An extravascular fluid transport system based on structural framework of fibrous connective tissues in human body.

OBJECTIVE: Interstitial fluid in extracellular matrices may not be totally fixed but partially flow through long-distance oriented fibrous connective tissues via physical mechanisms. We hypothesized there is a long-distance interstitial fluid transport network beyond vascular circulations. MATERIALS AND METHODS: We first used 20 volunteers to determine hypodermic entrant points to visualize long-distance extravascular pathway by MRI. We then investigated the extravascular pathways initiating from the point of thumb in cadavers by chest compressor. The distributions and structures of long-distance pathways from extremity ending to associated visceral structures were identified. RESULTS: Using fluorescent tracer, the pathways from right thumb to right atrium wall near chest were visualized in seven of 10 subjects. The cutaneous pathways were found in dermic, hypodermic and fascial tissues of hand and forearm. The perivascular pathways were along the veins of arm, axillary sheath, superior vena cava and into the superficial tissues on right atrium. Histological and micro-CT data showed these pathways were neither blood nor lymphatic vessels but long-distance oriented fibrous matrices, which contained the longitudinally assembled micro-scale fibres consistently from thumb to superficial tissues on right atrium. CONCLUSIONS: These data revealed the structural framework of the fibrous extracellular matrices in oriented fibrous connective tissues was of the long-distance assembled fibres throughout human body. Along fibres, interstitial fluid can systemically transport by certain driving-transfer mechanisms beyond vascular circulations.

Anatomy of the lymphatic drainage of the upper limb and breast and its role in lymphedema prevention after breast cancer treatment / Anatomía del drenaje linfático del miembro superior y de la mama y su papel en la prevención del linfedema después del tratamiento del cáncer de mama

The surgical treatment of breast cancer has been enhanced throughout the years in order to offer oncologically safer and more effective results with lower esthetic impact and fewer sequelae. The lymphedema of the upper limb is still an iatrogenic result of great incidence and morbidity after this treatment. A possible existence of independent breast and upper limb lymphatic pathways has become the issue of many researchers willing to minimize its occurrence. This review aims to compare the lymphatic pathways in the axilla described by traditional anatomy books and recently published articles about Axillary Reverse Mapping (ARM). With this purpose, a comparative table was made with the descriptions found in books and articles, a statistic table of the data collected, a flowchart of anastomoses among nodes and an analytical drawing of the most statistically mentioned drained areas. It was observed that there is great variability in the descriptions of drainage and anastomoses among the lymph nodes in the references used, so there should be a consensus of a universal description which also assembles possible anatomical variations. Furthermore, the findings brought about by recent studies show possible anastomoses among pathways and lymph nodes, however they have not been taken into consideration when ARM was initially proposed. Therefore, the axillary resection with the preservation of the posterior and lateral axillary lymph nodes is theoretically possible to avoid lymphedema of the upper limb, but the development of an updated universal description that involves all possible anatomical variations will provide a safer and more effective treatment.

El tratamiento quirúrgico del cáncer de mama ha mejorado a lo largo de los años con el fin de ofrecer resultados oncológicamente más seguros y eficaces con menor impacto estético y menos secuelas. El linfedema del miembro superior es todavía un resultado iatrogénico de gran incidencia y morbilidad después de este tratamiento. La posible existencia de vías linfáticas de mama y de los miembros superiores independientes se ha convertido en un tema central de muchas investigaciones para lograr minimizar su ocurrencia. Esta revisión tiene como objetivo comparar las vías linfáticas en la axila descritas en los libros de anatomía tradicionales con artículos recientemente publicados sobre Mapeo Reverso Axilar (MRA). Con este fin, se realizó un cuadro comparativo con las descripciones y un diagrama de flujo de las anastomosis entre los nodos, además de un dibujo analítico de las áreas drenadas estadísticamente más mencionadas. Se observó que existe una gran variabilidad en las descripciones sobre el drenaje y las anastomosis entre los nodos linfáticos, por lo que la descripción universal no debería ser considerada un consenso debido a que también presenta posibles variaciones anatómicas. Por otra parte, los resultados producidos por los estudios recientes muestran posibles anastomosis entre las vías y los nodos linfáticos, sin embargo, no se han tomado en consideración cuando se propuso inicialmente el MRA. Por lo tanto, la resección axilar con la preservación de la parte posterior y los nodos linfáticos axilares laterales es teóricamente posible para evitar el linfedema del miembro superior, pero el desarrollo de una descripción universal actualizada, que incluya todas las posibles variaciones anatómicas, proporcionará un tratamiento más seguro y eficaz.

Temporal Change of Alcian Blue-Stained Primo Vascular System in Lymph Vessels of Rats.

This study aims to investigate the temporal change of a vascular system now known as the primo vascular system (PVS). We used Alcian blue (AB) dye for imaging the distribution of the PVS in lymphatic vessels. The target lymph vessels were chosen as they are easily accessible from the skin, and long-term observation is possible with intact physiological conditions due to a minimal surgical procedure. AB solution was injected into the inguinal lymph node and the target lymph vessels were located along the superficial epigastric vessels. The imaging system allowed processing for extraction of images showing changes in the AB intensity of the visualized PVS components. This newly developed procedure can be used for further study on various dynamic processes of PVS in lymph vessels.

Observation of a Flowing Duct in the Abdominal Wall by Using Nanoparticles.

The primo vascular system (PVS) is being established as a circulatory system that corresponds to acupuncture meridians. There have been two critical questions in making the PVS accepted as a novel liquid flowing system. The first one was directly to show the flow of liquid in PVS and the second one was to explain why it was not observed in the conventional histological study of animal tissues. Flow in the PVS in the abdominal cavity was previously verified by injecting Alcian blue into a primo node. However, the tracing of the dye to other subsystems of the PVS has not been done. In the current work we injected fluorescent nanoparticles (FNPs) into a primo node and traced them along a primo vessel which was inside a fat tissue in the abdominal wall. Linea alba is a white middle line in the abdominal skin of a mammal and a band of fat tissue is located in parallel to the linea alba in the parietal side of the abdominal wall of a rat. In this fat band a primo vessel runs parallel to the prominent blood vessels in the fat band and is located just inside the parietal peritoneum. About the second question on the reason why the PVS was not in conventional histological study the current work provided the answer. Histological analysis with hematoxyline and eosine, Masson's trichrome, and Toluidine blue could not discriminate the primo vessel even when we knew the location of the PVS by the trace of the FNPs. This clearly explains why the PVS is hard to observe in conventional histology: it is not a matter of resolution but the contrast. The PVS has very similar structure to the connective tissues that surround the PVS. In the current work we propose a method to find the PVS: Observation of mast cell distribution with toluidine blue staining and the PN has a high density of mast cells, while the lymph node has low density.

Primo Vessel Stressed by Lipopolysaccharide in Rabbits.

For tracking the primo vascular system, we observed the primo vessels in vivo in situ using the lipopolysaccharide (LPS) response in the lymphatic vessels of a rabbit. Injection of LPS (200 µg/kg) into the lymph nodes resulted in greatly stained primo vessels, which were swollen in some cases. We were able to obtain comparative images through alcian blue and diaminobenzidine staining, which clearly showed different morphologies of the primo vessels. The mechanism causing the response of the primo vessels to the injected LPS is still unclear; however, these results might be a first attempt at giving an explanation of the function of the primo vascular system and identifying the changes in the structure and function of the primo vascular system in response to an external stimulus such as an injection of LPS.

Effective isolation of primo vessels in lymph using sound- and ultrasonic-wave stimulation.

The effects of stimulation with sound and ultrasonic waves of a specific bandwidth on the microdissection of primo vessels in lymphatic vessels of rabbit were investigated. The primo vessels stained with alcian-blue dye injected in the lymph nodes were definitely visualized and more easily isolated by sound-wave vibration and ultrasonic stimulation applied to rabbits at various frequencies and intensities. With sound wave at 7 Hz and ultrasonic waves at 2 MHz, the probability of detecting the primo vessels was improved to 90%; however, without wave stimulation the probability of discovering primo vessels was about 50% only. Sound and ultrasonic waves at specific frequency bands should be effective for microdissection of the primo vessels in the abdominal lymph of rabbit. We suggest that oscillation of the primo vessels by sound and ultrasonic waves may be useful to visualize specific primo structure, and wave vibration can be a very supportive process for observation and isolation of the primo vessels of rabbits.

Visualization of the primo vascular system afloat in a lymph duct.

Because of the potential roles of the primo vascular system (PVS) in cancer metastasis, immune function, and regeneration, understanding the molecular biology of the PVS is desirable. The current state of PVS research is comparable to that of lymph research prior to the advent of Lymphatic vessel endothelial hyaluronan receptor 1 (LYVE-1). There is very little knowledge of the molecular biology of the PVS due to difficulties in identifying and isolating primo endothelial cells. Present investigations rely on the morphology and the use of differential staining procedures to identify the PVS within tissues, making detailed molecular studies all but impossible. To overcome such difficulties, one may emulate the explosive development of lymph molecular biology. For this purpose, there is a need for a reliable method to obtain PVS specimens to initiate the molecular investigation. One of the most reliable methods is to detect the primo vessels and primo nodes afloat in the lymph flow. The protocols for observation of the PVS in the large lymph ducts in the abdominal cavity and the thoracic cavity were reported earlier. These methods require a laparectomy and skillful techniques. In this work, we present a protocol to identify and harvest PVS specimens from the lymph ducts connecting the inguinal and the axillary nodes, which are located entirely in the skin. Thus, the PVS specimen is more easily obtainable. This method is a stepping-stone toward development of a system to monitor migration of cancer cells in metastasis from a breast tumor to the axillary nodes, where cancer cells use the PVS as a survival rope in hostile lymph flow.

Isolation and morphological features of primo vessels in rabbit lymph vessels.

Until now, even though intensive research has been dedicated to the primo vascular system (PVS) during these years, no statistical data on primo vessels and primo vessels in lymph flow have been available. Recently, the general morphological features of primo vessels in lymph vessels around the abdominal aorta were identified from microdissections of tissues from New Zealand White rabbits, and with Alcian blue staining, primo vessels in lymphatic vessels could be definitely identified under a digital microscope. The micro-dissected specimens in situ reveal rod-shaped nuclei stained by Acridine orange. The blue-stained nuclei, which were distributed in a broken-lined stripe, formed a tube structure of about 20 µm in diameter. The distance between the nuclei of two cells on neighboring aligned stripes, which is also the diameter of the micro tube, was measured to be about 5∼10 µm. The average length of the primo vessels was 2.4 mm, with the longest being 5.6 mm. The average size of the primo vessel was 50 µm, and the average diameters of the primo and the lymph vessels were 26.0 µm and 258.5 µm, respectively. Occasionally, without the use of Alcian blue staining, milk-white transparent primo vessels were observed floating in lymph vessels. Thus, we suggest that the PVS might also have an important function connected with the lymph system. We also expect the traditional Korean meridian system to leave its invisible world during the last thousands of years and soon enter the visible scientific world.

Primo vessel inside a lymph vessel emerging from a cancer tissue.

Primo vessels were observed inside the lymph vessels near the caudal vena cava of a rabbit and a rat and in the thoracic lymph duct of a mouse. In the current work we found a primo vessel inside the lymph vessel that came out from the tumor tissue of a mouse. A cancer model of a nude mouse was made with human lung cancer cell line NCI-H460. We injected fluorescent nanoparticles into the xenografted tumor tissue and studied their flow in blood, lymph, and primo vessels. Fluorescent nanoparticles flowed through the blood vessels quickly in few minutes, and but slowly in the lymph vessels. The bright fluorescent signals of nanoparticles disappeared within one hour in the blood vessels but remained much longer up to several hours in the case of lymph vessels. We found an exceptional case of lymph vessels that remained bright with fluorescence up to 24 hours. After detailed examination we found that the bright fluorescence was due to a putative primo vessel inside the lymph vessel. This rare observation is consistent with Bong-Han Kim's claim on the presence of a primo vascular system in lymph vessels. It provides a significant suggestion on the cancer metastasis through primo vessels and lymph vessels.

Protocol for the observation of the primo vascular system in the lymph vessels of rabbits.

Molecular-level understanding of the structure and the functions of the lymphatic system has greatly enhanced the importance of this second circulation system, especially in connection with cancer metastasis and inflammation. Recently, a third circulatory system, the primo vascular system (PVS) was found in various parts of an animal's body, especially as threadlike structures floating in the lymphatic flow in lymph vessels. Although the medical significance of this emerging system will require much work in the future, at present, several important suggestions in connection with immune cells, stem cells, and cancer metastasis have already appeared. Experiments to observe the PVS in the lymph vessels near the caudal vena cava of rabbits and rats have been performed by several independent teams, but reproduction requires considerable skill and technical know-how. In this article, we provide a detailed protocol to detect the PVS inside the lymph vessels of a rabbit. Detection and isolation are the first steps in unraveling the physiological functions of the PVS, which awaits intensive research.

Study of novel threadlike structures on the intestinal fascia of dogs.

The primo-vascular system was visualized in the mesentery surrounding the small intestine of a dog using Trypan blue. This structure, which was first observed in a rat, formed a network as primo-vessel branches were joined to primo-nodes. Other characteristic features of the primo-vascular system, such as bundles of tubes with fibrous extracellular matrix in a primo-vessel and a broken-line alignment of rod-shaped nuclei along the primo-vessel, were observed. Blood vessels, lymph vessels, and primo-vessels were present in the same mesentery, and they could clearly be distinguished by histological differences.

Identification of lymphatics in the ciliary body of the human eye: a novel "uveolymphatic" outflow pathway.

Impaired aqueous humor flow from the eye may lead to elevated intraocular pressure and glaucoma. Drainage of aqueous fluid from the eye occurs through established routes that include conventional outflow via the trabecular meshwork, and an unconventional or uveoscleral outflow pathway involving the ciliary body. Based on the assumption that the eye lacks a lymphatic circulation, the possible role of lymphatics in the less well defined uveoscleral pathway has been largely ignored. Advances in lymphatic research have identified specific lymphatic markers such as podoplanin, a transmembrane mucin-type glycoprotein, and lymphatic vessel endothelial hyaluronan receptor-1 (LYVE-1). Lymphatic channels were identified in the human ciliary body using immunofluorescence with D2-40 antibody for podoplanin, and LYVE-1 antibody. In keeping with the criteria for lymphatic vessels in conjunctiva used as positive control, D2-40 and LYVE-1-positive lymphatic channels in the ciliary body had a distinct lumen, were negative for blood vessel endothelial cell marker CD34, and were surrounded by either discontinuous or no collagen IV-positive basement membrane. Cryo-immunogold electron microscopy confirmed the presence D2-40-immunoreactivity in lymphatic endothelium in the human ciliary body. Fluorescent nanospheres injected into the anterior chamber of the sheep eye were detected in LYVE-1-positive channels of the ciliary body 15, 30, and 45 min following injection. Four hours following intracameral injection, Iodine-125 radio-labeled human serum albumin injected into the sheep eye (n = 5) was drained preferentially into cervical, retropharyngeal, submandibular and preauricular lymph nodes in the head and neck region compared to reference popliteal lymph nodes (P < 0.05). These findings collectively indicate the presence of distinct lymphatic channels in the human ciliary body, and that fluid and solutes flow at least partially through this system. The discovery of a uveolymphatic pathway in the eye is novel and highly relevant to studies of glaucoma and other eye diseases.

Comparison of the characteristic features of Bonghan ducts, blood and lymphatic capillaries.

OBJECTIVE: To show that the characteristic morphological and ultrastructural features of a Bonghan corpuscle and duct presented here are consistent with the description given in the early reports of Bonghan Kim. MATERIALS AND METHODS: We compared the morphological aspects of Bonghan ducts with those of blood and lymphatic capillaries on the ultrastructural level to display the manifestly distinctive nature of the Bonghan system. RESULTS: The walls of the ductules were observed to be composed of a single layer of endothelial cells with characteristic rod-shaped nuclei and were not surrounded by a basal lamina or by accessory cells, such as pericytes or smooth muscle cells. The abluminal cell membranes of Bonghan ductules were not attached by anchoring filaments to the fibers of extracellular matrices as observed in lymphatic capillaries. The cytoplasmic processes of ductule endothelial cells appear to form overlapping and interdigitated interconnections which completely lack junctional elements. Although the cytoplasm of ductule endothelial cells contained a well-developed rough endoplasmic reticulum and many free ribosomes and polysomes, there was a relatively small number of pinocytotic vesicles and lacks specific organelles, such as Weibel-Palade bodies. CONCLUSIONS: The Bonghan corpuscles are specialized structures consisting of different types of immune cells randomly scattered as single cells in the matrix or clustered in follicle-like formations. Moreover, the Bonghan ductules in the corpuscle contain flowing immune cells and occasionally basophilic bodies.

Avances en el tratamiento kinésico del linfedema secundario en pacientes con cáncer de mama / Progress in Kinesic treatment of secondary lymphedema in patients with breast cancer

En el tratamiento del cáncer de mama, la complicación más frecuente que se presenta es el linfedema secundario del miembro superior. El linfedema es una enfermedad crónica provocada por una destrucción de la anatomía linfática normal. Aparece en un 20% -25% de los casos y hasta un 35% cuando la cirugía se asocia a radioterapia. Pese a los importantes avances existentes en el tratamiento de linfedema, no hay un consenso entre los investigadores respecto a la elección de la terapia adecuada, debido a la falta de criterios uniformes de diagnósticos, que se traducen en la derivación tardía de los pacientes hacia los profesionales especializados. El enfoque terapéutico actual consiste en la integración de equipos multidisciplinarios reunidos en un Programa Integral de Rehabilitación del Edema (PIR), el que reúne enfoques preventivos - a través de la educación del paciente - y conservadores tales como el Drenaje Linfático Manual, presoterapia secuencial intermitente, ejercicios específicos, vendaje multicapas y elementos de contención. Estos tratamientos si bien no buscan la cura del Linfedema, permiten minimizar las secuelas funcionales y mejorar la calidad de vida de las pacientes portadoras de esta patología.

In breast cancer treatment, the most common complication is the Secondary Lymphedema of the upper limb. Lymphedema is a chronic disease caused by the destruction of the normal lymphatic anatomy. It is produced in a 20-25% of the cases, and up to 35% when the surgery is associated to radiotherapy. Despite the important existing advances in the management of Iymphedema, there is no consensus between the investigators with respect to the election of the suitable therapy, due to the lack of uniform criteria of diagnosis, which are translated in a delayed derivation of the patients towards the specialized professionals. The present therapeutic approach, consists of the integration of multidisciplinary teams joined in an Integral Program of Rehabilitation of Edema (IPR), which gathers preventive approaches - through the education of the patient - and conservative approaches, such as the MDL (Manual Lymphatic Drainage), Intermittent Sequential Pressotherapy specific. Exercises, multi layer Bandage, and elements of containment. These treatments, although do not aim at the cure of the Lymphedema, allow the decrease of the functional sequels and improve the quality of life of the patients who carry this pathology.

[Anatomical visibility of the meridian information channels].

The "structure imperfectness of signal channel rule" put forward by the author of the present paper may provide a theoretical evidence for the systematicness of meridian information channel. The conclusion that no special structure of the meridian-collateral system has been found is likely to serve as a piece of counterevidence. According to the latest structural view, the development of capillaries, lymphatic vessels and nerves needs target cells-released inducible factors. In the initial phase of the development of the organism, the asymmetry of the interspaces among cells results in the production and arrangement imbalance of the sequential factors which make the capillaries, lymphatic vessels and nerves distribute sequentially in time and space. Meridian-collateral, following the "systemic statistic distribution rule", is a general expression of this distribution pattern. As a systematic structure, the meridian-collateral system distributes in an optimized way in the human body and has both orderly and compatible characteristics. The author thinks that the meridian-collateral information channel is anatomically visible in the time and spatial structure, and in its logical structure and compatibility during the process of growth. Hence, many techniques of delicate anatomy, quantitative anatomy, growth anatomy, comparative anatomy, tridimensional remodeling of living creature and dynamical remodeling of growth all should be used as the important tools for studying the meridian information channel. The theory and the anatomical techniques determine what you would finally find.

Direct lymphatic spreading route into the liver from the gallbladder: an animal experiment using pig.

In the special occasion that the physiological lymphatic flow is obstructed, gallbladder carcinoma (GBC) may spread into the liver via lymphatic route. Therefore, this study was conducted to find out the direct lymphatic route draining into the liver from the gallbladder using pigs with ligated cystic ducts. After injecting the carbon particle suspension (CH40) or the contrast medium (Lipiodol) into the subserosal layer of the gallbladder, the lymphatic route into the liver was examined both macroscopically and histologically. In controls, CH40 or Lipiodol drained along the cystic duct toward the hepatoduodenal ligament. After occlusion of cystic duct, CH40 was interrupted at the ligated point, and then spread into the liver nearby the gallbladder bed, running off to the liver hilus, toward the hepatoduodenal ligament. This route was confirmed by the Lipiodol drainage into the right median lobe of the liver, equivalent to the segments V and IV a in humans. We presented for the first time the emergence of lymphatic drainage from the gallbladder into the liver after the occlusion of physiological lymphatic route using pigs. This implies that the direct spread into the segments V and IV a of liver should be considered in the surgical treatment of advanced GBC.

[The theory of lymphangion and current approaches to the pathogenesis, diagnosis and treatment of lymphedema of the lower extremities]. / Teoriia limfangiona i sovremennye podkhody k patogenezu, diagnostike i lecheniiu limfedemy nizhnikh konechnostei.

The paper describes the results of examination carried out by the Sankt-Peterburg school of lymphologists, pertaining to the structure, physiological properties and function of lymphangions responsible for active lymph transport. The problems of the pathogenesis, diagnosis and treatment of lymphedema of the lower extremities used in clinical practice are reviewed from the standpoint of the new theory. The data obtained as a result of the clinico-morphofunctional studies allowed to delineate the stages of lymphedema as dependent on the degree of lymphocytic structure and function integrity. Based on the aforesaid the new approaches to the diagnosis and selection of the treatment methods for lymphedema of the lower extremities have been formulated.

Perivascular space: possible anatomical substrate for the meridian.

BACKGROUND: Despite the meridian system being an important concept in Traditional Chinese Medicine (TCM), modern biology and Western medical systems have failed to find an anatomic substrate. Since the 1960s, a variety of phenomena along meridians have been reported, among which quite a few suggest that along meridians there is a fluid pathway (but not blood vessels or lymphatics). On the other hand, perivascular space (PVS) has been demonstrated to be a body fluid pathway in addition to blood vessels and lymphatics in some mammalian tissues, such as brain, thymus, and lung. OBJECTIVES: The present study was designed to examine the relationship between PVS and the meridian. We studied characteristics of the tissues around the blood vessels along the Stomach Meridian of Foot-Yangming and the Gallbladder Meridian of Foot-shaoyang, with the goal of identifying anatomical structure corresponding to the meridian described in TCM. DESIGN AND RESULTS: Through perivascular dye injection and frozen section histology, we found that there is PVS around the blood vessels along the meridians, and it is a fluid pathway. Subsequent physiologic studies revealed that the PVS shows significantly greater electrical conductivity and significantly higher partial oxygen pressure (pO(2)) compared to medial and lateral tissues. CONCLUSIONS: The PVS along the meridians has properties offering good explanation for the meridian phenomena. The work sheds new light on the studies of meridians and may contribute to research on the mechanism of Chinese acupuncture.