Acute cerebral artery constriction in the spontaneously hypertensive rat following blood and plasma administration into the subarachnoid space

The purpose of the present study was to demonstrate, using a vascular casting technique, acute vasoconstrictive changes in the cerebral vasculature 1 h following whole-blood or plasma infusion into the subarachnoid space of conscious spontaneously hypertensive rats. Vascular casts from animals infused (over 20 min) with 0.45 ml of heparinized autologous arterial blood or plasma exhibited incomplete filling, while casts from saline-infused controls exhibited virtually no filling defects. Significant elevations in intracranial pressure were noted in blood, but not in plasma- or saline-infused rats. Two characteristic forms of constriction occurred, depending upon the vessel lumen diameter. Vessels with lumen diameters >100 µm were flattened longitudinally with deep endothelial nuclear imprints, while smaller vessels had focal circular constrictions resembling beads. Arterial cast filling terminated in vessels with lumen diameters from 70 to 20 µm with focal signs of constriction at or near the point of cast termination. The results indicate that the presence of both blood and plasma in the subarachnoid space produces acute small-artery constriction. This phenomenon is due to a noncellular blood component and does not correlate with increases in intracranial pressure.     

Nitric oxide and vascular reactivity in developing zebrafish, Danio rerio

We used a newly developed digital motion analysis video technique to study the effects of nitric oxide (NO) and epinephrine on the early larval arterial and venous vasculature of zebrafish. Application of the NO donor sodium nitroprusside resulted in a significant increase in both the venous and arterial vessel diameters, whereas N(G)-nitro-L-arginine methyl ester caused a significant decrease in the same diameters. Thus our results show that both the venous and arterial vasculature of the 5- and 6-day-old zebrafish larvae are influenced by endogenously produced NO. By use of immunohistochemistry, NO synthase immunoreactivity was demonstrated in endothelial cells of the dorsal vein. Local application of epinephrine onto the dorsal artery had no effect on vessel diameter. However, if the embryos were preincubated with N(omega)-nitro-L-arginine methyl ester, addition of epinephrine resulted in a significant reduction in both arterial and venous vessel diameters. Thus this study provides increasing evidence that before a functional autonomic innervation of the peripheral vascular system, vascular tone in larval tissue is regulated by a complex interaction of vasoactive substances that are produced locally by vascular endothelial cells.     

3-Dimensional Resin Casting and Imaging of Mouse Portal Vein or Intrahepatic Bile Duct System

In organs, the correct architecture of vascular and ductal structures is indispensable for proper physiological function, and the formation and maintenance of these structures is a highly regulated process. The analysis of these complex, 3-dimensional structures has greatly depended on either 2-dimensional examination in section or on dye injection studies. These techniques, however, are not able to provide a complete and quantifiable representation of the ductal or vascular structures they are intended to elucidate. Alternatively, the nature of 3-dimensional plastic resin casts generates a permanent snapshot of the system and is a novel and widely useful technique for visualizing and quantifying 3-dimensional structures and networks.A crucial advantage of the resin casting system is the ability to determine the intact and connected, or communicating, structure of a blood vessel or duct. The structure of vascular and ductal networks are crucial for organ function, and this technique has the potential to aid study of vascular and ductal networks in several ways. Resin casting may be used to analyze normal morphology and functional architecture of a luminal structure, identify developmental morphogenetic changes, and uncover morphological differences in tissue architecture between normal and disease states. Previous work has utilized resin casting to study, for example, architectural and functional defects within the mouse intrahepatic bile duct system that were not reflected in 2-dimensional analysis of the structure^1,2, alterations in brain vasculature of a Alzheimer''s disease mouse model³, portal vein abnormalities in portal hypertensive and cirrhotic mice⁴, developmental steps in rat lymphatic maturation between immature and adult lungs⁵, immediate microvascular changes in the rat liver, pancreas, and kidney in response in to chemical injury⁶.Here we present a method of generating a 3-dimensional resin cast of a mouse vascular or ductal network, focusing specifically on the portal vein and intrahepatic bile duct. These casts can be visualized by clearing or macerating the tissue and can then be analyzed. This technique can be applied to virtually any vascular or ductal system and would be directly applicable to any study inquiring into the development, function, maintenance, or injury of a 3-dimensional ductal or vascular structure.     

Bifurcation asymmetry of the porcine coronary vasculature and its implications on coronary flow heterogeneity

The branching pattern of the coronary arteries and veins is asymmetric, i.e., many small vessels branch off of a large trunk such that the two daughter vessels at a bifurcation are of unequal diameters and lengths. One important implication of the geometric vascular asymmetry is the dispersion of blood flow at a bifurcation, which leads to large spatial heterogeneity of myocardial blood flow. To document the asymmetric branching pattern of the coronary vessels, we computed an asymmetry ratio for the diameters and lengths of all vessels, defined as the ratio of the daughter diameters and lengths, respectively. Previous data from silicone elastomer cast of the entire coronary vasculature including arteries, arterioles, venules, and veins were analyzed. Data on smaller vessels were obtained from histological specimens by optical sectioning, whereas data on larger vessels were obtained from vascular casts. Asymmetry ratios for vascular areas, volumes, resistances, and flows of the various daughter vessels were computed from the asymmetry ratios of diameters and lengths for every order of mother vessel. The results show that the largest orders of arterial and venous vessels are most asymmetric and the degree of asymmetry decreases toward the smaller vessels. Furthermore, the diameter asymmetry at a bifurcation is significantly larger for the coronary veins (1.7-6.8 for sinus veins) than the corresponding arteries (1.5-5.8 for left anterior descending coronary artery) for orders 2-10, respectively. The reported diameter asymmetry at a bifurcation leads to significant heterogeneity of blood flow at a bifurcation. Hence, the present data quantify the dispersion of blood flow at a bifurcation and are essential for understanding flow heterogeneity in the coronary circulation.     

Venous valve anatomy and morphometry: studies on the duckling using vascular corrosion casting

The anatomy and morphometry of venous values associated with the vasculature of the head of the duckling were studied using vascular corrosion casting and scanning electron microscopy. All valves encountered were bicuspid, and casts typically exhibited slight expansions at valve sinuses and deep slits at the sites of valve leaflets. The locations, numbers, and orientations of endothelial nuclei on all surfaces of the valves were clearly revealed by imprints in the casting resin. Endothelial cell densities were significantly higher on the surfaces of valve leaflets (about 10 cells/1,000 micron2) than on other venous surfaces (about 7 cells/1,000 micron2). Endothelial nuclei on the medial surface of the valve leaflet were oriented parallel to the long axis of the vessel, whereas those on the lateral surface were oriented perpendicular to that axis. The close proximities of valves in some vessels and the presence of anomalies such as the sharing of leaflets by adjacent valves were readily demonstrated with the corrosion-casting techniques. These methods provide a useful means for studying the fine, three-dimensional details of venous valve anatomy.     

牦牛和黄牛的睾丸形态及其血管构筑特征

本实验旨在研究牦牛（Bos grunniens）和黄牛（B. taurus）的睾丸形态及其血管构筑和动脉管径特征,为牦牛睾丸在高原环境的生理适应性提供依据。从14头屠宰后的成年牦牛体内采集睾丸28枚,从9头成年黄牛体内采集睾丸18枚,测定其形态指标,利用血管铸型技术制作动静脉构筑标本,研究睾丸血管解剖学及主要动脉管径的特征。结果显示,牦牛和黄牛的睾丸、附睾的形态特征及动脉管径有差异,牦牛大部分睾丸动脉及其分支的管径与睾丸重的相对值极显著地高于黄牛（P < 0.01）,牦牛睾丸的主要动脉构筑特征与黄牛的相同,但其大的集合静脉数量较少,小静脉呈“编织袋”状紧密排布。研究认为,牦牛睾丸的血管解剖特征可为睾丸提供更为充足的血液,其相对发达的动脉血管和睾丸静脉“编织袋”状分布特点有利于睾丸的温度调节及精子成熟,可能是牦牛生殖器官适应高海拔环境的生理特征之一。     

Development and pathology of the hyaloid, choroidal and retinal vasculature

During embryogenesis, the development and differentiation of the eye requires the concomitant formation of the neural/glial elements along with a dense vascular network. The adult neural retina is supported by two distinct vascular systems, the proper retinal vessels and the choroidal vessels. The two beds differ not only in their pattern of embryonic differentiation, but also in their function in the adult organism. The retinal vasculature has barrier properties similar to those observed in the brain, whereas the choroidal vessels display a highly fenestrated phenotype. The hyaloid vasculature is a transient embryonic vascular bed which is complete at birth in mammals and regresses contemporaneously with the formation of the retinal vasculature. The dependence of the retina on its blood supply makes it highly vulnerable to any vascular changes and indeed ocular diseases, such as proliferative retinopathy, age-related macular degeneration and the hyperplastic primary vitreous, which are associated with abnormalities of the different vascular beds of the eye. A number of factors have been implicated in developmental and pathological changes in vessel formation and regression, including fibroblast growth factors, platelet-derived endothelial growth factor and vascular endothelial growth factor, among others. The purpose of this review is to describe and discuss new insights into the mechanisms and molecular cues involved in the development of the normal and pathological vascular systems of the eye. The characterization of the molecules and cell-cell interactions involved in the formation, stabilization and regression of new vessels has led to the identification of potential control points for therapeutic intervention.     

Glomerular bypass shunts in the kidney of the Atlantic hagfish,Myxine glutinosa

Summary The vascular pathways associated with the glomerulus of the Atlantic hagfish, Myxine glutinosa have been studied by scanning electron microscopy of corroded resin casts of the vasculature. Although the overall pattern of the renal vasculature did not differ from earlier reports, a previously unreported vascular pathway which arose from the renal artery and bypassed the glomerular capillaries in 28% of glomeruli was clearly demonstrated. Glomerular bypass shunts either ran to join the loose capillary network around Bowman's capsule and thereby drain into the network of vessels associated with the mesonephric duct (ureter), or ran directly into the ureteral system of vessels and subsequently into the posterior cardinal veins. Glomerular bypass shunts which theoretically permit renal arterial blood to bypass the process of filtration may play a role in the regulation of body fluid volume.     

Blood vessel occlusion by Cryptococcus neoformans is a mechanism for haemorrhagic dissemination of infection

Meningitis caused by infectious pathogens is associated with vessel damage and infarct formation, however the physiological cause is often unknown. Cryptococcus neoformans is a human fungal pathogen and causative agent of cryptococcal meningitis, where vascular events are observed in up to 30% of patients, predominantly in severe infection. Therefore, we aimed to investigate how infection may lead to vessel damage and associated pathogen dissemination using a zebrafish model that permitted noninvasive in vivo imaging. We find that cryptococcal cells become trapped within the vasculature (dependent on their size) and proliferate there resulting in vasodilation. Localised cryptococcal growth, originating from a small number of cryptococcal cells in the vasculature was associated with sites of dissemination and simultaneously with loss of blood vessel integrity. Using a cell-cell junction tension reporter we identified dissemination from intact blood vessels and where vessel rupture occurred. Finally, we manipulated blood vessel tension via cell junctions and found increased tension resulted in increased dissemination. Our data suggest that global vascular vasodilation occurs following infection, resulting in increased vessel tension which subsequently increases dissemination events, representing a positive feedback loop. Thus, we identify a mechanism for blood vessel damage during cryptococcal infection that may represent a cause of vascular damage and cortical infarction during cryptococcal meningitis.     

In Utero Intra-cardiac Tomato-lectin Injections on Mouse Embryos to Gauge Renal Blood Flow

The formation and perfusion of developing renal blood vessels (apart from glomeruli) are greatly understudied. As vasculature develops via angiogenesis (which is the branching off of major vessels) and vasculogenesis (de novo vessel formation), perfusion mapping techniques such as resin casts, in vivo ultrasound imaging, and micro-dissection have been limited in demonstrating the intimate relationships between these two processes and developing renal structures within the embryo. Here, we describe the procedure of in utero intra-cardiac ultrasound-guided FITC-labeled tomato lectin microinjections on mouse embryos to gauge the ontogeny of renal perfusion. Tomato lectin (TL) was perfused throughout the embryo and kidneys harvested. Tissues were co-stained for various kidney structures including: nephron progenitors, nephron structures, ureteric epithelium, and vasculature. Starting at E13.5 large caliber vessels were perfused, however peripheral vessels remained unperfused. By E15.5 and E17.5, small peripheral vessels as well as glomeruli started to become perfused. This experimental technique is critical for studying the role of vasculature and blood flow during embryonic development.     

Effects of cilazapril on the retinal vessels in spontaneously hypertensive rats: corrosion cast and scanning electron microscopic study

The effects of the long-term oral angiotensin-converting enzyme inhibitor, cilazapril, on retinal circulation in stroke-prone spontaneously hypertensive (SHR-SP) rats were assessed by scanning electron microscopy (SEM), corrosion casts and transmission electron microscopy (TEM). Two groups of 20 male SHR-SP rats were compared. One group was treated with 10 mg/kg/day of cilazapril from 4 to 40 weeks of age, and the other group received no treatment. A third group of male Wistar-Kyoto (WKY) rats served as age-matched controls. At regular intervals the rats were weighed, and their systolic blood pressure was measured. Cilazapril normalized systolic arterial pressure to 121+/-2.7 mm Hg (SD) in the treated SHR-SP rats. There was no significant difference in body weight between the two groups of SHR-SP. In the 40-week-old SHR-SP rats without treatment corrosion cast and SEM revealed hypertensive retinal vascular changes. In the 40-week-old SHR-SP rats treated with cilazapril, these changes were markedly decreased to the level seen in WKY rats. The differences in caliber of retinal capillaries between the treated SHR-SP and untreated SHR-SP rats were statistically significant (p<.0001). TEM in the cilazapril-treated SHR-SP rats revealed intact basement membranes (0.29+/-0.057 microm) of the endothelial cells and pericytes, but in the untreated SHR-SP rats the basement membrane was thickened (0.51+/-0.123 microm) (p<.0001) and the pericytes damaged. Our results show that the long-term administration of cilazapril decreased systolic arterial pressure to a nearly normal level and prevented hypertensive retinal vascular changes, probably by improving endothelial function. The effects of cilazapril on the retinal vasculature are described for the first time. SEM of corrosion casts is a valuable technique for showing the effects of some drugs on the vasculature easily, precisely and three-dimensionally.     

A Continuum Mathematical Model of the Developing Murine Retinal Vasculature

Angiogenesis, the process of new vessel growth from pre-existing vasculature, is crucial in many biological situations such as wound healing and embryogenesis. Angiogenesis is also a key regulator of pathogenesis in many clinically important disease processes, for instance, solid tumour progression and ocular diseases. Over the past 10–20 years, tumour-induced angiogenesis has received a lot of attention in the mathematical modelling community and there have also been some attempts to model angiogenesis during wound healing. However, there has been little modelling work of vascular growth during normal development. In this paper, we describe an in silico representation of the developing retinal vasculature in the mouse, using continuum mathematical models consisting of systems of partial differential equations. The equations describe the migratory response of cells to growth factor gradients, the evolution of the capillary blood vessel density, and of the growth factor concentration. Our approach is closely coupled to an associated experimental programme to parameterise our model effectively and the simulations provide an excellent correlation with in vivo experimental data. Future work and development of this model will enable us to elucidate the impact of molecular cues upon vasculature development and the implications for eye diseases such as diabetic retinopathy and neonatal retinopathy of prematurity.     

Retinal Arterioles Narrow with Increasing Duration of Anti-Retroviral Therapy in HIV Infection: A Novel Estimator of Vascular Risk in HIV?

Objectives

HIV infection is associated with an increased risk of age-related morbidity mediated by immune dysfunction, atherosclerosis and inflammation. Changes in retinal vessel calibre may reflect cumulative structural damage arising from these mechanisms. The relationship of retinal vessel calibre with clinical and demographic characteristics was investigated in a population of HIV-infected individuals in South Africa.

Methods

Case-control study of 491 adults ≥30 years, composed of 242 HIV-infected adults and 249 age- and gender-matched HIV-negative controls. Retinal vessel calibre was measured using computer-assisted techniques to determine mean arteriolar and venular diameters of each eye.

Results

The median age was 40 years (IQR: 35–48 years). Among HIV-infected adults, 87.1% were receiving highly active antiretroviral therapy (HAART) (median duration, 58 months), their median CD4 count was 468 cells/µL, and 84.3% had undetectable plasma viral load. Unadjusted mean retinal arteriolar diameters were 163.67±17.69 µm in cases and 161.34±17.38 µm in controls (p = 0.15). Unadjusted mean venular diameters were 267.77±18.21 µm in cases and 270.81±18.98 µm in controls (p = 0.07). Age modified the effect of retinal arteriolar and venular diameters in relation to HIV status, with a tendency towards narrower retinal diameters in HIV cases but not in controls. Among cases, retinal arteriolar diameters narrowed with increasing duration of HAART, independently of age (167.83 µm <3 years of HAART vs. 158.89 µm >6 years, p-trend = 0.02), and with a HIV viral load >10,000 copies/mL while on HAART (p = 0.05). HIV-related venular changes were not detected.

Conclusions

Narrowing of retinal arteriolar diameters is associated with HAART duration and viral load, and may reflect heightened inflammatory and pro-atherogenic states of the systemic vasculature. Measurement of retinal vascular calibre could be an innovative non-invasive method of estimating vascular risk in HIV-infected individuals.     

西藏小型猪部分脏器血管铸型标本制作

目的研究西藏小型猪内脏铸型,对西藏小型猪在相关生物医学研究上的应用提供参考。方法应用过氯乙烯和牙托粉单独或混合填充剂灌注方法制作西藏小型猪心脏、肝脏、脾脏、肺脏、肾脏和脑铸型研究其血管分布规律。结果西藏小型猪内脏铸型外观完整美观,保持正常解剖状态、位置,充分显示和暴露主干的分支分布,各管道饱满,管道粗细适当,标本色泽鲜艳。结论通过单独或混合灌注过氯乙烯和牙托粉填充剂可以成功制作西藏小型猪铸型标本,所做标本显示的脏器血管分布情况对相关生物医学研究具有重要应用价值。     

Glomerular bypass shunts and distribution of glomeruli in the kidney of the lesser spotted dogfish,Scyliorhinus caniculus

Summary Scanning electron microscopy of corroded resin casts of the renal vasculature of Scyliorhinus caniculus has revealed a novel vascular pathway arising from the afferent arteriole and bypassing the glomerulus. This glomerulus bypass shunt occurred in 36% of the glomerular casts examined. The shunt ran to join a peritubular network of capillaries and thereby offers the potential to vary the degree of glomerular perfusion and control the proportion of active glomeruli. In 29% of glomeruli two efferent arterioles drained the capillary knot. Glomeruli were located close to the dorsal margin of the posterior mass of the kidney, and towards the lateral edge of the anterior lobes of the kidney of female dogfish. In male dogfish, glomeruli were evenly distributed through the posterior mass of kidney, while in female dogfish 89% of glomeruli occurred in the posterior mass and 11% of glomeruli were located within the small anterior lobes.     

Computer-assisted measurement of vessel shape from 3T magnetic resonance angiography of mouse brain

Blood vessel morphology (vessel radius, branching pattern, and tortuosity) is altered by a multitude of diseases. Although murine models of human pathology are important to the investigation of many diseases, there are few publications that address quantitative measurements of murine vascular morphology. This report outlines methods of imaging mice in vivo using magnetic resonance angiograms obtained on a clinical 3T unit, of defining mouse vasculature from these images, and of quantifying measures of vessel shape. We provide examples of both healthy and diseased vasculature and illustrate how the approach can be used to assess pathology both visually and quantitatively. The method is amenable to the assessment of many diseases in both human beings and mice.     

Resin-casting: a method for investigating apoplastic spaces

A method has been developed in which liquid resin can be injected or infiltrated into spaces within a plant body, for example the lumens of vessels, fibers, and intercellular spaces. After polymerization of the resin, plant tissues are digested away with two solutions used in sequence: first, equal parts concentrated hydrogen peroxide and glacial acetic acid; second, concentrated sulfuric acid. Complete digestion renders three-dimensional casts of the spaces in the original tissue. These can be examined with scanning electron microscopy, light microscopy, or a dissecting microscope. Casts have such high fidelity and high resolution that details of pit canals, pit chambers, and perforation plates can be studied. Vessel casts over 15 cm long and revealing the details of several thousand constituent vessel elements have been obtained easily. Casts of the lumens of all cell types and of narrow intercellular spaces are obtained by prolonged infiltration with a low-viscosity resin solution. Alternatively, rapid, brief injection of the resin by vacuum produces casts predominantly of just those spaces that were open to the resin at the cut ends of the sample, for example the lumens of vessels and secretory ducts or the intercellular space network of an aerenchymatous parenchyma.     

Small molecule screen for compounds that affect vascular development in the zebrafish retina

Blood vessel formation in the vertebrate eye is a precisely regulated process. In the human retina, both an excess and a deficiency of blood vessels may lead to a loss of vision. To gain insight into the molecular basis of vessel formation in the vertebrate retina and to develop pharmacological means of manipulating this process in a living organism, we further characterized the embryonic zebrafish eye vasculature, and performed a small molecule screen for compounds that affect blood vessel morphogenesis. The screening of approximately 2000 compounds revealed four small molecules that at specific concentrations affect retinal vessel morphology but do not produce obvious changes in trunk vessels, or in the neuronal architecture of the retina. Of these, two induce a pronounced widening of vessel diameter without a substantial loss of vessel number, one compound produces a loss of retinal blood vessels accompanied by a mild increase of their diameter, and finally one other generates a severe loss of retinal vessels. This work demonstrates the utility of zebrafish as a screening tool for small molecules that affect eye vasculature and presents several compounds of potential therapeutic importance.     

Scaling of myocardial mass to flow and morphometry of coronary arteries.

There is no doubt that scaling relations exist between myocardial mass and morphometry of coronary vasculature. The purpose of this study is to quantify several morphological (diameter, length, and volume) and functional (flow) parameters of the coronary arterial tree in relation to myocardial mass. Eight normal porcine hearts of 117-244 g (mean of 177.5 +/- 32.7) were used in this study. Various coronary subtrees of the left anterior descending, right coronary, and left circumflex arteries were perfused at pressure of 100 mmHg with different colors of a polymer (Microfil) to obtain rubber casts of arterial trees corresponding to different regions of myocardial mass. Volume, diameter, and cumulative length of coronary arteries were reconstructed from casts to analyze their relationship to the perfused myocardial mass. Volumetric flow was measured in relationship with perfused myocardial mass. Our results show that arterial volume is linearly related to regional myocardial mass, whereas the sum of coronary arterial branch lengths, vessel diameters, and volumetric flow show an approximately 3/4, 3/8, and 3/4 power-law relationship, respectively, in relation to myocardial mass. These scaling laws suggest fundamental design principles underlying the structure-function relationship of the coronary arterial tree that may facilitate diagnosis and management of diffuse coronary artery disease.     

Blood velocity measurement in human conjunctival vessels

The bulbar conjunctiva is one of the few areas in which blood flow in the peripheral vasculature can be directly and noninvasively observed in the human. Although extensive literature exists describing morphological changes which correlate with a variety of systemic diseases in this vasculature, little quantitative data is available on hemodynamics in either normal or abnormal states. The hemodynamic data available are primarily subjective assessments of "low flow." Approaches to place the subjective assessment on more quantitative grounds have usually been based on photographic techniques that have intrinsic inadequacies. The objective of the work reported here was to develop a system capable of providing sequential blood velocity data potentially useful for providing quantitative information on blood flow and its change in the microvessels of the human conjunctiva. The method that has evolved uses a standard Zeiss slit-lamp to image a subject's conjunctival vessels by using a 1-inch Newvicon TV camera with an electronic magnification of 2x. The video image is simultaneously recorded on a video tape recorder (VTR) to an overall system magnification of approximately 4 microm/raster line. The data acquisition phase requires approximately 5 minutes of patient time, whereas the actual determination of blood velocity in individual vessels is done offline through a modification of the dual-slit videodensimetric method. Two independently controllable video cursors are placed axially over the vessel image with the VTR in the still-frame mode. For each consecutive video field, the position of two reference points on the vessel and the position of each cursor relative to these and to each other are encoded into a computer to track the moving image caused by normal eye movement. The computer then determines new cursor coordinates to ensure a constant position within the vessel. The electrical signals obtained for each cursor site and for each video field are cross-correlated to yield the average blood velocity over the sampled time interval. The system has been calibrated in vitro from 0.2 to 2.5 mm/sec, evaluated in experimental animals, and used to measure blood velocity (0.3 to 1.5 mm/sec) in human conjunctival venules with diameters ranging from 20 to 50 microm. At this writing, blood velocity has been recorded during a period of about 3 months in the same vessel of several postmyocardial infarction patients. Thus, the method appears suitable for determining sequential changes in small vessel blood flow in patients over extended periods of time.