A new method for the joint visualization of vascular structures and connective tissues: Corrosion casting and 1 N NaOH maceration

Corrosion casting combined with scanning electron microscopy (SEM) has been widely used to study the morphofunctional aspects of microcirculation in many organs. In this study, we present an optimization of the corrosion casting (CC) technique associating it with NaOH 1 N maceration method to obtain a clear visualization of the relationships existing between the microvascular architecture of an organ and its extracellular matrix. Briefly, experiments were performed macerating the tissue previously injected with a low viscosity acrylic resin in 1 N NaOH and then observing it at SEM. In this study, we present an application of this technique to better evaluate the extracellular components of the vascular wall in medium-sized and capillary vessels both in skin and in kidney. The results obtained yielded clear images of the three-dimensional layout of medium-sized and capillary vessels in comparison with the extracellular environment. Furthermore, detailed information was obtained on the three-dimensional layout of fibers constituting the walls of venules, arterioles, and capillaries. In addition, the tubular collagenic structures surrounding the excretory tubules of the kidney and the dermal glands of the skin were depicted and their relationships with their vascular supply described in detail. © 2006 Wiley-Liss, Inc

The fibrillar crimps of the sclera

Crimps are a typical feature of tendons and of some ligaments where they are responsible for the non-linear behaviour of the tissue at low strain values. Previous studies (Raspanti et al., 2005) shown that in tendons these visible crimps always correspond to a distinctive buckling and/or left-handed torsion of the collagen fibrils, subsequently named “fibrillar crimp” (Franchi et al, 2007). These planar crimps are exclusive of the collagen fibrils of tendons and ligaments, whose subfibrils run straight and parallel and tend to behave like stiff rods; by contrast, the collagen fibrils of most tissues, which are sometimes defined as “reticular fibrils” of “type III fibrils” and whose subfibrils follow an helical course, are almost infinitely flexible and can withstand even sharp U-turns without buckling. In the present study we investigated the rat sclera, whose ultrastructure is closely related to that of tendons and ligaments (Raspanti et al., 1992). The tissue was observed by light microscopy, high-resolution scanning electron microscopy and atomic force microscopy. The sclera appears made of flattened fascicles of large and heterogeneous collagen fibrils, running in all directions and often following a wavy course. The individual fascicles are identical in structure and appearance to those of tendon, as are the numerous crimps which can be easily observed along their fibrils. The presence of crimps in the sclera cannot be taken for granted because at variance with tendon, which is subject to intermittent load/recoil cycles, this tissue is held under continuous tension by the intraocular pressure (IOP). A typical intra-ocular pressure of 20-30 mmHg would induce in the rat sclera a tension varying from 50 to 80 kN/m2, and the elastic behaviour of the crimps may have a role in maintaining the shape of the eyeball. The crimps may therefore be fully functional, confirming this tissue as a sort of hemispherical tendon

Ultrastructural aspects of mineralization-induced modifications in turkey tendon

In all tendons the collagen fascicles follow a wavy course (actually a flattened lefthanded helix) forming visible crimps. Each crimp corresponds to a sharp bend and/ or an axial twisting of individual collagen fibrils (Raspanti et al., 2005; Franchi et al., 2010), and even once the fibril are straightened out a permanent local deformation remains visible, still revealing the original crimp location (Raspanti et al., 2005). The tendons of some birds represent a special case as they undergo a physiological process of gradual mineralization involving heavy modifications of the tissue architecture. In the present research, turkey tendons appeared to be more finely subdivided into thinner fascicles than most tendons; they contained a greater amount of cell-rich endotenon tissue as well as occasional nodules of cartilage-like matrix. The most striking finding, however, was the complete disappearance of the crimps in the calcified portions of the tendon, while they were present with the usual morphology in the non-mineralized portion. The mineralized fibrils ran perfectly straight, but the electron microscopy revealed traces of pre-existing crimps locked in the extended position by the mineralization process. The inorganic phase itself appeared composed of two different types of fine particles, respectively growing inside or around the collagen fibrils and looking as tightly packed fine needles or as larger platelets regularly arranged in relation with the D-period. The perifibrillar mineral could play a critical role in the mechanical coupling of adjoining fascicles and in the transmission of tensile loads along the tendon itself

Heparins and 2019-nCoV infection: a narrative review

Abstract. – OBJECTIVE: Patients with 2019-nCoV infection have a high risk to develop venous thrombotic events. Several guidelines recommend the use of either unfractionated heparin or low molecular weight heparins in preventing thrombotic events in these patients. However, results from clinical studies, so far published, reached controversial conclusions on heparin efficacy in this kind of patients since the incidence of venous thromboembolism remains high despite prophylaxis. This narrative review aims to provide an overview of the antiviral and anti-inflammatory properties of heparins and their efficacy and safety in SARSCoV-2 medical ward-patients. Moreover, anatomical findings and ongoing trials are also reported. Finally, this narrative review tries to explain why heparins fail to prevent venous thrombosis. MATERIALS AND METHODS: We searched for the most relevant published studies on heparins and 2019-nCoV infected patients using the MEDLINE electronic database in the period between January and December 2020. Articles were preliminarily defined as eligible if they: a) were in English language, b) enrolled 250 or more medical ward-patients and 100 or more ICU-patients, c) reported results on patients treated with heparins in a percentage of at least 70% and d) performed an objectively confirmed diagnosis of VTE. RESULTS: Data from medium to large scientific studies show that the incidence of venous thrombotic events in medical ward-patients with SARS-CoV-2 vary between 0% and 8.3%, while this rate is higher, from 6.2% to 49%, in Intensive Care Unit-patients. However, heparins reduce the mortality rate in these patients of about 50%. Histological findings show that thrombosis could affect capillaries, main and small-midsized vessels, and it is associated with diffuse alveolar damage. CONCLUSIONS: Heparins have anti-inflammatory and anti-viral properties, which may be of help in reducing mortality in SARS-CoV-2 patients. Failure of heparins at prophylactic dosages in preventing VTE, especially in ICU-patients, could be due to the severity of the disease. Data on the use of heparins in an early phase of the 2019-nCoV infection are still lacking

Microvascularization of the human digit as studied by corrosion casting

The aim of this study was to describe microcirculation in the human digit, focusing on the vascular patterns of its cutaneous and subcutaneous areas. We injected a functional supranumerary human thumb (Wassel type IV) with a low-viscosity acrylic resin through its digital artery. The tissues around the vessels were then digested in hot alkali and the resulting casts treated for scanning electron microscopy. We concentrated on six different areas: the palmar and dorsal side of the skin, the eponychium, the perionychium, the nail bed and the nail root. On the palmar side, many vascular villi were evident: these capillaries followed the arrangement of the fingerprint lines, whereas on the dorsal side they were scattered irregularly inside the dermal papillae. In the hypodermal layer of the palmar area, vascular supports of sweat glands and many arteriovenous anastomoses were visible, along with glomerular-shaped vessels involved in thermic regulation and tactile function. In the eponychium and perionychium, the vascular villi followed the direction of nail growth. In the face of the eponychium in contact with the nail, a wide-mesh net of capillaries was evident. In the nail bed, the vessels were arranged in many longitudinal trabeculae parallel to the major axis of the digit. In the root of the nail, we found many columnar vessels characterized by multiple angiogenic buttons on their surface

3D imaging of the osteon lacunar-canalicular system: an improved corrosion-casting method

The osteocyte lacunar-canalicular system has been the subject of several studies either by serial sectioning of embedded specimens, or by confocal laser scanning microscopy, or with FIB/SEM imaging. With all these techniques the persistence of the bone matrix hampers the visualization of the finer details of the canalicular network. In the present research the methyl-metacrylate casting technique was applied to human cortical bone in order to evidentiate the 3-D organization of the osteons lacunar-canalicular system. The MMA monomer infiltration into the vascular canals, and hence into the lacunar-canalicular system was driven by capillarity, helped by evaporation and the resulting negative pressure. The resin penetration was complete in the Haversian canal but limited to a depth of 4 – 5 layers of lacunae around the canal itself, suggesting that each secondary osteon is a close system limited by an impenetrable cementing line. The casts duplicated the shape, position and connections of the lacunae, and allowed an unrestricted exploration of lacunar-canalicular network without manipulations such as cutting or sawing. Two systems of canalicula could be distinguished: the equatorial, which connects osteocytes lying on the same concentrical level, and the radial, which interconnects different levels. The equatorial canalicula radiate from the lacunar border on a planar surface around the lacuna, forming straight and basically parallel canals; the radial have a predominant direction perpendicular to the equatorial plane. The mean length of the radial canalicula was 38.4 +/- 7.35 um in human osteons, while their mean diameter was 195.7 +/- 79.58 nm. The equatorial canalicula had an average diameter of 249.7 +/- 73.78 nm, significantly larger (p < 0.001) than the radial. The architecture of lacunar-canalicular network obtained by the MMA casts was compared with theoretical models. Our data suggests that the intracanalicular pericellular fluid is static and that the supply of ions, oxygen and metabolites to the osteocytes can be assured by the diffusion of these molecules into the static volume of the solution without the need to hypothesize a fluid flow within the canalicular system

Different eyes, different views. Scanning Electron Microscopy applied to forensic investigations

The Scanning Electron Microscope (SEM) is an extremely versatile instrument, essential in a wide array of applications in forensic analysis: for example, it is used to analyze gunshot residue, bullet fingerprints, bullet wipe or patterns around the bullet hole, to examine traces of foreign material embedded in or adhered to bullets (which can provides critical information in the trajectory reconstruction of spent bullets); to study environmental dusts, fibers (both natural and artificial) and to identify unknown small particles; to detect non visible blood stains; to analyze diatoms in drowning cases; and for ink and paper analysis. One central feature of SEM is its ability of providing both panoramic and highly magnified views of the same sample, giving an almost 3D view of the specimen. It is the ideal trait d’union between macroscopic information collected during autoptic or investigative activity and microscopic information obtained with the light microscope. Above all, SEM allows performing a progressive and targeted microdissection of the sample. In this presentation, a selected number of investigations are shown in order to illustrate through specific cases general purpose applications. An elderly man was killed with several blows of axe at the head. SEM investigation allowed us to reconstruct the sequence of the blows, to recognize the type of weapon, to determine how this latter was used and how sharp it was. These results allowed the police to reject the initial version of the suspected, which was eventually convicted of willful murder. A young man died with multiple traumatic and fulguration lesions. SEM analysis allowed us to perform a detailed study of the burnt tissue and to reconstruct the path of the electric discharge, concluding that the primary causa mortis was an accidental electrocution, which caused the subsequent trauma. A child died of a sudden, dramatic internal bleeding. The autopsy revealed that some time before she had swallowed a coin battery which had become lodged in the oesophagus. Here the decaying products of the battery caused an electro-chemical dissection of the oesophagus and, finally, of the descending aorta. The SEM analysis revealed the details of the progressive degeneration of the surrounding tissues