Lymph-derived chemokines direct early neutrophil infiltration in the lymph nodes upon Staphylococcus aureus skin infection.

A large number of neutrophils infiltrate the lymph node (LN) within 4 h after Staphylococcus aureus skin infection (4 h postinfection [hpi]) and prevent systemic S. aureus dissemination. It is not clear how infection in the skin can remotely and effectively recruit neutrophils to the LN. Here, we found that lymphatic vessel occlusion substantially reduced neutrophil recruitment to the LN. Lymphatic vessels effectively transported bacteria and proinflammatory chemokines (i.e., Chemokine [C-X-C motif] motif 1 [CXCL1] and CXCL2) to the LN. However, in the absence of lymph flow, S. aureus alone in the LN was insufficient to recruit neutrophils to the LN at 4 hpi. Instead, lymph flow facilitated the earliest neutrophil recruitment to the LN by delivering chemokines (i.e., CXCL1, CXCL2) from the site of infection. Lymphatic dysfunction is often found during inflammation. During oxazolone (OX)-induced skin inflammation, CXCL1/2 in the LN was reduced after infection. The interrupted LN conduits further disrupted the flow of lymph and impeded its communication with high endothelial venules (HEVs), resulting in impaired neutrophil migration. The impaired neutrophil interaction with bacteria contributed to persistent infection in the LN. Our studies showed that both the flow of lymph from lymphatic vessels to the LN and the distribution of lymph in the LN are critical to ensure optimal neutrophil migration and timely innate immune protection in S. aureus infection.

Flow-dependent regulation of rat mesenteric lymphatic vessel contractile response requires activation of endothelial TRPV4 channels.

OBJECTIVES: The objective of our study is to evaluate the involvement of the transient receptor potential vanilloid 4 (TRPV4) in the alteration of lymphatic pumping in response to flow and determine the signaling pathways involved. METHODS: We used immunofluorescence imaging and western blotting to assess TRPV4 expression in rat mesenteric lymphatic vessels. We examined inhibition of TRPV4 with HC067047, nitric oxide synthase (NOS) with L-NNA and cyclooxygenases (COXs) with indomethacin on the contractile response of pressurized lymphatic vessels to flow changes induced by a stepwise increase in pressure gradients, and the functionality of endothelial TRPV4 channels by measuring the intracellular Ca2+ response of primary lymphatic endothelial cell cultures to the selective agonist GSK1016790A. RESULTS: TRPV4 protein was expressed in both the endothelial and the smooth muscle layer of rat mesenteric lymphatics with high endothelial expression around the valve sites. When maintained under constant transmural pressure, most lymphatic vessels displayed a decrease in contraction frequency under conditions of flow and this effect was ablated through inhibition of NOS, COX or TRPV4. CONCLUSIONS: Our findings demonstrate a critical role for TRPV4 in the decrease in contraction frequency induced in lymphatic vessels by increases in flow rate via the production and action of nitric oxide and dilatory prostanoids.

X-ray irradiation negatively affects immune responses in the lymphatic network.

Immune checkpoint inhibitor therapy has been attracting attention as a new cancer treatment and is likely to be widely used in combination with radiotherapy. Therefore, examination of the effects of X-ray irradiation on sentinel lymph nodes and lymphatic vessels, which are involved in antigen presentation, is important for therapy. The hindlimbs of mice were irradiated with X-rays (total radiation doses: 2, 10, and 30 Gy), and X-ray computed tomography (CT) imaging was performed using 15-nm or 2-nm gold nanoparticles (AuNPs) as contrast agents on days 7, 14, and 28 after irradiation to evaluate the diameter of the collecting lymph vessels and lymph flow within the irradiated area. X-ray CT imaging data using 15-nm AuNPs on day 28 after irradiation showed that the diameter of the collecting lymph vessels was significantly larger in all irradiated groups compared to the control group (p ≤ 0.01). CT imaging with 2-nm AuNPs showed that lymphatic drainage was significantly reduced in the lymph nodes irradiated with 10 Gy and 30 Gy compared to the lymph nodes irradiated with 2 Gy (p ≤ 0.05). Additionally, immunohistochemical analyses were conducted to evaluate the area density and morphology of high endothelial venules (HEVs) in the lymph nodes, which are important vessels for naive T cells to enter the lymph nodes. The expression level of MECA-79, which specifically localized to HEVs, was significantly decreased in the 10 Gy and 30 Gy irradiation groups compared to the control group (p ≤ 0.05). There was a significant decrease in normal HEV morphology (p ≤ 0.05) and a significant increase in abnormal HEV morphology (p ≤ 0.05) in all irradiated groups. These results also showed that X-ray irradiation induced a time- and radiation dose-dependent increase in the diameter of the collecting lymph vessels, stagnation of intralymphatic lymph flow, and a reduction in the area density of HEVs and their abnormal morphology, demonstrating that X-ray irradiation affected the immune responses. Therefore, these findings suggest that X-ray irradiation to lymph nodes may impair the opportunity for antigen presentation in the lymph nodes, which is the key to cancer immunity, and that for this reason, it is important to carefully plan irradiation of sentinel lymph nodes and develop treatment strategies according to future treatment options.

Restoration of lymph flow by flap transfer can prevent severe lower extremity lymphedema after inguino-pelvic lymphadenectomy.

PURPOSE: Severe lymphedema is difficult to treat because of the associated extensive scar formation. Therefore, preventing scar formation might alleviate the severity of lymphedema following lymphadenectomy. In this study, we evaluated the usefulness of flap transfer, performed immediately after lymphadenectomy, for preventing scar formation. METHODS: Twenty-three patients with subcutaneous malignancy in a lower extremity, who underwent inguino-pelvic lymphadenectomy, were divided into groups based on whether flap transfer was performed. The severity of lymphedema was categorized according to the ratio of the circumference of the affected extremity to that of the unaffected extremity, as mild (< 20% increase in volume), moderate (20-40%), or severe (> 40%). RESULTS: In the 18 patients who underwent lymphadenectomy without flap transfer, lymphedema was classified as mild in 7, moderate in 7, and severe in 4. In the five patients who underwent lymphadenectomy with flap transfer, lymphedema was classified as mild in 4 and moderate in 1. This difference between the groups did not reach significance. CONCLUSIONS: The findings of this study suggest that flap transfer may help prevent scar formation and contribute to the restoration of lymph flow after lymphadenectomy.

GLP-1 (Glucagon-Like Peptide-1) Is Physiologically Relevant for Chylomicron Secretion Beyond Its Known Pharmacological Role.

[Figure: see text].

Insufficient hepatic uptake in pediatric patients on lymphoscintigraphy.

BACKGROUND: To demonstrate the association between pediatric outcomes and tracer hepatic uptake as a marker of systemic circulation in lymphoscintigraphy. METHODS: We included 31 initial lymphoscintigraphic studies. We compared the presence or absence of hepatic uptake between deceased and survived patients in both early and delayed images using Fisher's exact test. Kaplan-Meier survival analysis was performed based on imaging results, and these curves were compared using the log-rank test. The primary endpoint was death and the survival period was defined from the day of examination to the day of the last visit or death. RESULTS: Of 31 patients, six died. Hepatic uptake was significantly different in both early and delayed images (early images, died [with/without visualization] vs. survived [with/without visualization], 0/6 vs. 13/12, P = 0.028; delayed images, died [with/without visualization] vs. survived [with/without visualization], 2/4 vs. 22/3, P = 0.014) between deceased and survived patients. Survival periods were significantly different between the two groups with and without hepatic uptake in early and delayed images (with/without visualization in early imags = 1,177.1 ± 773.8 days/426.7 ± 419.8 days, P = 0.008 and with/without visualization in delayed images = 821.3 ± 738.0 days/467.4 ± 452.4 days, P = 0.003). CONCLUSIONS: Visualization of hepatic uptake in both early and delayed lymphoscintigraphy is associated with patient outcomes. Hepatic uptake could represent tracer inflow into the systemic circulation, indicating preservation of the connection between the lymphatic system and the systemic circulation. Physicians should evaluate these findings carefully on lymphoscintigraphy.

TRPV4 channels' dominant role in the temperature modulation of intrinsic contractility and lymph flow of rat diaphragmatic lymphatics.

The lymphatic system drains and propels lymph by extrinsic and intrinsic mechanisms. Intrinsic propulsion depends upon spontaneous rhythmic contractions of lymphatic muscles in the vessel walls and is critically affected by changes in the surrounding tissue like osmolarity and temperature. Lymphatics of the diaphragm display a steep change in contraction frequency in response to changes in temperature, and this, in turn, affects lymph flow. In the present work, we demonstrated in an ex vivo diaphragmatic tissue rat model that diaphragmatic lymphatics express transient receptor potential channels of the vanilloid 4 subfamily (TRPV4) and that their blockade by both the nonselective antagonist Ruthenium Red and the selective antagonist HC-067047 abolished the response of lymphatics to temperature changes. Moreover, the selective activation of TRPV4 channels by means of GSK1016790A mirrored the behavior of vessels exposed to increasing temperatures, pointing out the critical role played by these channels in sensing the temperature of the lymphatic vessels' environment and thus inducing a change in contraction frequency and lymph flow.NEW & NOTEWORTHY The present work addresses the putative receptor system that enables diaphragmatic lymphatics to change intrinsic contraction frequency and thus lymph flow according to the changes in temperature of the surrounding environment, showing that this role can be sustained by TRPV4 channels alone.

Adaptation of the hepatic transudation barrier to sinusoidal hypertension.

The role of the hepatic transudation barrier in determining ascites volume and protein content in chronic liver disease is poorly understood. Therefore, the purpose of the present study was to characterize how chronic sinusoidal hypertension impacts hepatic transudation barrier properties and the transudation rate. The suprahepatic inferior vena cava was surgically constricted, and animals were exposed to either short-term (SVH; 2-3 wk) or long-term venous hypertension (LVH; 5-6 wk). Compared with SVH, LVH resulted in lower peritoneal fluid pressure, ascites volume, and ascites protein concentration. The transudation barrier protein reflection coefficient was significantly higher, and the transudation barrier hydraulic conductivity, transudation rate, and transudate-to-lymph protein concentration ratio were significantly lower in LVH animals compared with SVH animals. The sensitivity of transudation rates to acute changes in interstitial fluid pressures was also significantly lower in LVH animals compared with SVH animals. In contrast, there was no detectable difference in hepatic lymph flow rate or sensitivity of lymph flow to acute changes in interstitial fluid pressures between SVH and LVH animals. Taken together, these data suggest that decreased hepatic transudation barrier permeability to fluid and protein and increased reflection coefficient led to a decrease in the hepatic contribution to ascites volume. The present work, to the best of our knowledge, is the first to quantify an anti-ascites adaptation of the hepatic transudation barrier in response to chronic hepatic sinusoidal hypertension.

Glucose and GLP-2 (Glucagon-Like Peptide-2) Mobilize Intestinal Triglyceride by Distinct Mechanisms.

OBJECTIVE: Dietary triglycerides are partially retained in the intestine within intracellular or extracellular compartments, which can be rapidly mobilized in response to several stimuli, including glucose and GLP-2 (glucagon-like peptide-2). To elucidate the mechanism of intestinal lipid mobilization, this study examined the patterns and time course of lymph flow and triglycerides after glucose and GLP-2 treatment in rats. Approach and Results: Lymph flow, triglyceride concentration, and triglyceride output were assessed in mesenteric lymph duct-cannulated rats in response to an intraduodenal (i.d.) lipid bolus followed 5 hours later by either (1) i.d. saline+intraperitoneal (i.p.) saline (placebo), (2) i.d. glucose plus i.p. saline, (3) i.d. saline+i.p. GLP-2, or (4) i.d. glucose+i.p. GLP-2. GLP-2 and glucose administered alone or in combination stimulated total triglyceride output to a similar extent, but the timing and pattern of stimulation differed markedly. Whereas GLP-2 rapidly increased lymph flow with no effect on lymph triglyceride concentration or triglyceride:apoB48 (apolipoprotein B48) ratio (a surrogate marker of chylomicron size) compared with placebo, glucose transiently decreased lymph flow followed by delayed stimulation of lymph flow and increased lymph triglyceride concentration and triglyceride:apoB48 ratio. CONCLUSIONS: Glucose and GLP-2 robustly enhanced intestinal triglyceride output in rats but with different effects on lymph flow, lymph triglyceride concentration, and chylomicron size. GLP-2 stimulated triglyceride output primarily by enhancing lymph flow with no effect on chylomicron size, whereas glucose mobilized intestinal triglycerides, stimulating secretion of larger chylomicrons. This suggests that these 2 stimuli mobilize intestinal lipid by different mechanisms.

Lymphatic Valves Separate Lymph Flow Into a Central Stream and a Slow-Moving Peri-Valvular Milieu.

The lymphatic system plays a pivotal role in the transport of fats, waste, and immune cells, while also serving as a metastatic route for select cancers. Using live imaging and particle tracking, we experimentally characterized the lymph flow field distal from the inguinal lymph node in the vicinity of normal bileaflet and malformed unileaflet intraluminal valves. Particle tracking experiments demonstrated that intraluminal lymphatic valves concentrate higher velocity lymph flow in the center of the vessel, while generating adjacent perivalvular recirculation zones. The recirculation zones are characterized by extended particle residence times and low wall shear stress (WSS) magnitudes in comparison to the rest of the lymphangion. A malformed unileaflet valve skewed lymph flow toward the endothelium on the vessel wall, generating a stagnation point and a much larger recirculation zone on the opposite wall. These studies define physical consequences of bileaflet and unileaflet intraluminal lymphatic valves that affect lymph transport and the generation of a heterogeneous flow field that affects the lymphatic endothelium nonuniformly. The characterized flow fields were recreated in vitro connecting different flow environments present in the lymphangion to a lymphatic endothelial cell (LEC) pro-inflammatory phenotype. Unique and detailed insight into lymphatic flow is provided, with potential applications to a variety of diseases that affect lymph transport and drug delivery.

[Efficacy of vacuum-gradient therapy in reducing orthodontic treatment time]. / Effekt uskoreniya ortodonticheskoi korrektsii zubochelyustnykh anomalii vakuum-gradientnoi terapiei.

The purpose of the study was to evaluate the efficacy of vacuum-gradient therapy in reducing orthodontic treatment time. MATERIALS AND METHODS: The study comprised 57 patients aged from 25 to 35 years (mean age 28.8±4.6 years) with teeth crowding divided into two groups. In group 1 (n=29) orthodontic correction was combined with vacuum-gradient therapy by means of modified «AVLT-Desna¼ device, while patients in group 2 (n=28) underwent orthodontic correction alone. Microhaemo- and lymphocirculation was studied by laser Doppler flowmetry. RESULTS: Lymph draining and microhaemocirculation are impaired by teeth crowding. Orthodontic correction influences on microhaemo- and lymphocirculation of periodontal tissue on tooth movement and improves mechanisms of haemodynamic regulation. CONCLUSIONS: The method of vacuum-gradient therapy combined with complex malocclusion correction allows to accelerate orthodontic treatment and to improve microcirculation in the moved teeth area.

Inhibition of Contraction Strength and Frequency by Wall Shear Stress in a Single-Lymphangion Model.

The phasic contractions of collecting lymphatic vessels are reduced in strength and occur at diminished frequency when a favorable pressure difference and the resulting antegrade flow create large fluid shear stresses at the luminal surface. This paper describes a minimal phenomenological model of this mechanism that is applied to a previously validated numerical model of a phasically contracting lymphangion. The parameters of the inhibition model are quantitatively matched to observations in isolated segments of rat lymphatic vessel, first for mesenteric lymphatics then for thoracic duct, and outcomes from the numerical model are then qualitatively compared with recent observations in isolated segments of rat thoracic duct.

KLF2-mediated disruption of PPAR-γ signaling in lymphatic endothelial cells exposed to chronically increased pulmonary lymph flow.

Lymphatic abnormalities associated with congenital heart disease are well described, yet the underlying mechanisms remain poorly understood. Using a clinically relevant ovine model of congenital heart disease with increased pulmonary blood flow, we have previously demonstrated that lymphatic endothelial cells (LECs) exposed in vivo to chronically increased pulmonary lymph flow accumulate ROS and have decreased bioavailable nitric oxide (NO). Peroxisome proliferator-activated receptor-γ (PPAR-γ), which abrogates production of cellular ROS by NADPH oxidase, is inhibited by Krüppel-like factor 2 (KLF2), a flow-induced transcription factor. We hypothesized that chronically increased pulmonary lymph flow induces a KLF2-mediated decrease in PPAR-γ and an accumulation of cellular ROS, contributing to decreased bioavailable NO in LECs. To better understand the mechanisms that transduce the abnormal mechanical forces associated with chronically increased pulmonary lymph flow, LECs were isolated from the efferent vessel of the caudal mediastinal lymph node of control ( n = 5) and shunt ( n = 5) lambs. KLF2 mRNA and protein were significantly increased in shunt compared with control LECs, and PPAR-γ mRNA and protein were significantly decreased. In control LECs exposed to shear forces in vitro, we found similar alterations to KLF2 and PPAR-γ expression. In shunt LECs, NADPH oxidase subunit expression was increased, and bioavailable NO was significantly lower. Transfection of shunt LECs with KLF2 siRNA normalized PPAR-γ, ROS, and bioavailable NO. Conversely, pharmacological inhibition of PPAR-γ in control LECs increased ROS equivalent to levels in shunt LECs at baseline. Taken together, these data suggest that one mechanism by which NO-mediated lymphatic function is disrupted after chronic exposure to increased pulmonary lymph flow is through altered KLF2-dependent PPAR-γ signaling, resulting in increased NADPH oxidase activity, accumulation of ROS, and decreased bioavailable NO. NEW & NOTEWORTHY Lymphatic endothelial cells, when exposed in vivo to chronically elevated pulmonary lymph flow in a model of congenital heart disease with increased pulmonary blood flow, demonstrate Krüppel-like factor 2-dependent disrupted peroxisome proliferator-activated receptor-γ signaling that results in the accumulation of reactive oxygen species and decreased bioavailable nitric oxide.

Neutrophil-lymphatic interactions during acute and chronic disease.

The lymphatic system aids in osmoregulation through tissue fluid transport, but is also designed to support communication between cells of the innate and adaptive immune systems. During inflammation, changes within the lymphatics can result in an altered response to infection. Neutrophils have been described as one key cell type that facilitates antigen capture and presentation within the lymphatic system, enabling an effective adaptive immune response. Disruption of neutrophil recruitment during inflammation, due to alterations in lymphatics, is a growing area of study due to their key role in infection resolution. In this review, we discuss the currently known methods by which neutrophils are recruited to the lymphatic system and what subsequent effects they have on resident and recruited cells within the lymph vessels and nodes. We also discuss the changes in neutrophil activation and recruitment during chronic inflammatory diseases and their relationship to lymphatic dysfunction.

Detection of alternative subpleural lymph flow pathways using indocyanine green fluorescence.

PURPOSE: Pulmonary lymphatic fluid predominately flows along the bronchi. However, there are reports suggesting that an alternative lymphatic pathway exist, which may result in skip metastases. The aim of this study was to evaluate the subpleural lymph flow in vivo using indocyanine green (ICG) fluorescence. METHODS: One hundred cases were enrolled. ICG was injected into the macroscopically healthy subpleural space. Intraoperative fluorescence images were then observed in real time. RESULTS: ICG fluorescence was observed moving through subpleural channels in 58/100 cases. ICG flowed into adjacent lobes over interlobar lines in 18 cases and flowed from the visceral pleura directly into the mediastinum in 5 cases. The frequency of mediastinal detection without hilar lymph node detection was significantly higher in the left lung compared to the right (p < 0.05). The subpleural lymph flow detection rates were significantly lower in patients with smoking pack-years ≥ 40 than those with < 40 (p < 0.05). CONCLUSIONS: The flow of lymphatic fluid directly into the mediastinum suggests one mechanism of skip metastasis. In addition, the reduction of the subpleural lymph flows in smokers with ≥ 40 pack-years suggests that smoking might modify lymph flow patterns. These findings may assist in selecting the optimal therapy for patients with possible skip metastasis.

Lymph flow guided irradiation of regional lymph nodes in patients with cervical cancer: Preliminary analysis of scintigraphic data.

PURPOSE: To evaluate patterns of lymph flow from primary lesions in patients with cervical cancer and to determine how useful for radiotherapy planning this information can be. MATERIALS AND METHODS: SPECT-CT visualization of sentinel (SLN) lymph nodes (LNs) was performed in 36 primary patients with IB-IIB cervical cancer. The acquisition started 120-240 min after 4 peritumoral injections of 99mTc-radiocolloids (150-300 MBq in 0.4-1 ml). We determined localization of LN with uptake of radiocolloids, type of lymph flow (mono-, bi-lateral) and lymph flow patterns (supraureteral paracervical, infraureteral paracervical and directly to para-aortic LNs). RESULTS: SLNs were visualized in 31 of 36 women. Bilateral lymph-flow was detected in 22 (71%), monolateral - in the other 9 (29%) cases. The distribution of SLNs was as follows: external iliac - 64.5%, internal iliac - 54.8%, obturator - 32.2%, common iliac - 35.5% and pre-sacral 3.2%. Para-aortic LNs were visualized in 5 (16.1%) patients. The supraureteral paracervical pattern of lymph flow was identified in 22, infraureteral paracervical - in 4 and their combination - in the other 5 women. CONCLUSION: Visualization of an individual pattern of lymph flow from primary cervical cancer can be considered as a promising tool for optimization of the volume of irradiated regional LNs.

Emerging trends in the pathophysiology of lymphatic contractile function.

Lymphatic contractile dysfunction is central to a number of pathologies that affect millions of people worldwide. Due to its critical role in the process of inflammation, a dysfunctional lymphatic system also compromises the immune response, further exacerbating a number of inflammation related diseases. Despite the critical physiological functions accomplished by the transport of lymph, a complete understanding of the contractile machinery of the lymphatic system lags far behind that of the blood vasculature. However, there has been a surge of recent research focusing on different mechanisms that underlie both physiological and pathophysiological aspects of lymphatic contractile function. This review summarizes those emerging paradigms that shed some novel insights into the contractile physiology of the lymphatics in normal as well as different disease states. In addition, this review emphasizes the recent progress made in our understanding of various contractile parameters and regulatory elements that contribute to the normal functioning of the lymphatics.

Interstitial IgG antibody pharmacokinetics assessed by combined in vivo- and physiologically-based pharmacokinetic modelling approaches.

KEY POINTS: For therapeutic antibodies, total tissue concentrations are frequently reported as a lump sum measure of the antibody in residual plasma, interstitial fluid and cells. In terms of correlating antibody exposure to a therapeutic effect, however, interstitial pharmacokinetics might be more relevant. In the present study, we collected total tissue and interstitial antibody biodistribution data in mice and assessed the composition of tissue samples aiming to correct total tissue measurements for plasma and cellular content. All data and parameters were integrated into a refined physiologically-based pharmacokinetic model for monoclonal antibodies to enable the tissue-specific description of antibody pharmacokinetics in the interstitial space. We found that antibody interstitial concentrations are highly tissue-specific and dependent on the underlying capillary structure but, in several tissues, they reach relatively high interstitial concentrations, contradicting the still-prevailing view that both the distribution to tissues and the interstitial concentrations for antibodies are generally low. ABSTRACT: For most therapeutic antibodies, the interstitium is the target space. Although experimental methods for measuring antibody pharmacokinetics (PK) in this space are not well established, thus making quantitative assessment difficult, the interstitial antibody concentration is assumed to be low. In the present study, we combined direct quantification of antibodies in the interstitial fluid with a physiologically-based PK (PBPK) modelling approach, with the aim of better describing the PK of monoclonal antibodies in the interstitial space of different tissues. We isolated interstitial fluid by tissue centrifugation and conducted an antibody biodistribution study in mice, measuring total tissue and interstitial concentrations in selected tissues. Residual plasma, interstitial volumes and lymph flows, which are important PBPK model parameters, were assessed in vivo. We could thereby refine the PBPK modelling of monoclonal antibodies, better interpret antibody biodistribution data and more accurately predict their PK in the different tissue spaces. Our results indicate that, in tissues with discontinuous capillaries (liver and spleen), interstitial concentrations are reflected by the plasma concentration. In tissues with continuous capillaries (e.g. skin and muscle), ∼50-60% of the plasma concentration is found in the interstitial space. In the brain and kidney, on the other hand, antibodies are restricted to the vascular space. Our data may significantly impact the interpretation of biodistribution data of monoclonal antibodies and might be important when relating measured concentrations to a therapeutic effect. By contrast to the view that the antibody distribution to the interstitial space is limited, using direct measurements and model-based data interpretation, we show that high antibody interstitial concentrations are reached in most tissues.

Temperature-dependent modulation of regional lymphatic contraction frequency and flow.

Lymph drainage and propulsion are sustained by an extrinsic mechanism, based on mechanical forces acting from the surrounding tissues against the wall of lymphatic vessels, and by an intrinsic mechanism attributable to active spontaneous contractions of the lymphatic vessel muscle. Despite being heterogeneous, the mechanisms underlying the generation of spontaneous contractions share a common biochemical nature and are thus modulated by temperature. In this study, we challenged excised tissues from rat diaphragm and hindpaw, endowed with spontaneously contracting lymphatic vessels, to temperatures from 24°C (hindpaw) or 33°C (diaphragmatic vessels) to 40°C while measuring lymphatic contraction frequency (fc) and amplitude. Both vessel populations displayed a sigmoidal relationship between fc and temperature, each centered around the average temperature of surrounding tissue (36.7 diaphragmatic and 32.1 hindpaw lymphatics). Although the slope factor of the sigmoidal fit to the fc change of hindpaw vessels was 2.3°C·cycles-1·min-1, a value within the normal range displayed by simple biochemical reactions, the slope factor of the diaphragmatic lymphatics was 0.62°C·cycles-1·min-1, suggesting the added involvement of temperature-sensing mechanisms. Lymph flow calculated as a function of temperature confirmed the relationship observed on fc data alone and showed that none of the two lymphatic vessel populations would be able to adapt to the optimal working temperature of the other tissue district. This poses a novel question whether lymphatic vessels might not adapt their function to accommodate the change if exposed to a surrounding temperature, which is different from their normal condition.NEW & NOTEWORTHY This study demonstrates to what extent lymphatic vessel intrinsic contractility and lymph flow are modulated by temperature and that this modulation is dependent on the body district that the vessels belong to, suggesting a possible functional misbehavior should lymphatic vessels be exposed to a chronically different temperature.

Evaluation of lymph flow patterns in splenic flexural colon cancers using laparoscopic real-time indocyanine green fluorescence imaging.

PURPOSE: The treatment of splenic flexural colon cancer is not standardized because the lymphatic drainage is variable. The aim of this study is to evaluate the lymph flow at the splenic flexure. METHODS: From July 2013 to January 2016, consecutive patients of the splenic flexural colon cancer with a preoperative diagnosis of N0 who underwent laparoscopic surgery were enrolled. Primary outcome is frequency of the direction of lymph flow from splenic flexure. We injected indocyanine green (2.5 mg) into the submucosal layer around the tumor and observed lymph flow using the laparoscopic near-infrared camera system in 30 min after injection. RESULTS: Thirty-one patients were enrolled in this study. The lymph flow was visualized in 31 patients (100 %) without any complications. No case exhibited lymph flow in both the left colic artery (LCA) and left branch of the middle colic artery (lt-MCA) areas. There were 19 cases (61.3 %) with lymph flow directed to the area of the root of the inferior mesenteric vein (IMV), regardless of the presence of the left accessory aberrant colic artery. Lymph node metastases were observed in six cases (19.4 %), and all of the involved lymph nodes existed in lymph flow areas determined by real-time indocyanine green fluorescence imaging. CONCLUSIONS: The findings of the lymph flow pattern of splenic flexure suggest that lymph node dissection at the root of the IMV area is important, and it may be not necessary to ligate both the lt-MCA and LCA, at least in cases without widespread lymph node metastases.