence of LLD in CC survivors.

AIMS: Lymphatic vessel density (LVD) for the evaluation of tumor metastasis and prognosis remains controversial. The aim of this study was to elucidate the association between tumor cells and lymphatic vessels, and evaluate LVD in oral squamous cell carcinoma (OSCC). MAIN METHODS: 128 OSCC cases were used to determine the expression of lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1) and vascular endothelial growth factor C (VEGF-C). Mann-Whitney or Kruskal-Wallis tests were employed to analyze the association between clinicopathological data and intratumoral LVD (ILVD), peritumoral LVD (PLVD), and VEGF-C; comparisons between ILVD and PLVD were made with t-test. Correlations between LVD and VEGF-C were analyzed by Spearman's correlation coefficient. Disease-specific survival curves were obtained with Kaplan-Meier method and compared using the log-rank test. Cox multiple regression was used to clarify the independent effect of clinicopathological data on clinical outcome. KEY FINDINGS: Tumor tissues were positively stained with LYVE-1 and VEGF-C. Both tumor metastasis and recurrence were associated with ILVD. A significant association between ILVD and VEGF-C expression was observed (P < 0.05). A significant association between high ILVD and poor disease-specific survival was observed (P < 0.05). SIGNIFICANCE: This study showed that ILVD was significantly associated with increased lymphatic metastasis, tumor recurrence, and reduced disease-specific survival in patients with OSCC. ILVD could be an indicator to predict the prognosis of OSCC.

Lymphatic remodelling in the hypoxic tumour microenvironment (TME) is critically involved in the metastasis of cervical squamous cell carcinoma (CSCC); however, its underlying mechanisms remain unclear. Here, we uncovered a novel lymphatic pattern in the hypoxic TME, wherein lymphatic vessels (LVs) are encapsulated by tumour-associated macrophages (TAMs) to form an interconnected network. We describe these aggregates as LVEM (LVs encapsulated by TAMs) considering their advantageous metastatic capacity and active involvement in early lymph node metastasis (LNM). Mechanistic investigations revealed that interleukin-10 (IL-10) derived from hypoxic TAMs adjacent to LVs was a prerequisite for lymphangiogenesis and LVEM formation through its induction of Sp1 upregulation in lymphatic endothelial cells (LECs). Interestingly, Sp1(high) LECs promoted the transactivation of C-C motif chemokine ligand 1 (CCL1) to facilitate TAM and tumour cell recruitment, thereby forming a positive feedback loop to strengthen the LVEM formation. Knockdown of Sp1 or blockage of CCL1 abrogated LVEM and consequently attenuated LNM. Notably, CSCC(non-LNM) is largely devoid of hypoxic TAMs and the resultant LVEM, which might explain its metastatic delay. These findings identify a novel and efficient metastasis-promoting lymphatic pattern in the hypoxic TME, which might provide new targets for anti-metastasis therapy and prognostic assessment.

The anatomical spread of lymph node (LN) metastasis is of practical importance in the surgical management of colon cancer (CC). We examined the effect of KRAS, BRAF and microsatellite instability (MSI) on LN count and anatomical spread pattern in stage III CC. We determined KRAS, BRAF and MSI status from stage III CC patients. Biomarker status was correlated with LN count and anatomical spread pattern, which was classified as sequential or skipped. Relapse-free survival (RFS) was estimated using Kaplan-Meier method, and correlations were assessed using log-rank and Cox regression analyses. We analyzed 369 stage III CC patients. The proportion of KRAS mutant (mt), BRAF mt, and MSI-high (H) were 44.2% (163/344), 6.8% (25/344), and 6.8% (25/344), respectively. The mean number of metastatic LN was higher in microsatellite-stable (MSS) compared to MSI patients (3.5 vs. 2.7, p = 0.0406), while no differences were observed according to KRAS or BRAF status. Interestingly, patients with BRAF mt and MSI-H were less likely to harbor skipped metastatic LN (9.3% vs 20% and 4% vs 10.5% compared to BRAF wild type (wt) and MSS, respectively), while KRAS status did not predict anatomical spread pattern. Patients with KRAS wt and MSI-H showed superior RFS compared to KRAS mt and MSS patients, respectively, whereas BRAF status did not affect RFS. Differences exist in the anatomical pattern of invaded LN according to the molecular status of stage III CC. Patients with MSI-H CC have less invaded and skipped LN, suggesting a tailored surgical approach is possible.

The aim of this study was to examine whether lymphatic invasion in papillary thyroid carcinoma (PTC) occurs when tumour-associated macrophages (TAMs) injure lymphatic vessels together with cancer cells. While there was no difference in the lymphatic vessel density in PTC and follicular thyroid carcinoma (FTC), the number of TAMs around the lymphatic vessels was increased in PTC compared to that in FTC. In particular, TAMs were observed together with cancer cells in lymphatic invasive lesions, and the number of M2 cells inside and outside the lymphatic vessels showed a significant correlation. MMP-2 mRNA was expressed in nonneoplastic stromal cells as well as cancer cells, and double immunofluorescence staining confirmed M2 positivity. Consequently, this study reveals that M2 TAMs around lymphatic vessels within the tumour border of PTC may be associated with the lymphatic invasion of cancer cells. This study represents a step forward in elucidating the mechanism of lymphatic invasion.

Head and neck squamous cell carcinoma (HNSCC) metastasizes to the locoregional lymph nodes at high rates and is related to poor clinical outcomes. However, the mechanism by which cancer cells migrate to the lymph nodes is unclear. To address this, we established a conditioned medium culture system for HNSCC cells and lymphatic endothelial cells (LECs) and investigated their crosstalk. Stimulation with tumor-conditioned medium (TCM) activated LECs, resulting in a robust increase in cell proliferation to induce lymphatic hyperplasia. Further, stimulation of HNSCC cells with activated LEC Conditioned media (TCM-LEC CM) induced cell invasion. Among various chemokines, CXCL5 promoted the invasion of TCM-LEC CM-treated HNSCC cells. The level of CXCL5 protein was higher in cancer tissues than those in normal tissues from HNSCC patients. Furthermore, treatment with SB225002, a CXCR2 (CXCL5 receptor) inhibitor, resulted in decreased lymph node metastasis in vivo. In conclusion, inhibition of CXCL5-CXCR2 signaling between cancer cells and LECs suppresses cancer cell invasion and metastasis in vitro and in vivo. This novel therapeutic strategy might be a practical approach to the clinical management of HNSCC.

The principal issue derived from thyroid cancer is its high propensity to metastasize to the lymph node. Aberrant exprssion of long non-coding RNAs have been extensively reported to be significantly correlated with lymphatic metastasis of thyroid cancer. However, the clinical significance and functional role of lncRNA-MAPK8IP1P2 in lymphatic metastasis of thyroid cancer remain unclear. Here, we reported that MAPK8IP1P2 was downregulated in thyroid cancer tissues with lymphatic metastasis. Upregulating MAPK8IP1P2 inhibited, while silencing MAPK8IP1P2 enhanced anoikis resistance in vitro and lymphatic metastasis of thyroid cancer cells in vivo. Mechanistically, MAPK8IP1P2 activated Hippo signaling by sponging miR-146b-3p to disrupt the inhibitory effect of miR-146b-3p on NF2, RASSF1, and RASSF5 expression, which further inhibited anoikis resistance and lymphatic metastasis in thyroid cancer. Importantly, miR-146b-3p mimics reversed the inhibitory effect of MAPK8IP1P2 overexpression on anoikis resistance of thyroid cancer cells. In conclusion, our findings suggest that MAPK8IP1P2 may serve as a potential biomarker to predict lymphatic metastasis in thyroid cancer, or a potential therapeutic target in lymphatic metastatic thyroid cancer.

BACKGROUND: Zosteriform skin metastasis (ZSM) is rare, and its etiology is not well understood. ZSM is possibly derived from the retrograde movement of cancer cells through the lymphatic vessels during disease development. However, it has been difficult to demonstrate it, as no specific findings have been observed. CASE PRESENTATION: A 68-year-old man presented to our department with neck lymphadenopathy. After detailed examinations, squamous cell lung carcinoma (cT2aN3M1c) was diagnosed. Although cisplatin combined with gemcitabine was administered, his cancerous lymphangiopathy was exacerbated, and ZSM was observed on his right chest. Pembrolizumab was initiated as a second-line chemotherapy; however, the patient died 7 months after the initial presentation. In this case, fluorodeoxyglucose-positron emission tomography indicated the presence of skin metastasis and cancerous lymphangiopathy. Similarly, after performing an autopsy, tumor-cell filled lymph ducts were observed in the right subclavian and the cutaneous lymphatic vessel from the right hilar lymph nodes. CONCLUSIONS: To the best of our knowledge, this is the first study to demonstrate that the localization of ZSM in the cutaneous lymphatics was caused by the retrograde movement of cancer cells through the lymphatic vessels, using radiographical and pathological analysis. In addition, fluorodeoxyglucose-positron emission tomography may help predict skin metastasis induced by cancerous lymphangiopathy.

Triple negative breast cancer (TNBC) is a significant clinical problem to which immunotherapeutic strategies have been applied with limited success. Using the syngeneic E0771 TNBC mouse model, this work explores the potential for antitumor CD8(+) T cell immunity to be primed extratumorally in lymphoid tissues and therapeutically leveraged. CD8(+) T cell viability and responses within the tumor microenvironment (TME) were found to be severely impaired, effects coincident with local immunosuppression that is recapitulated in lymphoid tissues in late stage disease. Prior to onset of a locally suppressed immune microenvironment, however, CD8(+) T cell priming within lymph nodes (LN) that depended on tumor lymphatic drainage remained intact. These results demonstrate tumor-draining LNs (TdLN) to be lymphoid tissue niches that support the survival and antigenic priming of CD8(+) T lymphocytes against lymph-draining antigen. The therapeutic effects of and CD8(+) T cells response to immune checkpoint blockade were furthermore improved when directed to LNs within the tumor-draining lymphatic basin. Therefore, TdLNs represent a unique potential tumor immunity reservoir in TNBC for which strategies may be developed to improve the effects of ICB immunotherapy.

OBJECTIVE: to validate the proposed N descriptor revision on a large cohort of patients and assess the impact of tumour location on the distribution pattern of lymph node metastases for patients with NSCLC. METHODS: This is a retrospective review of a consecutive series of patients who had anatomical lung resections. Systematic lymph node dissection was done for all patients. RESULTS: Between January 2009 and December 2019 2566 patients had surgical resection for NSCLC. 448 patients (17.5%) had histologically confirmed lymph node metastases: 257 (57.4%) had pN1 and 191 pN2. Median age of the study population was 69.1 years. Overall survival (OS) for study population was 37.3 months with 5-year survival rate of 35.7%. The survival analysis of the N subgroups showed the pN2 patients had a median OS of 27.9 months vs. 41.7 months for pN1 patients (p = 0.013). Analysis as per the new proposal of the N subgroups N1a vs N1b vs N2a1 vs N2a2 vs N2b showed that median survival OS was 41.7 vs. 39.2 mo vs. 33.3 mo vs. 28.9 mo vs. 24.6 mo (p = 0.099). There was statistically significant difference in survival between N2 patients with skip metastasis and N2 patients without skip metastases: OS 32.2 (95% CI: 16.8–47.6) months vs. 24.2 months (p = 0.024). On multivariate analysis only pathological N (p = 0.011) and the new proposed N classification (p = 0.006) were independent prognostic factors for survival. CONCLUSIONS: N1 and N2 disease are heterogeneous groups and require further stratification. The number of N2 lymph node stations involved and the presence or not of N1 disease translated to significant differences in survival and therefore have to be included in N staging.

Solid tumors are composed of tumor cells and stromal cells including lymphatic endothelial cells (LEC), which are mainly viewed as cells forming lymphatic vessels involved in the transport of metastatic and immune cells. We here reveal a new mechanism by which tumor exposed-LEC (teLEC) exert mitogenic effects on tumor cells. Our conclusions are supported by morphological and molecular changes induced in teLEC that in turn enhance cancer cell invasion in 3D cultures and tumor cell proliferation in vivo. The characterization of teLEC secretome by RNA-Sequencing and cytokine array revealed that interleukine-6 (IL6) is one of the most modulated molecules in teLEC, whose production was negligible in unexposed LEC. Notably, neutralizing anti-human IL6 antibody abrogated teLEC-mediated mitogenic effects in vivo, when LEC were mixed with tumor cells in the ear sponge assay. We here assign a novel function to teLEC that is beyond their role of lymphatic vessel formation. This work highlights a new paradigm, in which teLEC exert “fibroblast-like properties”, contribute in a paracrine manner to the control of tumor cell properties and are worth considering as key stromal determinant in future studies.

Lymph node metastasis (LNM), a critical prognostic determinant in cancer patients, is critically influenced by the presence of numerous heterogeneous cancer-associated fibroblasts (CAFs) in the tumor microenvironment. However, the phenotypes and characteristics of the various pro-metastatic CAF subsets in cervical squamous cell carcinoma (CSCC) remain unknown. Here, we describe a CAF subpopulation with elevated periostin expression (periostin(+) CAFs), located in the primary tumor sites and metastatic lymph nodes, that positively correlated with LNM and poor survival in CSCC patients. Mechanistically, periostin(+) CAFs impaired lymphatic endothelial barriers by activating the integrin-FAK/Src-VE-cadherin signaling pathway in lymphatic endothelial cells and consequently enhanced metastatic dissemination. In contrast, inhibition of the FAK/Src signaling pathway alleviated periostin-induced lymphatic endothelial barrier dysfunction and its related effects. Notably, periostin(−) CAFs were incapable of impairing endothelial barrier integrity, which may explain the occurrence of CAF-enriched cases without LNM. In conclusion, we identified a specific periostin(+) CAF subset that promotes LNM in CSCC, mainly by impairing the lymphatic endothelial barriers, thus providing the basis for potential stromal fibroblast-targeted interventions that block CAF-dependent metastasis.

Renal clear cell carcinoma (ccRCC), is an inflammation-related malignancy with poor therapeutic outcome. Interferon-induced transmembrane protein 2 (IFITM2), an inflammation related gene, is reported to promote tumor progression via inducing cytokine release and lymphatic metastasis. However, IFITM2's role in ccRCC remains unclear. In this study, we aimed to explore the role of IFITM2 in ccRCC. In vitro studies displayed overexpressed IFITM2 level in tumor tissues, while analysis of 538 cases from TCGA unveiled the correlation of upregulated-IFITM2 with shorter survival. Migration and invasion of ccRCC were inhibited following the downregulation of IFITM2. Cocultured with IFITM2-silenced ccRCC cells, human lymphatic endothelial cells were inhibited in proliferation, migration and tube formation, indicating that lymphangiognesis was contributed by IFITM2 expression. Taken together, IFITM2 promotes ccRCC progression by inducing malignant characteristics and lymphatic metastasis. Therefore, IFITM2 represents a promising novel target for therapy and effective prediction of malignancy of ccRCC.

Cancer-associated lymphatic endothelial cells (LECs) are an active barrier to the effector arm of the anti-tumor immune response; however, it remains unclear how LECs become immunosuppressive in the tumor microenvironment (TME). Exosomal microRNAs (miRNAs) have recently been implicated in intercellular crosstalk within the TME. Here, we report a mechanistic model via which cervical cancer-secreted, exosome-encapsulated microRNA (miR)-1468-5p promotes lymphatic PD-L1 upregulation and lymphangiogenesis to impair T cell immunity. Subsequently, exosomal miR-1468-5p epigenetically activates the JAK2/STAT3 pathway in LECs by directly targeting homeobox containing 1 (HMBOX1) in the SOCS1 promoter, activating an immunosuppressive program that allows cancer cells to escape anti-cancer immunity. Furthermore, clinical data reveal that high serum exosomal miR-1468-5p levels correlate with TME immunosuppressive status and poor prognosis in cervical cancer (CCa) patients. Taken together, our results suggest that cancer-secreted exosomal miR-1468-5p instructs LECs to form an integrated immunosuppressive TME component and may be a prognostic biomarker and therapeutic target for CCa.

Central conducting lymphatics (CCLs) disorders represent a broad spectrum of clinical entities ranging from self-limiting traumatic leaks treated by conservative strategies, to complex lymphatic circulation abnormalities that are progressive and unresponsive to currently available treatments. Dynamic contrast-enhanced magnetic resonance lymphangiography (DCMRL) performed by intranodal injection of gadolinium-based contrast material is a recently developed technique which allows a minimally invasive evaluation of the CCL abnormalities providing a dynamic assessment of lymph flow and its pathways. In our institution, DCMRL is performed after bilateral cannulation of inguinal lymph nodes, using a MR protocol which includes volumetric 3D T2-SPACE (sampling perfection with application-optimized contrasts using different flip-angle evolution) and free-breathing respiratory navigated sequence and TWIST (time-resolved angiography with Interleaved stochastic trajectories) MR angiography sequence, during intranodal injection of paramagnetic contrast medium. Although DCMRL applications in clinical practice are still improving, a minimally invasive assessment of lymphatic pathways is particularly important both in pediatric patients with primitive lymphatic system disorders and in children with complex congenital heart disease associated with CCL impairment.

Lymphatic malformations are congenital alterations of normal embryonic lymphatic development. We present a case of a premature 7-week-old male with a large central conducting lymphatic malformation and significant abdominal chylorrhea. He was successfully treated with combined endolymphatic and surgical approaches. To the authors' knowledge, this is the first case to be described.

Background: Extensive lymphatic malformations (LM) may cause substantial morbidity. The mTOR (mammalian target of rapamycin) inhibitor sirolimus shows promise for treating vascular anomalies, though response assessment is not standardized. Objective: To retrospectively characterize changes on MRI in extensive LM in children treated with sirolimus. Methods: Twenty-five children treated with sirolimus for extensive LM were included. Baseline MRI was defined as MRI closest to therapy initiation; follow-up MRI was defined as most recent MRI while on therapy. Two pediatric radiologists independently determined MRI lesion volume by tracing lesion contours on all slices (normalized to patient body mass index) and signal by placing an ROI on lesions' dominant portion (normalized to CSF signal), on baseline and follow-up T2-weighted sequences. Inter-reader agreement was determined, and values averaged for further analysis. Volume and signal changes were compared with patient, lesion, and therapy characteristics. Results: Mean (+/-SD) interval between sirolimus initiation and follow-up MRI was 22.1+/-13.8 months. Mean lesion volume index on baseline and follow-up MRI was 728 mL/m2 +/- 970 mL/m2 and 345 mL/m2+/-501 mL/m2, respectively (p < .001). Ninety-two percent demonstrated a decrease in volume index >10% (mean volume change −46.4%+/-28.2%). Volume change was inversely correlated with age (r = −0.466, p = .02). Mean volume change was −64.7%+/-25.4% in children under 2 years old versus −32.0%+/-21.6% in remaining children (p = .008). Mean volume change was −58.1%+/-24.0% for craniocervical lesions versus −35.5%+/-28.2% for body and extremity lesions (p = .03). Mean lesion signal ratio on baseline and follow-up MRI was 0.81+/-0.29, and 0.59+/-0.26, respectively (p < .001). Mean signal ratio change was −23.8%+/-22.7%. Volume and signal changes were moderately correlated (r = 0.469; p = .02). Volume and signal changes were not associated with sex, lesion subtype, sirolimus serum concentration, or interval between sirolimus initiation and follow-up MRI (p > .05). Inter-reader agreement for volume index change was excellent [intraclass correlation coefficient (ICC) = 0.983] and for signal ratio change was moderate-good (ICC = 0.764). Conclusion: Sirolimus treatment for extensive LM in children is associated with significant reductions in volume and signal on T2-weighted MRI. The volume decrease is greater in younger children and craniocervical lesions. Clinical Impact: The results may facilitate development of standardized MRI-based criteria for assessing pharmacotherapy response of vascular malformations.

Lymphangiomatosis is a rare benign proliferation and dilation of the lymphatic channels that can involve multiple organs with a variety of clinical presentations. We report a case of generalized lymphangiomatosis in a 16-year-old male who presented with a subcutaneous swelling and pain over his lower back area. The patient was diagnosed of generalized lymphangiomatosis with involvement of soft tissues, pulmonary, thoracic and lumbosacral vertebrae, and spinal canal which subsequently confirmed by cytology. In this case report, we aim to discuss radiological features of the relevant differentials, and the patient's symptoms. We also briefly discuss the treatment options as well as prognosis.

Abdominal lymphatic malformations (LM) are rare congenital malformations of the lymphatic system, representing only 2% of all LM in newborns. They may arise from intra-abdominal solid organs (such as the liver, pancreas, kidneys, spleen, adrenal glands, and gastrointestinal tract), mesentery, omentum, and retroperitoneum. Mesenteric LM are the most commonly seen, with retroperitoneal LM being the second most common. Fetal abdominal LM could be associated with karyotypic or other abnormalities, including skin edema, hydrops fetalis, and polyhydramnios, and prenatal diagnosis and perinatal counseling for these LM are important. Prenatal ultrasound (US) and magnetic resonance imaging (MRI) have led to an increased diagnosis of abdominal LM and improved monitoring and intervention postnatally. This article provides an overview of fetal abdominal LM, including the prenatal diagnoses, differential diagnoses, comprehensive illustrations of the imaging findings, treatments, and fetal outcomes.

Lymphatic malformations (LMs) are rare congenital anomalies. LMs are often refractory to standard treatments, including surgical resection, debulking, and sclerotherapy. Use of sildenafil, a phosphodiesterase-5 inhibitor, for treatment of pediatric LMs has been reported with demonstrated benefit to some patients. This case series reports treatment of three patients (aged 14–37 years) suffering from complicated or refractory LMs with a low-dose oral phosphodiesterase-5 inhibitor, resulting in significant clinical improvement.

OBJECTIVES: To investigate the prevalence of arm-involvement in Klippel-Trenaunay syndrome (KTS)-patients and to describe the venous anatomy and/or venous aberrations present in the arm, and if possible, their relationship to complaints (pain, congestion and thromboembolic events). METHODS: A retrospective cohort-study was performed with data from medical records of a large KTS-cohort (n = 173) from a tertiary referral center. Within this cohort, a descriptive study (n = 12) was performed on the KTS-patients with arm involvement and who had been examined with Colour Duplex Ultrasonography (CDU). RESULTS: Our KTS-cohort (n = 173) comprised 43 patients (24.9%) with arm-involvement; in nineteen patients (11.0%) the arm was the only affected limb. Of those KTS patients investigated with CDU, 9 out of 12 (75%) had an aberrant venous anatomy. CONCLUSION: Future research needs to clarity whether the complaints of KTS-patients in general are caused by an aberrant venous anatomy, coagulation alterations and/or other factors. Herein, the KTS-arms may play an important role.