Functional analysis in vivo of engineered valved venous conduit with decellularized matrix and two bone marrow-derived progenitors in sheep.

Tissue engineering has been considered a promising approach for creating grafts to replace autologous venous valves. Here, ovine bone marrow-derived endothelial progenitor cells (EPCs) and multipotent adult progenitor cells (MAPCs) were harvested and then loaded into decellularized venous matrix to create tissue-engineered (TE) valved vein. Subsequently, the ovine femoral veins containing the valve were removed and replaced by TE grafts or acellular matrix only. The morphology and function were analysed for up to 1 year by ultrasonography, angiography, H&E staining and scanning electron microscopy (SEM). The differentiation of seeded cells was traced immunofluorochemically. The results showed that decellularized venous matrix could initially and feebly attract endogenous cells, but failed afterwards and were insufficient to restore valve function. On the contrary, the seeded cells differentiated into endothelial cells (ECs) in vivo and formed a monolayer endothelium, and smooth muscle cells within the scaffold therefore produced TE grafts comparable to the native vein valve. This TE graft remained patent and sufficient after implantation into the venous circuit of the ovine lower extremity for at least 6 months. Unfortunately, cells seeded on the luminal surface and both sides of the leaflets lost their biological functions at 12 months, resulting in thrombosis formation and leading to complete occlusion of the TE grafts and impotent venous valves. These findings suggest that this TE valved venous conduit can function physiologically in vivo in the medium term. Before translating this TE venous valve into clinical practice, the durability should be improved and thrombogenicity should be suppressed. Copyright © 2016 John Wiley & Sons, Ltd.

Biocompatibility evaluation of tissue-engineered valved venous conduit by reseeding autologous bone marrow-derived endothelial progenitor cells and multipotent adult progenitor cells into heterogeneous decellularized venous matrix.

Clinical treatment of chronic deep venous insufficiency remains difficult despite the availability of various therapies. Previous experimental efforts have demonstrated that the tissue-engineered valvedvenous conduit (TEVV) is a promising option to replace the damaged venous valve. The aim of the present study was to evaluate the TEVV by reseeding bone marrow-derived endothelial progenitor cells and multipotent adult progenitor cells into acellular matrix according to International Standard ISO10993, and to clarify their interactions with blood, the local effect after implantation both in vitro and vivo, and immunogenicity. The results showed that the 2-cm long TEVV did not cause haemolysis in vitro and remained patent without thrombosis formation in vivo. However, the luminal surface of TEVV was partially covered by multilayer cells. Compared with the native ovine femoral vein segment, the TEVV beneath the mouse skin produced significant mononuclear cell infiltration, with serum interleukin-6 and tumour necrosis factor-α similar to normal. The TEVV maintained its structural integrity, while the native ovine femoral vein segments fell apart at postoperative week nine. The TEVV implantation did not change serum immunoglobulin G. In addition, the seeds and extracts of the scaffold did not affect the proliferation of mouse lymphocytes. These findings suggest that the histocompatibility, haemocompatibility and immunogenicity of this TEVV are acceptable owing to complete removal of the cellular components of autologous seeds and residues of chemical regents, thus providing an experimental basis for further clinical translation. Copyright © 2014 John Wiley & Sons, Ltd.

Architectural properties of the neuromuscular compartments in selected forearm skeletal muscles.

The purposes f this study were to (i) explore the possibility of splitting the selected forearm muscles into separate compartments in human subjects; (ii) quantify the architectural properties of each neuromuscular compartment; and (iii) discuss the implication of these properties in split tendon transfer procedures. Twenty upper limbs from 10 fresh human cadavers were used in this study. Ten limbs of five cadavers were used for intramuscular nerve study by modified Sihler's staining technique, which confirmed the neuromuscular compartments. The other 10 limbs were included for architectural analysis of neuromuscular compartments. The architectural features of the compartments including muscle weight, muscle length, fiber length, pennation angle, and sarcomere length were determined. Physiological cross-sectional area and fiber length/muscle length ratio were calculated. Five of the selected forearm muscles were ideal candidates for splitting, including flexor carpi ulnaris, flexor carpi radials, extensor carpi radialis brevis, extensor carpi ulnaris and pronator teres. The humeral head of pronator teres contained the longest fiber length (6.23 ± 0.31 cm), and the radial compartment of extensor carpi ulnaris contained the shortest (2.90 ± 0.28 cm). The ulnar compartment of flexor carpi ulnaris had the largest physiological cross-sectional area (5.17 ± 0.59 cm(2)), and the ulnar head of pronator teres had the smallest (0.67 ± 0.06 cm(2)). Fiber length/muscle length ratios of the neuromuscular compartments were relatively low (average 0.27 ± 0.09, range 0.18-0.39) except for the ulnar head of pronator teres, which had the highest one (0.72 ± 0.05). Using modified Sihler's technique, this research demonstrated that each compartment of these selected forearm muscles has its own neurovascular supply after being split along its central tendon. Data of the architectural properties of each neuromuscular compartment provide insight into the 'design' of their functional capability. In addition to improving our understanding of muscle anatomy and function, elucidation of forearm neuromuscular compartments architecture may ultimately provide information useful for selection of muscle subdivisions used in tendon transfer.

Applied Anatomy of the Femoral Veins in Macaca fascicularis / Anatomía Aplicada de las Venas Femorales en Macaca fascicularis

The aim was to understand the anatomical features of the venous valve in Macaca fascicularis and to compare it with that of humans. The bilateral lower limbs (24 limbs from 12 animals) of Macaca fascicularis cadavers were dissected, and the femoral veins (FVs) were equally divided into distal, intermediate, and proximal sections. The external diameter of the FV in each section was measured. The venous valves were observed microscopically and stained with hematoxylin and eosin as well as trichrome. Data describing the human venous valve were collected from the current literature. No great saphenous veins were found among the 24 lower limbs from the Macaca fascicularis cadavers. The external diameters of the FVs in the distal, intermediate, and proximal sections were 3.53 ± 0.37 mm, 3.42 ± 0.55 mm, and 3.37 ± 0.54 mm, respectively. In most cases, there was one venous bivalve located in the FV approximately 0-2.71 mm below the junction of the FV and the deep femoral vein. Endothelium covered the luminal and sinusal surfaces of the leaflets. Abundant collagen fibers were found under the endothelial cells beneath the luminal surface of the leaflets. An elastin fiber network was located under the sinus endothelial surface. Smooth muscle cells in the FV extend to the edge of the valve. The venous valve of Macaca fascicularis is similar to that of humans, both morphologically and histologically. However, there is only one venous bivalve and no great saphenous vein in Macaca fascicularis.

El objetivo fue comprender las características anatómicas de la válvula venosa en Macaca fascicularis y compararla con la de los humanos. Fueron disecados bilateralmente los miembros pélvicos (24 miembros de 12 animales) de cadáveres de Macaca fascicularis; las venas femorales (VF) fueron divididas en secciones distal, media y proximal. Se midió el diámetro externo de las VFs en cada sección. Las válvulas venosas se observaron microscópicamente y se tiñeron con H-E y tricrómico. Los datos para describir la válvula venosa humana se obtuvieron desde la literatura. No se encontraron venas safenas magnas entre los 24 miembros inferiores. Los diámetros externos de las VFs en las secciones distal, media y proximal fueron 3,53±0,37 mm, 3,42 mm±0,55, y 3,37±0,54 mm, respectivamente. En la mayoría de los casos, hubo vena bivalva situada aproximadamente 0-2,71 mm debajo de la unión de la VF y la vena femoral profunda. El endotelio cubrió las superficies luminal y sinusal. Se observaron abundantes fibras de colágeno en las células endoteliales bajo la superficie luminal de las válvulas. Una red de fibras de elastina se encontró bajo la superficie del seno endotelial. Las células musculares lisas en las VFs se extiendían hasta el margen de la válvula. La válvula venosa del Macaca fascicularis es similar a la de los seres humanos, morfológica e histológicamente. Sin embargo, sólo hubo una vena bivalvular, y no se observaron venas safenas en Macaca fascicularis.

Two methods can simultaneously display both intramuscular nerves and blood vessels.

BACKGROUND: The arrangement and relationship of intramuscular nerves and blood vessels are critical to clinical physicians, but the majority of previous studies could not reflect them precisely. METHODS: In method 1, after rabbits were perfused with barium sulfate liquid silica gel, the specimen muscle was isolated and subjected to Sihler staining. The specimen muscles then underwent optical photography and molybdenum target radiography. The obtained photograph and the radiograph were then overlapped together to draw a map of intramuscular nerves and blood vessels. In method 2, after rabbits and child cadavers were perfused with semitransparent red liquid silica gel, the specimen muscle was isolated and subjected to Sihler staining. Finally, the specimen muscles were placed on an x-ray film viewer for photography. RESULTS: Both methods had ideal outcomes. Through digital subtraction angiography, a distribution map of intramuscular nerves and blood vessels could be obtained in method 1, whereas in the photographs taken through method 2, the intramuscular nerves were counterstained with dark blue and the intramuscular blood vessels were stained red, which was more precise and direct. In method 2, specimens could be made into stereoscopic models. CONCLUSIONS: These two methods that can simultaneously display intramuscular nerves and blood vessels have been significantly improved and lead to a good result. They also have their own advantages. When a muscle studied is small or flat, method 2 is recommended because of its conciseness and convenience. If the muscle is large, method 1 is applicable.

Anatomic study of fabella and its surrounding structures in a Chinese population.

PURPOSE: To explore the anatomic features of the fabella and its relationship with the common peroneal nerve and the fabellofibular ligament, so as to provide anatomical evidence for clinical diagnosis and treatment of fabella diseases in a Chinese population. METHODS: Sixty-one formalin-fixed knee specimens were obtained for anatomic dissection. Structural features of the fabella were investigated by radiological and histological tests. RESULTS: There were 53 cases (86.89%) with fabellae in the lateral head of the gastrocnemius muscle, including 34 bony ones (55.74%), whereas only 6 cases had fabellae in the medial head (9.84%). The fabellae were accompanied by common peroneal nerves on their surfaces in 11 cases (20.8%), and the presence of the fabella was not generally predictive of a fabellofibular ligament. As much as 57.9% of the cartilage fabellae were not visualized on radiograph. The structure of the ossified fabella is similar to a typical long bone. CONCLUSIONS: Fabellae were mainly present in the lateral head of the gastrocnemius muscle in a large proportion of the Chinese population. More than half of the cartilage fabellae were not visualized on radiograph. Its clinical significance could not be ignored by physicians and anatomists.

Anatomic study of popliteus complex of the knee in a Chinese population.

This study explored the anatomic relationship of the popliteus complex (PC) and collected anatomic data of PC in a Chinese population. The anatomic study was performed using 81 formalin-fixed knees. The femoral attachment of the popliteus tendon could be classified into three types with respect to femoral attachment of the lateral collateral ligament. The popliteofibular ligament presented as ligament (87.7%) or fascia (12.3%), originating from the musculotendinous junction of the popliteus muscle or just proximal to it. Given the great variability of the posterolateral structures, reconstructive surgeons should design a method based on individual anatomic features, rather than a fixed reconstructive surgery.

Profiling of innervations of mimetic muscles in fresh human cadavers using a modified Sihler's technique.

The nerve map of the human face, although important in guiding facial surgery, has not been well defined. In this study we applied a modified Sihler's technique to profile intramuscular innervation of human mimetic muscles. Six fresh cadaveric heads were used. The intramuscular distribution of the facial nerve in human mimetic muscles was visualized using a modified Sihler's technique. Modified Sihler's staining revealed a three-dimensional picture of the clearly purple-black intramuscular facial and sensory nerves. The nerve branching patterns of both facial halves were asymmetrical. None of the fine nerve branches crossed over the midline. The facial nerve branches divided into secondary rami and formed a mesh-like plexus before entering the target muscles at a right angle. The modified Sihler's technique can profile intramuscular innervation of human mimetic muscles. Our nerve map of the face offers valuable guidance for facial reanimation surgery, facial cosmetic surgery, and parotid surgery.

Quantitative anatomical study of male pelvic autonomic plexus and its clinical potential in rectal resection.

The pelvic autonomic nerves innervate the pelvic viscera, and carry a high risk of damage during surgery. This high risk has been ascribed to the complex interrelationship of pelvic paravisceral structures and the difficulty in identifying particular structures, despite the fact that the anatomic characteristics of the pelvic autonomic plexus have been well documented. We dissected ten male embalmed adult cadavers with particular attention to the quantitative parameters of the pelvic plexus and its subsidiary plexus. The right inferior hypogastric plexus and its rectal branch were found to be significantly longer and wider than the left one, while the transverse diameter of the vesical and prostatic branches of the left side was significantly larger the right. The inferior mesenteric plexus gave off fibers directly to form the pelvic plexus in four of 20 hemipelves (20%). In the side-by-side comparison, the distance to midpoint of the sacral promontory of the left rectal plexus was significantly longer than that of the right, whereas the maximum length (the length of the longest nerve fiber from origin to corresponding organ) of the left vesical plexus was significantly shorter than that of the right. Additionally, the craniocaudal and dorsoventral diameters of the right pelvic autonomic plexus were significantly shorter those of the left. The quantitative parameters relating to the pelvic autonomic plexuses not only can enhance our understanding of its anatomy and function, but can also be used as references for surgical procedures and robot-assisted surgery.

Intramuscular innervations of muscle flaps that are commonly used in clinical settings.

PURPOSE: Abductor hallucis, latissimus dorsi, gracilis, rectus abdominis, sartorius and pectoralis minor are muscle flaps that are commonly used in clinic, but their intramuscular innervation has seldom been systematically investigated. METHODS: Five Chinese fresh human cadavers were included in the study and abductor hallucis, latissimus dorsi, gracilis, rectus abdominis, sartorius and pectoralis muscles were dissected. After gross anatomy measurement, the specimens were then stained by Sihler's staining technique. Intramuscular innervation was observed and the number as well as distribution was recorded. RESULTS: Intramuscular nerves were clearly visualized by Sihler's staining technique. Based on the shape and muscle-tendon morphology, Lim et al. in Muscle Nerve 29:523-530, 2004 grouped the muscles into trapezoidal-shaped (type I), spindle-shaped (type II), and combination-shaped (type III). According to Lim's study the abductor hallucis was a type IIb muscle and was divided into two compartments by the distal tendon. Latissimus dorsi was a type I muscle, divided into 3-4 compartments by intramuscular nerve branches. Gracilis was a type IIa muscle and the distal part was divided into two compartments by intramuscular nerve branches. Rectus abdominis was a type III muscle and the four bellies comprised four compartments, each of which could be designated as a subunit. Sartorius was a type II muscle and it could be divided into 2-3 compartments along the long axis. Pectoralis minor was a type I muscle that was divided into two compartments by extramuscular terminal nerves. CONCLUSIONS: The six muscles are divided into several compartments by the tendon or nerve branches, and all of them make good donor tissue for muscle compartment transfer in reconstructive surgery.

Neurovascular details about forearm muscles: applications in their clinical use in functional muscular transfer.

PURPOSE: The concept of muscle subunits has been introduced for limb reconstruction and functional conservation of donor site, yet it requires thorough understanding of neurovascular anatomy of muscles. The present study provides neurovascular details of forearm skeletal muscles. METHODS: Twenty-eight forearms were dissected totally, ten for observing extramuscular nerve distribution, ten for observing intramuscular nerve distribution by modified Sihler's technique, and the other eight for observing intramuscular artery distribution by aqueous barium sulfate infusion. The forearm muscles were classified into three different types according to Lym's classification. Numbers of extramuscular and intramuscular nerves were counted and compared between the types. Intramuscular vascular distribution was also classified into three different types according to Mathes' method. Intramuscular vascular distribution was compared with nerve distribution as well. RESULTS: There were also some variations in the composition of muscular branches. Numbers of intramuscular nerve branches were significantly higher in muscle types IIa and III. Morphology and intramuscular neurovascular distribution of type II muscles are easy to split into two independent parts for functional muscular flap transfer according to morphology and intramuscular neurovascular distribution. CONCLUSIONS: The present study elucidated that flexor carpi ulnaris and radialis, extensor carpi ulnaris, and carpi radialis brevis were suitable for functional muscular transfer from several different insights, but the roles of pronator teres and supinator remained to be controversial. This study might be very helpful to plastic surgeons and anatomists.

Morphological characteristics of the anterior ethmoidal artery in ethmoid roof and endoscopic localization.

OBJECTIVES: To provide anatomical data to help identify and locate the anterior ethmoidal artery (AEA) precisely during endoscopic procedures. METHOD: We dissected 15 adult cadaver heads, which provided 30 specimens, to study morphological characteristics, courses, and several types of variations. RESULTS: We found the average diameter of the AEA to be 0.80 +/- 0.24 mm. In 85.7% of the cases, the artery was seen between the second and third lamella. Other locations were over the roof of the frontal recess cells (10.7%) and the roof of the posterior ethmoid sinus (3.6%). The AEA ran parallel to the ethmoid roof and formed a slight curve. When viewed from the superior side, the angle formed by the long axis of the artery and the lamina papyracea was 60.5 degrees +/- 16.4 degrees. In 83.3% of the cases, the anterior ethmoidal canal (AEC) was identified as a separate canal, and in 16.7% the canal was embedded in the ethmoid roof. In 10 of the 30 cases (33.3%), the AEC presented some degree of dehiscence. CONCLUSION: As a result of these dissections, we found that the AEA's course in the ethmoid roof varies. The morphological characteristics-that the AEA runs parallel to the ethmoid roof, forming a slight posterolateral to anteromedial curve as it passes from the orbit to the cribriform plate-are the most reliable factors used to identify the artery during surgery.

[Anatomic study of intramuscular nerve and blood vessel in forearm muscles: an anatomical study].

OBJECTIVE: To study the distribution of intramuscular nerve and blood vessels in forearm muscles and to discuss the possibility of dividing the forearm muscles into independent functional units. METHODS: (1) The muscles were dissected in 10 forearms from 5 fresh adult human cadavers and stained with the Sihler's nerve staining; (2) The blood vessels were studied in eight forearm muscles from 4 fresh adult human cadavers with irrigation of a mixture of 30% barium sulfate and gelatin from brachial artery and then X-photographed. All pictures were compared to study the intramuscular distribution of nerve and blood vessels. RESULTS: The intramuscular nerve branches were stained purple-black and visualized clearly. The muscles were classified into three types according to the distribution characters of intramuscular nerve and blood vessels. And the types of muscles could be further subdivided into a and b subtypes. CONCLUSION: According to the neurovascular distribution, the forearm muscles in type II a and type III a can be divided into independent function units for muscle functional transplantation.

Observation and measurements of long thoracic nerve: a cadaver study and clinical consideration.

Long thoracic nerve (LTN) is an important nerve originating from cervical nerve roots. It varies a lot in origins and branches, which lead to several clinical problems, such as diagnosis, prophylaxis and treatment of LTN injury. LTN was dissected in 38 cadavers in the present study. Origin, level of union, branches, sites where nerve entered the muscle, length of nerve trunk and branches as well as transverse diameter were documented. Different derivations of LTN were observed, and C4-7, C5-7, C5 and C7, C5-7, C5-8, C6 and C7, and branch from C6 was the most important components of LTN. After evolution, LTN trunk was composed by superior and inferior trunks at scalenus muscle or the three superior slips level. Branches of LTN traveled on the surface of the six superior slips of anterior serratus muscle and then penetrated through the inferior slips without correlation between different branches. Mean length of trunk of LTN is 111.73 (30.08) mm, axis of cross section was 2.27x0.96 mm at the union level and 1.91 x 0.68 mm at the end branch. Each slip was innervated by 1-4 branches of LTN. The observation and measurement data described in our study presented some variations and could provide clinicians with important information on diagnosis, prophylaxis and treatment of LTN injury and pursuing more suitable muscle flaps for reconstruction operation.

[Anatomy of buccal and marginal mandibular branches of facial nerve and its clinical significance].

OBJECTIVE: To study the course and distribution of buccal and marginal mandibular branches of facial nerve, and its relevance to the treatment of facial paralysis and the protection of facial nerve during surgery. METHODS: 12 cadaver heads were dissected (24 specimens). The course of the buccal and marginal mandibular branch and the interconnections between them were observed. The relationship of buccal branch to parotid duct, marginal mandibular branch to the inferior border of mandible were studied. With modified Sihler's staining technique, the distribution of facial nerve branches in innervated mimetic muscles was displayed. These anatomic relationships mentioned above were further confirmed during the operation of 40 patients with facial paralysis. RESULTS: Parotid duct had a constant surface landmark. Buccal branch mainly consisted of 2-3 ramifications in 87.5% of the specimens, while marginal mandibular branch was double or single in 95.9% of the specimens. The buccal branch coursed within the distance between 10.7 mm above and 9.3 mm below the parotid duct, and innervated mimetic muscles of midface. The marginal mandibular branch coursed within the distance between 13.4 mm above and 4.8 mm below the lower border of mandible, crossed superiorly the facial artery and innervated mimetic muscles of lower lip. CONCLUSIONS: There is a close relationship of buccal branch to parotid duct and marginal mandibular branch to facial artery and lower border of mandible. With modified Sihler's staining technique, the original 3-dimensional picture of the intramuscular nerve distribution in human mimetic muscles.