The lymphatic system acts to return fluid from the interstitial space back into the blood circulation. In normal conditions, lymphangions, the segment of lymphatic vessel in between valves, cyclically contract and can pump lymph from low pressure tissues to the higher-pressure veins of the neck. With edema, however, this pressure gradient can reverse, and the role of contraction is less clear. Like ventricles, lymphangions are sensitive to both preload and afterload. Unlike ventricles, lymphangions are arranged in series, so that the outlet pressure of one lymphangion becomes the inlet pressure of another. Anything that alters the relative timing and frequency of adjacent lymphangions alters both preload and afterload of each lymphangion and thus mean lymph flow. To explore the effect of timing and frequency of contraction on lymph flow, we developed a computational model of a lymphatic vessel with lymphangions described by the classic description of time-varying elastance. When pumping up a pressure gradient, as in normal conditions, or when pumping down a pressure gradient, as in some cases of edema, we found that flow was optimized when the lymphangions in the vessel were pumping with a very short time delay between their cycles, and the flow was reduced when the time delay between the contractions was reduced to zero. However, a difference in frequency between adjacent lymphangions alters instantaneous flow but does not affect mean flow. These results suggest an important role for the timing of the contraction in optimizing lymph flow. However, a difference in frequencies between adjacent lymphangions has little effect on altering lymph flow, suggesting that tight control of lymphangion coordination may not be critical for lymphatic function.

The lymphatic system acts to return fluid from the interstitial space back into the blood circulation. In normal conditions, lymphangions, the segment of lymphatic vessel in between valves, cyclically contract and can pump lymph from low pressure tissues to the higher-pressure veins of the neck. With edema, however, this pressure gradient can reverse, and the role of contraction is less clear. Like ventricles, lymphangions are sensitive to both preload and afterload. Unlike ventricles, lymphangions are arranged in series, so that the outlet pressure of one lymphangion becomes the inlet pressure of another. Anything that alters the relative timing and frequency of adjacent lymphangions alters both preload and afterload of each lymphangion and thus mean lymph flow. To explore the effect of timing and frequency of contraction on lymph flow, we developed a computational model of a lymphatic vessel with lymphangions described by the classic description of time-varying elastance. When pumping up a pressure gradient, as in normal conditions, or when pumping down a pressure gradient, as in some cases of edema, we found that flow was optimized when the lymphangions in the vessel were pumping with a very short time delay between their cycles, and the flow was reduced when the time delay between the contractions was reduced to zero. However, a difference in frequency between adjacent lymphangions alters instantaneous flow but does not affect mean flow. These results suggest an important role for the timing of the contraction in optimizing lymph flow. However, a difference in frequencies between adjacent lymphangions has little effect on altering lymph flow, suggesting that tight control of lymphangion coordination may not be critical for lymphatic function.

Although lymphatic vessel behavior is analogous to hearts (e.g. systole and diastole) and blood vessels (e.g. basal tone), hearts and blood vessels have fundamentally different contractile properties. While summation during contraction is minimized in the heart, summation is necessary for tonic contraction in blood vessels. Because lymphatic vessel behavior mimics cardiac and vascular behavior, we hypothesized that above a critical contraction frequency there is significant summation, evidenced by significantly increased diastolic active tension (i.e. basal tone). We used an isovolumic, controlled-flow preparation to examine the interaction of contraction cycle-time with contraction frequency. Using segments of isolated lymphatic vessels (~1 cm in length and 3-4 mm in diameter) from bovine mesentery, we measured transmural pressure and diameter for end-diastole and end-systole during spontaneous contractions for 10 volume steps. We found time between contractions (beat-to-beat period) decreases with increasing diameter, and total contraction time (vessel twitch length, 11.08 ± 1.54 s) slightly increases with increasing diameter. At the intersection of these relationships, there is a critical period, below which the vessel does not have time to fully relax. Above the diameter at the critical period, diastolic active tension (end-diastolic minus passive vessel tension) significantly increases with increases in diameter (309 to 562% change in slope, p<0.0001), and, below the critical period, diastolic active tension increases with decreases in beat-to-beat period (712 to 2208% change in slope, p<0.0014). Because this transition occurs within a physiological range, it suggests summation may be crucial for lymphatic vessel function as a pump and a conduit.

Although lymphatic vessel behavior is analogous to hearts (e.g. systole and diastole) and blood vessels (e.g. basal tone), hearts and blood vessels have fundamentally different contractile properties. While summation during contraction is minimized in the heart, summation is necessary for tonic contraction in blood vessels. Because lymphatic vessel behavior mimics cardiac and vascular behavior, we hypothesized that above a critical contraction frequency there is significant summation, evidenced by significantly increased diastolic active tension (i.e. basal tone). We used an isovolumic, controlled-flow preparation to examine the interaction of contraction cycle-time with contraction frequency. Using segments of isolated lymphatic vessels (~1 cm in length and 3-4 mm in diameter) from bovine mesentery, we measured transmural pressure and diameter for end-diastole and end-systole during spontaneous contractions for 10 volume steps. We found time between contractions (beat-to-beat period) decreases with increasing diameter, and total contraction time (vessel twitch length, 11.08 ± 1.54 s) slightly increases with increasing diameter. At the intersection of these relationships, there is a critical period, below which the vessel does not have time to fully relax. Above the diameter at the critical period, diastolic active tension (end-diastolic minus passive vessel tension) significantly increases with increases in diameter (309 to 562% change in slope, p<0.0001), and, below the critical period, diastolic active tension increases with decreases in beat-to-beat period (712 to 2208% change in slope, p<0.0014). Because this transition occurs within a physiological range, it suggests summation may be crucial for lymphatic vessel function as a pump and a conduit.

The lymphatic system is responsible for several vital roles in human body, one of which is maintaining fluid and protein balance. There is no central pump in the lymphatic system and the transport of fluid against gravity and adverse pressure gradient is maintained by the extrinsic and intrinsic pumping mechanisms. Any disruption of the lymphatic system due to trauma or injury can lead to edema. There is no cure for lymphedema partly because the knowledge of the function of the lymphatic system is lacking. Thus, a well-developed model of the lymphatic system is crucial to improve our understanding of its function. Here we used a lumped parameter approach to model a chain of lymphangions in series. Equations of conservation of mass, conservation of momentum, and vessel wall force balance were solved for each lymphangion computationally. Due to the lack of knowledge of the parameters describing the system in the literature, more accurate measurements of these parameters should be pursued to advance the model. Because of the difficulty of the isolated vessel and in-situ experiments, we performed a parameter sensitivity analysis to determine the parameters that affect the system most strongly. Our results showed that more accurate estimations of active contractile force and physiologic features of lymphangions, such as length/diameter ratios, should be pursued in future experiments. Also further experiments are required to refine the valve behavior and valve parameters.

The histologic distinction of follicular patterned lesions of thyroid, that is follicular adenoma, follicular carcinoma, and the follicular variant of papillary thyroid carcinoma can be extremely difficult. The differential diagnostic criteria regarding nuclear features of papillary thyroid carcinoma are subjective, resulting in high interobserver variability. Although papillary thyroid carcinoma metastasizes mainly via lymphatic vessels, whereas follicular carcinoma spreads mostly hematogenously, there are no data regarding utility of objective quantitative criteria such as lymphatic and general blood vessel density for the differential diagnosis of these lesions. In this study, 35 follicular patterned lesions of thyroid (14 follicular adenomas, 10 follicular carcinomas, and 11 of the follicular variant of papillary thyroid carcinomas) were evaluated immunohistochemically. An assessment of intra- and peritumoral lymphatic vessel density was performed using novel lymphatic endothelium-specific marker D2-40, and the intra- and peritumoral general vessel density was determined by the panendothelial marker CD31. There were no significant differences in the intra- and/or peritumoral general vessel densities, and peritumoral lymphatic vessel densities among follicular adenoma, follicular carcinoma and the follicular variant of papillary thyroid carcinoma. In contrast, the intratumoral lymphatic vessel density was significantly higher in the follicular variant of papillary thyroid carcinoma than in either follicular adenoma or follicular carcinoma (34.63, 15.04, and 0.11 respectively; P<0.0001). The results of the study show that intratumoral lymphatic vessel density may serve as a useful tool in the differential diagnosis of follicular patterned lesions of thyroid. © 2005 USCAP, Inc All rights reserved.

Os tumores carcinóides típicos broncopulmonares são proliferações de células neuroendócrinas. Foram consideradas como adenomas e acreditava-se que não tinham potencial para disseminação hematogênica e linfática. Porém, a ocorrência de metástase linfática e hematogênica acontece em um quinto dos indivíduos acometidos por essa patologia. A variação no comportamento clínico dos carcinóides broncopulmonares torna imperativa a realização de pesquisas que visem à melhor compreensão dessa doença. É fundamental determinar a agressividade e o risco individual da ocorrência de metástase linfática e hematogênica para que se possa oferecer um tratamento individualizado para cada binômio doente-doença. A classificação atual divide os tumores carcinóides, conforme o grau histológico de malignidade em típico e atípico, agrupando as neoplasias de acordo com o índice mitótico, relação volumétrica núcleo/citoplasma, presença ou ausência de necrose, pleomorfismo nuclear e invasão vascular. Esta análise, porém, é realizada em espécimes histológicos corados pela hematoxilina-eosina, técnica tradicional consagrada, mas que não permite avaliar processos biomoleculares relacionados ao potencial maligno das células que já podem estar presentes e não serem detectados pelo método. Em tumores carcinóides vários estudos já foram realizados na tentativa de identificar o potencial proliferativo de células que ainda não apresentam figuras de mitose, como PCNA, p53, Ki-67, o processo apoptótico (Bcl-2, Bax e Bak), fibras do sistema colágeno e elástico e angiogênese. Entretanto, a linfangiogênese nunca foi estudada. Na última década várias moléculas funcionais e constitucionais que são expressas especificamente nas células do endotélio ou nos podócitos dos vasos linfáticos foram identificadas, como o VEGF-C, VEGFR-3 e o LYVE-1, possibilitando a melhor compreensão da linfangiogênese. Estudamos a imunomarcação dessas estruturas no carcinóide típico. Pela primeira vez no Brasil, a quantificação de vasos linfáticos foi realizada usando o LYVE1 como marcador. Apesar do uso de vários bloqueios de sítios inespecíficos não foi possível quantificar a expressão do VEGF-C e VEGFR-3 em carcinóides típicos, pois não encontramos controle interno negativo. Houve diferença significante entre as médias da idade em relação ao gênero. Não houve diferença significante entre as médias do diâmetro e número de linfonodos acometidos em relação ao gênero. Em relação ao grupo com e sem metástase encontramos difenca significante em relação ao diâmetro e ao comprometimento da margem. Não houve diferença da mediana do número de vasos linfáticos corados por mil células entre os grupos sem e com metástase linfática. Por regressão logística identificamos o diâmetro do tumor primário como uma variável independente preditiva do risco de metástase hematogênica e o diâmetro do tumor primário e a localização central ou periférica como variáveis independentes preditivas do risco de qualquer metástase (linfática ou hematogênica). O número de vasos linfáticos corados por mil células não foi identificado pelo modelo de regressão logistica como uma variável independente preditiva do risco individual de metástase linfática. Conclui-se que há correlação do diâmetro do tumor com o potencial de metástase hematogênica e há correlação entre diâmetro e localização do tumor primário e a ocorrência de metástase linfática ou hematogênica. A quantificação da imunoexpressão do LYVE-1 não demonstrou correlação. Outras técnicas devem ser estudadas e empregadas para identificar a importância da linfangiogênese no carcinóide típico. / Typical pulmonary carcinoids are neuroendocrine cells proliferations and they were former considered lung adenomas with no hematogenic or lymphatic metastatic potential. However, it is known that up to 20% of patients develop metastatic disease. It is mandatory that new studies be developed due to the variation in clinical presentation of these patients. It is also required that the individual risk of lymphatic and hematogenic metastasis be determined in order to individualize the patients treatment. Pulmonary carcinoids are classified according to hystologic grade. The current classification includes hystologic grade, presence or absence of necrosis, nuclear pleomorphism, and vascular invasion. This classification is based on Hematoxylin and Eosin stain and this technique can not assess biomolecular processes related to malignant potential. Trying to identify the malignant potential of the carcinoid tumors some studies have already been designed to identify some proteins as PCNA, p53, Ki-67, apoptosis proteins (Bcl-2, Bax and Bak), collagen and elastic fibers as well as angiogenic process. However, the lymphangiogenic mechanism has never been evaluated in typical pulmonary carcinoid tumors. Recently some molecules (VEGF-C, VEGFR-3 and LYVE-1) that are specifically expressed in the endothelium of the lymphatic vessels have been identified. These findings have improved the lymphangiogenic mechanism comprehension. This study used the immunohistochemical technique to identify VEGF-C, VEGFR-3 and LYVE-1 in 182 patients submitted to surgical procedures to treat Typical pulmonary carcinoid tumors. Lymphatic metastasis were diagnosed in 23 of 182 patients and 17 of 182 patients were identified with hematogenic metastasis. Futhermore, this study was the first reported one which has tried to quantify the lymphatic vessels using the LYVE-1 as an immunohistochemical marker. This study could not assess VEGF-C and VEGFR-e expression in Typical pulmonary carcinoids since an internal negative control could not be determined. There was a statistical difference between the median age and gender. There was no statistical difference between the median diameter and the number of positive lymph nodes related to the gender. This study demonstrated a statistical difference between the diameter and positive margins related to the group of patients that have developed metastatic disease and the group of patients with no metatastatic disease. There was no difference between the group of patients that have developed metastatic disease and the group of patients with no metatastatic disease according to the median number of lymphatic vessels stained. Based on logistic regression this study demonstrated that there is a predictive risk of developing hematogenic metastasis related to the diameter of the tumor. The predictive risk of the lymphatic metastasis was not improved by the number of the immunohistochemical stained lymphatic vessels, according to the logistic regression model. The immunohistochemical expression of LYVE-1 has not demonstrated statistical correlation between the parameters studied. Other than immnuhistochemical techniques are required to improve the comprehension of the lymphangiogenic mechanism involved in the Typical pulmonary carcinoid tumor

Os tumores carcinóides típicos broncopulmonares são proliferações de células neuroendócrinas. Foram consideradas como adenomas e acreditava-se que não tinham potencial para disseminação hematogênica e linfática. Porém, a ocorrência de metástase linfática e hematogênica acontece em um quinto dos indivíduos acometidos por essa patologia. A variação no comportamento clínico dos carcinóides broncopulmonares torna imperativa a realização de pesquisas que visem à melhor compreensão dessa doença. É fundamental determinar a agressividade e o risco individual da ocorrência de metástase linfática e hematogênica para que se possa oferecer um tratamento individualizado para cada binômio doente-doença. A classificação atual divide os tumores carcinóides, conforme o grau histológico de malignidade em típico e atípico, agrupando as neoplasias de acordo com o índice mitótico, relação volumétrica núcleo/citoplasma, presença ou ausência de necrose, pleomorfismo nuclear e invasão vascular. Esta análise, porém, é realizada em espécimes histológicos corados pela hematoxilina-eosina, técnica tradicional consagrada, mas que não permite avaliar processos biomoleculares relacionados ao potencial maligno das células que já podem estar presentes e não serem detectados pelo método. Em tumores carcinóides vários estudos já foram realizados na tentativa de identificar o potencial proliferativo de células que ainda não apresentam figuras de mitose, como PCNA, p53, Ki-67, o processo apoptótico (Bcl-2, Bax e Bak), fibras do sistema colágeno e elástico e angiogênese. Entretanto, a linfangiogênese nunca foi estudada. Na última década várias moléculas funcionais e constitucionais que são expressas especificamente nas células do endotélio ou nos podócitos dos vasos linfáticos foram identificadas, como o VEGF-C, VEGFR-3 e o LYVE-1, possibilitando a melhor compreensão da linfangiogênese. Estudamos a imunomarcação dessas estruturas no carcinóide típico. Pela primeira vez no Brasil, a quantificação de vasos linfáticos foi realizada usando o LYVE1 como marcador. Apesar do uso de vários bloqueios de sítios inespecíficos não foi possível quantificar a expressão do VEGF-C e VEGFR-3 em carcinóides típicos, pois não encontramos controle interno negativo. Houve diferença significante entre as médias da idade em relação ao gênero. Não houve diferença significante entre as médias do diâmetro e número de linfonodos acometidos em relação ao gênero. Em relação ao grupo com e sem metástase encontramos difenca significante em relação ao diâmetro e ao comprometimento da margem. Não houve diferença da mediana do número de vasos linfáticos corados por mil células entre os grupos sem e com metástase linfática. Por regressão logística identificamos o diâmetro do tumor primário como uma variável independente preditiva do risco de metástase hematogênica e o diâmetro do tumor primário e a localização central ou periférica como variáveis independentes preditivas do risco de qualquer metástase (linfática ou hematogênica). O número de vasos linfáticos corados por mil células não foi identificado pelo modelo de regressão logistica como uma variável independente preditiva do risco individual de metástase linfática. Conclui-se que há correlação do diâmetro do tumor com o potencial de metástase hematogênica e há correlação entre diâmetro e localização do tumor primário e a ocorrência de metástase linfática ou hematogênica. A quantificação da imunoexpressão do LYVE-1 não demonstrou correlação. Outras técnicas devem ser estudadas e empregadas para identificar a importância da linfangiogênese no carcinóide típico. / Typical pulmonary carcinoids are neuroendocrine cells proliferations and they were former considered lung adenomas with no hematogenic or lymphatic metastatic potential. However, it is known that up to 20% of patients develop metastatic disease. It is mandatory that new studies be developed due to the variation in clinical presentation of these patients. It is also required that the individual risk of lymphatic and hematogenic metastasis be determined in order to individualize the patients treatment. Pulmonary carcinoids are classified according to hystologic grade. The current classification includes hystologic grade, presence or absence of necrosis, nuclear pleomorphism, and vascular invasion. This classification is based on Hematoxylin and Eosin stain and this technique can not assess biomolecular processes related to malignant potential. Trying to identify the malignant potential of the carcinoid tumors some studies have already been designed to identify some proteins as PCNA, p53, Ki-67, apoptosis proteins (Bcl-2, Bax and Bak), collagen and elastic fibers as well as angiogenic process. However, the lymphangiogenic mechanism has never been evaluated in typical pulmonary carcinoid tumors. Recently some molecules (VEGF-C, VEGFR-3 and LYVE-1) that are specifically expressed in the endothelium of the lymphatic vessels have been identified. These findings have improved the lymphangiogenic mechanism comprehension. This study used the immunohistochemical technique to identify VEGF-C, VEGFR-3 and LYVE-1 in 182 patients submitted to surgical procedures to treat Typical pulmonary carcinoid tumors. Lymphatic metastasis were diagnosed in 23 of 182 patients and 17 of 182 patients were identified with hematogenic metastasis. Futhermore, this study was the first reported one which has tried to quantify the lymphatic vessels using the LYVE-1 as an immunohistochemical marker. This study could not assess VEGF-C and VEGFR-e expression in Typical pulmonary carcinoids since an internal negative control could not be determined. There was a statistical difference between the median age and gender. There was no statistical difference between the median diameter and the number of positive lymph nodes related to the gender. This study demonstrated a statistical difference between the diameter and positive margins related to the group of patients that have developed metastatic disease and the group of patients with no metatastatic disease. There was no difference between the group of patients that have developed metastatic disease and the group of patients with no metatastatic disease according to the median number of lymphatic vessels stained. Based on logistic regression this study demonstrated that there is a predictive risk of developing hematogenic metastasis related to the diameter of the tumor. The predictive risk of the lymphatic metastasis was not improved by the number of the immunohistochemical stained lymphatic vessels, according to the logistic regression model. The immunohistochemical expression of LYVE-1 has not demonstrated statistical correlation between the parameters studied. Other than immnuhistochemical techniques are required to improve the comprehension of the lymphangiogenic mechanism involved in the Typical pulmonary carcinoid tumor

<p>Subcutaneous administration has become a common approach for drug delivery of biotherapeutics, such as monoclonal antibodies, which is achieved mainly by absorption through the lymphatic system. This dissertation focuses on the computational modeling of the fluid flow and solute transport in the skin tissue and the quantitative investigation of lymphatic uptake. First, the various mechanisms governing drug transport and lymphatic uptake of biotherapeutics through subcutaneous injection are investigated quantitatively through high-fidelity numerical simulations, including lymphatic drainage, blood perfusion, binding, and metabolism. The tissue is modeled as a homogeneous porous medium using both a single-layered domain and a multi-layered domain, which includes the epidermis, dermis, hypodermis (subcutaneous tissue), and muscle layers. A systematic parameter study is conducted to understand the roles of different properties of the tissue in terms of permeability, porosity, and vascular permeability. The role of binding and metabolism on drug absorption is studied by varying the binding parameters for different macromolecules after coupling the transport equation with a pharmacokinetic equation. The interstitial pressure plays an essential role in regulating the absorption of unbound drug proteins during the injection, while the binding and metabolism of drug molecules reduce the total free drugs. </p> <p>  </p> <p>The lymphatic vessel network is essential to achieve the functions of the lymphatic system. Thus, the drug transport and lymphatic uptake through a three-dimensional hybrid discrete-continuum vessel network in the skin tissue are investigated through high-fidelity numerical simulations. The explicit heterogeneous vessel network is embedded into the continuum model to investigate the role of explicit heterogeneous vessel network in drug transport and absorption. The solute transport across the vessel wall is investigated under various transport conditions. The diffusion of the drug solutes through the explicit vessel wall affects the drug absorption after the injection, while the convection under large interstitial pressure dominates the drug absorption during the injection. The effect of diffusion cannot be captured by the previously developed continuum model. Furthermore, the effects of injection volume and depth on the lymphatic uptake are investigated in a multi-layered domain. The injection volume significantly affects lymphatic uptake through the heterogeneous vessel network, while the injection depth has little influence. At last, the binding and metabolism of drug molecules are studied to bridge the simulation to the experimentally measured drug clearance. </p> <p><br></p> <p>Convective transport of drug solutes in biological tissues is regulated by the interstitial fluid pressure, which plays a crucial role in drug absorption into the lymphatic system through the subcutaneous (SC) injection.  An approximate continuum poroelasticity model is developed to simulate the pressure evolution in the soft porous tissue during an SC injection. This poroelastic model mimics the deformation of the tissue by introducing the time variation of the interstitial fluid pressure. The advantage of this method lies in its computational time efficiency and simplicity, and it can accurately model the relaxation of pressure. The interstitial fluid pressure obtained using the proposed model is validated against both the analytical and the numerical solution of the poroelastic tissue model. The decreasing elasticity elongates the relaxation time of pressure, and the sensitivity of pressure relaxation to elasticity decreases with the hydraulic permeability, while the increasing porosity and permeability due to deformation alleviate the high pressure. </p> <p><br></p> <p>At last, an improved Kedem-Katchalsky model is developed to study solute transport across the lymphatic vessel network, including convection and diffusion in the multi-layered poroelastic tissue with a hybrid discrete-continuum vessel network embedded inside. The effect of different drug solutes with different Stokes radii and different structures of the lymphatic vessel network, such as fractal trees and Voronoi structure, on the lymphatic uptake is investigated. The drug solute with a small size has a larger partition coefficient and diffusivity across the openings of the lymphatic vessel wall, which favors drug absorption. The Voronoi structure is found to be more efficient in lymphatic uptake. </p> <p><br></p> <p>The systematic and quantitative investigation of subcutaneous absorption based on high-fidelity numerical simulations can provide guidance on the optimization of drug delivery systems and is valuable for the translation of bioavailability from the pre-clinical species to humans. We provide a novel approach to studying the diffusion and convection of drug molecules into the lymphatic system by developing the hybrid discrete-continuum vessel network. The study of the solute transport across the discrete lymphatic vessel walls further improves our understanding of lymphatic uptake. The novel and time-efficient computational model for solute transport across the lymphatic vasculature connects the microscopic properties of the lymphatic vessel membrane to macroscopic drug absorption. The comprehensive hybrid vessel network model developed here can be further used to improve our understanding of the diseases caused by the disturbed lymphatic system, such as lymphedema, and provide insights into the treatment of diseases caused by the malfunction of lymphatics.</p>